Direct and indirect effects of pathogenic bacteria on the integrity of intestinal barrier

Abstract

Bacterial translocation is a pathological process involving migration of pathogenic bacteria across the intestinal barrier to enter the systemic circulation and gain access to distant organs. This phenomenon has been linked to a diverse range of diseases including inflammatory bowel disease, pancreatitis, and cancer. The intestinal barrier is an innate structure that maintains intestinal homeostasis. Pathogenic infections and dysbiosis can disrupt the integrity of the intestinal barrier, increasing its permeability, and thereby facilitating pathogen translocation. As translocation represents an essential step in pathogenesis, a clear understanding of how barrier integrity is disrupted and how this disruption facilitates bacterial translocation could identify new routes to effective prophylaxis and therapy. In this comprehensive review, we provide an in-depth analysis of bacterial translocation and intestinal barrier function. We discuss currently understood mechanisms of bacterial–enterocyte interactions, with a focus on tight junctions and endocytosis. We also discuss the emerging concept of bidirectional communication between the intestinal microbiota and other body systems. The intestinal tract has established ‘axes’ with various organs. Among our regulatory systems, the nervous, immune, and endocrine systems have been shown to play pivotal roles in barrier regulation. A mechanistic understanding of intestinal barrier regulation is crucial for the development of personalized management strategies for patients with bacterial translocation-related disorders. Advancing our knowledge of barrier regulation will pave the way for future research in this field and novel clinical intervention strategies.

Introduction

The human intestine contains an enormous and diverse microbiota, predominantly comprising bacteria. The symbiotic relationship between humans and their intestinal bacteria profoundly affects multiple aspects of quality of life. The intestinal microbiota contributes to food digestion, nutrient absorption, and protection against pathogenic infections.^1–3 Pathogenic infection and infection-induced microbial dysbiosis can permeabilize the intestinal barrier, allowing bacterial translocation exiting the intestine. ⁴ Such translocation contributes to inflammatory and immunity-related diseases including colitis, liver cirrhosis, pancreatitis, and rheumatoid arthritis. ⁵ The intestinal barrier is a complex structure that separates the lumen from the host’s interior, protecting against pathogenic microbial invasion while facilitating the absorption of nutrients, electrolytes, and solutes, thereby maintaining the host’s health status.^2,6,7 When multiple defences fail, translocated bacteria can spread through adjacent lymph nodes or circulating blood to distant organs, leading to fatal bacteremia, sepsis, and tumorigenesis. ⁸ Pathogenic bacteria can also evade immune responses upon exposure to immune cells.^8,9 This barrier is also regulated by the enteric nervous system, inflammatory responses, external antigen stimulation, metabolic disorder, and autoimmunity.^4,10 Understanding mechanisms of bacterial translocation mainly concerns elucidating how bacteria damage and become translocated across the intestinal barrier. Owing to the complexity of involved regulatory factors and the diversity of pathogenic bacteria, our understanding of bacterial translocation is still very limited. People with different dietary habits, ages, and health conditions display extremely varied microbiota composition, making statistical data analysis challenging. A better understanding of bacterial translocation will clarify its role in pathogenesis and facilitate the prevention and treatment of intestinal-derived infections. This review focuses on how pathogenic bacteria increase intestinal barrier permeability and mediate bacterial translocation, thereby providing possible targets for the treatment of bacterial translocation.

