Gram-Positive and Gram-Negative Probiotic Extracellular Vesicles: Mechanisms of Immune Modulation and Therapeutic Opportunities in Atopic Dermatitis

Vipul Wayal; Mey-Fann Lee; Jiu-Yao Wang; Yi-Hsing Chen

doi:10.1007/s12016-026-09201-2

. 2026 Sep 11;69(1):76. doi: 10.1007/s12016-026-09201-2

Gram-Positive and Gram-Negative Probiotic Extracellular Vesicles: Mechanisms of Immune Modulation and Therapeutic Opportunities in Atopic Dermatitis

Vipul Wayal ¹, Mey-Fann Lee ², Jiu-Yao Wang ^3,^4,^5,^✉, Yi-Hsing Chen ^1,^5,^6,^✉

PMCID: PMC13569578 PMID: 42726367

Abstract

Atopic dermatitis (AD) is a prevalent chronic inflammatory skin disease characterized by epidermal barrier dysfunction, type 2 immune skewing, and microbiome dysbiosis. Although live probiotics can modulate the gut-skin axis, their clinical use is limited by variable efficacy and safety concerns. Probiotic extracellular vesicles (PEVs) have emerged as cell-free nanotherapeutics that retain microbial bioactivity while circumventing the risks associated with live bacteria. However, the comparative immunological roles of vesicles from Gram-positive and Gram-negative probiotics remain incompletely understood. This review systematically compares Gram-negative outer membrane vesicles (OMVs) and Gram-positive membrane vesicles (MVs) in the context of AD. OMVs, enriched with lipopolysaccharide (LPS) and other bioactive cargo, mainly signal through toll-like receptor 4 (TLR4) to regulate innate immunity, strengthen antimicrobial defense, and modulate inflammation. In contrast, Gram-positive MVs, containing lipoteichoic acid (LTA), peptidoglycan (PG), proteins, and small RNAs, preferentially activate TLR2-linked pathways that support immune tolerance, regulatory T cell (Treg) responses, and barrier repair. Both PEV classes demonstrate significant potential in preclinical models by attenuating inflammation, improving epithelial integrity, and restoring microbiome balance. Despite this potential, clinical translation requires standardized isolation protocols, scalable manufacturing, rigorous safety testing, and regulatory alignment. Advances in microbial engineering and EV design may enable targeted, microbiota-directed therapies for AD.

Keywords: Atopic dermatitis, Extracellular vesicles, Gut-skin axis, Immunomodulation, Outer membrane vesicles, Toll-like receptors

Introduction

Atopic dermatitis (AD) represents the most prevalent chronic inflammatory skin disorder globally, characterized by intense pruritus, erythematous lesions, and profound disruption of the skin barrier function, which significantly affects patients’ quality of life [1, 2]. The estimated global prevalence of AD is approximately 2.6%, with 10–30% of children and around 2–10% of adults affected [2–5]. Clinically, AD is characterized by a chronic relapsing course with age-dependent morphological patterns and varying severity. In children, lesions typically involve the face and extensor surfaces, whereas in adults, flexural areas are commonly affected. Beyond cutaneous manifestations, persistent pruritus, sleep disturbance, and increased susceptibility to local and systemic infections contribute to significant psychosocial and socioeconomic burden [6].

AD involves a complex and multifactorial interplay of genetic predisposition, immune dysregulation, environmental triggers, and cutaneous dysbiosis. AD pathophysiology is very complex and predominantly driven by a Th2-skewed immune response with elevated levels of interleukin-4 (IL-4), IL-13, and increased immunoglobulin E (IgE) production, alongside dysregulation of skin barrier components, proteins including filaggrin (FLG) and loricrin (LOR), and lipids like ceramides [7–11]. Recent studies revealed that the disease extends beyond localized skin inflammation. The gut-skin axis has been considered a critical regulator of AD progression and potential therapeutic target, as supporting evidence demonstrates concurrent dysbiosis of both cutaneous and gut microbiota in affected individuals [11–15]. Several clinical studies have explored the therapeutic potential of probiotics and probiotic-derived metabolites in mitigating AD severity. Probiotics are proposed to exert immunomodulatory effects by restoring microbial balance, enhancing epithelial barrier integrity, and modulating Th1/Th2 immune responses [15–20]. Emerging evidence suggests that probiotic supplementation may help reduce the risk of AD development, particularly when administered during early life, and may also alleviate disease severity in both children and adult populations [21]. However, clinical outcomes remain heterogeneous, likely due to differences in probiotic strains, dosage, treatment duration, and host-specific factors [22, 23].

Current treatments for AD include corticosteroids, calcineurin inhibitors, systemic immunosuppressants, PDE4 (phosphodiesterase 4) inhibitors, Janus Kinase (JAK) inhibitors and biologics targeting IL-4/IL-13 signaling. While these treatments are effective, they often carry potential side effects and fail to achieve sustained remission in many patients, underscoring the need for novel therapeutic approaches with improved safety profiles and efficacy [2, 24–28]. Extracellular vesicles (EVs) are nanoscale, membrane-bound structures released by cells that facilitate intercellular communication through the transfer of bioactive cargo. They play a critical role in host–microbe interactions by modulating immune responses and maintaining tissue homeostasis [29–31]. These functional properties have led to an increase in scientific interest in bacterial EVs, especially in atopic diseases. Despite this growing interest, a critical gap remains in understanding the differential immunological properties of EVs derived from Gram-positive and Gram-negative probiotic bacteria. These bacterial classes exhibit distinct cell wall architectures that directly influence EV biogenesis, composition, and host interactions. Gram-negative bacteria produce outer membrane vesicles (OMVs) enriched with lipopolysaccharides (LPS), phospholipids, and periplasmic components, whereas Gram-positive bacteria release membrane vesicles (MVs) containing lipoteichoic acids (LTA), peptidoglycan (PG) fragments, and cytoplasmic constituents. These distinct envelope-derived components contribute to differential host recognition and downstream immune responses [32, 33].

Emerging evidence suggests that Gram-positive and Gram-negative probiotic EVs (PEVs) differentially regulate key immune pathways in AD, including NF-κB, mitogen-activated protein kinase (MAPK), and inflammasome signaling, thereby influencing Th1, Th2, Th17, and regulatory T cell (Treg) responses [34–36]. Gram-negative EVs often exhibit stronger immunostimulatory effects due to LPS, whereas Gram-positive EVs tend to induce more controlled immunomodulation, potentially offering a safer profile [37, 38]. In addition to immune regulation, PEVs contribute to skin barrier restoration by promoting keratinocyte differentiation, enhancing tight junction integrity, and supporting lipid synthesis such as ceramides in AD [39–42].

From a translational perspective, EV-based therapies offer advantages over live probiotics, including improved safety, stability, and reduced infection risk [37, 43]. However, challenges such as variability in isolation methods, lack of standardization, and limited clinical evidence regarding optimal dosing and long-term safety remain significant barriers to clinical application [44, 45]. In this context, this review aims to provide a comprehensive comparative analysis of Gram-positive and Gram-negative PEVs, specifically focusing on their biogenesis, molecular composition, and immunomodulatory mechanisms in AD. We further discuss current preclinical and clinical evidence, highlight therapeutic potential, and identify key challenges and future directions for the development of EV-based interventions in AD.

Biogenesis and Molecular Composition: A Comparative View

EVs derived from probiotic bacteria represent a conserved yet structurally diverse mechanism of intercellular communication. Their biogenesis and molecular composition are fundamentally dictated by the architectural differences between Gram-positive and Gram-negative bacterial cell envelopes. These structural disparities directly influence vesicle formation pathways, cargo selection, and ultimately their immunological interactions within the host, particularly in inflammatory conditions such as AD [46–48]. Gram-negative bacteria possess a dual-membrane system consisting of an inner cytoplasmic membrane, thin PG layer, and outer membrane (OM) enriched with LPS. In contrast, Gram-positive bacteria lack an OM and are characterized by a thick, multilayered PG cell wall with LTA and teichoic acids [33]. These intrinsic differences impose distinct constraints and mechanisms for vesicle biogenesis. At the molecular level, EV cargo includes proteins, lipids, nucleic acids (DNA, RNA), metabolites, small molecules and cell wall-associated components [49]. However, qualitative and quantitative composition differ markedly between Gram-positive and Gram-negative probiotic bacterial EVs.

Gram-Negative Probiotic OMVs Structure, Biogenesis and Composition

Pathogenic and non-pathogenic Gram-negative bacteria release OMVs through a process fundamentally linked to their distinctive double-membrane architecture. OMVs are spherical bilayered nanostructures (20–300 nm in diameter) derived from OM and contain LPS, PG, outer membrane proteins (OMPs), inner membrane proteins, periplasmic space components, and cytoplasmic constituents [50].

The biogenesis of OMVs involves multiple mechanisms as shown in Fig. 1: (A) Periplasmic stress-driven membrane blebbing, where accumulation of misfolded proteins, PG fragments and mislocalized envelope components generates local envelope stress and promotes outward membrane curvature, resulting in OM blebbing. (B) LPS assembly defects & lipid remodeling, in which disruptions in LPS biosynthesis, transport, or OM lipid homeostasis disturb lipid asymmetry, causing localized membrane expansion and curvature that favors OMV formation. (C) Disruption of OM-PG tethering through weakened interactions involving structural linker proteins, such as Braun’s lipoprotein (Lpp) and OmpA, allowing the untethered membrane microdomains to spontaneously bulge and pinch off into the extracellular space. (D) Explosive Cell Lysis, where the bacterial cell undergoes sudden lysis triggered by severe envelope stress and/or the action of phage-encoded enzymes (e.g., endolysins) that degrade PG layer. This rupture of cell envelope causes membrane fragments to rapidly self-assemble into multiple OMVs. Ultimately, the relative contribution and dominance of these mechanisms are largely species-specific and influence the cargo composition and immunomodulatory potential of resulting OMVs [51–54].

The asymmetric composition of the OM, with LPS on the outer leaflet and phospholipids on the inner leaflet, is critical for OMV formation and host immune recognition [55]. A recent landmark study demonstrated that OMVs isolated from the non-pathogenic probiotic Escherichia coli Nissle 1917 (EcN) possess potent immunomodulatory capabilities. Specifically, they induce M1 macrophage polarization through profound metabolic remodeling, underscoring the potential of EcN OMVs as targeted immunomodulators for therapeutic applications [56].

The molecular cargo of OMVs is highly diverse, encompassing lipids, LPS, small-molecule metabolites, PG fragments, nucleic acids, and various protein classes. Specifically, these nanosized vesicles transport a broad class of OMPs, periplasmic proteins, and effector molecules, including adhesins, invasins, and functional enzymes. Among the most abundantly enriched proteins are structural OMPs, such as OmpA, OmpC, and OmpF, alongside periplasmic proteins like AcrA and alkaline phosphatase [33]. Furthermore, a recent comprehensive proteomic characterization of OMVs derived from the probiotic strain E. coli Nissle 1917 (EcN), utilizing 1D SDS-PAGE coupled with highly sensitive LC-MS/MS, identified 192 vesicular proteins with high confidence. Notably, 18 of these proteins were encoded by EcN strain-specific genes, while 57 constituents were frequently identified in pathogen-derived OMVs [57].

Lipids constitute major structural components of Gram-negative bacterial OMVs. Previous studies have demonstrated that OMVs from enterotoxigenic E. coli are primarily composed of glycerophospholipids, including phosphatidylglycerol, phosphatidylethanolamine (PE), and cardiolipin. These lipid species play critical roles in maintaining membrane architecture and facilitating the characteristic curvature required for OMV biogenesis [52, 58]. Among them, phosphatidylglycerol and PE are consistently reported as the predominant lipid constituents of OMVs. Similar lipidomic analyses of OMVs from other Gram-negative bacterial species have likewise identified PE as a major membrane component, highlighting its conserved structural importance in OMV formation and stability [59–61]. A recent study suggests that variations in phospholipid composition and their relative proportions can significantly affect membrane biophysical properties, including fluidity, thickness, and lipid stacking. These alterations are thought to contribute to the stabilization of OMV structure and morphology. Furthermore, subtle differences in lipid composition between the bacterial OM and OMVs may facilitate membrane curvature, thereby promoting vesicle biogenesis and outward budding during OMV formation [61].

Gram-negative bacterial OMVs possess the ability to encapsulate and transport genetic material, including DNA and various RNA species such as rRNA, tRNA, mRNA, and various classes of small RNAs (sRNAs). Both surface-associated and intraluminal DNA have been identified within OMVs [60, 62]. Studies have shown that following DNase treatment to remove externally bound DNA, the remaining luminal nucleic acid content consists predominantly of protected plasmid DNA, indicating that encapsulation within OMVs confers substantial resistance to enzymatic degradation. This protective capacity enhances the stability and persistence of genetic material in extracellular environments [62, 63]. Furthermore, diverse forms of luminal DNA have been reported in OMVs derived from a wide range of bacterial species, highlighting their potential role in horizontal gene transfer, bacterial communication, and environmental adaptation [64]. In addition to DNA, emerging evidence indicates that OMVs carry various RNA species that may be transferred to host cells through endocytic pathways or interact directly with host cell membranes. These OMV-associated RNAs are thought to contribute to host-pathogen communication, modulate immune responses, and further support the role of OMVs as efficient nucleic acid delivery systems with potential applications in vaccine development and RNA-based therapeutics [65].

In addition to lipids, proteins, and nucleic acids, OMVs also contain a variety of other biologically active components, including small cellular metabolites, PG fragments, and enzymatic molecules. These constituents further contribute to the functional diversity of OMVs by modulating bacterial physiology and influencing host–microbe interactions. Collectively, the heterogeneous cargo of OMVs reflects their multifaceted roles in intercellular communication, immune regulation, and environmental adaptation [66, 67]. The primary molecular constituents of prominent Gram-negative probiotic OMVs, alongside their mechanistic significance, are summarized in Table 1.

Table 1.

Major molecular constituents identified in OMVs derived from different Gram-negative probiotic bacteria and their biological functions

Bacterial strain	Major OMV components	Key biological functions	References
Escherichia coli Nissle 1917 (EcN)	OmpA, OmpC, OmpF, AcrA/B, murein hydrolases; LPS, PE, phosphatidylglycerol; metabolites and nucleic acids	Macrophage activation and antimicrobial responses; epithelial barrier reinforcement; regulation of inflammatory and systemic immune pathways	[56, 68–76]
Akkermansia muciniphila	Amuc_1100; branched-chain diacyl PE; immunomodulatory sRNAs	Gut barrier protection; immune homeostasis; attenuation of macrophage-driven inflammation	[77–82]
Bacteroides fragilis	Polysaccharide A; SusC/RagA transporters; glycosidases and proteases	Treg induction, Th17 suppression, and microbiota homeostasis	[83, 84]
Bacteroides thetaiotaomicron	SusC/SusD/SusE/SusF/SusG complexes; phospholipids; nucleic acids	Polysaccharide utilization, nutrient metabolism, and microbial cross-feeding	[84, 85]
Parabacteroides goldsteinii	SusC/RagA proteins; alkaline phosphatase; endoglycosidase; attenuated LPS; sphingolipids; metabolites; sRNAs	Gut–skin and gut–lung immunomodulation; suppression of inflammatory signaling; microbiota remodeling	[86–88]
Odoribacter splanchnicus	LOS; penta-acylated lipid A; sulfonolipids; sphingolipids	Intestinal barrier repair and suppression of inflammatory and NLRP3-associated responses	[89, 90]

Open in a new tab

Gram-Positive Probiotic MVs Structure and Biogenesis

Gram-positive probiotic strains secrete MVs that exhibit a fundamentally distinct structural and biochemical architecture compared to the OMVs of Gram-negative bacteria. Typically ranging from 20 to 400 nm in diameter, these MVs lack the LPS-containing OM characteristic of Gram-negative OMVs. Instead, their vesicular membrane is highly enriched with LTA as a major surface immunomodulatory molecule, intricately packaged alongside PG fragments and a complex repertoire of cell wall-associated and cytoplasmic effector molecules; therefore, they are commonly referred to as cytoplasmic membrane vesicles (CMVs) [33, 51, 91]. Historically, CMVs were overlooked due to restrictive thickness of Gram-positive PG layer. However, emerging evidence indicates that CMV biogenesis is facilitated by localized remodeling and degradation of this structural barrier, although the precise molecular mechanisms governing continuous CMV production remain incompletely understood [91, 92].

Recent advances in super-resolution microscopy have revealed that MV precursors initially pinch off from the cytoplasmic membrane and temporarily accumulate within the inner wall zone (IWZ) [93]. From the IWZ, these precursors are released via three proposed structural mechanisms: (1) localized membrane blebbing through transient pores generated by autolysin-mediated PG remodeling; (2) progressive cell wall degradation facilitating vesicle escape; and (3) explosive cell lysis driven by elevated turgor pressure [93]. Complementing these observations, bubbling cell death (BCD) has emerged as a prominent physiological model for MV formation [46]. In the BCD pathway, abundant MV production is intrinsically linked to terminal cell lysis. This process is driven by extensive PG hydrolysis, which is mediated by prophage-encoded endolysin and/or triggered by genotoxic stress (e.g., antibiotics, UV exposure, prophage activation). This further generates localized structural weak points that ultimately lead to explosive bacterial envelope rupture and excessive MV release [46, 94].

Given the diversity and complexity of proposed Gram-positive MV biogenesis pathways, Fig. 2 summarizes two representative mechanisms including localized membrane blebbing and BCD-associated MV production that currently have strongest conceptual and experimental support. Together, these mechanisms highlight the potential continuum between regulated MV secretion and stress-associated vesicle generation in Gram-positive bacteria.

The composition and size distribution of MVs heavily depend on the parent bacterial species and specific environmental growth conditions. Recent comprehensive proteomic analyses demonstrate that Lactobacillus species produce MVs with a highly diverse molecular cargo including various proteins, PG, lipids, nucleic acids and metabolites, specifically enriched in cell wall-associated proteins, metabolic enzymes, biogenesis-related factors, and cell wall hydrolases. This specialized proteomic cargo enables these nanoscale vesicles to replicate the potent immunomodulatory and gut-protective effects of their parental strains, notably by attenuating host inflammation and suppressing NF-κB signaling pathways across the intestinal barrier [95]. A recent proteomic analysis of Lactobacillus-derived MVs (L. plantarum, L. fermentum, and L. gasseri) was conducted using liquid chromatography-tandem mass spectrometry (LC-MS/MS). The identified vesicular proteins were predominantly associated with core biological functions, including catalytic activity, glycolysis, gluconeogenesis, and cell wall organization. Notably, motif enrichment analysis revealed a significant concentration of serine-rich proteins uniquely packaged within MVs of L. plantarum and L. fermentum [96].

Studies have identified that MVs from L. plantarum and L. reuteri contain an exclusive pool of proteins that are significantly enriched in membrane-bound and cell surface-associated proteins relative to total whole-cell proteome. Subcellular localization profiling of this vesicular cargo demonstrates that 56.5% of identified proteins originate from the cytoplasm, while the remaining 43.5% comprise membrane and secreted proteins. Functionally, most of these constituents are broadly categorized as metabolic enzymes, proteases, nucleic acid-binding proteins, transmembrane transporters, and ribosomal structural components. Crucially, these specific MVs also encapsulate a distinct subset of key immunoregulatory proteins, most notably glucosyltransferase (gtfG), serine protease (degP), 60 kDa chaperonin (groL), elongation factor Tu (EF-Tu), and inositol polyphosphate phosphatase, which further support their role in mediating host intestinal immune homeostasis [97, 98].

Beyond this extensive vesicular proteome, current evidence demonstrates that immunomodulatory activity of Lactobacillus-derived MVs is also fundamentally driven by their lipid and nucleic acid constituents. Investigations reveal that these MVs are enriched with Gram-positive-specific bacterial LTA, which serves as a critical surface immunogen. Unlike the LPS of Gram-negative bacterial OMVs, vesicular LTA exerts potent immunomodulatory functions without inherent endotoxicity. Mechanistically, LTA acts as a strong agonist for TLR2, driving profound anti-inflammatory responses primarily through the induction of IL-10 production, a phenomenon that distinctly differentiates the tolerogenic profile of MV-associated LTA from classical pro-inflammatory pathways activated by Gram-negative bacterial LPS [99, 100].

Furthermore, vesicular RNAs have been identified as highly active signaling mediators. For instance, sRNAs consisting of 5 S, 16 S, 23 S rRNA, and tRNA packaged within L. plantarum MVs have been shown to dose-dependently inhibit LPS-induced cellular inflammation [101]. Similarly, RNAs derived from L. gasseri MVs actively attenuate delayed-type hypersensitivity by inhibiting CD4⁺ T cell proliferation via MyD88-dependent signaling pathway [102]. Collectively, these findings indicate that the anti-inflammatory properties of probiotic MVs are not dictated by a single molecule, but rather by the synergistic action of specific intraluminal proteins, lipids, and functional RNAs. The core molecular constituents and corresponding mechanistic functions of major Gram-positive probiotic MVs are summarized in Table 2.