The intestinal barrier and bacterial translocation

The intestinal barrier is a fundamental innate defense system that regulates the intestinal environment and protects against invasion by pathogenic bacteria. It incorporates multiple barriers, including mechanical, chemical, microbial, and immune. The mechanical barrier, also known as the physical barrier, depends on the structures of the intestinal epithelium, lamina propria, and muscular mucosae. ¹¹ The intestinal epithelium is composed of several cell types, including Enterocytes, Goblet Cells, Paneth Cells, Cupped Cells, Enteroendocrine Cells, Microfold (M) cells, and tuft cells, ¹² collectively referred to as intestinal epithelial cells (IECs). ¹³ An important role of IECs is to maintain intestinal barrier integrity, which allows solvent, electrolytes, and nutrients to pass through and impedes access to pathogens. ¹⁴ IECs contribute to the physical and chemical barriers against microbial infection by secreting mucus, digestive juices, mucopolysaccharides, glycoproteins, glycolipids, and other antibacterial agents. ¹⁴ Mucus-secreting goblet cells are interspersed among the intestinal mucosal epithelial cells, and a translucent layer of mucus is continuously distributed on the surface of the intestinal mucosa, which competes for IEC binding sites, preventing bacteria from adhering to the intestinal epithelium. Intestinal antimicrobial substances, including bile, mucopolysaccharides, lysozymes, and glycoproteins, also play roles in maintaining intestinal barrier function. ¹⁵ IECs are continuously generated and replaced every 4–5 days from intestinal stem cells (ISCs) through renewal and migration. ¹⁶ Aged IECs undergo apoptosis and slough off into the intestinal lumen. ISCs residing in the crypts and expressing leucine-rich repeat-containing G-protein-coupled receptor 5 (Lgr5) are responsible for the steady replenishment of surface-resident cells. This replenishment is vital for intestinal homeostasis and protection against pathogenic bacterial invasion. ¹⁷ A structurally and functionally undamaged barrier is foundational to intestinal health. Impairment of the intestinal barrier increases host susceptibility to various infectious and inflammatory diseases. Bacterial translocation indicates increased permeability of the monolayer cell barrier of the intestinal epithelium.

The concept of translocation of intestinal pathogenic bacteria was first proposed by Wolochow et al. ¹⁸ Bacterial translocation is now generally acknowledged that the microbiota initially colonizes the intestine, where endotoxins, peptidoglycans, and/or metabolites penetrate the intestinal mucosal barrier and reach distant organs and tissues. ¹⁹ Bacterial translocation may occur during microbial dysbiosis, intestinal barrier damage, and systemic immune suppression. A wide range of other diseases have also been linked to bacterial translocation, including depression, anxiety, malignancies, heart failure, and cardiopulmonary bypass.^20–24 Bifidobacteria induce necrotic enterocolitis and increase intestinal permeability before the onset of disease, indicating that intestinal permeability can be enhanced before symptomatic disease is observed. ²⁵ Intestinal pathogen diseases are not always triggered solely by altered tight junction integrity. Abnormal immune responses within the host may also increase disease predisposition by increasing tissue inflammation. Pathogenic bacterial translocation occurs mainly through tight junctions and endocytosis but can also induce a neuroimmune response through necrosis or disorders of IEC renewal. Interactions between immune cells and IECs play a significant role in maintaining intestinal barrier integrity. Defects in the intestinal barrier are often accompanied by inflammation, which recruits immune cells including immature dendritic cells, neutrophils, macrophages, and myeloid-derived suppressor cells.^8,26–28 Myeloid-derived cells possess the ability to harbor pathogenic bacteria, which may be a way by which translocated bacteria evade immune clearance.^29,30 Altered immune cell frequencies can increase intestinal barrier permeability. Pathogenic bacteria cross through the mucosal epithelium, causing various intestine-associated diseases, including inflammatory bowel disease (IBD) and pancreatitis, and cancer. ³¹ For example, in patients with IBD, the concentration of total bacterial DNA in intestinal tissues and blood samples is increased, suggesting the occurrence of bacterial translocation and microbial dysbiosis ³² It has also been shown that the ruin of the intestinal barrier leads to bacteria translocation where they are thought not to be present. Through the Apc^min mouse infection with Helicobacter hepaticus, it was found that H. hepaticus DNA was present in the mammary gland, but not in the uninfected Apc^min mouse, and the incidence of breast cancer was higher than the uninfected mouse. MDSC were found to migrate from the gut to the mammary gland, and the reinfusion of MDSC from the infected group to the uninfected group increased mammary tumor number, suggesting that MDSC may play a synergistic role in H. hepaticus metastasis and promoting tumorigenesis. ⁸ Colitis and gut dysbiosis reduce the barrier permeability, allowing the presence of gut-derived bacteria and lipopolysaccharides (LPS) in liver,^33,34 which recruited the CXCR2⁺ PMN-MDSCs formatting an immunosuppressive microenvironment in liver, thereby promoting the development of hepatocellular carcinoma. ³⁵ Bacteria in the tumor microenvironment may be associated with gene expression and pathway activity. ³⁶ Removed intratumoral bacteria could significantly reduce lung metastasis of breast cancer, but without affecting primary tumor growth. Further confirmed that circulating tumor cells harboring intratumoral bacteria enhanced their resistance to fluid shear stress, thereby promoting survival. ³⁷ Nowadays, more and more diseases are found to be related to bacterial translocation, and more evidence is still needed to prove it.