Table 2.

Major molecular constituents identified in MVs derived from different Gram-positive probiotic bacteria and their biological functions

Bacterial strain	Major MV components	Key biological functions	References
Lactiplantibacillus plantarum	Serine-rich proteins; ABC transporters; surface-associated proteins; GAPDH; LTA; sRNAs	Anti-inflammatory activity; macrophage/Treg modulation; protection against AD-associated inflammation	[39, 91, 96, 97, 100, 101]
Limosilactobacillus reuteri	gtfG, DegP, GroL, EF-Tu; AMPs and proteases; glycerolipids and sphingolipids; reuterin	Immune homeostasis; antimicrobial and antibiofilm activity; tissue repair	[91, 95, 98, 103]
Bifidobacterium longum	ABC transporters; quorum-sensing and solute-binding proteins; mucin-binding proteins; LTA	Immune modulation, mast-cell regulation, and improvement of AD-like barrier dysfunction	[91, 104–107]
Lacticaseibacillus rhamnosus	LTA; cell-wall and transport proteins; ribosomal proteins; miRNAs	TLR2-associated immune modulation, IL-10 induction, and wound repair	[99, 108, 109]
Lactobacillus gasseri	Bacteriocins; phage-associated proteins; lysozyme; APF-2; LTA; RNA	Host–microbe interaction, antimicrobial activity, and suppression of T-cell-mediated inflammation	[102, 110, 111]
Lactococcus lactis	Metabolic enzymes; phage-associated proteins; nisin Z	Immune regulation and antibacterial activity	[112–114]
Lacticaseibacillus casei	Cell-wall hydrolases; lysozyme; P45, p40, p75; LTA; muramidases	Antibiofilm activity, EGFR-mediated repair, and anti-inflammatory effects	[115–119]
Lactobacillus acidophilus	SplA, GAPDH, mucus-binding protein, bacteriocin, FmtB, ABC transporter ATPase	Antibacterial activity and balanced macrophage immunomodulation	[115, 120]

Open in a new tab

Distinct Immunomodulatory Mechanisms of OMVs and MVs in AD

The differential immunomodulatory properties of PEVs are largely determined by their surface-associated molecules and pattern recognition receptors (PRRs) they engage, thereby influencing the balance between pro-inflammatory and anti-inflammatory immune responses [37]. The therapeutic potential of PEVs in AD is based on their ability to counteract allergic inflammation and restore epithelial barrier function [37]. Throughout the gut-skin axis, antigen-presenting cells (APCs) and keratinocytes are continuously exposed to microbial vesicles. Because Gram-negative and Gram-positive EVs possess distinct cell envelope components, they preferentially engage different sets of host PRRs. This receptor-ligand specificity directs downstream cellular responses, thereby regulating NF-κB activation, inflammasome assembly, and T-cell differentiation [39, 47]. Unraveling these distinct signaling pathways is essential for translating EV biology into effective, cell-free therapeutics capable of resolving AD pathogenesis.

PEVs–immune cell communication occurs via a two-tiered process involving initial surface-receptor engagement and subsequent cellular internalization. Surface-exposed lipid and glycan constituents (e.g., LPS on OMVs; LTA and PG fragments on MVs) serve as primary contact ligands that trigger surface PRRs (TLR4/MD-2 and TLR2/TLR6) on dendritic cells (DCs) and macrophages [37, 67, 121]. Concurrently, intact vesicles are internalized through clathrin-mediated endocytosis, macropinocytosis, or lipid-raft-dependent pathways, enabling vesicular escape and the delivery of luminal cargo directly into host cytosol [71, 99]. Within the host cell, protected luminal proteins such as L. casei p40/p75 or EcN hydrolases and functional small RNAs (sRNAs/miRNAs) access intracellular sensors, including NOD1/2, caspase-11, and endosomal TLRs to post-transcriptionally regulate cytokine expression and suppress NF-κB/MAPK signaling [73, 101, 118].

However, whether lipids, proteins, or RNAs act as the “dominant” therapeutic mediator remains a central, unresolved question in the field. Current evidence indicates that EV bioactivity is rarely driven by a single isolated component, but rather through a multi-component synergy where the outer lipid-protein envelope protects and delivers labile nucleic acids and enzymes to target intracellular compartments [63, 65]. Importantly, current mechanistic evidence remains largely correlative; most published studies evaluate phenotypic outcomes using whole, unseparated EV isolates rather than fractionated cargos (e.g., protease-digested, RNase-treated, or delipidated vesicles) or bacterial gene-knockout mutants. Consequently, distinguishing vesicle-specific effects from co-purified molecular complexes remains a critical experimental gap that requires rigorous loss-of-function validation in future studies [37].

Gram-Negative OMVs: LPS-Driven Immune Polarization

LPS is a primary immunomodulatory component of Gram-negative probiotic OMVs. It is a potent pathogen-associated molecular pattern (PAMP) molecule composed of lipid A, core hydrophilic oligosaccharide, and O-antigen, recognized primarily by TLR4 in complex with myeloid differentiation factor 2 (MD-2). LPS engages the TLR4/MD-2 complex, triggering both MyD88-dependent (NF-κB and AP-1) and MyD88-independent (IRF3-mediated type I interferon production) signaling pathways [121]. This signaling primes canonical NLRP3 inflammasome activation, which can subsequently be triggered by secondary danger signals. However, probiotic-derived OMVs are distinct from their pathogenic counterparts; specific structural modifications to their lipid A and O-antigens tend to promote an anti-inflammatory cytokine profile over pro-inflammatory mediators. While they effectively attenuate Th2-associated cytokine responses, their overall mechanism suggests immune regulation alongside acute antimicrobial defense. Aligned with this, a previous study demonstrated that EcN-derived OMVs significantly promoted cellular proliferation, phagocytic functions, and immune-related enzymatic activity [68]. Stimulation with appropriate concentrations of EcN-OMVs induced robust anti-inflammatory responses (upregulation of IL-10) rather than pro-inflammatory cytokines (IL-6 and TNF-α). Furthermore, these OMVs also actively modulated production of Th1-polarizing (IL-12) and Th2-polarizing (IL-4) cytokines [68]. This inherent capacity to modulate the Th1/Th2 balance is highly relevant to AD, as it suggests that probiotic OMVs may help correct allergic immune dysregulation while simultaneously enhancing macrophage-mediated antibacterial defense.

The immunomodulatory potency of Gram-negative OMVs largely depends on their high LPS content; experimental evidence indicates that OMVs can present LPS in a much more biologically active configuration than purified, cell-free LPS. For instance, OMVs derived from the probiotic E. coli O83 (EcO83) exhibit an LPS-to-protein ratio approximately five times higher than standard cellular fractions, making them highly potent TLR4 agonists [122]. The role of LPS in allergic diseases, including atopic eczema and asthma, presents a well-documented paradox. While acute, high-dose exposure to pathogenic endotoxins can exacerbate inflammatory flares, continuous environmental exposure to bacterial LPS (such as farm dust) is strongly associated with protection against allergic sensitization [122, 123], a cornerstone of the hygiene hypothesis. This protective, anti-allergic effect is primarily driven by calibrated LPS-mediated TLR4 activation.

Prior in vivo studies have demonstrated that prophylactic administration of EcO83 effectively reduces allergic airway inflammation in a strictly TLR4-dependent manner [123]. Further confirming this mechanism, neutralizing the LPS on EcO83-OMVs using polymyxin B markedly attenuates OMVs’ ability to trigger immune-priming cytokines, such as IL-6 and TNF-α, in host cells [122]. These findings may also be relevant to AD, which shares a similar Th2-dominant immunopathology with allergic asthma. This LPS paradox highlights the critical advantage of using probiotic OMVs. Rather than functioning solely as a pro-inflammatory endotoxin, OMV-associated LPS is presented within a natural vesicular context that enables controlled engagement of TLR4. Consequently, OMV-associated LPS may promote immune polarization away from pathogenic Th2 responses, thereby recapitulating the protective effects proposed by hygiene hypothesis and potentially mitigating AD-associated inflammation.

The immunological impact of OMV-associated LPS remains a subject of debate, as it has been reported to exert both beneficial immunomodulatory and pro-inflammatory effects depending on its bacterial source and biological context. However, current evidence suggests that LPS associated with probiotic-derived OMVs may contribute to attenuation of atopic diseases [122]. Therefore, although probiotic-derived OMVs represent a promising therapeutic strategy for AD, further in vivo studies are needed to elucidate the complex role of OMV-associated LPS and optimize these vesicles for safe and effective anti-allergic therapies. To achieve this, future studies should distinguish OMV-specific effects from those mediated by free LPS using TLR4-deficient models or LPS-deficient bacterial mutants [124]. In addition, successful clinical translation will require bioengineering approaches, such as genetic modification of the highly inflammatory lipid A moiety, to reduce endotoxicity while preserving the beneficial immunomodulatory properties of OMVs [124].

Gram-Positive MVs: LTA-Mediated Immune Tolerance

The immunomodulatory effects of Gram-positive MVs are largely attributed to LTA, PG, lipoproteins, and other cell wall-associated proteins. Among these, LTA is a major PAMP and is predominantly recognized by TLR2, typically in association with TLR6 and its co-receptor CD14 [47, 125]. Activation of these receptors initiates MyD88-dependent signaling, resulting in controlled activation of NF-κB and MAPK pathways. However, unlike MVs from cutaneous pathogens (such as S. aureus) that trigger severe barrier disruption and inflammatory flares, commensal and probiotic Gram-positive MVs deliver tolerogenic signals that support tissue repair and promote keratinocyte differentiation, illustrating how non-pathogenic vesicles can competitively restore cutaneous immune homeostasis [39, 47].

Beyond innate immune regulation, probiotic-derived MVs also profoundly influence adaptive immunity, primarily through interactions with DCs, which serve as key mediators linking innate immune sensing to T-cell activation. Recent evidence suggests that the host immune response to LTA is highly dose-dependent, providing a mechanistic basis for the balance between commensal tolerance and pathogenic inflammation [126]. At low concentrations, which resemble the physiological release of LTA-bearing MVs from probiotic bacteria. Notably, low-dose LTA suppresses DC activation, limits effector T-cell (Teff) responses, and attenuates the production of pro-inflammatory cytokines induced by other potent stimuli, including LPS-mediated TLR4 activation [126]. In contrast, high concentrations of LTA, such as those associated with extensive S. aureus colonization during AD exacerbations, promote the upregulation of C-type lectin receptors on DCs. This shift overrides the tolerogenic effects of TLR2 signaling, resulting in enhanced Teff activation and exacerbation of cutaneous inflammation and epidermal barrier dysfunction [126].

Consequently, LTA-bearing probiotic MVs represent a promising therapeutic strategy for restoring immune homeostasis. However, precise immunological activity of LTA is heavily influenced by its structural heterogeneity across different bacterial species, underscoring the critical importance of careful strain selection for therapeutic applications [127]. To fully harness these immunomodulatory properties, future studies employing purified LTA preparations, TLR2-deficient in vivo models, and genetically engineered probiotic strains will be essential. These approaches will definitively distinguish LTA-specific effects from those mediated by other MV-associated cargo, facilitating the development of optimized, safe, cell-free therapeutics.

In addition to regulating innate and adaptive immune responses, Gram-positive probiotic MVs contribute to the maintenance of epithelial barrier integrity by promoting keratinocyte differentiation and reinforcing tight junctions, thereby limiting allergen penetration and microbial invasion [100, 128]. Furthermore, their nanoscale size facilitates systemic distribution beyond intestinal lumen, enabling these MVs to reach distant organ sites such as the skin and thereby modulate host physiology via the gut-skin axis [129]. This translocation potential of MVs to interact with immune and epithelial cells at remote organs, influencing inflammatory signaling and skin barrier function. The specific local and systemic mechanisms driving this gut-skin axis crosstalk in AD are detailed in the following section.

Gut-Skin Axis: Systemic Reach of PEVs in Context to AD

The therapeutic efficacy of PEVs extends beyond GI tract through the gut-skin axis, a bidirectional communication network linking intestinal microbiota with cutaneous immune homeostasis [12]. Following oral administration or endogenous release within the intestinal lumen, probiotic-derived EVs interact with intestinal epithelial cells and mucosal immune cells, where they regulate local immune responses and reinforce epithelial barrier integrity [70, 72, 95]. Owing to their nanoscale size (approximately 20–400 nm), EVs can traverse the intestinal mucus layer and epithelial barrier through transcellular uptake or paracellular transport before entering the systemic circulation [130]. This unique biodistribution enables probiotic-derived vesicles to deliver bioactive cargos to distant organs such as the skin, thereby extending their immunomodulatory functions beyond the gut.

Emerging evidence well supports the role of the gut-skin axis in AD pathophysiology, where intestinal dysbiosis and “leaky gut” are intrinsically linked to the onset, severity, and persistence of cutaneous inflammation [12, 14]. In this context, the systemic delivery of PEVs represents a powerful mechanism for simultaneously treating both ends of this axis. By traversing the gut barrier into the systemic circulation, orally administered PEVs can exert synchronized dual-compartment actions: reinforcing gut epithelial tight junctions and suppressing mucosal cytokine cascades, while simultaneously delivering lipid mediators, ceramides, and tolerogenic cues to distant cutaneous tissues [41, 131]. Furthermore, emerging evidence indicates that PEVs can modulate host–microbiome neuroendocrine communication such as systemic serotonin metabolism, thereby addressing both cutaneous inflammatory manifestations and neuro-sensory (pruritic) burden of AD via the gut–brain–skin axis [41]. Supporting this systemic inter-organ communication, a recent study demonstrated that Akkermansia muciniphila-derived EVs cross biological barriers to bidirectionally regulate tryptophan metabolism and restore central serotonin levels, effectively resolving systemic inflammatory and neuro-cognitive deficits [132]. This capacity to deliver metabolic regulators to distant organs further underscores the profound systemic reach of PEVs.

In addition to immunomodulation, PEVs contribute directly to the restoration of epidermal barriers. Recent studies suggest that PEVs enhance the expression of structural proteins such as FLG, LOR, claudin-1 (CLDN1), occludin (OCLN), and ZO-1, thereby reducing transepidermal water loss (TEWL) and limiting allergen penetration [37, 42]. Furthermore, EV-mediated stimulation of host AMP production, including cathelicidin (LL-37) and β-defensins, enhances innate cutaneous defense and restricts colonization by S. aureus, a major contributor to AD exacerbation [133]. Collectively, these coordinated effects support a model in which PEVs function as natural nanoscale messengers that transmit beneficial microbial signals from gut to skin, simultaneously restoring epithelial barrier integrity and immune tolerance to attenuate AD (Fig. 3). Mechanistically, Gram-negative OMVs predominantly mediate controlled TLR4 signaling to promote immune tolerance, regulate Th1/Th2 balance, and induce AMP production, whereas Gram-positive MVs primarily activate tolerogenic TLR2 signaling to promote Treg responses, suppress excessive Teff activation, and enhance keratinocyte differentiation. Together, these complementary mechanisms reinforce epithelial barrier function, restore cutaneous immune homeostasis, and mitigate AD progression.

Fig. 3 — Proposed mechanisms underlying probiotic-derived Gram-positive MV- and Gram-negative OMV-mediated modulation of the gut–skin axis in AD. Following intestinal release and systemic translocation, PEVs regulate distinct PRR-mediated signaling pathways that promote immune tolerance, reinforce epidermal barrier integrity, enhance antimicrobial defenses, and collectively restore cutaneous homeostasis, thereby alleviating AD. (Created in https://BioRender.com.)

Comparative Synthesis of Gram-Positive MVs and Gram-Negative OMVs

Taken together, the evidence discussed above indicates that Gram-positive MVs and Gram-negative OMVs share the capacity to modulate host immunity and epithelial barrier function but differ substantially in their envelope-derived molecular cues and vesicular cargo. Gram-negative OMVs are characterized by LPS and OM-associated components, whereas Gram-positive MVs contain LTA, PG-derived components, lipoproteins, proteins, and nucleic acids [33, 37, 47, 91, 99, 121, 125]. These compositional differences contribute to distinct patterns of innate and adaptive immune regulation, with probiotic MVs frequently associated with regulatory or anti-inflammatory responses, whereas OMVs can produce stronger innate immune activation together with strain- and dose-dependent immunoregulatory effects [37, 39, 56, 68, 100, 122].

Both vesicle classes have also been implicated in epithelial barrier protection, although the molecular mechanisms and experimental evidence vary among bacterial strains and disease models [39–42, 100, 107, 128, 133]. A side-by-side synthesis of these structural, immunological, barrier-related, and translationally relevant characteristics is provided in Table 3. Importantly, these differences should be interpreted as context-dependent properties rather than as absolute distinctions between Gram-positive and Gram-negative vesicles.

Table 3.

Critical comparative synthesis of Gram-positive probiotic MVs and Gram-negative probiotic OMVs in AD

Evaluation parameter	Gram-positive probiotic MVs	Gram-negative probiotic OMVs	Critical comparative implication
Envelope architecture & origin	Cytoplasmic membrane-derived vesicles released through the thick PG layer; formation involves membrane remodeling, autolysin-associated PG degradation, and stress-associated pathways	Outer-membrane-derived vesicles associated with membrane blebbing, envelope stress, OM–PG interactions, and other biogenesis mechanisms	Distinct envelope architectures influence vesicle composition, biogenesis, and host interactions [33, 46, 51, 91–94]
Cargo complexity	LTA, PG-derived components, lipoproteins, membrane/cell-wall proteins, metabolites, DNA and RNA	LPS, OMPs, periplasmic proteins, phospholipids, metabolites, DNA and RNA	Both are complex, multifunctional cargo carriers; cargo composition is strongly strain- and condition-dependent [49, 56–67, 95–98]
Dominant immune-recognition pathways	Primarily TLR2/TLR6-associated recognition of LTA and other Gram-positive surface components	Primarily TLR4/MD-2 recognition of LPS, with additional PRR and inflammasome-related sensing depending on cargo and cell type	Distinct PAMP profiles provide different routes for innate immune modulation [47, 99–102, 121, 125–127]
Immunological profile	Frequently associated with regulatory/tolerogenic responses, including IL-10 production, modulation of effector T cells, and macrophage reprogramming	Can produce strong innate immune activation, antimicrobial responses, macrophage modulation, and Th1/Th2 rebalancing; effects depend on strain, dose, and LPS structure	Neither class is uniformly anti-inflammatory or pro-inflammatory; biological activity is context-dependent [37, 39, 56, 68, 100, 122]
Barrier-related effects	Promotion of keratinocyte differentiation and enhancement of barrier-associated proteins such as FLG, LOR, and related junctional components	Enhancement of epithelial/tight-junction integrity and antimicrobial defense, including modulation of ZO-1, occludin, and host antimicrobial responses	Both platforms can support barrier restoration, but the underlying mechanisms and evidence differ among models [39–42, 70–73, 100, 107, 128, 134, 135]
Safety considerations	Lack of LPS-associated endotoxin; however, LTA and other bioactive components remain immunologically active and require strain- and dose-specific evaluation	LPS/lipid-A-associated inflammatory activity represents an additional safety consideration and may require dose optimization or vesicle engineering	Safety should be established for each vesicle preparation rather than inferred solely from Gram classification [37, 43–45, 122–124]
Translational considerations	Production, purification, characterization, reproducibility, and potency standardization remain important development requirements	In addition to general EV manufacturing requirements, control of LPS-associated activity and vesicle composition is particularly important	Both platforms require standardized production and quality-control strategies before clinical application [37, 44, 45]

Open in a new tab

Beyond broad differences between Gram-positive and Gram-negative architectures, EV cargo composition and immunomodulatory potency exhibit profound strain-level heterogeneity. Even within the same taxonomic class or genus, distinct probiotic strains express unique genomic repertoires, enzymatic machineries, and surface antigens that dictate vesicle bioactivity [33, 46, 51, 91]. Among Gram-negative probiotics, while E. coli strains (e.g., EcN, EcO83) harbor canonical hexa-acylated lipid A that potently triggers TLR4 signaling [56, 68, 121–123], other commensals such as P. goldsteinii and O. splanchnicus assemble under-acylated (penta-acylated) lipid A and novel sphingolipids that suppress NLRP3 inflammasomes without triggering hyperinflammation [86–90].