Mechanisms of bacterial translocation

Intestinal barrier defects

Paracellular routes

Intestinal barrier integrity is based on the presence of a series of intercellular junctions composed of apical junction complexes and desmosomes. ³⁸ The former includes Tight Junctions (TJs) and Adherens Junctions (AJs). TJs play crucial roles in preventing paracellular transit. Solutes and small molecules can be transported paracellularly through TJs; however, proteins, lipids, and peptides cannot.^39,40 TJ proteins include transmembrane proteins, such as occludin, claudin, JAM-A (JAM-A), and intracellular scaffold proteins, such as zonula occludins (ZO) and tricellsin. ⁴¹ These proteins interact with the cytoskeleton, resulting in complex structures. ⁴² To maintain barrier integrity, TJ proteins act in concert with intracellular signaling and membrane-spanning proteins.^43,44 Occludin was the first discovered tight junction protein. ⁴⁵ TJ barrier function depends heavily on occludin for structural integrity. A close relationship has been observed between occludin expression and barrier properties. ⁴⁶ Claudins have diverse functions. Barrier function is strengthened by claudins -1, -3, -4, -5, and -8, and weakened by claudins -2, -7, -10, and -23. ⁴⁷ To date, the roles of the other TJ proteins remain unclear. AJs are formed by the transmembrane proteins E-cadherin and nectin, and intracellular components including p120-catenin, α-catenin and β-catenin. ³⁸ Nectin is an adhesion molecule, and afadin is an actin filament-binding protein. Afadin links nectin with the actin cytoskeleton, while catenin links E-cadherin to the actin cytoskeleton. ⁴⁸

Under pathological conditions, TJs and AJS can be disrupted. Bacteria, toxins, LPS, and enzymes are likely to breach the intestinal barrier through paracellular pathways to facilitate bacterial translocation. Pathogenic bacteria frequently manipulate paracellular barriers using LPS, virulence factors, and enzymes, to directly downregulate the expression of TJ and AJ proteins and activate pro-inflammatory cytokines that disrupt tight junctions or recruit immune cells to disrupt the barrier. Bacteria utilize multiple mechanisms to alter permeability, leading to progressive changes concurrent with infection progression.

Campylobacter jejuni secretes a serine protease, high-temperature requirement A (htrA), that downregulates claudin-8 to breach the intestinal barrier.^49–51C. jejuni, co-infected with non-invasive bacterial strains facilitates barrier crossing by the non-invasive bacteria, ⁵¹ which can be attributed to opening of IEC junctions local to C. jejuni infection. Fusobacterium nucleatum upregulates myosin light chain phosphorylation and triggers actomyosin contraction, which distributes ZO-1, occludin, and E-cadherin to increase barrier permeability. ⁵² Colonic epithelial barrier dysfunction is exacerbated by C. concisus activating M1-macrophage by reducing the expression of occludin and tricellsulin in the tricellular proteins of three-cell TJs. ⁵³ Helicobacter hepaticus downregulates ZO-1 and Claudin-1, decreases numbers of lysozyme-positive immune cells in the colonic submucosa, and damages the intestinal barrier in Il-17a^-/- mice. ⁵⁴ Salmonella serovar virulence relies on Salmonella plasmid virulence (Spv) genes. Through F-actin rearrangements and suppression of protein kinase C signaling, SpvB rearranges claudin-1, occludin, and E-cadherin, which play key roles in impairing the intestinal epithelial barrier and facilitating bacterial translocation. ⁵⁵ Listeria monocytogenes secretes the Listeria adhesion protein which interacts with the host cell receptor heat shock protein 60. This process activates the NF-κB pathway, expediting the opening of the epithelial barrier mediated by myosin light-chain kinase (MLCK) via cellular redistribution of junctional proteins, including claudin-1, occludin, and E-cadherin, thereby promoting bacterial translocation.^56,57 In addition, LPS decreases occludin and ZO-1 expression and activates Ca²⁺-activated Cl^- channels and epithelial Na⁺ channels, thereby depolarizing the apical membrane and initiating ZO-1 tyrosine phosphorylation.^58,59 LPS effects depend on mast cell degranulation. ⁶⁰ Both can disrupt barrier integrity, significantly increasing paracellular permeability. Infection triggers release of the proinflammatory cytokine tumor necrosis factor α (TNF-α).^8,61,62 TNF-α stimulates barrier permeability by activating MLCK II and inducing occludin endocytosis.^63,64