Similarly, B. fragilis OMVs selectively present capsular Polysaccharide A (PSA) to induce tolerogenic DCs and Tregs via TLR2 rather than TLR4 [83, 84]. Among Gram-positive probiotics, L. plantarum MVs utilize specialized serine-rich proteins and regulatory sRNAs to promote M2b macrophage polarization [39, 96, 100, 101], whereas L. casei MVs rely on vesicle-bound p40 and p75 muramidases to transactivate epidermal growth factor receptor (EGFR) signaling for barrier repair [117–119], and B. longum MVs express unique solute- and mucin-binding proteins that induce mast cell apoptosis [104–107]. Consequently, the therapeutic efficacy and safety profile of PEVs are strictly strain-specific and cannot be broadly generalized or extrapolated across species [37, 44, 45].

Preclinical Evidence and Clinical Translation

The available evidence for PEVs in AD remains predominantly preclinical and spans multiple experimental levels. In vitro studies using keratinocytes, macrophages, DCs, and other immune-cell systems provide mechanistic evidence of cellular and molecular effects, whereas animal studies provide evidence of efficacy within complex physiological and disease contexts. Emerging 3D skin, gut-on-a-chip, and related microphysiological models offer an intermediate level of human-relevant evidence but remain experimental [131, 134, 135]. Importantly, clinical studies directly evaluating probiotic-derived EVs in AD are currently lacking; therefore, reported therapeutic benefits should be interpreted as preclinical evidence that supports further investigation rather than established clinical efficacy.

Efficacy in In Vitro and In Vivo Models

To elucidate the cellular mechanisms underlying EV-mediated immune regulation, numerous in vitro studies have focused on the interactions between PEVs and APCs, particularly macrophages and DCs [37]. For instance, Ma et al. demonstrated that EcN-OMVs induce M1 macrophage polarization by metabolically reprogramming. Furthermore, their findings indicate that specific metabolites encapsulated within EcN-OMVs, such as succinate, stearic acid, and BCAAs, directly drive macrophage proliferation, phagocytosis, and migration. Ultimately, this study establishes a strong theoretical foundation for utilizing the metabolic and immunomodulatory cargo of probiotic OMVs in targeted therapeutic applications [56]. Aligned with this in vitro model, another study by Zheng et al. demonstrated that A. muciniphila-derived EVs (Akk EVs), exert profound anti-inflammatory and barrier-modulating effects across both in vitro and in vivo models. For the in vitro model, this study utilized an LPS-induced RAW264.7 macrophage model to comprehensively validate the specific anti-inflammatory effects. To translate these findings in vivo, they studied protective effects of Akk EVs in a murine colitis model; a three-week oral administration to C57BL/6J male mice resulted in significant attenuation of disease severity, notably inhibiting weight loss and colon shortening. Ultimately, this dual-model approach highlighted that Akk EVs protect against intestinal inflammation by reinforcing the epithelial barrier, rebalancing the immune response, and modulating the local microbiota, thereby underscoring their potential as advanced postbiotic therapeutics [82].

Within the specific context of AD, a comprehensive study by Kim et al. demonstrated that L. plantarum-derived EVs effectively protect against AD-like skin inflammation induced by pathogenic S. aureus EVs [39]. To validate the therapeutic and preventive efficacy of these EVs, this study used both in vitro and in vivo models. In vitro, immortalized human epidermal keratinocytes (HaCaT cells) and macrophages were utilized to evaluate protective cellular responses. To translate these findings in vivo, researchers established an AD model in NC/Nga mice using a combination of tape-stripping and topical application of S. aureus EVs. Exposure to these pathogenic EVs rapidly induced severe AD-like skin inflammation, characterized by epidermal thickening, dermal infiltration of mast cells and eosinophils, and elevated cutaneous production of IL-4, IL-5, IFN-γ, and IL-17. In contrast, intervention with L. plantarum EVs successfully mitigated these hallmark pathological features of AD [39].

Building on these immunomodulatory mechanisms, in vitro assays utilizing human THP-1 and primary skin-derived cells demonstrated that EVs from L. plantarum (KCCM11179P) actively drive monocyte differentiation into anti-inflammatory M2 macrophage phenotype [100]. EV exposure enhanced cellular adherence, upregulated key regulatory markers (CD14, ICAM-1, and CD209), and triggered the release of M2b-specific tissue-repair chemokines (CCL1, CCL20, and CXCL2). These in vitro findings suggest that probiotic EVs can promote an anti-inflammatory macrophage phenotype and may contribute to immune regulation in the context of AD [100].

Expanding upon these localized cellular mechanisms, a distinct study by Kim et al. provided crucial insights into how B. longum and its derived EVs (BLEVs) modulate skin barrier function [107]. The in vitro study demonstrated that direct application of BLEVs to differentiated human primary epidermal keratinocytes (HEKs) effectively restored the expression of essential epidermal barrier proteins (FLG and LOR). To explore systemic gut-skin efficacy, researchers utilized a highly translatable hairless mouse model (Crl: SKH1-hrhr). Although the in vivo arm of the study evaluated oral administration of parent B. longum strain, which successfully attenuated DNCB-induced cutaneous inflammation, normalized transepidermal water loss (TEWL), and suppressed systemic IgE and Th2 cytokines, the parallel in vitro findings with BLEVs are equally significant. These findings demonstrate that BLEVs can directly modulate epidermal barrier-associated proteins in vitro, while the parallel in vivo findings with the parent B. longum strain provide complementary evidence for the therapeutic relevance of this probiotic system [107].

In addition to APCs, Tregs are crucial for suppressing allergen-specific IgE production and maintaining Th1/Th2 balance. Using a human in vitro model, researchers evaluated the capacity of B. bifidum (LMG13195) MVs to drive Treg differentiation. By co-culturing monocyte-derived DCs with naïve CD4⁺ T cells, they demonstrated that B. bifidum MVs potently promoted the polarization of naïve CD4⁺ T cells into functional CD25^highFOXP3^highCD127^−/low Tregs. Furthermore, MV-primed DCs induced the highest secretion of anti-inflammatory IL-10 relative to pro-inflammatory cytokines (IFN-γ, TNF-α, and IL-17). These findings suggest that B. bifidum MVs represent a promising cell-free adjuvant for allergen-specific immunotherapy by enhancing immune tolerance and potentially improving therapeutic efficacy [134].

While conventional two-dimensional (2D) cell cultures and murine models have been instrumental in advancing AD research, their translational relevance is limited by interspecies immunological differences and their inability to fully recapitulate the complex architecture and physiology of human skin. Addressing these limitations has driven the development of advanced human-relevant bioengineering platforms, including three-dimensional (3D) skin equivalents and organ-on-a-chip (OOC) technologies [135]. By integrating microfluidic systems with 3D multicellular constructs, OOC platforms dynamically replicate key physiological features of human skin, including tissue stratification, vascular perfusion, and cell–cell communication. Consequently, these platforms provide a more physiologically relevant and predictive model for investigating AD pathogenesis and evaluating next-generation therapeutics [135].

Demonstrating the utility of microfluidic platforms, a recent study by Lee et al. employed an LPS-induced gut-on-a-chip model to evaluate the therapeutic efficacy of L. plantarum HY7714-derived EVs against intestinal inflammation and skin aging [131]. This 3D microphysiological system enabled real-time assessment of epithelial morphology and barrier integrity, revealing that EV treatment repaired inflammation-induced structural damage, restored tight junction integrity, and suppressed inflammatory signaling. These findings highlight the value of organ-on-a-chip technology as a physiologically relevant platform for investigating the mechanisms of probiotic EVs and their therapeutic potential in modulating the gut-skin axis [131]. Collectively, these in vitro, in vivo, and emerging microfluidic studies provide complementary preclinical evidence supporting further investigation of PEVs for AD and related inflammatory or barrier-defect conditions. Future development of integrated gut–skin-on-a-chip platforms may further advance mechanistic understanding by enabling simultaneous evaluation of intestinal and cutaneous responses within a single physiologically relevant system. Together, these advances position PEVs as promising candidates for future translational research, with clinical studies representing the next essential step toward therapeutic application in AD.

Critical Evaluation and Methodological Limitations of Preclinical Evidence

While published preclinical studies demonstrate promising proof-of-concept efficacy, several critical methodological caveats must be acknowledged when interpreting these data for clinical translation:

Limitations of Murine AD Models

Most in vivo evidence relies on acute, chemically induced murine models (e.g., topical DNCB, oxazolone, or mechanical tape-stripping). While these models reproduce acute Th2 inflammation and epidermal barrier disruption, they fail to fully recapitulate the chronic, relapsing course, polygenic susceptibility (e.g., hereditary FLG null mutations), and complex cutaneous microbiome dysbiosis seen in human AD patients. Furthermore, rodent skin possesses a substantially thinner stratum corneum, higher follicular density, and distinct immune architecture compared to human skin, which may artificially facilitate topical EV penetration and inflate observed therapeutic efficacy [135].

Overreliance on Simplified 2D In Vitro Monocultures

A significant proportion of mechanistic data is derived from immortalized monocultures (e.g., HaCaT, RAW 264.7, THP-1). These reductionist systems lack the multicellular complexity, mechanical tension, air-liquid interface, and dynamic immune-epithelial crosstalk inherent to living human tissue [135].

Conflation of Whole Bacteria with Purified EV Efficacy

In several preclinical reports (e.g., B. longum studies), isolated EVs were validated in vitro, while in vivo efficacy was demonstrated exclusively using the live parent bacterial strain [107]. Extrapolating in vivo benefits of the whole bacterium to its derived extracellular vesicles without direct, head-to-head in vivo EV administration represents an evidential gap that requires direct confirmation.

Lack of Pharmacokinetics and Dose-Optimization Studies

Current preclinical studies predominantly test single, arbitrarily selected high-dose EV regimens over short experimental windows (typically 1 to 3 weeks). There is a critical scarcity of rigorous dose-response curves, tissue retention kinetics, pharmacokinetic clearance data, and long-term safety evaluations for repeated topical or oral administration [44, 45]. Addressing these methodological gaps through standardized, human-relevant 3D organotypic cultures and chronic models is essential prior to clinical progression [135].

From Bench to Bedside: Clinical Prospects of PEVs in AD

Although clinical studies specifically evaluating PEVs in AD are currently lacking, clinical evidence from live probiotics and other postbiotic interventions provides indirect support for the broader therapeutic relevance of microbiome-based approaches. However, these findings cannot be directly extrapolated to PEVs because EV composition, pharmacokinetics, dosing, and biological activity may differ substantially from those of live probiotics or other microbial products [136, 137]. However, clinical outcomes remain inconsistent. A recent meta-analysis of randomized, placebo-controlled trials showed that while certain topical probiotics, particularly Vitreoscilla filiformis-based formulations, improved SCORAD (Scoring AD) indices, the overall pooled therapeutic benefit was not statistically significant [136]. Similarly, a pilot study of a topical L. lactis lysate cream demonstrated excellent tolerability but failed to achieve significant clinical efficacy [138]. These variable outcomes highlight the limitations of conventional topical probiotic formulations and emphasize the need for more standardized, mechanism-based approaches.

In this context, PEVs represent a promising next-generation postbiotic platform that overcomes the inherent physical and safety limitations of earlier biotherapeutics. For AD patients with compromised epidermal barriers, applying live probiotics carries theoretical risks of opportunistic infection or unpredictable microbial colonization. As a cell-free alternative, PEVs largely circumvent these safety concerns [37]. Furthermore, unlike bulky live bacteria that remain confined to skin surface, the structurally intact, nanoscale lipid-bilayer architecture of PEVs may facilitate penetration through the compromised epidermal barrier characteristic of AD, enabling delivery of bioactive cargo to resident skin cells [37, 139]. Despite these advantages, several manufacturing, bioengineering, and regulatory challenges must still be overcome before PEV-based therapies can be translated into clinical practice.

Translational Challenges and Future Perspectives

Although preclinical research into PEVs-based AD treatments shows promising potential, the field is still in its early phase of progression from proof-of-concept studies toward clinical application. Currently, this transition is constrained by several interconnected scientific, technical, and regulatory challenges. A comprehensive evaluation of these limitations, together with a clear definition of future research directions, is essential to advance PEVs from experimental models into standardized and clinically viable therapeutics.

Manufacturing Scalability and Production Efficiency

The large-scale production of PEVs represents one of the primary barriers to clinical translation. Under physiological conditions, probiotic bacteria naturally release EVs at relatively low levels, making industrial-scale production difficult. Recent advances in bacterial engineering have partially overcome this limitation through the development of hypervesiculating strains generated by modifying genes involved in the overexpression of OM proteases (particularly in Gram-negative bacteria), PG remodeling, or membrane stability. These engineered strains have demonstrated substantial increases in EV production while largely preserving their biological activity [140].

Despite these advances, scalable manufacturing of EVs remains a significant challenge. Ultracentrifugation (UC), the most widely used laboratory isolation technique, is labor-intensive, time-consuming, and unsuitable for large-scale manufacturing. Moreover, prolonged high-speed centrifugation may damage EVs structures or promote their aggregation, resulting in low EVs recovery rates. To overcome this issue, emerging technologies such as density-gradient centrifugation (DGC), tangential flow filtration (TFF), cross-flow filtration, and continuous bioprocessing offer more scalable alternatives for EVs production with improved recovery and reproducibility [141]. Nevertheless, optimization of fermentation conditions, bacterial growth parameters, purification workflows, and storage stability remain necessary to ensure high production yields while maintaining EVs integrity and biological function [140].

In addition to upstream yield, storage stability and batch-to-batch reproducibility represent formidable hurdles for industrial translation. Bacterial EVs in aqueous suspension are prone to membrane fusion, leakage of intraluminal cargo, and spontaneous aggregation over time or during repeated freeze–thaw cycles. While storage at − 80 °C preserves structural integrity for laboratory research, commercial distribution demands stable formulations. Developing optimized lyophilization (freeze-drying) protocols using non-toxic lyoprotectants/cryoprotectants (such as trehalose, sucrose, or mannitol) is essential to maintain vesicle size, membrane fluidity, and biological potency at refrigerated (4 °C) or room temperatures. Furthermore, biological variability is inherent to bacterial fermentation and may arise from subtle fluctuations in growth phase (exponential vs. stationary), nutrient availability, and membrane lipid turnover which frequently results in batch-to-batch heterogeneity in cargo packaging. Establishing automated bioreactor controls and strict upstream standard operating procedures will be required to guarantee consistent, reproducible vesicle yields [140].

Quality Control and Standardization

The absence of standardized methodologies for PEV isolation, characterization, and quality assessment remains a major obstacle to reproducibility and comparability across preclinical and translational studies. Different isolation techniques including UC, size-exclusion chromatography, DGC, and TFF yield vesicle preparations with marked differences in purity, particle recovery, and macromolecular cargo composition, confounding direct comparisons between published reports [142]. In addition, rigorous assessment of preparation purity is exceptionally critical for PEVs because bacterial fermentation broths contain abundant non-vesicular contaminants, including cell wall and outer membrane fragments, flagellin, unencapsulated nucleic acids, secreted proteins, and complex culture media constituents that readily co-isolate with intact vesicles. If not stringently separated, these bioactive impurities can obscure the true biological mechanisms of vesicles, introduce batch variability, and provoke off-target inflammatory responses [140].

To improve scientific rigor, the International Society for Extracellular Vesicles (ISEV) established the Minimal Information for Studies of Extracellular Vesicles (MISEV2023) guidelines, providing an essential global framework for reporting EV isolation, biochemical characterization, and cargo analysis [143]. However, while MISEV2023 offers vital overarching standards primarily established around eukaryotic vesicles, dedicated consensus guidelines specifically tailored for bacterial and probiotic-derived EVs are still lacking. Unlike mammalian exosomes, bacterial vesicles feature unique structural envelopes (e.g., LPS-rich outer membranes or LTA-embedded cytoplasmic membranes), distinct biogenesis pathways, and broad size distributions (20–400 nm). Establishing consensus guidelines specifically addressing bacterial EV-specific markers, membrane purity standards, and clearance thresholds for microbial culture artifacts represents a critical unmet need in the field.

For clinical translation under current good manufacturing practice (cGMP) standards, comprehensive quality control (QC) frameworks must establish well-defined critical quality attributes and standardized batch release criteria. This requires multi-modal orthogonal characterization, integrating nanoparticle tracking analysis (NTA) or dynamic light scattering (DLS) for particle size distribution and concentration, cryogenic transmission electron microscopy (Cryo-TEM) for vesicular morphology and membrane integrity, and quantitative purity metrics such as particle-to-protein and particle-to-lipid ratios. Furthermore, because bacterial EVs can exceed 200 nm in diameter, conventional terminal 0.22 μm sterile filtration can lead to membrane shear, vesicle rupture, or selective exclusion of larger therapeutic subfractions, underscoring the necessity of validated closed-system aseptic processing and rigorous bioburden monitoring. Finally, moving beyond physical characterization, the clinical pipeline demands the development of standardized, disease-relevant bioassays to quantitatively determine the functional immunomodulatory and barrier-fortifying potency of each manufactured PEV batch prior to clinical application [143–145].

Pharmacokinetics, Biodistribution, Safety, and Immunogenicity

Limited understanding of the pharmacokinetic behavior of PEVs remains a major barrier to their clinical translation. Fundamental questions regarding penetration through the stratum corneum, biodistribution within inflamed skin, tissue retention, cellular uptake, and metabolic clearance remain largely unanswered [139]. Because AD requires chronic and potentially repeated treatment, comprehensive evaluation of repeated-dose exposure, chronic toxicity, off-target effects, and potential alterations of the resident skin microbiome will be essential for establishing a clinically acceptable safety profile [139].

EV origin is an important consideration in safety assessment; however, safety cannot be inferred solely from Gram classification. Gram-positive MVs lack LPS-associated endotoxin, but their biological activity remains dependent on strain, vesicular cargo, dose, purity, and exposure duration [99, 126, 127]. Gram-negative OMVs require additional consideration of LPS-associated inflammatory activity, particularly with increasing dose or repeated administration [122, 123]. Although controlled exposure to OMV-associated LPS may exert beneficial immunomodulatory effects, excessive or prolonged exposure may increase the risk of endotoxin-associated toxicity and unwanted inflammatory responses. Accordingly, dose optimization and appropriate monitoring of vesicle-associated endotoxin activity will be important for the development of Gram-negative OMV-based therapies [60, 122, 123].

Safety concerns are therefore relevant to both EV classes. Their complex and strain-dependent cargo may influence immunogenicity, biodistribution, and off-target biological effects. In addition, inadequate purification may result in co-isolation of non-vesicular bacterial components, including cellular and membrane fragments, soluble proteins, nucleic acids, and residual culture-derived materials, potentially affecting product consistency, interpretation of biological activity, and safety [140, 142]. Consequently, absence of LPS in Gram-positive MVs should not be interpreted as evidence of an intrinsically risk-free platform, because other bioactive components, including LTA and bacterial proteins, can remain immunologically active in a strain- and dose-dependent manner [99, 126, 127]. For both platforms, repeated administration also warrants assessment of cumulative toxicity, persistent or unintended immune activation, altered tissue distribution, and possible effects on the resident microbiome. Overall, clinical development will require preparation-specific assessment of composition, purity, potency, dose, route of administration, biodistribution, immunogenicity, and long-term safety rather than reliance on general assumptions based on EV origin.

Regulatory Challenges

PEVs occupy a unique regulatory position as complex, cell-free biological products whose heterogeneous composition and manufacturing processes directly influence product quality, potency, and batch-to-batch consistency. This makes it difficult to classify PEVs within existing pharmaceutical or biological regulatory frameworks. Consequently, the clinical translation of PEVs is hindered not only by technical challenges but also by the absence of dedicated regulatory pathways. To date, no major regulatory authority, including U.S. Food and Drug Administration (FDA), European Medicines Agency (EMA), or Therapeutic Goods Administration (TGA), has established specific technical evaluation guidelines for EV-based therapeutics, and no EV product has yet received full market approval [144, 145].

The need for clear regulatory oversight is further underscored by FDA public safety warnings regarding the unauthorized marketing of unlicensed exosome products [139]. Therefore, successful clinical translation of PEVs will require close collaboration among researchers, manufacturers, and regulatory agencies to establish EV-specific regulatory frameworks that incorporate stringent cGMP standards, standardized quality attributes, validated potency assays, and comprehensive long-term safety monitoring.

Delivery Strategies for Topical PEV Therapy

Efficient delivery of PEVs to diseased skin remains a major technological challenge for their clinical application in AD. The highly organized structure of stratum corneum represents a formidable physical barrier that limits EVs penetration following topical administration, potentially reducing local bioavailability and therapeutic efficacy [139]. Therefore, developing delivery systems that enhance skin penetration while preserving PEV stability and biological activity represents an important priority. Emerging formulation strategies, including hydrogels, creams, lipid-based carriers, dissolvable microneedle patches, and bioadhesive platforms, may improve EVs retention, controlled release, and localized therapeutic effects [146]. In addition, physical permeation-enhancing approaches such as microneedles, ultrasound, and iontophoresis provide promising strategies to facilitate PEV transport across the skin barrier [146]. Ultimately, utilizing these advanced delivery platforms will maximize targeted topical efficacy while simultaneously minimizing off-target systemic exposure.

Recent advances in artificial intelligence (AI) and machine learning have driven new predictive modeling and personalized medicine approaches, demonstrating potential for predicting skin permeability, optimizing topical formulations, and supporting personalized therapeutic strategies through the integration of large molecular and clinical datasets [147]. Although these approaches have not yet been extensively applied to PEV-based therapies, they may facilitate the rational design of PEV formulations and improve topical delivery for AD. Future advances in EV surface engineering may further enhance skin targeting, penetration efficiency, and disease-specific delivery, thereby improving the clinical potential of PEV-based therapeutics.

Future Research Priorities

To bridge the gap between compelling preclinical proof-of-concept studies and successful clinical translation, future research on PEVs in AD must focus on the following interconnected scientific and clinical priorities:

Defining Major Knowledge Gaps

Several fundamental questions remain unresolved. These include how bacterial strain, growth conditions, and vesicle biogenesis determine cargo selection; which vesicular components are responsible for specific immunological and barrier-related effects; and how individual cargo components interact to produce strain-specific biological responses. Further studies should also distinguish vesicle-mediated activity from effects caused by free LPS, LTA, proteins, nucleic acids, or other co-isolated bacterial components. Clarifying the mechanisms underlying intestinal uptake, systemic trafficking, skin targeting, and cargo delivery will be essential for establishing well-defined mechanisms of action [95, 139–143].

Standardized Experimental Approaches

Future studies should use harmonized experimental designs that allow meaningful comparison across bacterial strains and EV platforms. This includes standardized isolation and characterization procedures, consistent normalization of EV dose, reporting of particle concentration and purity, comprehensive cargo characterization, and validated functional potency assays. Standardized reporting should also distinguish particle-based dosing from protein- or lipid-based normalization and clearly define the biological model, administration route, exposure duration, and relevant controls. Such harmonization will improve reproducibility and enable more reliable cross-study comparisons [142, 143].

Selection of Appropriate Preclinical and Human-Relevant Models

The choice of experimental model is critical for translational interpretation. Established murine AD models, including NC/Nga and chemically induced models, remain valuable for evaluating inflammation, barrier dysfunction, immune responses, and systemic effects. However, differences between murine and human skin architecture and immune regulation limit direct extrapolation to patients. Future research should therefore compare PEV efficacy across complementary animal models rather than relying on a single model and should incorporate human 3D skin equivalents, organ-on-a-chip systems, and other microphysiological platforms to improve human relevance [135]. These approaches may also facilitate simultaneous evaluation of intestinal and cutaneous responses in gut–skin axis studies.

Biomarker and Patient-Stratification Strategies

Biomarker development should become an integral component of future PEV research. Candidate biomarkers may include immune signatures, epidermal barrier-associated molecules, microbiome features, EV biodistribution parameters, and pharmacodynamic responses. Integrating microbiome profiling, multi-omics analyses, and clinical phenotyping may enable identification of patient subgroups most likely to benefit from specific PEV preparations. Crucially, recent machine learning analyses of clinical datasets reveal that gut microbiota-derived EVs form a distinct taxonomic entity entirely separate from the conventional gut microbiota [148]. Therefore, future clinical studies must profile microbiota-derived EVs independently, as standard fecal microbiome composition does not accurately reflect the actual EV-mediated communication between the microbiome and the host [148]. Such approaches could ultimately support mechanism-based patient stratification and improve monitoring of therapeutic response and safety [139, 149].

Engineering Strategies for Defined and Targeted PEVs

Bioengineering approaches may enable the development of more consistent and disease-specific PEV products. Future strategies should focus on controlling vesicle production, cargo composition, immunomodulatory activity, tissue targeting, and safety while preserving vesicle integrity and biological function. Hypervesiculating strains may improve production efficiency, whereas synthetic biology and CRISPR-based engineering could enable more precise control of vesicular cargo or surface properties [140, 149]. Engineering strategies should nevertheless be accompanied by rigorous evaluation of unintended changes in vesicle composition, host immune activation, and long-term safety.

Priorities for Future Clinical Trials

Clinical translation should proceed in a staged manner after adequate preclinical characterization and manufacturing standardization. Early-phase clinical studies should initially focus on well-characterized PEV preparations with defined composition, reproducible potency, and acceptable toxicological profiles. Key objectives should include evaluation of dose, route of administration, pharmacokinetics, biodistribution, tolerability, immunogenicity, and preliminary clinical activity using validated AD severity and barrier-related endpoints. Randomized controlled trials will subsequently be required to determine efficacy relative to standard or placebo-controlled treatments, with biomarker-based assessments incorporated where appropriate. Because direct clinical studies of PEVs in AD are currently lacking, early trials should emphasize safety and pharmacological characterization before large-scale efficacy studies. Long-term follow-up will also be essential because AD is a chronic disease requiring repeated treatment.

Collectively, these priorities provide a roadmap for moving PEV research from heterogeneous preclinical studies toward standardized, mechanism-informed, and clinically evaluable therapeutic platforms (Fig. 4). Successful translation will depend on integrating rigorous EV characterization, appropriate disease models, preparation-specific safety assessment, biomarker-guided development, rational engineering, scalable manufacturing, and carefully phased clinical evaluation.

Fig. 4 — Translational roadmap and future perspectives for the clinical application of PEV-based therapeutics in AD. The schematic highlights the major challenges and future directions for PEVs clinical translation, including scalable manufacturing, rigorous quality control (QC) and standardization, safety evaluation, advanced topical delivery strategies, and next-generation engineering approaches to enable safe and effective clinical application. (Created in https://BioRender.com.)

Conclusion

PEVs represent a transformative conceptual shift in microbiome-based medicine, evolving from empirical live-microbe interventions toward defined, cell-free nanotherapeutics for AD. By deciphering the architectural disparities between bacterial envelopes, recent research has illuminated distinct mechanistic paradigms: Gram-negative OMVs utilize controlled TLR4 signaling and metabolic cargo to reinforce innate antimicrobial defense and rebalance Th1/Th2 immunity, whereas Gram-positive MVs preferentially engage tolerogenic TLR2 pathways to expand regulatory T cells, suppress inflammatory cascades, and directly upregulate epidermal barrier proteins through local and gut–skin axis signaling.

Despite these compelling preclinical advances, substantial translational hurdles separate laboratory findings from clinical reality. Current evidence remains almost exclusively confined to reductionist 2D cell cultures and acute murine models that do not fully mirror the chronic, polygenic, and relapsing nature of human AD. Furthermore, the clinical pipeline is constrained by significant chemistry, manufacturing, and controls challenges—including low physiological yields, batch-to-batch cargo heterogeneity, storage instability, and the lack of consensus quality-control frameworks specifically tailored to bacterial vesicles. Addressing the narrow therapeutic window of LPS-containing OMVs and establishing rigorous bioburden and purity standards remain mandatory prerequisites before human administration.

Looking forward, the realistic clinical translation of PEVs will depend on targeted bioengineering and well-staged clinical trial designs. Early-phase clinical application will likely center on standardized, topical formulations for mild-to-moderate disease before advancing to systemic therapies. Leveraging synthetic biology to generate detoxified, hypervesiculating bacterial strains, alongside adopting human-relevant 3D organotypic platforms and biomarker-guided patient stratification, will be essential to realize the potential of PEVs as safe, consistent, and mechanism-informed therapeutics for AD.

Acknowledgements

All figures in this manuscript were created using BioRender.com.

Author Contributions

Conceptualization of the review, V.W. and Y.-H.C; literature survey and wrote the manuscript, V.W.; visualization (figures and table), V.W.; project administration, Y.-H.C.; critical scientific advice and reviewed the manuscript, M.-F.L., J.-Y.W and Y.-H.C. All authors reviewed and approved the final version of the submitted manuscript.

Funding

This work was supported by the China Medical University Hospital, Taichung, Taiwan.

Data Availability

No datasets were generated or analysed during the current study.

Declarations

Ethical Approval

Not applicable.

Competing Interests

The authors declare no competing interests.

Footnotes

Publisher’s Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Contributor Information

Jiu-Yao Wang, Email: 008910@tool.caaumed.org.tw.

Yi-Hsing Chen, Email: yhchen008@cmu.edu.tw.