In addition, pathogenic bacteria and their virulence factors can directly damage TJ and AJ proteins. Group A Streptococcus (GAS) can translocate through the intestinal barrier. Streptococcal pyrogenic exotoxins B and S cleave E-cadherin to promote GAS translocation.^65,66Clostridium perfringens secretes pore-forming delta-toxin, which stimulates a-disintegrin and metalloprotease 10, causing E-cadherin cleavage ⁶⁷ and an enterotoxin that targets claudin -9 and -4 also causes barrier damage.^68,69Vibrio cholerae hemagglutinin/protease and Aeromonas hydrophila pore-forming aerolysin can directly cleave occludin, damaging the barrirer. ⁷⁰ Aeromonas sobria serine protease (ASP), secreted by A. sobria, degrades tight junction components including ZO-1, 2, 3, and claudin-7, and facilitates bacterial translocation through the intestinal epithelial cell line, T84. ¹¹ In addition, ASP destroys components including nectin-2 and afadin. ⁷¹ Haderer et al. ⁷² reported that a novel bacterial protease produced by Escherichia coli and Proteus mirabilis in patients with spontaneous bacterial peritonitis was responsible for the cleavage of E-cadherin structures.

Transcellular route

Microbiota-Mediated Translocation can occur via Epithelial Endocytosis. Antibiotic-resistant commensal bacteria spread via epithelial transcytosis as a result of microbial dysbiosis. ⁷³ M cells are important entrances for bacterial translocation. M cells are specialized epithelial cells in the mucosal immune system that are scattered among the epithelial cells of the intestinal mucosa. ⁷⁴ M cells sample pathogens in contact with their apical membranes, enclose them in vesicles, and deliver them to the immune system through the basolateral membrane. ⁷⁵ Pathogenic bacteria use M cells as a portal of entry to invade the host and cause infection. ⁷⁶ Transcytosis subversion via M cells may be pivotal for evasion of adaptive immunity by intracellular pathogens. ⁷⁷

In addition to opening the TJ barrier, L. monocytogenes crosses the intestinal barrier by undergoing transcytosis. The bacterial invasion protein internalin A interacts with M cells in the Peyer’s patch and the E-cadherin receptor, prompting goblet cell exocytosis and epithelial extrusion. ⁵⁶ Yersinia pseudotuberculosis (Yptb) exhibiting low type three secretion system (T3SS) expression shows higher transcytosis than Yptb exhibiting high expression. ⁷⁸ Yersinia outer proteins as effectors of T3SS hinder the uptake function of M cells in early-stage infections. ⁷⁹ Therefore, Yptb alters the interplay between the immune system and IECs to subvert intestinal barrier function. Moreover, Salmonella typhimurium invades the intestinal barrier via M cells.^80,81 Invasion by S. typhimurium involves epithelium-sampling lamina propria phagocytes as well as type III secretion system-2-dependent epithelium traversal and basolateral exit. S. typhimurium type III effector protein SopB induces epithelial-mesenchymal transition (EMT) of follicle-associated epithelial enterocytes into M cells by inducing the EMT-regulating transcription factor Slug. Oral Brucella abortus infects the host via uptake through the prion protein PrP (C) on the apical surface of M cells. ⁸² All of these factors enable barrier penetration.