References

1.Yue C, Zhou H, Wang X, Yu J, Hu Y, Zhou P et al (2024) Atopic dermatitis: pathogenesis and therapeutic intervention. MedComm 5:e70029. 10.1002/mco2.70029 [DOI] [PMC free article] [PubMed] [Google Scholar]
2.Hansen-Sackey EB, Hartono S (2025) Atopic Dermatitis: Pathophysiology and Emerging Treatments. Allergies 5:40. 10.3390/allergies5040040 [DOI] [Google Scholar]
3.Moreiras-Arias N, Nieto-Fontarigo JJ, Salgado FJ, González-Vilas D, Paredes-Suárez C, Combo-García E et al (2026) Novel therapeutic strategies for atopic dermatitis: biomarker modulation and clinical implications. A systematic review. Clin Rev Allergy Immunol 69:4. 10.1007/s12016-025-09129-z [DOI] [PMC free article] [PubMed] [Google Scholar]
4.Kolb L, Villarreal LA, Sathe NC (2026) Atopic dermatitis. In: StatPearls [Internet] StatPearls Publishing Treasure Island (FL). https://www.ncbi.nlm.nih.gov/books/NBK448071/. Accessed 27 May 2026 [PubMed]
5.Tian J, Zhang D, Yang Y, Huang Y, Wang L, Yao X et al (2023) Global epidemiology of atopic dermatitis: a comprehensive systematic analysis and modelling study. Br J Dermatol 190:55–61. 10.1093/bjd/ljad339 [DOI] [PubMed] [Google Scholar]
6.Lyons JJ, Milner JD, Stone KD (2015) Atopic dermatitis in children: clinical features, pathophysiology, and treatment. Immunol Allergy Clin North Am 35:161–183. 10.1016/j.iac.2014.09.008 [DOI] [PMC free article] [PubMed] [Google Scholar]
7.Bai R, Zheng Y, Dai X (2025) Atopic dermatitis: diagnosis, molecular pathogenesis, and therapeutics. Mol Biomed 6:71. 10.1186/s43556-025-00313-3 [DOI] [PMC free article] [PubMed] [Google Scholar]
8.Langan SM, Irvine AD, Weidinger S (2020) Atopic dermatitis. Lancet 396:345–360. 10.1016/S0140-6736(20)31286-1 [DOI] [PubMed] [Google Scholar]
9.Borici S (2025) Atopic dermatitis and food allergy in children. Acta Paediatr 114:3096–3107. 10.1111/apa.70283 [DOI] [PubMed] [Google Scholar]
10.Baidya A, Mabalirajan U (2025) Pathogenesis and management of atopic dermatitis: insights into epidermal barrier dysfunction and immune mechanisms. Explor Asthma Allergy 3:100973. 10.37349/eaa.2025.100973 [DOI] [Google Scholar]
11.Zhong L, Zhou X, Su J, Zhang Y, Zhang D, Wan H (2026) Microbiome dysbiosis and therapeutic restoration in atopic dermatitis. Front Cell Infect Microbiol 16:1693905. 10.3389/fcimb.2026.1693905 [DOI] [PMC free article] [PubMed] [Google Scholar]
12.Micu AE, Popescu IA, Halip IA, Mocanu M, Vâță D, Hulubencu AL et al (2026) From gut dysbiosis to skin inflammation in atopic dermatitis: Probiotics and the gut–skin axis—clinical outcomes and microbiome implications. Int J Mol Sci 27:365. 10.3390/ijms27010365 [DOI] [PMC free article] [PubMed] [Google Scholar]
13.Zhang Z, Wang R, Li M, Lu M (2025) Current insights and trends in atopic dermatitis and microbiota interactions: a systematic review and bibliometric analysis. Front Microbiol 16:1613315. 10.3389/fmicb.2025.1613315 [DOI] [PMC free article] [PubMed] [Google Scholar]
14.Lee SY, Lee E, Park YM, Hong SJ (2018) Microbiome in the gut-skin axis in atopic dermatitis. Allergy Asthma Immunol Res 10:354–362. 10.4168/aair.2018.10.4.354 [DOI] [PMC free article] [PubMed] [Google Scholar]
15.Zhao Y, Yu C, Zhang J, Yao Q, Zhu X, Zhou X (2025) The gut–skin axis: Emerging insights in understanding and treating skin diseases through gut microbiome modulation (Review). Int J Mol Med 56:210. 10.3892/ijmm.2025.5651 [DOI] [PMC free article] [PubMed] [Google Scholar]
16.Mazziotta C, Tognon M, Martini F, Torreggiani E, Rotondo JC (2023) Probiotics mechanism of action on immune cells and beneficial effects on human health. Cells 12:184. 10.3390/cells12010184 [DOI] [PMC free article] [PubMed] [Google Scholar]
17.Anania C, Brindisi G, Martinelli I, Bonucci E, D’Orsi M, Ialongo S et al (2022) Probiotics function in preventing atopic dermatitis in children. Int J Mol Sci 23:5409. 10.3390/ijms23105409 [DOI] [PMC free article] [PubMed] [Google Scholar]
18.Lim A, Baek J, Seo M, Kim K-y, Kim S, Kwak W et al (2025) Atopic dermatitis-alleviating effects of lactiplantibacillus plantarum LRCC5195 paraprobiotics through microbiome modulation and safety assessment via genomic characterization and in vitro analysis. Sci Rep 15:30545. 10.1038/s41598-025-16102-5 [DOI] [PMC free article] [PubMed] [Google Scholar]
19.Sung M, Sim S, Lim A, Moon JS, Jeon J, Heo K et al (2025) BCL-2 multi-strain probiotics for immunomodulation in vitro and in vivo alleviation of atopic dermatitis. Microorganisms 13:1950. 10.3390/microorganisms13081950 [DOI] [PMC free article] [PubMed] [Google Scholar]
20.Kang H-J, Kim G-C, Lee C-G, Park S, Sharma G, Verma R et al (2021) Probiotics-derived metabolite ameliorates skin allergy by promoting differentiation of FOXP₃⁺regulatory T cells. J Allergy Clin Immunol 147:1517–1521. 10.1016/j.jaci.2020.11.040 [DOI] [PubMed] [Google Scholar]
21.Zhong L, Su J, Zhou X, Wan H (2025) Probiotics supplements for the prevention of atopic dermatitis in children: an umbrella review. Front Nutr 12:1587348. 10.3389/fnut.2025.1587348 [DOI] [PMC free article] [PubMed] [Google Scholar]
22.Xue X, Yang X, Shi X, Deng Z (2023) Efficacy of probiotics in pediatric atopic dermatitis: A systematic review and meta-analysis. Clin Transl Allergy 13:e12283. 10.1002/clt2.12283 [DOI] [PMC free article] [PubMed] [Google Scholar]
23.Liscano Y, Muñoz Morales D, Suarez Daza F, Vidal Cañas S, Martinez Guevara D, Artunduaga Cañas E (2025) Microbial interventions for inflammatory skin diseases: A systematic review and meta-analysis of atopic dermatitis and psoriasis. Microorganisms 13:2416 [DOI] [PMC free article] [PubMed] [Google Scholar]
24.Mirali S, Drucker AM (2025) New topical and systemic treatments for atopic dermatitis. Clin Exp Allergy 55:1057–1069. 10.1111/cea.70136 [DOI] [PMC free article] [PubMed] [Google Scholar]
25.Biliński K, Rakoczy K, Karwowska A, Cichy O, Wojno A, Wojno A et al (2025) Anti-inflammatory therapies for atopic dermatitis: A new era in targeted treatment. J Clin Med 14:5053. 10.3390/jcm14145053 [DOI] [PMC free article] [PubMed] [Google Scholar]
26.Jeskey J, Kurien C, Blunk H, Sehmi K, Areti S, Nguyen D et al (2024) Atopic dermatitis: A review of diagnosis and treatment. J Pediatr Pharmacol Ther 29:587–603. 10.5863/1551-6776-29.6.587 [DOI] [PMC free article] [PubMed] [Google Scholar]
27.Ali S, Ion A, Orzan OA, Bălăceanu-Gurău B (2024) Emerging treatments and new vehicle formulations for atopic dermatitis. Pharmaceutics 16:1425. 10.3390/pharmaceutics16111425 [DOI] [PMC free article] [PubMed] [Google Scholar]
28.Elahi N, Astaneh ME, Ai J, Rizwan M (2024) Atopic dermatitis treatment: A comprehensive review of conventional and novel bioengineered approaches. Int J Biol Macromol 282:137083. 10.1016/j.ijbiomac.2024.137083 [DOI] [PubMed] [Google Scholar]
29.Zaborowski MP, Balaj L, Breakefield XO, Lai CP (2015) Extracellular vesicles: Composition, biological relevance, and methods of study. Bioscience 65:783–797. 10.1093/biosci/biv084 [DOI] [PMC free article] [PubMed] [Google Scholar]
30.Repici A, Piraino G, Wolfe V, Kaplan J, Nakamura T, Zingarelli B (2025) Role of extracellular vesicles as mediators of cell communication and novel biomarkers in sepsis. J Clin Med 14:6649. 10.3390/jcm14186649 [DOI] [PMC free article] [PubMed] [Google Scholar]
31.Premchandani T, Umekar M, Tatode A, Taksande J, Khan R, Faizan M et al (2025) Extracellular vesicles in host–pathogen interactions: Roles of exosomes and bacterial outer membrane vesicles in immunity and microbial communication. Bacteria 4:63. 10.3390/bacteria4040063 [DOI] [Google Scholar]
32.Yu Y-j, Wang X-h, Fan G-C (2018) Versatile effects of bacterium-released membrane vesicles on mammalian cells and infectious/inflammatory diseases. Acta Pharmacol Sin 39:514–533. 10.1038/aps.2017.82 [DOI] [PMC free article] [PubMed] [Google Scholar]
33.Kim JH, Lee J, Park J, Gho YS (2015) Gram-negative and Gram-positive bacterial extracellular vesicles. Semin Cell Dev Biol 40:97–104. 10.1016/j.semcdb.2015.02.006 [DOI] [PubMed] [Google Scholar]
34.Melo-Marques I, Cardoso SM, Empadinhas N (2024) Bacterial extracellular vesicles at the interface of gut microbiota and immunity. Gut Microbes 16:2396494. 10.1080/19490976.2024.2396494 [DOI] [PMC free article] [PubMed] [Google Scholar]
35.El-Saadony MT, Saad AM, Sitohy M, Alkafaas SS, Dladla M, Ghosh S et al (2026) Probiotics and human health: biological activities, nutritional aspects, immunomodulatory properties, applications, and future perspectives - a comprehensive review. Front Immunol 16:1713426. 10.3389/fimmu.2025.1713426 [DOI] [PMC free article] [PubMed] [Google Scholar]
36.Ma T, Shen X, Shi X, Sakandar HA, Quan K, Li Y et al (2023) Targeting gut microbiota and metabolism as the major probiotic mechanism - An evidence-based review. Trends Food Sci Technol 138:178–198. 10.1016/j.tifs.2023.06.013 [DOI] [Google Scholar]
37.Kozakiewicz D, Razim A, Schabussova I, Górska S (2025) Immunomodulatory properties of probiotic extracellular vesicles- do they have the potential to fill the gap in therapeutic strategies for allergies? Cell Commun Signal 23:549. 10.1186/s12964-025-02532-1 [DOI] [PMC free article] [PubMed] [Google Scholar]
38.Suri K, D’Souza A, Huang D, Bhavsar A, Amiji M (2023) Bacterial extracellular vesicle applications in cancer immunotherapy. Bioact Mater 22:551–566. 10.1016/j.bioactmat.2022.10.024 [DOI] [PMC free article] [PubMed] [Google Scholar]
39.Kim MH, Choi SJ, Choi HI, Choi JP, Park HK, Kim EK et al (2018) Lactobacillus plantarum-derived extracellular vesicles protect atopic dermatitis induced by Staphylococcus aureus-derived extracellular vesicles. Allergy Asthma Immunol Res 10:516–532. 10.4168/aair.2018.10.5.516 [DOI] [PMC free article] [PubMed] [Google Scholar]
40.Baek J, Cho S, Lee G, Ki H, Kim SY, Choi GM et al (2025) Modulation of moisturizing and barrier related molecular markers by extracellular vesicles derived from Leuconostoc mesenteroides DB-21 isolated from camellia japonica flower. Curr Issues Mol Biol 47:1022. 10.3390/cimb47121022 [DOI] [PMC free article] [PubMed] [Google Scholar]
41.Choi H, Kwak M-J, Choi Y, Kang AN, Mun D, Eor JY et al (2025) Extracellular vesicles of Limosilactobacillus fermentum SLAM216 ameliorate skin symptoms of atopic dermatitis by regulating gut microbiome on serotonin metabolism. Gut Microbes 17:2474256. 10.1080/19490976.2025.2474256 [DOI] [PMC free article] [PubMed] [Google Scholar]
42.Cho Y-H, Kim J-W, Kim N, Kim H-S, Jang J-H, Bae J-T et al (2025) Lactobacillus brevis-derived exosomes enhance skin barrier integrity by upregulating key barrier-related proteins. Clin Cosmet Investig Dermatol 18:1151–1162. 10.2147/ccid.s512793 [DOI] [PMC free article] [PubMed] [Google Scholar]
43.Shi Y, Li S, Yang S (2025) Probiotic-derived extracellular vesicles: a novel weapon against viral infections. Biosci Microbiota Food Health 44:245–250. 10.12938/bmfh.2025-027 [DOI] [PMC free article] [PubMed] [Google Scholar]
44.Atoom AM, Hamed-Ahmed M, Al-Hasnaawei S, Malathi H, Maharana L, Nanda A et al (2026) Therapeutic potential and translational challenges of extracellular vesicles in neonatal medicine. Bioeng Transl Med 11:e70093. 10.1002/btm2.70093 [DOI] [PMC free article] [PubMed] [Google Scholar]
45.Primorac D, Brlek P, Bulić L, Hrvatin N, Škaro V, Projić P et al (2026) Therapeutic potential of extracellular vesicles: From biogenesis, isolation and molecular characterization to addressing translational gaps and regulatory barriers. Int J Mol Sci 27:1676. 10.3390/ijms27041676 [DOI] [PMC free article] [PubMed] [Google Scholar]
46.Wang Y, Duan C, Yu X (2025) Extracellular vesicles of Gram-negative and Gram-positive probiotics. J Microbiol 63:e2506005. 10.71150/jm.2506005 [DOI] [PMC free article] [PubMed] [Google Scholar]
47.Liu BD, Akbar R, Oliverio A, Thapa K, Wang X, Fan GC (2024) Bacterial extracellular vesicles in the regulation of inflammatory response and host-microbe interactions. Shock 61:175–188. 10.1097/shk.0000000000002252 [DOI] [PMC free article] [PubMed] [Google Scholar]
48.Kurata A, Uegaki K (2025) Recent advances in understanding the role of extracellular vesicles from probiotics in intestinal immunity signaling. Biochem Soc Trans 53:419–429. 10.1042/bst20240150 [DOI] [PMC free article] [PubMed] [Google Scholar]
49.Zhang Y, Liang F, Zhang D, Qi S, Liu Y (2023) Metabolites as extracellular vesicle cargo in health, cancer, pleural effusion, and cardiovascular diseases: An emerging field of study to diagnostic and therapeutic purposes. Biomed Pharmacother 157:114046. 10.1016/j.biopha.2022.114046 [DOI] [PubMed] [Google Scholar]
50.Avila-Calderón ED, Ruiz-Palma MdS, Aguilera-Arreola MG, Velázquez-Guadarrama N, Ruiz EA, Gomez-Lunar Z et al (2021) Outer membrane vesicles of Gram-negative bacteria: An outlook on biogenesis. Front Microbiol 12:557902. 10.3389/fmicb.2021.557902 [DOI] [PMC free article] [PubMed] [Google Scholar]
51.Zhang L, He Y, Wang G, Sun J, Chen Y, Wang Z (2026) Bacterial membrane vesicles: Biogenesis, functions, and emerging biotechnological applications. Microorganisms 14:689. 10.3390/microorganisms14030689 [DOI] [PMC free article] [PubMed] [Google Scholar]
52.Jan AT (2017) Outer Membrane Vesicles (OMVs) of Gram-negative bacteria: A perspective update. Front Microbiol 8:1053. 10.3389/fmicb.2017.01053 [DOI] [PMC free article] [PubMed] [Google Scholar]
53.Schwechheimer C, Kuehn MJ (2015) Outer-membrane vesicles from Gram-negative bacteria: biogenesis and functions. Nat Rev Microbiol 13:605–619. 10.1038/nrmicro3525 [DOI] [PMC free article] [PubMed] [Google Scholar]
54.Zhao X, Wei Y, Bu Y, Ren X, Dong Z (2025) Review on bacterial outer membrane vesicles: structure, vesicle formation, separation and biotechnological applications. Microb Cell Fact 24:27. 10.1186/s12934-025-02653-9 [DOI] [PMC free article] [PubMed] [Google Scholar]
55.Giordano NP, Cian MB, Dalebroux ZD (2020) Outer membrane lipid secretion and the innate immune response to Gram-negative bacteria. Infect Immun 88:e00920–e00919. 10.1128/iai.00920-19 [DOI] [PMC free article] [PubMed] [Google Scholar]
56.Ma D, Zhang Y, Zhang J, Shi J, Gao S, Long F et al (2025) Outer membrane vesicles derived from probiotic Escherichia coli Nissle 1917 promote metabolic remodeling and M1 polarization of RAW264.7 macrophages. Front Immunol 16:1501174. 10.3389/fimmu.2025.1501174 [DOI] [PMC free article] [PubMed] [Google Scholar]
57.Aguilera L, Toloza L, Giménez R, Odena A, Oliveira E, Aguilar J et al (2014) Proteomic analysis of outer membrane vesicles from the probiotic strain Escherichia coli Nissle 1917. Proteomics 14:222–229. 10.1002/pmic.201300328 [DOI] [PubMed] [Google Scholar]
58.Horstman AL, Kuehn MJ (2000) Enterotoxigenic Escherichia coli secretes active heat-labile enterotoxin via outer membrane vesicles. J Biol Chem 275:12489–12496. 10.1074/jbc.275.17.12489 [DOI] [PMC free article] [PubMed] [Google Scholar]
59.Sartorio MG, Valguarnera E, Hsu FF, Feldman MF (2022) Lipidomics analysis of outer membrane vesicles and elucidation of the inositol phosphoceramide biosynthetic pathway in Bacteroides thetaiotaomicron. Microbiol Spectr 10:e0063421. 10.1128/spectrum.00634-21 [DOI] [PMC free article] [PubMed] [Google Scholar]
60.Yang J, Wang T, Zheng Y, Wei Y, Tao J, Chen Z et al (2026) Outer membrane vesicles: Versatile nanocarriers for therapeutic delivery and immune modulation. Acta Pharm Sin B 16:1914–1942. 10.1016/j.apsb.2026.01.033 [DOI] [PMC free article] [PubMed] [Google Scholar]
61.Zhang M, Zhao X, Tang M, Zou W (2026) Bacterial outer membrane vesicles: Research advances from biogenesis mechanisms to engineered applications. Membr (Basel) 16:208. 10.3390/membranes16060208 [DOI] [PMC free article] [PubMed] [Google Scholar]
62.Abolhasani FS, Vaghefinanekaran N, Yarahmadi A, Akrami S, Mirmahdavi S, Yousefi MH et al (2025) Outer membrane vesicles in gram-negative bacteria and its correlation with pathogenesis. Front Immunol 16:1541636. 10.3389/fimmu.2025.1541636 [DOI] [PMC free article] [PubMed] [Google Scholar]
63.Bitto NJ, Chapman R, Pidot S, Costin A, Lo C, Choi J et al (2017) Bacterial membrane vesicles transport their DNA cargo into host cells. Sci Rep 7:7072. 10.1038/s41598-017-07288-4 [DOI] [PMC free article] [PubMed] [Google Scholar]
64.Dell’Annunziata F, Folliero V, Giugliano R, De Filippis A, Santarcangelo C, Izzo V et al (2021) Gene transfer potential of outer membrane vesicles of Gram-negative bacteria. Int J Mol Sci 22:5985. 10.3390/ijms22115985 [DOI] [PMC free article] [PubMed] [Google Scholar]
65.Mosca K, Turbant F, Achouak W, Wien F, Arluison V (2026) RNAs associated with bacterial outer membrane vesicles: Structural insights into surface composition. J Extracell Vesicles 15:e70306. 10.1002/jev2.70306 [DOI] [PMC free article] [PubMed] [Google Scholar]
66.Marinacci B, Krzyżek P, Pellegrini B, Turacchio G, Grande R (2023) Latest update on outer membrane vesicles and their role in horizontal gene transfer: A mini-review. Membr (Basel) 13:860. 10.3390/membranes13110860 [DOI] [PMC free article] [PubMed] [Google Scholar]
67.Liang J, Li M, Xu J, Chen S (2026) Bacterial extracellular vesicles in the strategic interplay between pathogens and hosts. Microorganisms 14:1362. 10.3390/microorganisms14061362 [DOI] [PMC free article] [PubMed] [Google Scholar]
68.Hu R, Lin H, Li J, Zhao Y, Wang M, Sun X et al (2020) Probiotic Escherichia coli Nissle 1917-derived outer membrane vesicles enhance immunomodulation and antimicrobial activity in RAW264.7 macrophages. BMC Microbiol 20:268. 10.1186/s12866-020-01953-x [DOI] [PMC free article] [PubMed] [Google Scholar]
69.Li N, Xin H, Deng K (2025) Separation and characterization of heterogeneity among various sizes of outer membrane vesicles derived from the probiotic Escherichia coli Nissle 1917. Membranes 15:141. 10.3390/membranes15050141 [DOI] [PMC free article] [PubMed] [Google Scholar]
70.Han L, Liu X-W, Li W-X, Zang T, Li Z-P, Deng Y et al (2024) Innovative probiotic Escherichia coli Nissle 1917-derived outer membrane vesicles coupled microspheres demonstrate anti-inflammatory and barrier-enhancing properties in colitis. Chem Eng J 487:150711. 10.1016/j.cej.2024.150711 [DOI] [Google Scholar]
71.Cañas MA, Giménez R, Fábrega MJ, Toloza L, Baldomà L, Badia J (2016) Outer membrane vesicles from the probiotic Escherichia coli Nissle 1917 and the commensal ECOR12 enter intestinal epithelial cells via clathrin-dependent endocytosis and elicit differential effects on DNA damage. PLoS ONE 11:e0160374. 10.1371/journal.pone.0160374 [DOI] [PMC free article] [PubMed] [Google Scholar]
72.Yu L, Zhang H, Shi C, Zhou Q, Li J, Lu B et al (2026) Targeted intestinal barrier repair via probiotic-derived engineered outer membrane vesicles: A 3A1M strategy with antioxidant, anti-inflammatory, anti-ferroptotic, and microbiome modulation effects. Acta Pharm Sin B 16:2527–2552. 10.1016/j.apsb.2026.01.031 [DOI] [PMC free article] [PubMed] [Google Scholar]
73.Cañas M-A, Fábrega M-J, Giménez R, Badia J, Baldomà L (2018) Outer membrane vesicles from probiotic and commensal Escherichia coli activate NOD1-mediated immune responses in intestinal epithelial cells. Front Microbiol 9:498. 10.3389/fmicb.2018.00498 [DOI] [PMC free article] [PubMed] [Google Scholar]
74.Shi J, Ma D, Gao S, Long F, Wang X, Pu X et al (2023) Probiotic Escherichia coli Nissle 1917-derived outer membrane vesicles modulate the intestinal microbiome and host gut-liver metabolome in obese and diabetic mice. Front Microbiol 14:1219763. 10.3389/fmicb.2023.1219763 [DOI] [PMC free article] [PubMed] [Google Scholar]
75.Xie J, Dai X-j, Li Q, Zhang W, Nie X, Ji H et al (2026) Escherichia coli Nissle 1917 alleviates Alzheimer’s disease in mice through OmpA-containing outer membrane vesicles. Cell Rep Med 7:102781. 10.1016/j.xcrm.2026.102781 [DOI] [PMC free article] [PubMed] [Google Scholar]
76.Rodas-Pazmiño K, Pazmiño-Gómez B, Cagua-Montaño L, Valle-Asan S, Acosta-Farías M, Fajardo-Aguilar PJ et al (2026) Strain-specific microRNA reprogramming of human dendritic cells by probiotic and commensal Escherichia coli outer membrane vesicles. Microorganisms 14:323. 10.3390/microorganisms14020323 [DOI] [PMC free article] [PubMed] [Google Scholar]
77.Wang L, Tang L, Feng Y, Zhao S, Han M, Zhang C et al (2020) A purified membrane protein from Akkermansia muciniphila or the pasteurised bacterium blunts colitis associated tumourigenesis by modulation of CD8(+) T cells in mice. Gut 69:1988–1997. 10.1136/gutjnl-2019-320105 [DOI] [PMC free article] [PubMed] [Google Scholar]
78.Mo C, Lou X, Xue J, Shi Z, Zhao Y, Wang F et al (2024) The influence of Akkermansia muciniphila on intestinal barrier function. Gut Pathog 16:41. 10.1186/s13099-024-00635-7 [DOI] [PMC free article] [PubMed] [Google Scholar]
79.Ottman N, Reunanen J, Meijerink M, Pietilä TE, Kainulainen V, Klievink J et al (2017) Pili-like proteins of Akkermansia muciniphila modulate host immune responses and gut barrier function. PLoS ONE 12:e0173004. 10.1371/journal.pone.0173004 [DOI] [PMC free article] [PubMed] [Google Scholar]
80.Bae M, Cassilly CD, Liu X, Park SM, Tusi BK, Chen X et al (2022) Akkermansia muciniphila phospholipid induces homeostatic immune responses. Nature 608:168–173. 10.1038/s41586-022-04985-7 [DOI] [PMC free article] [PubMed] [Google Scholar]
81.Toyofuku M, Schild S, Kaparakis-Liaskos M, Eberl L (2023) Composition and functions of bacterial membrane vesicles. Nat Rev Microbiol 21:415–430. 10.1038/s41579-023-00875-5 [DOI] [PubMed] [Google Scholar]
82.Zheng T, Hao H, Liu Q, Li J, Yao Y, Liu Y et al (2023) Effect of extracellular vesicles derived from Akkermansia muciniphila on intestinal barrier in colitis mice. Nutrients 15:4722. 10.3390/nu15224722 [DOI] [PMC free article] [PubMed] [Google Scholar]
83.Zakharzhevskaya NB, Vanyushkina AA, Altukhov IA, Shavarda AL, Butenko IO, Rakitina DV et al (2017) Outer membrane vesicles secreted by pathogenic and nonpathogenic Bacteroides fragilis represent different metabolic activities. Sci Rep 7:5008. 10.1038/s41598-017-05264-6 [DOI] [PMC free article] [PubMed] [Google Scholar]
84.Elhenawy W, Debelyy MO, Feldman MF (2014) Preferential packing of acidic glycosidases and proteases into Bacteroides outer membrane vesicles. mBio 5:e00909–e00914. 10.1128/mbio.00909-14 [DOI] [PMC free article] [PubMed] [Google Scholar]
85.Shipman JA, Berleman JE, Salyers AA (2000) Characterization of four outer membrane proteins involved in binding starch to the cell surface of Bacteroides thetaiotaomicron. J Bacteriol 182:5365–5372. 10.1128/jb.182.19.5365-5372.2000 [DOI] [PMC free article] [PubMed] [Google Scholar]
86.Du A, Chen J, Xie Z, Yang Y, Chen C-B, Lu C-W et al (2025) Multi-omics analysis of outer membrane vesicles from P. goldsteinii in a psoriasis mouse model. Life Sci 377:123776. 10.1016/j.lfs.2025.123776 [DOI] [PubMed] [Google Scholar]
87.Su D, Li M, Xie Y, Xu Z, Lv G, Jiu Y et al (2025) Gut commensal bacteria Parabacteroides goldsteinii-derived outer membrane vesicles suppress skin inflammation in psoriasis. J Control Release 377:127–145. 10.1016/j.jconrel.2024.11.014 [DOI] [PubMed] [Google Scholar]
88.Zhu W, Han L, He L, Wei S, Li J, Xin L et al (2026) Parabacteroides goldsteinii-derived outer membrane vesicles alleviate acute lung injury via modulation of bile acid metabolism. J Nanobiotechnol 24:390. 10.1186/s12951-026-04288-3 [DOI] [PMC free article] [PubMed] [Google Scholar]
89.Hiippala K, Barreto G, Burrello C, Diaz-Basabe A, Suutarinen M, Kainulainen V et al (2020) Novel Odoribacter splanchnicus strain and its outer membrane vesicles exert immunoregulatory effects in vitro. Front Microbiol 11:575455. 10.3389/fmicb.2020.575455 [DOI] [PMC free article] [PubMed] [Google Scholar]
90.Zhuang J, Zhuang Z, Chen B, Yang Y, Chen H, Guan G (2025) Odoribacter splanchnicus-derived extracellular vesicles alleviate inflammatory bowel disease by modulating gastrointestinal inflammation and intestinal barrier function via the NLRP3 inflammasome suppression. Mol Med 31:56. 10.1186/s10020-025-01063-2 [DOI] [PMC free article] [PubMed] [Google Scholar]
91.Verbunt J, Stassen FRM (2025) Probiotic membrane vesicles: emerging tools for disease treatment. Microbiome Res Rep 4:25. 10.20517/mrr.2025.20 [DOI] [PMC free article] [PubMed] [Google Scholar]
92.Arce-Rodríguez A, De la Peña Noya J, Borrero-de Acuña JM (2026) Bacterial extracellular vesicles in biomedical research and clinical translation. Microb Biotechnol 19:e70350. 10.1111/1751-7915.70350 [DOI] [PMC free article] [PubMed] [Google Scholar]
93.Jeong D, Kim MJ, Park Y, Chung J, Kweon HS, Kang NG et al (2022) Visualizing extracellular vesicle biogenesis in gram-positive bacteria using super-resolution microscopy. BMC Biol 20:270. 10.1186/s12915-022-01472-3 [DOI] [PMC free article] [PubMed] [Google Scholar]
94.Toyofuku M, Nomura N, Eberl L (2019) Types and origins of bacterial membrane vesicles. Nat Rev Microbiol 17:13–24. 10.1038/s41579-018-0112-2 [DOI] [PubMed] [Google Scholar]
95.Lee S, Jo E, Lee E-Y, Choi D (2026) Proteomics of extracellular vesicles derived from Lactobacillus: Implications in gut immune homeostasis and therapeutic potential. Biomed Pharmacother 194:118955. 10.1016/j.biopha.2025.118955 [DOI] [PubMed] [Google Scholar]
96.Lee B-H, Chen Y-Z, Shen T-L, Pan T-M, Hsu W-H (2023) Proteomic characterization of extracellular vesicles derived from lactic acid bacteria. Food Chem 427:136685. 10.1016/j.foodchem.2023.136685 [DOI] [PubMed] [Google Scholar]
97.Bajic SS, Cañas MA, Tolinacki M, Badia J, Sánchez B, Golic N et al (2020) Proteomic profile of extracellular vesicles released by Lactiplantibacillus plantarum BGAN8 and their internalization by non-polarized HT29 cell line. Sci Rep 10:21829. 10.1038/s41598-020-78920-z [DOI] [PMC free article] [PubMed] [Google Scholar]
98.Hu R, Lin H, Wang M, Zhao Y, Liu H, Min Y et al (2021) Lactobacillus reuteri-derived extracellular vesicles maintain intestinal immune homeostasis against lipopolysaccharide-induced inflammatory responses in broilers. J Anim Sci Biotechnol 12:25. 10.1186/s40104-020-00532-4 [DOI] [PMC free article] [PubMed] [Google Scholar]
99.Champagne-Jorgensen K, Mian MF, McVey Neufeld KA, Stanisz AM, Bienenstock J (2021) Membrane vesicles of Lacticaseibacillus rhamnosus JB-1 contain immunomodulatory lipoteichoic acid and are endocytosed by intestinal epithelial cells. Sci Rep 11:13756. 10.1038/s41598-021-93311-8 [DOI] [PMC free article] [PubMed] [Google Scholar]
100.Kim W, Lee EJ, Bae IH, Myoung K, Kim ST, Park PJ et al (2020) Lactobacillus plantarum-derived extracellular vesicles induce anti-inflammatory M2 macrophage polarization in vitro. J Extracell Vesicles 9:1793514. 10.1080/20013078.2020.1793514 [DOI] [PMC free article] [PubMed] [Google Scholar]
101.Yamasaki-Yashiki S, Kawashima F, Saika A, Hosomi R, Kunisawa J, Katakura Y (2024) RNA-based anti-inflammatory effects of membrane vesicles derived from Lactiplantibacillus plantarum. Foods 13:967. 10.3390/foods13060967 [DOI] [PMC free article] [PubMed] [Google Scholar]
102.Yoshida A, Yamada K, Yamazaki Y, Sashihara T, Ikegami S, Shimizu M et al (2011) Lactobacillus gasseri OLL2809 and its RNA suppress proliferation of CD4(+) T cells through a MyD88-dependent signalling pathway. Immunology 133:442–451. 10.1111/j.1365-2567.2011.03455.x [DOI] [PMC free article] [PubMed] [Google Scholar]
103.Chen Y, Huang X, Liu A, Fan S, Liu S, Li Z et al (2024) Lactobacillus Reuteri vesicles regulate mitochondrial function of macrophages to promote mucosal and cutaneous wound healing. Adv Sci (Weinh) 11:e2309725. 10.1002/advs.202309725 [DOI] [PMC free article] [PubMed] [Google Scholar]
104.Mandelbaum N, Zhang L, Carasso S, Ziv T, Lifshiz-Simon S, Davidovich I et al (2023) Extracellular vesicles of the Gram-positive gut symbiont Bifidobacterium longum induce immune-modulatory, anti-inflammatory effects. NPJ Biofilms Microbiomes 9:30. 10.1038/s41522-023-00400-9 [DOI] [PMC free article] [PubMed] [Google Scholar]
105.Kim JH, Jeun EJ, Hong CP, Kim SH, Jang MS, Lee EJ et al (2016) Extracellular vesicle-derived protein from Bifidobacterium longum alleviates food allergy through mast cell suppression. J Allergy Clin Immunol 137:507–516e8. 10.1016/j.jaci.2015.08.016 [DOI] [PubMed] [Google Scholar]
106.Nishiyama K, Takaki T, Sugiyama M, Fukuda I, Aiso M, Mukai T et al (2020) Extracellular vesicles produced by Bifidobacterium longum export mucin-binding proteins. Appl Environ Microbiol 86:e01464–e01420. 10.1128/aem.01464-20 [DOI] [PMC free article] [PubMed] [Google Scholar]
107.Kim S, Han SY, Lee J, Kim NR, Lee BR, Kim H et al (2022) Bifidobacterium longum and galactooligosaccharide improve skin barrier dysfunction and atopic dermatitis-like skin. Allergy Asthma Immunol Res 14:549–564. 10.4168/aair.2022.14.5.549 [DOI] [PMC free article] [PubMed] [Google Scholar]
108.Vuong N, Alomia M, Byne A, Gade P, Philipson TR, Alhammad RI et al (2025) Lactobacillus rhamnosus-derived extracellular vesicles influence calcium deposition in a model of breast cancer intraductal calcium stress. iScience 28:112538. 10.1016/j.isci.2025.112538 [DOI] [PMC free article] [PubMed] [Google Scholar]
109.Wang J, Li X, Zhao X, Yuan S, Dou H, Cheng T et al (2024) Lactobacillus rhamnosus GG-derived extracellular vesicles promote wound healing via miR-21-5p-mediated re-epithelization and angiogenesis. J Nanobiotechnol 22:644. 10.1186/s12951-024-02893-8 [DOI] [PMC free article] [PubMed] [Google Scholar]
110.Artuyants A, Hong J, Dauros-Singorenko P, Phillips A, Simoes-Barbosa A (2024) Lactobacillus gasseri and Gardnerella vaginalis produce extracellular vesicles that contribute to the function of the vaginal microbiome and modulate host–Trichomonas vaginalis interactions. Mol Microbiol 122:357–371. 10.1111/mmi.15130 [DOI] [PubMed] [Google Scholar]
111.Shiraishi T, Yokota S, Sato Y, Ito T, Fukiya S, Yamamoto S et al (2018) Lipoteichoic acids are embedded in cell walls during logarithmic phase, but exposed on membrane vesicles in Lactobacillus gasseri JCM 1131(T). Benef Microbes 9:653–662. 10.3920/bm2017.0124 [DOI] [PubMed] [Google Scholar]
112.Lee DH, Park HK, Lee HR, Sohn H, Sim S, Park HJ et al (2022) Immunoregulatory effects of Lactococcus lactis-derived extracellular vesicles in allergic asthma. Clin Transl Allergy 12:e12138. 10.1002/clt2.12138 [DOI] [PMC free article] [PubMed] [Google Scholar]
113.Liu Y, Tempelaars MH, Boeren S, Alexeeva S, Smid EJ, Abee T (2022) Extracellular vesicle formation in Lactococcus lactis is stimulated by prophage-encoded holin-lysin system. Microb Biotechnol 15:1281–1295. 10.1111/1751-7915.13972 [DOI] [PMC free article] [PubMed] [Google Scholar]
114.Wang Y, Hu L, Gao B, Yang R, Huang Y, Li P et al (2025) Lactococcus lactis-derived extracellular vesicles: A novel nanodelivery system enhance the properties of nisin Z. Food Chem 489:144776. 10.1016/j.foodchem.2025.144776 [DOI] [PubMed] [Google Scholar]
115.Dean SN, Leary DH, Sullivan CJ, Oh E, Walper SA (2019) Isolation and characterization of Lactobacillus-derived membrane vesicles. Sci Rep 9:877. 10.1038/s41598-018-37120-6 [DOI] [PMC free article] [PubMed] [Google Scholar]
116.da Silva Barreira D, Laurent J, Lourenço J, Novion Ducassou J, Couté Y, Guzzo J et al (2023) Membrane vesicles released by Lacticaseibacillus casei BL23 inhibit the biofilm formation of Salmonella Enteritidis. Sci Rep 13:1163. 10.1038/s41598-023-27959-9 [DOI] [PMC free article] [PubMed] [Google Scholar]
117.Domínguez Rubio AP, Martínez JH, Martínez Casillas DC, Coluccio Leskow F, Piuri M, Pérez OE (2017) Lactobacillus casei BL23 produces microvesicles carrying proteins that have been associated with its probiotic effect. Front Microbiol 8:1783. 10.3389/fmicb.2017.01783 [DOI] [PMC free article] [PubMed] [Google Scholar]
118.Bäuerl C, Coll-Marqués JM, Tarazona-González C, Pérez-Martínez G (2020) Lactobacillus casei extracellular vesicles stimulate EGFR pathway likely due to the presence of proteins P40 and P75 bound to their surface. Sci Rep 10:19237. 10.1038/s41598-020-75930-9 [DOI] [PMC free article] [PubMed] [Google Scholar]
119.Kuhn T, Aljohmani A, Frank N, Zielke L, Mehanny M, Laschke MW et al (2024) A cell-free, biomimetic hydrogel based on probiotic membrane vesicles ameliorates wound healing. J Control Release 365:969–980. 10.1016/j.jconrel.2023.12.011 [DOI] [PubMed] [Google Scholar]
120.Lonngi Sosa CD, González Díaz FR, Álvarez HR, Vargas Ruiz A, Higuera Piedrahita RI, de la Cruz Cruz HA et al (2026) Membrane vesicles from Lactobacillus acidophilus promote superior cytokine modulation and antimicrobial signaling compared with their whole cells in RAW 264.7 macrophages. Int J Mol Sci 27:2764. 10.3390/ijms27062764 [DOI] [PMC free article] [PubMed] [Google Scholar]
121.Mazgaeen L, Gurung P (2020) Recent advances in lipopolysaccharide recognition systems. Int J Mol Sci 21:379. 10.3390/ijms21020379 [DOI] [PMC free article] [PubMed] [Google Scholar]
122.Schmid AM, Razim A, Wysmołek M, Kerekes D, Haunstetter M, Kohl P et al (2023) Extracellular vesicles of the probiotic bacteria E. coli O83 activate innate immunity and prevent allergy in mice. Cell Commun Signal 21:297. 10.1186/s12964-023-01329-4 [DOI] [PMC free article] [PubMed] [Google Scholar]
123.Zwicker C, Sarate P, Drinić M, Ambroz K, Korb E, Smole U et al (2018) Prophylactic and therapeutic inhibition of allergic airway inflammation by probiotic Escherichia coli O83. J Allergy Clin Immunol 142:1987–1990e7. 10.1016/j.jaci.2018.07.029 [DOI] [PubMed] [Google Scholar]
124.Tiku V, Tan M-W (2021) Host immunity and cellular responses to bacterial outer membrane vesicles. Trends Immunol 42:1024–1036. 10.1016/j.it.2021.09.006 [DOI] [PubMed] [Google Scholar]
125.Blanc L, Castanier R, Mishra AK, Ray A, Besra GS, Sutcliffe I et al (2013) Gram-positive bacterial lipoglycans based on a glycosylated diacylglycerol lipid anchor are microbe-associated molecular patterns recognized by TLR2. PLoS ONE 8:e81593. 10.1371/journal.pone.0081593 [DOI] [PMC free article] [PubMed] [Google Scholar]
126.Saito S, Lin S-H, Wu H-Y (2019) The unique immunoregulatory function of Staphylococcus aureus lipoteichoic acid in dendritic cells. J 2:326–339. 10.3390/j2030022 [DOI] [Google Scholar]
127.Furnari S, Ciantia R, Garozzo A, Furneri PM, Fuochi V (2025) Lactobacilli-derived microbe-associated molecular patterns (MAMPs) in host immune modulation. Biomolecules 15:1609. 10.3390/biom15111609 [DOI] [PMC free article] [PubMed] [Google Scholar]
128.Domínguez Rubio AP, D’Antoni CL, Piuri M, Pérez OE (2022) Probiotics, their extracellular vesicles and infectious diseases. Front Microbiol 13:864720. 10.3389/fmicb.2022.864720 [DOI] [PMC free article] [PubMed] [Google Scholar]
129.Yuan S, Jin M, Zhang Y, Zhang M, Yuan M, Ding X et al (2025) Extracellular vesicles in skin: Biological function and therapeutic potential. Int J Nanomed 20:11211–11233. 10.2147/ijn.s548986 [DOI] [PMC free article] [PubMed] [Google Scholar]
130.Donoso-Meneses D, Figueroa-Valdés AI, Khoury M, Alcayaga-Miranda F (2023) Oral administration as a potential alternative for the delivery of small extracellular vesicles. Pharmaceutics 15:716. 10.3390/pharmaceutics15030716 [DOI] [PMC free article] [PubMed] [Google Scholar]
131.Lee H, Lee Y-H, Hong D-K, Mo S-J, Jeon S, Park S-D et al (2024) Targeting inflammation and skin aging via the gut–skin axis: The role of Lactiplantibacillus plantarum HY7714-derived extracellular vesicles. Microorganisms 12:2466. 10.3390/microorganisms12122466 [DOI] [PMC free article] [PubMed] [Google Scholar]
132.Chen X, Li Q, Zhang W, Xu Y, Nie X, Wang X et al (2026) Akkermansia muciniphila-derived extracellular vesicles alleviate colitis-related cognitive impairment via tryptophan metabolic reprogramming of the gut–brain axis. Gut Microbes 18:2611546. 10.1080/19490976.2025.2611546 [DOI] [PMC free article] [PubMed] [Google Scholar]
133.Nakatsuji T, Chen TH, Narala S, Chun KA, Two AM, Yun T et al (2017) Antimicrobials from human skin commensal bacteria protect against Staphylococcus aureus and are deficient in atopic dermatitis. Sci Transl Med 9:eaah4680. 10.1126/scitranslmed.aah4680 [DOI] [PMC free article] [PubMed] [Google Scholar]
134.López P, González-Rodríguez I, Sánchez B, Gueimonde M, Margolles A, Suárez A (2012) Treg-inducing membrane vesicles from Bifidobacterium bifidum LMG13195 as potential adjuvants in immunotherapy. Vaccine 30:825–829. 10.1016/j.vaccine.2011.11.115 [DOI] [PubMed] [Google Scholar]
135.Li L, Si W, Jin Z, Jin W (2026) Research progress on construction technology of 3D human skin models and its application prospects in dermatology. Int J Mol Sci 27:1808. 10.3390/ijms27041808 [DOI] [PMC free article] [PubMed] [Google Scholar]
136.Flint E, Ahmad N, Rowland K, Hildebolt C, Raskin D (2024) Topical probiotics reduce atopic dermatitis severity: A systematic review and meta-analysis of double-blind, randomized, placebo-controlled trials. Cureus 16:e70001. 10.7759/cureus.70001 [DOI] [PMC free article] [PubMed] [Google Scholar]
137.Flores Rodríguez JC, Morán Ortega MJ, Jimenez Ordoñez AC, Rojas López AK, Leyva MC, Romero HV (2026) Postbiotics in dermatology: A literature review of emerging topical therapies for acne, rosacea, and eczema. Cureus 18:e105440. 10.7759/cureus.105440 [DOI] [PMC free article] [PubMed] [Google Scholar]
138.Salo V, Remitz A, Lauerma A, Salava A (2026) Topical probiotic Lactobacillus lactis treatment in atopic dermatitis: a placebo-controlled pilot study on tolerability and efficacy. Front Med (Lausanne) 13:1694229. 10.3389/fmed.2026.1694229 [DOI] [PMC free article] [PubMed] [Google Scholar]
139.Tang W-Z, Liao C-Y, Xu H-Y, Huang W-T, Wu Z-X, Chen T-Y et al (2026) Extracellular vesicles in atopic dermatitis: unraveling pathogenic mediators and engineering therapeutic vectors. Front Immunol 17:1846799. 10.3389/fimmu.2026.1846799 [DOI] [PMC free article] [PubMed] [Google Scholar]
140.Cannizzaro F, Gallo A, La Scala S, Gallo G, Faddetta T (2026) Bacterial extracellular vesicles in biotechnology: Current challenges and strategies for production enhancement. Fermentation 12:86. 10.3390/fermentation12020086 [DOI] [Google Scholar]
141.Li B, Wu W, Xu W, Qian H, Ji C (2025) Advances of extracellular vesicles isolation and detection frontier technology: from heterogeneity analysis to clinical application. J Nanobiotechnol 23:678. 10.1186/s12951-025-03768-2 [DOI] [PMC free article] [PubMed] [Google Scholar]
142.Choi D, Lee EY (2025) Standardizing bacterial extracellular vesicle purification: A call for consensus. J Microbiol Biotechnol 35:e2506011. 10.4014/jmb.2506.06011 [DOI] [PMC free article] [PubMed] [Google Scholar]
143.Welsh JA, Goberdhan DCI, O’Driscoll L, Buzas EI, Blenkiron C, Bussolati B et al (2024) Minimal information for studies of extracellular vesicles (MISEV2023): From basic to advanced approaches. J Extracell Vesicles 13:e12404. 10.1002/jev2.12404 [DOI] [PMC free article] [PubMed] [Google Scholar]
144.Rudge C, McClelland R, Chrzanowski W, Davis RL (2026) The regulatory landscape for extracellular vesicle therapies: Australian context and future directions. Extracell Vesicles Circ Nucl Acids 7:292–301. 10.20517/evcna.2025.129 [DOI] [PMC free article] [PubMed] [Google Scholar]
145.Xu G, Jin J, Fu Z, Wang G, Lei X, Xu J et al (2025) Extracellular vesicle-based drug overview: research landscape, quality control and nonclinical evaluation strategies. Signal Transduct Target Ther 10:255. 10.1038/s41392-025-02312-w [DOI] [PMC free article] [PubMed] [Google Scholar]
146.Wen T, Gu Y, Liu G (2026) Extracellular vesicles in wearable delivery systems for cosmeceutical applications. Adv Healthc Mater 26:e71379. 10.1002/adhm.71379 [DOI] [PMC free article] [PubMed] [Google Scholar]
147.Albayati N, Talluri SR, Dholaria N, Michniak-Kohn B (2025) AI-Driven innovation in skin kinetics for transdermal drug delivery: Overcoming barriers and enhancing precision. Pharmaceutics 17:188. 10.3390/pharmaceutics17020188 [DOI] [PMC free article] [PubMed] [Google Scholar]
148.Kaisanlahti A, Turunen J, Hekkala J, Mishra S, Karikka S, Amatya SB et al (2025) Gut microbiota-derived extracellular vesicles form a distinct entity from gut microbiota. mSystems 10:e0031125. 10.1128/msystems.00311-25 [DOI] [PMC free article] [PubMed] [Google Scholar]
149.Tang Z-Y-E, Yang X-l, Zhou Y-W, Zou H, Li Y, Wu H et al (2026) Advances in microbial extracellular vesicles: synthetic biology platforms and medical applications. Extracell Vesicles Circ Nucl Acids 7:945–972. 10.20517/evcna.2025.103 [DOI] [PMC free article] [PubMed] [Google Scholar]

Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Data Availability Statement

No datasets were generated or analysed during the current study.

[CR1] 1.Yue C, Zhou H, Wang X, Yu J, Hu Y, Zhou P et al (2024) Atopic dermatitis: pathogenesis and therapeutic intervention. MedComm 5:e70029. 10.1002/mco2.70029 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR2] 2.Hansen-Sackey EB, Hartono S (2025) Atopic Dermatitis: Pathophysiology and Emerging Treatments. Allergies 5:40. 10.3390/allergies5040040 [DOI] [Google Scholar]

[CR3] 3.Moreiras-Arias N, Nieto-Fontarigo JJ, Salgado FJ, González-Vilas D, Paredes-Suárez C, Combo-García E et al (2026) Novel therapeutic strategies for atopic dermatitis: biomarker modulation and clinical implications. A systematic review. Clin Rev Allergy Immunol 69:4. 10.1007/s12016-025-09129-z [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR4] 4.Kolb L, Villarreal LA, Sathe NC (2026) Atopic dermatitis. In: StatPearls [Internet] StatPearls Publishing Treasure Island (FL). https://www.ncbi.nlm.nih.gov/books/NBK448071/. Accessed 27 May 2026 [PubMed]

[CR5] 5.Tian J, Zhang D, Yang Y, Huang Y, Wang L, Yao X et al (2023) Global epidemiology of atopic dermatitis: a comprehensive systematic analysis and modelling study. Br J Dermatol 190:55–61. 10.1093/bjd/ljad339 [DOI] [PubMed] [Google Scholar]

[CR6] 6.Lyons JJ, Milner JD, Stone KD (2015) Atopic dermatitis in children: clinical features, pathophysiology, and treatment. Immunol Allergy Clin North Am 35:161–183. 10.1016/j.iac.2014.09.008 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR7] 7.Bai R, Zheng Y, Dai X (2025) Atopic dermatitis: diagnosis, molecular pathogenesis, and therapeutics. Mol Biomed 6:71. 10.1186/s43556-025-00313-3 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR8] 8.Langan SM, Irvine AD, Weidinger S (2020) Atopic dermatitis. Lancet 396:345–360. 10.1016/S0140-6736(20)31286-1 [DOI] [PubMed] [Google Scholar]

[CR9] 9.Borici S (2025) Atopic dermatitis and food allergy in children. Acta Paediatr 114:3096–3107. 10.1111/apa.70283 [DOI] [PubMed] [Google Scholar]

[CR10] 10.Baidya A, Mabalirajan U (2025) Pathogenesis and management of atopic dermatitis: insights into epidermal barrier dysfunction and immune mechanisms. Explor Asthma Allergy 3:100973. 10.37349/eaa.2025.100973 [DOI] [Google Scholar]

[CR11] 11.Zhong L, Zhou X, Su J, Zhang Y, Zhang D, Wan H (2026) Microbiome dysbiosis and therapeutic restoration in atopic dermatitis. Front Cell Infect Microbiol 16:1693905. 10.3389/fcimb.2026.1693905 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR12] 12.Micu AE, Popescu IA, Halip IA, Mocanu M, Vâță D, Hulubencu AL et al (2026) From gut dysbiosis to skin inflammation in atopic dermatitis: Probiotics and the gut–skin axis—clinical outcomes and microbiome implications. Int J Mol Sci 27:365. 10.3390/ijms27010365 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR13] 13.Zhang Z, Wang R, Li M, Lu M (2025) Current insights and trends in atopic dermatitis and microbiota interactions: a systematic review and bibliometric analysis. Front Microbiol 16:1613315. 10.3389/fmicb.2025.1613315 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR14] 14.Lee SY, Lee E, Park YM, Hong SJ (2018) Microbiome in the gut-skin axis in atopic dermatitis. Allergy Asthma Immunol Res 10:354–362. 10.4168/aair.2018.10.4.354 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR15] 15.Zhao Y, Yu C, Zhang J, Yao Q, Zhu X, Zhou X (2025) The gut–skin axis: Emerging insights in understanding and treating skin diseases through gut microbiome modulation (Review). Int J Mol Med 56:210. 10.3892/ijmm.2025.5651 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR16] 16.Mazziotta C, Tognon M, Martini F, Torreggiani E, Rotondo JC (2023) Probiotics mechanism of action on immune cells and beneficial effects on human health. Cells 12:184. 10.3390/cells12010184 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR17] 17.Anania C, Brindisi G, Martinelli I, Bonucci E, D’Orsi M, Ialongo S et al (2022) Probiotics function in preventing atopic dermatitis in children. Int J Mol Sci 23:5409. 10.3390/ijms23105409 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR18] 18.Lim A, Baek J, Seo M, Kim K-y, Kim S, Kwak W et al (2025) Atopic dermatitis-alleviating effects of lactiplantibacillus plantarum LRCC5195 paraprobiotics through microbiome modulation and safety assessment via genomic characterization and in vitro analysis. Sci Rep 15:30545. 10.1038/s41598-025-16102-5 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR19] 19.Sung M, Sim S, Lim A, Moon JS, Jeon J, Heo K et al (2025) BCL-2 multi-strain probiotics for immunomodulation in vitro and in vivo alleviation of atopic dermatitis. Microorganisms 13:1950. 10.3390/microorganisms13081950 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR20] 20.Kang H-J, Kim G-C, Lee C-G, Park S, Sharma G, Verma R et al (2021) Probiotics-derived metabolite ameliorates skin allergy by promoting differentiation of FOXP₃⁺regulatory T cells. J Allergy Clin Immunol 147:1517–1521. 10.1016/j.jaci.2020.11.040 [DOI] [PubMed] [Google Scholar]