Cell apoptosis

Pathogenic bacteria cause epithelial cell death in numerous ways, through activation of receptors leading to membrane lysis or mitochondrial dysfunction-mediated cell death. Clostridium difficile TcdA-induced enteritis depends on prostaglandin E2, which upregulates the Fas/FasL ligand to induce apoptosis in colonocytes. ⁸³ Kim and Kim proved that prostaglandin E2, via the EP1 receptor, upregulates BCL-2 homologous antagonist/killer (Bak), which could result in apoptosis in mitochondrial outer membrane pores. ⁸⁴ Additionally, toxin B (tcdB) could induce cell apoptosis and increase levels of IL-6 and TNF-α in fetal human colon epithelial cells through the AKT/FOXO3/PPM1B pathway. ⁸⁵ In colonocytes, TcdA, and TcdB disassemble actin microfilaments via glucosylation of Rho family proteins, causing epithelial cells to die and causing TJs to open. ⁸⁶ Cronobacter sakazakii increases intestinal cAMP and protein kinase A phosphorylation following IEC apoptosis after infection. ⁸⁷ EPEC induces intestinal epithelial cell line T84 apoptosis through oligonucleosome formation, PARP cleavage, and activation of caspases-3, -6, -8, and -9. ⁸⁸ Serapio-Palacios and Navarro-Garcia reported that the serine protease motif of EPEC-secreted protein C, an autotransporter protein, can cleave procaspase-3, stimulating the mitochondria-mediated apoptotic pathway. ⁸⁹ Moreover, MAP, the effector of EPEC T3SS, triggers dissolution of the host mitochondrial membrane potential, followed by Ca²⁺ efflux into the cytoplasm and activation of metalloproteinase domain-containing protein 10, which stimulates a mitogen-activated protein kinase cascade, ultimately causing cell apoptosis. ⁹⁰ Vibrio vulnificus secretes many virulence factors, including VvhA and VvpM. Recombinant (r) VvhA protein activates PKCα and ERK/JNK leading to an increase in production of reactive oxygen species, eventually stimulating NF-κB-dependent mitochondrial cell death. ⁹¹ Furthermore, VvpM activates extracellular signal-regulated kinase (ERK signaling, caspases-9, -3, and cytochrome release to induce IEC apoptosis. ⁹²

Klebsiella oxytoca colonization of the intestine plays a role in antibiotic-associated hemorrhagic colitis.^93–95K. oxytoca produces enterotoxins including tilivalline (TV) and tilimycin (TM) during colitis. Targets that induce cell apoptosis differ between TV and TM. TV causes mitotic arrest by binding tubulin and stabilizing microtubules, whereas TM induces strand breaks in DNA by activating damage repair mechanisms. ⁹⁶ SpvB mediates the accumulation of receptor-interacting protein kinase 3 (RIPK3) by reducing its degradation in an autophagy-dependent manner. RIPK3 increases MLKL phosphorylation and promotes necroptosis in IECs. ⁹⁷

Repair and renewal disorder of the intestinal barrier

Maintenance of the intestinal barrier relies on rigorous regulation of IEC death and renewal, and the intestinal epithelium can be almost entirely renewed every 4–5 days.^16,98 This tremendous capacity for renewal relies on ISCs residing at the bottom of the crypts. ¹⁷ ISCs include crypt base columnar cells (CBCs) and cells at position +4. ⁹⁹ The CBC-specific gene Lgr5 has been identified as a vital marker of active ISCs. ¹⁰⁰ Lgr5⁺ ISCs can generate all types of intestinal cells. ¹⁷ Several signaling pathways contribute to regulating ISC activity. Wnt and Notch signaling are considered the major driving pathways that contribute to intestinal maintenance, proliferation of ISCs, and differentiation of post-mitotic cells. Wnt/β-catenin signaling is required for physiological functions such as stem cell maintenance. ¹⁰¹ Interference with this signaling pathway leads to ISC functional disorder. Notch signaling targets CBC stem cells, and Notch inhibition causes a reduction in CBC number. ¹⁰²

Bacteria impair intestinal epithelial repair, increasing intestinal barrier permeability and bacterial translocation. They can induce barrier defects that cause lesions in the intestinal barrier, and can also affect intestinal epithelium repair and renewal by impacting ISCs. ¹⁰³ ISCs coexist with bacteria, placing bacteria in close contact with ISCs and affecting ISC proliferation and differentiation.^100,104Lm and C. difficile directly damage ISCs by reducing the number of Lgr5⁺ stem cells, which greatly influences intestinal integrity.^105,106 Some bacteria also impair the intestinal barrier by disturbing signaling pathways. For instance, Enterotoxigenic Escherichia coli produces heat-stable enterotoxins that suppress Fzd7 expression and downregulate the Wnt/β-catenin signaling pathway to inhibit ISC exposure. ¹⁰⁷ Conversely, S. typhimurium protein, AvrA, activates the Wnt/β-catenin pathway and S. typhimurium infection upregulates the level of p53 in vivo, which promotes the progression of intestinal adenomas. ¹⁰⁸ L. monocytogenes infection affects the differentiation of ISCs into paneth cells. ¹⁰⁵ In summary, pathogenic bacteria can not only directly decrease the number of ISCs to inhibit their function but can also interfere with the signaling pathway to disturb ISC activity.

The microbiota-immune-endocrine-nervous-axis

As the intestine has the abundant lymphatic tissue and nervous tissue interacting with the microbiota, intestinal barrier permeability is regulated by various internal and external factors. ¹⁰⁹ Nerve fibers permeate the intestinal tissue, forming the enteric nervous system. The enteric nervous system (ENS) is composed of intestinal peripheral neuronal cells, including sympathetic, parasympathetic, and sensory neurons, which convey signals from the central nervous system (CNS) to the gut and induce mutual communication.^110,111 The ENS, which coordinates symbiotic bacterial colonization and pathogenic microbe infections, plays an important role in regulating mucosal immunity, contributing to inflammation through the regulation of immunomodulatory neuropeptides, neurotransmitters, and inflammatory cytokines.^6,10,112,113 The intestine and CNS are constantly involved in bidirectional communication, and the intestinal microbiota play a critical role in this communication.^1,114,115 The intestinal microbiota has the ability to interact with the ENS. Interfacing with the vagus nerve can relay signals of peripheral inflammation to the CNS, and can secrete signal peptides or other molecules to the gut, modulating the composition of the microbiota.^116,117 The microbiota can also generate neuroactive metabolites which can travel in the circulation to the CNS. ¹¹⁸ However, neural pathways are not the only bidirectional pathways involved. The gut is rich in lymphoid tissue, and its mucosal immune system constantly communicates with the gut microbiota to modulate inflammation and immune functions. The metabolic capacity of the intestinal flora also significantly affects the endocrine environment of the body. The hypothalamic-pituitary-adrenal (HPA) axis which regulates the endocrine system, is in constant communication with the flora.^{117,119–121} In this setting, microbiota, immune system, endocrine system, and CNS function as a mutually coordinating inter-organ communication network to foster healthy metabolism, mental health, and neuroimmune regulation. This network could be called ‘The Microbiota-Immune-Endocrine-Brain-Axis’. Increased intestinal permeability caused by pathogenic infection, which activates immune-inflammatory responses resulting from reciprocal interactions between the intestinal tract and microbiota, has important implications for brain function, metabolism, and mental health.^4,122

In the presence of pathogenic bacteria, both humans and animals are susceptible to dysbiosis, LPS-induced depression, anxiety, and cognitive decline.^4,123 The microbiota involved in microglial homeostasis and metabolism appear to be key to the nervous system and several behavioral phenotypes.^2,124–126 The two-way regulation of nerves and flora, psychological stress, including depression, anxiety, and possible feedback, causes the intestinal barrier to become more permeable, mediating flora translocation. A likely mechanism is sympathetic activation, which leads to increased intercellular permeability in the gut.^127,128 Translocated flora can negatively impact mucosal immunity and immune-inflammatory responses.^{2,4,117,118,129,130} TLR4 recognizes LPS, which is widely expressed in intestinal tissues. ¹³¹ LPS has been reported to promote immune-inflammatory responses by activating Toll-like transmembrane receptor (TLR4).^4,132 LPS also activates RAGE, leading to endothelial hyperpermeability. ¹³³ Pathogens may directly interact with the intestinal epithelium, causing bacterial translocation, and subsequent intestinal inflammation. The local immune-inflammatory microenvironment reduces barrier integrity and leads to bacterial translocation. Microbial dysbiosis caused by pathogenic infections may mediate stress feedback, leading to increased intestinal barrier permeability. Additionally, toxins secreted by pathogenic bacteria can simultaneously regulate neuroimmunity. C. difficile produces two cytotoxic and enterotoxic proteins, toxin A (TcdA) and toxin B (TcdB), which are its two most potent virulence toxins. ¹³⁴ MIP-2 and IL-8 are released from IECs in response to toxin A exposure.^135,136 Cytokines activate macrophages and enteric nerves to release substance P, calcitonin gene-related peptide, and neurotensin, resulting in interaction between IECs and immune cells, and amplification of the inflammatory response. ⁸⁶ The intestinal microbiota is essential for development and function of the HPA axis. Imbalances in the HPA axis caused by the intestinal microbiota can affect the endocrine system and behavior, including anxiety or depression.^120,121,137 Stress-induced increased intestinal permeability allows bacteria and bacterial antigens to cross the epithelial barrier and activate mucosal immune responses, which in turn alters microbiome composition and enhances HPA drive. Psychological factors can also increase the efficacy of the intestinal barrier by inducing mast cells and corticotropin-releasing hormone eosinophils.^138,139 Corticotropin-releasing hormone can also return to stimulate mast cell degranulation to release TNF-α and protease. ¹⁴⁰ Moreover, increased levels of specific proinflammatory cytokines such as IFN-γ, TNF-α, IL-1β, and IL-6, via activation of HPA axis can disrupt TJ protein function, thereby resulting in bacterial translocation (Figure 1). ⁴