[CR21] 21.Zhong L, Su J, Zhou X, Wan H (2025) Probiotics supplements for the prevention of atopic dermatitis in children: an umbrella review. Front Nutr 12:1587348. 10.3389/fnut.2025.1587348 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR22] 22.Xue X, Yang X, Shi X, Deng Z (2023) Efficacy of probiotics in pediatric atopic dermatitis: A systematic review and meta-analysis. Clin Transl Allergy 13:e12283. 10.1002/clt2.12283 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR23] 23.Liscano Y, Muñoz Morales D, Suarez Daza F, Vidal Cañas S, Martinez Guevara D, Artunduaga Cañas E (2025) Microbial interventions for inflammatory skin diseases: A systematic review and meta-analysis of atopic dermatitis and psoriasis. Microorganisms 13:2416 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR24] 24.Mirali S, Drucker AM (2025) New topical and systemic treatments for atopic dermatitis. Clin Exp Allergy 55:1057–1069. 10.1111/cea.70136 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR25] 25.Biliński K, Rakoczy K, Karwowska A, Cichy O, Wojno A, Wojno A et al (2025) Anti-inflammatory therapies for atopic dermatitis: A new era in targeted treatment. J Clin Med 14:5053. 10.3390/jcm14145053 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR26] 26.Jeskey J, Kurien C, Blunk H, Sehmi K, Areti S, Nguyen D et al (2024) Atopic dermatitis: A review of diagnosis and treatment. J Pediatr Pharmacol Ther 29:587–603. 10.5863/1551-6776-29.6.587 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR27] 27.Ali S, Ion A, Orzan OA, Bălăceanu-Gurău B (2024) Emerging treatments and new vehicle formulations for atopic dermatitis. Pharmaceutics 16:1425. 10.3390/pharmaceutics16111425 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR28] 28.Elahi N, Astaneh ME, Ai J, Rizwan M (2024) Atopic dermatitis treatment: A comprehensive review of conventional and novel bioengineered approaches. Int J Biol Macromol 282:137083. 10.1016/j.ijbiomac.2024.137083 [DOI] [PubMed] [Google Scholar]

[CR29] 29.Zaborowski MP, Balaj L, Breakefield XO, Lai CP (2015) Extracellular vesicles: Composition, biological relevance, and methods of study. Bioscience 65:783–797. 10.1093/biosci/biv084 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR30] 30.Repici A, Piraino G, Wolfe V, Kaplan J, Nakamura T, Zingarelli B (2025) Role of extracellular vesicles as mediators of cell communication and novel biomarkers in sepsis. J Clin Med 14:6649. 10.3390/jcm14186649 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR31] 31.Premchandani T, Umekar M, Tatode A, Taksande J, Khan R, Faizan M et al (2025) Extracellular vesicles in host–pathogen interactions: Roles of exosomes and bacterial outer membrane vesicles in immunity and microbial communication. Bacteria 4:63. 10.3390/bacteria4040063 [DOI] [Google Scholar]

[CR32] 32.Yu Y-j, Wang X-h, Fan G-C (2018) Versatile effects of bacterium-released membrane vesicles on mammalian cells and infectious/inflammatory diseases. Acta Pharmacol Sin 39:514–533. 10.1038/aps.2017.82 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR33] 33.Kim JH, Lee J, Park J, Gho YS (2015) Gram-negative and Gram-positive bacterial extracellular vesicles. Semin Cell Dev Biol 40:97–104. 10.1016/j.semcdb.2015.02.006 [DOI] [PubMed] [Google Scholar]

[CR34] 34.Melo-Marques I, Cardoso SM, Empadinhas N (2024) Bacterial extracellular vesicles at the interface of gut microbiota and immunity. Gut Microbes 16:2396494. 10.1080/19490976.2024.2396494 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR35] 35.El-Saadony MT, Saad AM, Sitohy M, Alkafaas SS, Dladla M, Ghosh S et al (2026) Probiotics and human health: biological activities, nutritional aspects, immunomodulatory properties, applications, and future perspectives - a comprehensive review. Front Immunol 16:1713426. 10.3389/fimmu.2025.1713426 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR36] 36.Ma T, Shen X, Shi X, Sakandar HA, Quan K, Li Y et al (2023) Targeting gut microbiota and metabolism as the major probiotic mechanism - An evidence-based review. Trends Food Sci Technol 138:178–198. 10.1016/j.tifs.2023.06.013 [DOI] [Google Scholar]

[CR37] 37.Kozakiewicz D, Razim A, Schabussova I, Górska S (2025) Immunomodulatory properties of probiotic extracellular vesicles- do they have the potential to fill the gap in therapeutic strategies for allergies? Cell Commun Signal 23:549. 10.1186/s12964-025-02532-1 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR38] 38.Suri K, D’Souza A, Huang D, Bhavsar A, Amiji M (2023) Bacterial extracellular vesicle applications in cancer immunotherapy. Bioact Mater 22:551–566. 10.1016/j.bioactmat.2022.10.024 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR39] 39.Kim MH, Choi SJ, Choi HI, Choi JP, Park HK, Kim EK et al (2018) Lactobacillus plantarum-derived extracellular vesicles protect atopic dermatitis induced by Staphylococcus aureus-derived extracellular vesicles. Allergy Asthma Immunol Res 10:516–532. 10.4168/aair.2018.10.5.516 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR40] 40.Baek J, Cho S, Lee G, Ki H, Kim SY, Choi GM et al (2025) Modulation of moisturizing and barrier related molecular markers by extracellular vesicles derived from Leuconostoc mesenteroides DB-21 isolated from camellia japonica flower. Curr Issues Mol Biol 47:1022. 10.3390/cimb47121022 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR41] 41.Choi H, Kwak M-J, Choi Y, Kang AN, Mun D, Eor JY et al (2025) Extracellular vesicles of Limosilactobacillus fermentum SLAM216 ameliorate skin symptoms of atopic dermatitis by regulating gut microbiome on serotonin metabolism. Gut Microbes 17:2474256. 10.1080/19490976.2025.2474256 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR42] 42.Cho Y-H, Kim J-W, Kim N, Kim H-S, Jang J-H, Bae J-T et al (2025) Lactobacillus brevis-derived exosomes enhance skin barrier integrity by upregulating key barrier-related proteins. Clin Cosmet Investig Dermatol 18:1151–1162. 10.2147/ccid.s512793 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR43] 43.Shi Y, Li S, Yang S (2025) Probiotic-derived extracellular vesicles: a novel weapon against viral infections. Biosci Microbiota Food Health 44:245–250. 10.12938/bmfh.2025-027 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR44] 44.Atoom AM, Hamed-Ahmed M, Al-Hasnaawei S, Malathi H, Maharana L, Nanda A et al (2026) Therapeutic potential and translational challenges of extracellular vesicles in neonatal medicine. Bioeng Transl Med 11:e70093. 10.1002/btm2.70093 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR45] 45.Primorac D, Brlek P, Bulić L, Hrvatin N, Škaro V, Projić P et al (2026) Therapeutic potential of extracellular vesicles: From biogenesis, isolation and molecular characterization to addressing translational gaps and regulatory barriers. Int J Mol Sci 27:1676. 10.3390/ijms27041676 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR46] 46.Wang Y, Duan C, Yu X (2025) Extracellular vesicles of Gram-negative and Gram-positive probiotics. J Microbiol 63:e2506005. 10.71150/jm.2506005 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR47] 47.Liu BD, Akbar R, Oliverio A, Thapa K, Wang X, Fan GC (2024) Bacterial extracellular vesicles in the regulation of inflammatory response and host-microbe interactions. Shock 61:175–188. 10.1097/shk.0000000000002252 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR48] 48.Kurata A, Uegaki K (2025) Recent advances in understanding the role of extracellular vesicles from probiotics in intestinal immunity signaling. Biochem Soc Trans 53:419–429. 10.1042/bst20240150 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR49] 49.Zhang Y, Liang F, Zhang D, Qi S, Liu Y (2023) Metabolites as extracellular vesicle cargo in health, cancer, pleural effusion, and cardiovascular diseases: An emerging field of study to diagnostic and therapeutic purposes. Biomed Pharmacother 157:114046. 10.1016/j.biopha.2022.114046 [DOI] [PubMed] [Google Scholar]

[CR50] 50.Avila-Calderón ED, Ruiz-Palma MdS, Aguilera-Arreola MG, Velázquez-Guadarrama N, Ruiz EA, Gomez-Lunar Z et al (2021) Outer membrane vesicles of Gram-negative bacteria: An outlook on biogenesis. Front Microbiol 12:557902. 10.3389/fmicb.2021.557902 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR51] 51.Zhang L, He Y, Wang G, Sun J, Chen Y, Wang Z (2026) Bacterial membrane vesicles: Biogenesis, functions, and emerging biotechnological applications. Microorganisms 14:689. 10.3390/microorganisms14030689 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR52] 52.Jan AT (2017) Outer Membrane Vesicles (OMVs) of Gram-negative bacteria: A perspective update. Front Microbiol 8:1053. 10.3389/fmicb.2017.01053 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR53] 53.Schwechheimer C, Kuehn MJ (2015) Outer-membrane vesicles from Gram-negative bacteria: biogenesis and functions. Nat Rev Microbiol 13:605–619. 10.1038/nrmicro3525 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR54] 54.Zhao X, Wei Y, Bu Y, Ren X, Dong Z (2025) Review on bacterial outer membrane vesicles: structure, vesicle formation, separation and biotechnological applications. Microb Cell Fact 24:27. 10.1186/s12934-025-02653-9 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR55] 55.Giordano NP, Cian MB, Dalebroux ZD (2020) Outer membrane lipid secretion and the innate immune response to Gram-negative bacteria. Infect Immun 88:e00920–e00919. 10.1128/iai.00920-19 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR56] 56.Ma D, Zhang Y, Zhang J, Shi J, Gao S, Long F et al (2025) Outer membrane vesicles derived from probiotic Escherichia coli Nissle 1917 promote metabolic remodeling and M1 polarization of RAW264.7 macrophages. Front Immunol 16:1501174. 10.3389/fimmu.2025.1501174 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR57] 57.Aguilera L, Toloza L, Giménez R, Odena A, Oliveira E, Aguilar J et al (2014) Proteomic analysis of outer membrane vesicles from the probiotic strain Escherichia coli Nissle 1917. Proteomics 14:222–229. 10.1002/pmic.201300328 [DOI] [PubMed] [Google Scholar]

[CR58] 58.Horstman AL, Kuehn MJ (2000) Enterotoxigenic Escherichia coli secretes active heat-labile enterotoxin via outer membrane vesicles. J Biol Chem 275:12489–12496. 10.1074/jbc.275.17.12489 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR59] 59.Sartorio MG, Valguarnera E, Hsu FF, Feldman MF (2022) Lipidomics analysis of outer membrane vesicles and elucidation of the inositol phosphoceramide biosynthetic pathway in Bacteroides thetaiotaomicron. Microbiol Spectr 10:e0063421. 10.1128/spectrum.00634-21 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR60] 60.Yang J, Wang T, Zheng Y, Wei Y, Tao J, Chen Z et al (2026) Outer membrane vesicles: Versatile nanocarriers for therapeutic delivery and immune modulation. Acta Pharm Sin B 16:1914–1942. 10.1016/j.apsb.2026.01.033 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR61] 61.Zhang M, Zhao X, Tang M, Zou W (2026) Bacterial outer membrane vesicles: Research advances from biogenesis mechanisms to engineered applications. Membr (Basel) 16:208. 10.3390/membranes16060208 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR62] 62.Abolhasani FS, Vaghefinanekaran N, Yarahmadi A, Akrami S, Mirmahdavi S, Yousefi MH et al (2025) Outer membrane vesicles in gram-negative bacteria and its correlation with pathogenesis. Front Immunol 16:1541636. 10.3389/fimmu.2025.1541636 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR63] 63.Bitto NJ, Chapman R, Pidot S, Costin A, Lo C, Choi J et al (2017) Bacterial membrane vesicles transport their DNA cargo into host cells. Sci Rep 7:7072. 10.1038/s41598-017-07288-4 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR64] 64.Dell’Annunziata F, Folliero V, Giugliano R, De Filippis A, Santarcangelo C, Izzo V et al (2021) Gene transfer potential of outer membrane vesicles of Gram-negative bacteria. Int J Mol Sci 22:5985. 10.3390/ijms22115985 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR65] 65.Mosca K, Turbant F, Achouak W, Wien F, Arluison V (2026) RNAs associated with bacterial outer membrane vesicles: Structural insights into surface composition. J Extracell Vesicles 15:e70306. 10.1002/jev2.70306 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR66] 66.Marinacci B, Krzyżek P, Pellegrini B, Turacchio G, Grande R (2023) Latest update on outer membrane vesicles and their role in horizontal gene transfer: A mini-review. Membr (Basel) 13:860. 10.3390/membranes13110860 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR67] 67.Liang J, Li M, Xu J, Chen S (2026) Bacterial extracellular vesicles in the strategic interplay between pathogens and hosts. Microorganisms 14:1362. 10.3390/microorganisms14061362 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR68] 68.Hu R, Lin H, Li J, Zhao Y, Wang M, Sun X et al (2020) Probiotic Escherichia coli Nissle 1917-derived outer membrane vesicles enhance immunomodulation and antimicrobial activity in RAW264.7 macrophages. BMC Microbiol 20:268. 10.1186/s12866-020-01953-x [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR69] 69.Li N, Xin H, Deng K (2025) Separation and characterization of heterogeneity among various sizes of outer membrane vesicles derived from the probiotic Escherichia coli Nissle 1917. Membranes 15:141. 10.3390/membranes15050141 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR70] 70.Han L, Liu X-W, Li W-X, Zang T, Li Z-P, Deng Y et al (2024) Innovative probiotic Escherichia coli Nissle 1917-derived outer membrane vesicles coupled microspheres demonstrate anti-inflammatory and barrier-enhancing properties in colitis. Chem Eng J 487:150711. 10.1016/j.cej.2024.150711 [DOI] [Google Scholar]

[CR71] 71.Cañas MA, Giménez R, Fábrega MJ, Toloza L, Baldomà L, Badia J (2016) Outer membrane vesicles from the probiotic Escherichia coli Nissle 1917 and the commensal ECOR12 enter intestinal epithelial cells via clathrin-dependent endocytosis and elicit differential effects on DNA damage. PLoS ONE 11:e0160374. 10.1371/journal.pone.0160374 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR72] 72.Yu L, Zhang H, Shi C, Zhou Q, Li J, Lu B et al (2026) Targeted intestinal barrier repair via probiotic-derived engineered outer membrane vesicles: A 3A1M strategy with antioxidant, anti-inflammatory, anti-ferroptotic, and microbiome modulation effects. Acta Pharm Sin B 16:2527–2552. 10.1016/j.apsb.2026.01.031 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR73] 73.Cañas M-A, Fábrega M-J, Giménez R, Badia J, Baldomà L (2018) Outer membrane vesicles from probiotic and commensal Escherichia coli activate NOD1-mediated immune responses in intestinal epithelial cells. Front Microbiol 9:498. 10.3389/fmicb.2018.00498 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR74] 74.Shi J, Ma D, Gao S, Long F, Wang X, Pu X et al (2023) Probiotic Escherichia coli Nissle 1917-derived outer membrane vesicles modulate the intestinal microbiome and host gut-liver metabolome in obese and diabetic mice. Front Microbiol 14:1219763. 10.3389/fmicb.2023.1219763 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR75] 75.Xie J, Dai X-j, Li Q, Zhang W, Nie X, Ji H et al (2026) Escherichia coli Nissle 1917 alleviates Alzheimer’s disease in mice through OmpA-containing outer membrane vesicles. Cell Rep Med 7:102781. 10.1016/j.xcrm.2026.102781 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR76] 76.Rodas-Pazmiño K, Pazmiño-Gómez B, Cagua-Montaño L, Valle-Asan S, Acosta-Farías M, Fajardo-Aguilar PJ et al (2026) Strain-specific microRNA reprogramming of human dendritic cells by probiotic and commensal Escherichia coli outer membrane vesicles. Microorganisms 14:323. 10.3390/microorganisms14020323 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR77] 77.Wang L, Tang L, Feng Y, Zhao S, Han M, Zhang C et al (2020) A purified membrane protein from Akkermansia muciniphila or the pasteurised bacterium blunts colitis associated tumourigenesis by modulation of CD8(+) T cells in mice. Gut 69:1988–1997. 10.1136/gutjnl-2019-320105 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR78] 78.Mo C, Lou X, Xue J, Shi Z, Zhao Y, Wang F et al (2024) The influence of Akkermansia muciniphila on intestinal barrier function. Gut Pathog 16:41. 10.1186/s13099-024-00635-7 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR79] 79.Ottman N, Reunanen J, Meijerink M, Pietilä TE, Kainulainen V, Klievink J et al (2017) Pili-like proteins of Akkermansia muciniphila modulate host immune responses and gut barrier function. PLoS ONE 12:e0173004. 10.1371/journal.pone.0173004 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR80] 80.Bae M, Cassilly CD, Liu X, Park SM, Tusi BK, Chen X et al (2022) Akkermansia muciniphila phospholipid induces homeostatic immune responses. Nature 608:168–173. 10.1038/s41586-022-04985-7 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR81] 81.Toyofuku M, Schild S, Kaparakis-Liaskos M, Eberl L (2023) Composition and functions of bacterial membrane vesicles. Nat Rev Microbiol 21:415–430. 10.1038/s41579-023-00875-5 [DOI] [PubMed] [Google Scholar]

[CR82] 82.Zheng T, Hao H, Liu Q, Li J, Yao Y, Liu Y et al (2023) Effect of extracellular vesicles derived from Akkermansia muciniphila on intestinal barrier in colitis mice. Nutrients 15:4722. 10.3390/nu15224722 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR83] 83.Zakharzhevskaya NB, Vanyushkina AA, Altukhov IA, Shavarda AL, Butenko IO, Rakitina DV et al (2017) Outer membrane vesicles secreted by pathogenic and nonpathogenic Bacteroides fragilis represent different metabolic activities. Sci Rep 7:5008. 10.1038/s41598-017-05264-6 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR84] 84.Elhenawy W, Debelyy MO, Feldman MF (2014) Preferential packing of acidic glycosidases and proteases into Bacteroides outer membrane vesicles. mBio 5:e00909–e00914. 10.1128/mbio.00909-14 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR85] 85.Shipman JA, Berleman JE, Salyers AA (2000) Characterization of four outer membrane proteins involved in binding starch to the cell surface of Bacteroides thetaiotaomicron. J Bacteriol 182:5365–5372. 10.1128/jb.182.19.5365-5372.2000 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR86] 86.Du A, Chen J, Xie Z, Yang Y, Chen C-B, Lu C-W et al (2025) Multi-omics analysis of outer membrane vesicles from P. goldsteinii in a psoriasis mouse model. Life Sci 377:123776. 10.1016/j.lfs.2025.123776 [DOI] [PubMed] [Google Scholar]

[CR87] 87.Su D, Li M, Xie Y, Xu Z, Lv G, Jiu Y et al (2025) Gut commensal bacteria Parabacteroides goldsteinii-derived outer membrane vesicles suppress skin inflammation in psoriasis. J Control Release 377:127–145. 10.1016/j.jconrel.2024.11.014 [DOI] [PubMed] [Google Scholar]