Figure 1.

Pathogens cause defects in the intestinal barrier to translocate or disrupt the self-renewal of ISCs located at crypts, resulting in barrier renewal disorders. Some bacteria exploit M cells and goblet cells as their portal of entry through endocytosis. Pathogenic bacteria and their productions regulate nervous, endocrine and immune systems to regulate inflammatory responses, leading to barrier hyperpermeability. The entire regulatory process is bidirectional, and the regulatory network can be called ‘the nervous-endocrine-immune systems axis’. In addition, the immune cells phagocytose pathogenic bacteria which produce inflammatory factors and then enter the circulatory system.

Clinical treatment

Reshaping the gut microbial microenvironment is an important strategy for preventing and treating bacterial translocation. In this review, the mechanisms of barrier integrity regulation reveal promising ways to alleviate disease symptoms associated with bacterial translocation. Reinforcement of the intestinal barrier has become increasingly attractive with the identification and development of novel therapeutic targets.

Numerous studies have demonstrated the efficacy of probiotics and prebiotics in ameliorating microbial dysbiosis and enhancing intestinal barrier integrity to abrogate bacterial translocation. Using one type of probiotic alone or different probiotics simultaneously can strengthen the intestinal barrier. For instance, Lactobacillus species exert well-documented beneficial effects on the intestinal barrier.^141–149 The concurrent use of different probiotics can alleviate dextran Sulfate sodium salt-induced colitis by rectifying microbial dysbiosis and compromised barrier function.^150,151 Fecal bacterial transplantation to treat intestinal barrier defects has received considerable attention. A randomized controlled study involving 70 individuals that investigated whether probiotics can reduce bacterial translocation and cause changes in the intestinal flora found that probiotics reduced the count of live intestinal bacteria in the blood. Reducing translocation with probiotic intervention occurred late in the intervention at 16 weeks. ¹⁵² Other reports indicate that the intestinal barrier and translocation levels do not change significantly after probiotic intervention.^153,154 Owing to the different observation times and interventions, it is challenging to ascertain the promise of probiotics or fecal microbiota transplantation therapy. An increased number of larger cohort studies are required to fill this gap. We continue to face difficulties in sample collection and analysis, as well as a lack of high-efficiency targeted strains in this field. The virulence of the transplanted bacteria and possible infection require more attention and clinical studies to support it. ¹⁵⁵ Natural derivatives of many plants, including emodin, matrine, galangin, Kaempferol, and Pinocembrin are displaying promising bioactivities.^156–160 Traditional Chinese medicines, including aqueous extracts of Paeoniae Radix Alba (Paeonia lactiflora Pall), herbal medicines including Scutellaria-coptis, Astragalus membranaceus, and Scutellaria baicalensis Georgi, ameliorate DSS-induced colitis in mice by tuning the intestinal physical barrier, immune responses, and microbiota composition.^161–164 Gas-based therapeutics have seen rapid recent development, and some have displayed high therapeutic efficiency and biosafety. H₂ alleviates colitis by reprograming colonocyte metabolism and reinforcing the intestinal barrier. ¹⁶⁵ Additionally, the intake of small molecules such as Butyrate,^166–168 Brain orexin, ¹⁶⁹ vitamins A and D,^170,171 and Galactooligosaccharides^172–174 has improved intestinal barrier function. A high-fat diet, hypertension, and hyperglycemia drive intestinal barrier dysfunction and increase risk of enteric infection.^175–177 Antihyperglycemic Agents (e.g. metformin) can improve intestinal barrier function.^178–181 Therapies targeting ISCs have also been developed. The environmental sensor, aryl hydrocarbon receptor, protects from barrier damage by guarding the stem cell niche to maintain intestinal barrier integrity. ¹⁸² An important research direction for cancer treatment is to engineer bacterial vaccine carriers by utilizing bacterial characteristics that foster translocation. ¹⁸³ Further research is required to better understand the role of intestinal barrier integrity in various diseases and therapeutic interventions. By elucidating the signaling pathways that regulate barrier integrity, novel barrier-restoring agents that are critical for deciphering novel treatment strategies and potent approaches for disease treatment can be identified. Early diagnosis, intervention, or prevention of bacterial translocation could provide a new avenue for the treatment of intestinal-derived infections.