[CR88] 88.Zhu W, Han L, He L, Wei S, Li J, Xin L et al (2026) Parabacteroides goldsteinii-derived outer membrane vesicles alleviate acute lung injury via modulation of bile acid metabolism. J Nanobiotechnol 24:390. 10.1186/s12951-026-04288-3 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR89] 89.Hiippala K, Barreto G, Burrello C, Diaz-Basabe A, Suutarinen M, Kainulainen V et al (2020) Novel Odoribacter splanchnicus strain and its outer membrane vesicles exert immunoregulatory effects in vitro. Front Microbiol 11:575455. 10.3389/fmicb.2020.575455 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR90] 90.Zhuang J, Zhuang Z, Chen B, Yang Y, Chen H, Guan G (2025) Odoribacter splanchnicus-derived extracellular vesicles alleviate inflammatory bowel disease by modulating gastrointestinal inflammation and intestinal barrier function via the NLRP3 inflammasome suppression. Mol Med 31:56. 10.1186/s10020-025-01063-2 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR91] 91.Verbunt J, Stassen FRM (2025) Probiotic membrane vesicles: emerging tools for disease treatment. Microbiome Res Rep 4:25. 10.20517/mrr.2025.20 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR92] 92.Arce-Rodríguez A, De la Peña Noya J, Borrero-de Acuña JM (2026) Bacterial extracellular vesicles in biomedical research and clinical translation. Microb Biotechnol 19:e70350. 10.1111/1751-7915.70350 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR93] 93.Jeong D, Kim MJ, Park Y, Chung J, Kweon HS, Kang NG et al (2022) Visualizing extracellular vesicle biogenesis in gram-positive bacteria using super-resolution microscopy. BMC Biol 20:270. 10.1186/s12915-022-01472-3 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR94] 94.Toyofuku M, Nomura N, Eberl L (2019) Types and origins of bacterial membrane vesicles. Nat Rev Microbiol 17:13–24. 10.1038/s41579-018-0112-2 [DOI] [PubMed] [Google Scholar]

[CR95] 95.Lee S, Jo E, Lee E-Y, Choi D (2026) Proteomics of extracellular vesicles derived from Lactobacillus: Implications in gut immune homeostasis and therapeutic potential. Biomed Pharmacother 194:118955. 10.1016/j.biopha.2025.118955 [DOI] [PubMed] [Google Scholar]

[CR96] 96.Lee B-H, Chen Y-Z, Shen T-L, Pan T-M, Hsu W-H (2023) Proteomic characterization of extracellular vesicles derived from lactic acid bacteria. Food Chem 427:136685. 10.1016/j.foodchem.2023.136685 [DOI] [PubMed] [Google Scholar]

[CR97] 97.Bajic SS, Cañas MA, Tolinacki M, Badia J, Sánchez B, Golic N et al (2020) Proteomic profile of extracellular vesicles released by Lactiplantibacillus plantarum BGAN8 and their internalization by non-polarized HT29 cell line. Sci Rep 10:21829. 10.1038/s41598-020-78920-z [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR98] 98.Hu R, Lin H, Wang M, Zhao Y, Liu H, Min Y et al (2021) Lactobacillus reuteri-derived extracellular vesicles maintain intestinal immune homeostasis against lipopolysaccharide-induced inflammatory responses in broilers. J Anim Sci Biotechnol 12:25. 10.1186/s40104-020-00532-4 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR99] 99.Champagne-Jorgensen K, Mian MF, McVey Neufeld KA, Stanisz AM, Bienenstock J (2021) Membrane vesicles of Lacticaseibacillus rhamnosus JB-1 contain immunomodulatory lipoteichoic acid and are endocytosed by intestinal epithelial cells. Sci Rep 11:13756. 10.1038/s41598-021-93311-8 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR100] 100.Kim W, Lee EJ, Bae IH, Myoung K, Kim ST, Park PJ et al (2020) Lactobacillus plantarum-derived extracellular vesicles induce anti-inflammatory M2 macrophage polarization in vitro. J Extracell Vesicles 9:1793514. 10.1080/20013078.2020.1793514 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR101] 101.Yamasaki-Yashiki S, Kawashima F, Saika A, Hosomi R, Kunisawa J, Katakura Y (2024) RNA-based anti-inflammatory effects of membrane vesicles derived from Lactiplantibacillus plantarum. Foods 13:967. 10.3390/foods13060967 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR102] 102.Yoshida A, Yamada K, Yamazaki Y, Sashihara T, Ikegami S, Shimizu M et al (2011) Lactobacillus gasseri OLL2809 and its RNA suppress proliferation of CD4(+) T cells through a MyD88-dependent signalling pathway. Immunology 133:442–451. 10.1111/j.1365-2567.2011.03455.x [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR103] 103.Chen Y, Huang X, Liu A, Fan S, Liu S, Li Z et al (2024) Lactobacillus Reuteri vesicles regulate mitochondrial function of macrophages to promote mucosal and cutaneous wound healing. Adv Sci (Weinh) 11:e2309725. 10.1002/advs.202309725 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR104] 104.Mandelbaum N, Zhang L, Carasso S, Ziv T, Lifshiz-Simon S, Davidovich I et al (2023) Extracellular vesicles of the Gram-positive gut symbiont Bifidobacterium longum induce immune-modulatory, anti-inflammatory effects. NPJ Biofilms Microbiomes 9:30. 10.1038/s41522-023-00400-9 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR105] 105.Kim JH, Jeun EJ, Hong CP, Kim SH, Jang MS, Lee EJ et al (2016) Extracellular vesicle-derived protein from Bifidobacterium longum alleviates food allergy through mast cell suppression. J Allergy Clin Immunol 137:507–516e8. 10.1016/j.jaci.2015.08.016 [DOI] [PubMed] [Google Scholar]

[CR106] 106.Nishiyama K, Takaki T, Sugiyama M, Fukuda I, Aiso M, Mukai T et al (2020) Extracellular vesicles produced by Bifidobacterium longum export mucin-binding proteins. Appl Environ Microbiol 86:e01464–e01420. 10.1128/aem.01464-20 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR107] 107.Kim S, Han SY, Lee J, Kim NR, Lee BR, Kim H et al (2022) Bifidobacterium longum and galactooligosaccharide improve skin barrier dysfunction and atopic dermatitis-like skin. Allergy Asthma Immunol Res 14:549–564. 10.4168/aair.2022.14.5.549 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR108] 108.Vuong N, Alomia M, Byne A, Gade P, Philipson TR, Alhammad RI et al (2025) Lactobacillus rhamnosus-derived extracellular vesicles influence calcium deposition in a model of breast cancer intraductal calcium stress. iScience 28:112538. 10.1016/j.isci.2025.112538 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR109] 109.Wang J, Li X, Zhao X, Yuan S, Dou H, Cheng T et al (2024) Lactobacillus rhamnosus GG-derived extracellular vesicles promote wound healing via miR-21-5p-mediated re-epithelization and angiogenesis. J Nanobiotechnol 22:644. 10.1186/s12951-024-02893-8 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR110] 110.Artuyants A, Hong J, Dauros-Singorenko P, Phillips A, Simoes-Barbosa A (2024) Lactobacillus gasseri and Gardnerella vaginalis produce extracellular vesicles that contribute to the function of the vaginal microbiome and modulate host–Trichomonas vaginalis interactions. Mol Microbiol 122:357–371. 10.1111/mmi.15130 [DOI] [PubMed] [Google Scholar]

[CR111] 111.Shiraishi T, Yokota S, Sato Y, Ito T, Fukiya S, Yamamoto S et al (2018) Lipoteichoic acids are embedded in cell walls during logarithmic phase, but exposed on membrane vesicles in Lactobacillus gasseri JCM 1131(T). Benef Microbes 9:653–662. 10.3920/bm2017.0124 [DOI] [PubMed] [Google Scholar]

[CR112] 112.Lee DH, Park HK, Lee HR, Sohn H, Sim S, Park HJ et al (2022) Immunoregulatory effects of Lactococcus lactis-derived extracellular vesicles in allergic asthma. Clin Transl Allergy 12:e12138. 10.1002/clt2.12138 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR113] 113.Liu Y, Tempelaars MH, Boeren S, Alexeeva S, Smid EJ, Abee T (2022) Extracellular vesicle formation in Lactococcus lactis is stimulated by prophage-encoded holin-lysin system. Microb Biotechnol 15:1281–1295. 10.1111/1751-7915.13972 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR114] 114.Wang Y, Hu L, Gao B, Yang R, Huang Y, Li P et al (2025) Lactococcus lactis-derived extracellular vesicles: A novel nanodelivery system enhance the properties of nisin Z. Food Chem 489:144776. 10.1016/j.foodchem.2025.144776 [DOI] [PubMed] [Google Scholar]

[CR115] 115.Dean SN, Leary DH, Sullivan CJ, Oh E, Walper SA (2019) Isolation and characterization of Lactobacillus-derived membrane vesicles. Sci Rep 9:877. 10.1038/s41598-018-37120-6 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR116] 116.da Silva Barreira D, Laurent J, Lourenço J, Novion Ducassou J, Couté Y, Guzzo J et al (2023) Membrane vesicles released by Lacticaseibacillus casei BL23 inhibit the biofilm formation of Salmonella Enteritidis. Sci Rep 13:1163. 10.1038/s41598-023-27959-9 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR117] 117.Domínguez Rubio AP, Martínez JH, Martínez Casillas DC, Coluccio Leskow F, Piuri M, Pérez OE (2017) Lactobacillus casei BL23 produces microvesicles carrying proteins that have been associated with its probiotic effect. Front Microbiol 8:1783. 10.3389/fmicb.2017.01783 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR118] 118.Bäuerl C, Coll-Marqués JM, Tarazona-González C, Pérez-Martínez G (2020) Lactobacillus casei extracellular vesicles stimulate EGFR pathway likely due to the presence of proteins P40 and P75 bound to their surface. Sci Rep 10:19237. 10.1038/s41598-020-75930-9 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR119] 119.Kuhn T, Aljohmani A, Frank N, Zielke L, Mehanny M, Laschke MW et al (2024) A cell-free, biomimetic hydrogel based on probiotic membrane vesicles ameliorates wound healing. J Control Release 365:969–980. 10.1016/j.jconrel.2023.12.011 [DOI] [PubMed] [Google Scholar]

[CR120] 120.Lonngi Sosa CD, González Díaz FR, Álvarez HR, Vargas Ruiz A, Higuera Piedrahita RI, de la Cruz Cruz HA et al (2026) Membrane vesicles from Lactobacillus acidophilus promote superior cytokine modulation and antimicrobial signaling compared with their whole cells in RAW 264.7 macrophages. Int J Mol Sci 27:2764. 10.3390/ijms27062764 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR121] 121.Mazgaeen L, Gurung P (2020) Recent advances in lipopolysaccharide recognition systems. Int J Mol Sci 21:379. 10.3390/ijms21020379 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR122] 122.Schmid AM, Razim A, Wysmołek M, Kerekes D, Haunstetter M, Kohl P et al (2023) Extracellular vesicles of the probiotic bacteria E. coli O83 activate innate immunity and prevent allergy in mice. Cell Commun Signal 21:297. 10.1186/s12964-023-01329-4 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR123] 123.Zwicker C, Sarate P, Drinić M, Ambroz K, Korb E, Smole U et al (2018) Prophylactic and therapeutic inhibition of allergic airway inflammation by probiotic Escherichia coli O83. J Allergy Clin Immunol 142:1987–1990e7. 10.1016/j.jaci.2018.07.029 [DOI] [PubMed] [Google Scholar]

[CR124] 124.Tiku V, Tan M-W (2021) Host immunity and cellular responses to bacterial outer membrane vesicles. Trends Immunol 42:1024–1036. 10.1016/j.it.2021.09.006 [DOI] [PubMed] [Google Scholar]

[CR125] 125.Blanc L, Castanier R, Mishra AK, Ray A, Besra GS, Sutcliffe I et al (2013) Gram-positive bacterial lipoglycans based on a glycosylated diacylglycerol lipid anchor are microbe-associated molecular patterns recognized by TLR2. PLoS ONE 8:e81593. 10.1371/journal.pone.0081593 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR126] 126.Saito S, Lin S-H, Wu H-Y (2019) The unique immunoregulatory function of Staphylococcus aureus lipoteichoic acid in dendritic cells. J 2:326–339. 10.3390/j2030022 [DOI] [Google Scholar]

[CR127] 127.Furnari S, Ciantia R, Garozzo A, Furneri PM, Fuochi V (2025) Lactobacilli-derived microbe-associated molecular patterns (MAMPs) in host immune modulation. Biomolecules 15:1609. 10.3390/biom15111609 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR128] 128.Domínguez Rubio AP, D’Antoni CL, Piuri M, Pérez OE (2022) Probiotics, their extracellular vesicles and infectious diseases. Front Microbiol 13:864720. 10.3389/fmicb.2022.864720 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR129] 129.Yuan S, Jin M, Zhang Y, Zhang M, Yuan M, Ding X et al (2025) Extracellular vesicles in skin: Biological function and therapeutic potential. Int J Nanomed 20:11211–11233. 10.2147/ijn.s548986 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR130] 130.Donoso-Meneses D, Figueroa-Valdés AI, Khoury M, Alcayaga-Miranda F (2023) Oral administration as a potential alternative for the delivery of small extracellular vesicles. Pharmaceutics 15:716. 10.3390/pharmaceutics15030716 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR131] 131.Lee H, Lee Y-H, Hong D-K, Mo S-J, Jeon S, Park S-D et al (2024) Targeting inflammation and skin aging via the gut–skin axis: The role of Lactiplantibacillus plantarum HY7714-derived extracellular vesicles. Microorganisms 12:2466. 10.3390/microorganisms12122466 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR132] 132.Chen X, Li Q, Zhang W, Xu Y, Nie X, Wang X et al (2026) Akkermansia muciniphila-derived extracellular vesicles alleviate colitis-related cognitive impairment via tryptophan metabolic reprogramming of the gut–brain axis. Gut Microbes 18:2611546. 10.1080/19490976.2025.2611546 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR133] 133.Nakatsuji T, Chen TH, Narala S, Chun KA, Two AM, Yun T et al (2017) Antimicrobials from human skin commensal bacteria protect against Staphylococcus aureus and are deficient in atopic dermatitis. Sci Transl Med 9:eaah4680. 10.1126/scitranslmed.aah4680 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR134] 134.López P, González-Rodríguez I, Sánchez B, Gueimonde M, Margolles A, Suárez A (2012) Treg-inducing membrane vesicles from Bifidobacterium bifidum LMG13195 as potential adjuvants in immunotherapy. Vaccine 30:825–829. 10.1016/j.vaccine.2011.11.115 [DOI] [PubMed] [Google Scholar]

[CR135] 135.Li L, Si W, Jin Z, Jin W (2026) Research progress on construction technology of 3D human skin models and its application prospects in dermatology. Int J Mol Sci 27:1808. 10.3390/ijms27041808 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR136] 136.Flint E, Ahmad N, Rowland K, Hildebolt C, Raskin D (2024) Topical probiotics reduce atopic dermatitis severity: A systematic review and meta-analysis of double-blind, randomized, placebo-controlled trials. Cureus 16:e70001. 10.7759/cureus.70001 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR137] 137.Flores Rodríguez JC, Morán Ortega MJ, Jimenez Ordoñez AC, Rojas López AK, Leyva MC, Romero HV (2026) Postbiotics in dermatology: A literature review of emerging topical therapies for acne, rosacea, and eczema. Cureus 18:e105440. 10.7759/cureus.105440 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR138] 138.Salo V, Remitz A, Lauerma A, Salava A (2026) Topical probiotic Lactobacillus lactis treatment in atopic dermatitis: a placebo-controlled pilot study on tolerability and efficacy. Front Med (Lausanne) 13:1694229. 10.3389/fmed.2026.1694229 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR139] 139.Tang W-Z, Liao C-Y, Xu H-Y, Huang W-T, Wu Z-X, Chen T-Y et al (2026) Extracellular vesicles in atopic dermatitis: unraveling pathogenic mediators and engineering therapeutic vectors. Front Immunol 17:1846799. 10.3389/fimmu.2026.1846799 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR140] 140.Cannizzaro F, Gallo A, La Scala S, Gallo G, Faddetta T (2026) Bacterial extracellular vesicles in biotechnology: Current challenges and strategies for production enhancement. Fermentation 12:86. 10.3390/fermentation12020086 [DOI] [Google Scholar]

[CR141] 141.Li B, Wu W, Xu W, Qian H, Ji C (2025) Advances of extracellular vesicles isolation and detection frontier technology: from heterogeneity analysis to clinical application. J Nanobiotechnol 23:678. 10.1186/s12951-025-03768-2 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR142] 142.Choi D, Lee EY (2025) Standardizing bacterial extracellular vesicle purification: A call for consensus. J Microbiol Biotechnol 35:e2506011. 10.4014/jmb.2506.06011 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR143] 143.Welsh JA, Goberdhan DCI, O’Driscoll L, Buzas EI, Blenkiron C, Bussolati B et al (2024) Minimal information for studies of extracellular vesicles (MISEV2023): From basic to advanced approaches. J Extracell Vesicles 13:e12404. 10.1002/jev2.12404 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR144] 144.Rudge C, McClelland R, Chrzanowski W, Davis RL (2026) The regulatory landscape for extracellular vesicle therapies: Australian context and future directions. Extracell Vesicles Circ Nucl Acids 7:292–301. 10.20517/evcna.2025.129 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR145] 145.Xu G, Jin J, Fu Z, Wang G, Lei X, Xu J et al (2025) Extracellular vesicle-based drug overview: research landscape, quality control and nonclinical evaluation strategies. Signal Transduct Target Ther 10:255. 10.1038/s41392-025-02312-w [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR146] 146.Wen T, Gu Y, Liu G (2026) Extracellular vesicles in wearable delivery systems for cosmeceutical applications. Adv Healthc Mater 26:e71379. 10.1002/adhm.71379 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR147] 147.Albayati N, Talluri SR, Dholaria N, Michniak-Kohn B (2025) AI-Driven innovation in skin kinetics for transdermal drug delivery: Overcoming barriers and enhancing precision. Pharmaceutics 17:188. 10.3390/pharmaceutics17020188 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR148] 148.Kaisanlahti A, Turunen J, Hekkala J, Mishra S, Karikka S, Amatya SB et al (2025) Gut microbiota-derived extracellular vesicles form a distinct entity from gut microbiota. mSystems 10:e0031125. 10.1128/msystems.00311-25 [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR149] 149.Tang Z-Y-E, Yang X-l, Zhou Y-W, Zou H, Li Y, Wu H et al (2026) Advances in microbial extracellular vesicles: synthetic biology platforms and medical applications. Extracell Vesicles Circ Nucl Acids 7:945–972. 10.20517/evcna.2025.103 [DOI] [PMC free article] [PubMed] [Google Scholar]

PERMALINK

Gram-Positive and Gram-Negative Probiotic Extracellular Vesicles: Mechanisms of Immune Modulation and Therapeutic Opportunities in Atopic Dermatitis

Vipul Wayal

Mey-Fann Lee

Jiu-Yao Wang

Yi-Hsing Chen

Abstract

Introduction

Biogenesis and Molecular Composition: A Comparative View

Gram-Negative Probiotic OMVs Structure, Biogenesis and Composition

Fig. 1.

Table 1.

Gram-Positive Probiotic MVs Structure and Biogenesis

Fig. 2.

Table 2.

Distinct Immunomodulatory Mechanisms of OMVs and MVs in AD

Gram-Negative OMVs: LPS-Driven Immune Polarization

Gram-Positive MVs: LTA-Mediated Immune Tolerance

Gut-Skin Axis: Systemic Reach of PEVs in Context to AD

Fig. 3.

Comparative Synthesis of Gram-Positive MVs and Gram-Negative OMVs

Table 3.

Preclinical Evidence and Clinical Translation

Efficacy in In Vitro and In Vivo Models

Critical Evaluation and Methodological Limitations of Preclinical Evidence

Limitations of Murine AD Models

Overreliance on Simplified 2D In Vitro Monocultures

Conflation of Whole Bacteria with Purified EV Efficacy

Lack of Pharmacokinetics and Dose-Optimization Studies

From Bench to Bedside: Clinical Prospects of PEVs in AD

Translational Challenges and Future Perspectives

Manufacturing Scalability and Production Efficiency

Quality Control and Standardization

Pharmacokinetics, Biodistribution, Safety, and Immunogenicity

Regulatory Challenges

Delivery Strategies for Topical PEV Therapy

Future Research Priorities

Defining Major Knowledge Gaps

Standardized Experimental Approaches

Selection of Appropriate Preclinical and Human-Relevant Models

Biomarker and Patient-Stratification Strategies

Engineering Strategies for Defined and Targeted PEVs

Priorities for Future Clinical Trials

Fig. 4.

Conclusion

Acknowledgements

Author Contributions

Funding

Data Availability

Declarations

Ethical Approval

Competing Interests

Footnotes

Contributor Information

References

Associated Data

Data Availability Statement

ACTIONS

PERMALINK

RESOURCES

Similar articles

Cited by other articles

Links to NCBI Databases