Future perspectives

Clinical strain profiling, typically from stool samples, is commonly used to analyse intestinal microbiota. However, discrepancies in the analysis have been documented compared to samples taken directly from the intestinal contents. To facilitate the reproducibility and comparability of microbiota between different sites of the digestive tract, it is imperative to further develop sample collection and analysis methods. The ability to identify and annotate bacterial strains is critical for detecting bacterial translocation events and determining translocation mechanisms. Identifying high-risk strains, such as Enterotoxigenic Bacteroides fragilis, Helicobacter hepaticus, and Listeria before translocation may prevent disease exacerbation. Most steps in strain identification and annotation require standardized and accurate reference genomes. However, many strains lack high-quality references, leading to identification failures. In addition, bacterial strains entering the intestinal microenvironment in vivo undergo genomic changes, which is a significant obstacle to identifying translocated bacteria. It is thus important to improve current methods of strain annotation, identification, and tracking. Combinatorial use of multiomics may offer breakthroughs in the detection of translocated bacteria and identification of their translocation mechanisms. By combining omics technologies, such as 16S, metagenomics, and metabolomics, more microenvironment-specific characteristics of bacteria can be obtained, which can help us learn from different angular analyses to compare the characteristics of the bacteria, identify and annotate bacteria, and greatly improve accuracy rates.

To date, most studies have focused on the effects of a single or a few species of bacteria local to specific body tissues. Understanding host-microbial interactions is imperative to understand the underlying mechanisms of intestine-derived diseases. To understand how bacteria translocate and how these new environments affect strain behavior, we must understand what factors enable these microbes to translocate. The general mechanisms by which pathogenic bacteria foster flora translocation, including direct action on the intestinal barrier or passing through the microbiota-immune-endocrine-nervous axis, have been elucidated above. The complexity of these regulatory mechanisms makes it difficult to generalize them through a single theory or target. The bidirectional regulatory mechanism of the microbiota-immune-endocrine-nervous axis may underlie a positive feedback loop that further amplifies the translocation of the flora. Currently, an increasing number of researchers are realizing the impacts of psychological status on the intestinal tract. External stress may make patients more prone to eating disorders, unclean food intake, and pathogenic bacterial infections. Pathogen infection may cooperate with stress psychology to further damage the intestinal barrier and translocate bacteria.

Current treatment plans incorporating human experiments and probiotic (fecal bacteria) transplantation still exhibit disadvantages of poor stability, limited curative outcomes, and large individual differences. Symptoms including diarrhea and nausea may also occur. If the patient has severe intestinal leakage and flora translocation events, rash introduction of foreign flora may aggravate the condition or even cause bacteremia. Probiotic transplantation therapy may take a long time to enter the clinic, and it is also necessary to investigate the safety of probiotic transplantation replacement therapy for some patients with severe barrier impairment. Much work is still needed to prove reliable safety and efficacy.

Translocated bacteria are often found circulating in blood, and in distant organs, indicating that translocated bacteria not only exert local impacts but also trigger distal system diseases. An increasing number of immune cells is associated with the spread of translocated bacteria into the circulatory system. Little is known about the immune cells that carry translocated bacteria or help them to evade immune responses.