The Versatile Roles of Exosomes in Neurodegenerative Disorders: From Pathological Mechanism and Diagnostic Biomarkers to Therapeutic Application

Abstract

Neurodegenerative diseases are characterized by cognitive or motor impairments resulting from the progressive degeneration of neurons or myelin sheaths. Currently, the majority of these disorders remain incurable, leading to significant detriments in patients’ quality of life and imposing substantial economic burdens. Exosomes are a type of extracellular vesicles secreted by cells into the extracellular space, typically range from 30 to 150 nanometers in diameter. These vesicles have been identified in a variety of cell types and can participate in various physiological and pathological processes, including intercellular transport of biomolecules, signal transduction, immune modulation, tissue repair, facilitation of tumor metastasis, and evasion of immune surveillance by cancer cells. In recent years, the potential applications of exosomes in neurodegenerative diseases have garnered increasing attention. Their roles encompass early diagnostic biomarker functions, vehicles for drug delivery and therapeutic agents, mediators in the propagation of pathogenic proteins, and regulators of neuroinflammatory responses. In this review, we try to aims to synthesize recent progress in exosome isolation techniques, elucidate the pathogenesis of neurodegenerative diseases, including Alzheimer’s disease (AD), Parkinson’s disease (PD), Amyotrophic lateral sclerosis (ALS), and Huntington’s disease (HD), and examine the diagnostic and therapeutic applications of exosomes within these conditions. Furthermore, it addresses current limitations in exosome-based applications and explores potential strategies to overcome these challenges. The insights of this review may provide the research directions and guidance for future related researchers in these fields.

Introduction

Neurodegenerative diseases are a type of disease characterized by progressive loss of neuronal structure or function, which subsequently leads to functional impairments.1 Currently, neurodegenerative diseases have been identified as comprising Alzheimer’s disease (AD), Parkinson’s disease (PD), amyotrophic lateral sclerosis (ALS), Huntington’s disease (HD), and frontotemporal dementia (FTD).2 Initial manifestations of neurodegenerative diseases often involve memory deficits or reduced motor coordination, which progressively worsen over time, ultimately impairing daily living activities and resulting in severe cognitive decline or motor dysfunction.3–5 AD is currently recognized as one of the diseases with the highest incidence rate of neurodegenerative diseases.6 It is estimated that approximately 416 million individuals worldwide are affected by AD-related conditions, including 32 million diagnosed with AD dementia, 6.9 million with prodromal AD, and 315 million in the preclinical stage of AD. This accounts for 22% of the global population aged 50 years and older.7 Aging has been identified as the primary risk factor for AD, and evidence has demonstrated that the incidence rate of AD is increased rapidly accompanied with aging.8 PD has emerged as the second most common neurodegenerative disease worldwide, prevalent neurodegenerative disorder globally, with aging recognized as a significant risk factor contributing to its development.9,10 In contrast, the incidence rates of amyotrophic lateral sclerosis (ALS), Huntington’s disease (HD), and FTD remain comparatively low, with familial history and genetic factors identified as the primary etiological contributors to these conditions.11–14 In recent years, the worldwide incidence of neurodegenerative diseases has risen markedly, largely attributable to population aging, thereby imposing substantial economic burdens on societies worldwide.15

Exosomes are a class of nanoscale extracellular vesicles, ranging from 30 to 150 nm in diameter, that are endogenously secreted by cells.16–18 These vesicles possess a lipid bilayer membrane structurally analogous to that of cellular membranes, and their surfaces display antigens similar to those found on cells, which facilitates evasion from immune recognition and clearance during systemic circulation.19–21 Furthermore, exosomes have been demonstrated to encapsulate a variety of bioactive molecules, including proteins, lipids, RNAs, and DNAs, thereby playing a critical role in intercellular communication and serving as vehicles for drug delivery. The lipid bilayer membrane structure of exosomes can effectively protect these bioactive cargos from enzymatic degradation in the circulatory system, thereby enhancing their stability.22,23 Additionally, as naturally secreted nanoscale vesicles, exosomes exhibit inherent biocompatibility and low immunogenicity,24 which contributes to their prolonged circulation time and reduced clearance by the immune system, underscoring their favorable safety and tolerability profiles.25,26 The immunomodulatory and tissue repair capabilities of exosomes further underscore their therapeutic potential in the treatment of various pathologies, including neurodegenerative diseases.27

Crucially, the inherent biological properties that render exosomes promising platforms for therapeutic and diagnostic applications may also contribute to disease progression. Exosomes are capable of safeguarding and transporting proteins, lipids, and nucleic acids among neurons, microglia, astrocytes, and peripheral cells. In the context of oxidative stress, mitochondrial dysfunction, abnormal protein aggregation, or neuroinflammation, alterations in exosome secretion and cargo composition can facilitate the intercellular transmission of pathogenic proteins, such as amyloid-β, α-synuclein, and phosphorylated tau. Consequently, exosomes may exhibit dual functions in neurodegenerative diseases: acting both as pathological mediators that contribute to disease propagation, and as carriers for neuroprotective molecules or therapeutic compounds.

In this review, we summarize the recent progress in exosome isolation techniques, the underlying mechanisms of neurodegenerative disease pathogenesis, and the diagnostic and therapeutic applications of exosomes in neurodegenerative diseases. Furthermore, we critically assess the reproducibility and translational potential of existing studies, highlighting current limitations in the field and proposing potential strategies to overcome these challenges. It is our hope that this review will offer a balanced perspective and serve as a valuable resource for guiding future research.

Study Design and Methodology

This review collected the research articles published between 2021 and 2026 from the following databases: PubMed database, Scopus database, Embase database, ScienceDirect database, and Web of Science. The search strategy employed in this study incorporated the following keywords: “neurodegenerative disease”, “exosomes”, “pathogenesis”, “diagnosis”, and “treatment”. Inclusion criteria for selecting publications encompassed the following: articles published within the specified timeframe (2021–2026); The literature under review included original research, peer-reviewed papers and review articles; The languages of the literature were published in English. The selected studies was divided into the following three categories: (1) clinical studies, which include human-focused research such as clinical trials, observational studies, and case reports; (2) animal research, comprising basic research that utilizing animal models; (3) cell research, involving in vitro studies conducted on cell lines or primary cells. Exclusion criteria eliminated conference abstracts, commentaries, and editorials from consideration.

Isolation and Characteristics of Exosomes

Exosomes are one type of special lipid bilayer vesicles that encapsulate a diverse array of bioactive molecules.28 The isolation and purification of exosomes constitute a critical preliminary step in their application and investigation, as these processes directly influence the preservation of their structural integrity and functional properties.29 Currently, several methodologies are employed for exosome isolation, including ultracentrifugation, size exclusion chromatography, ultrafiltration, polymer precipitation, immunoaffinity capture, and microfluidic technology.30 Among these, ultracentrifugation remains the most traditional and extensively utilized technique.31 The basic principle of this method is to utilize the differences in density and size between exosomes and other cellular debris, proteins, and other impurities through the application of incrementally increasing centrifugal forces to achieve sedimentation. A notable advantage of ultracentrifugation is its broad applicability across a wide range of sample types.32 However, this technique may induce vesicle aggregation, structural damage, inconsistent recovery rates, and co-isolation of non-vesicular contaminants. Therefore, its widespread use does not necessarily indicate that it is the most appropriate method for clinical translation. To facilitate a comprehensive understanding of the relative merits and limitations of various exosome isolation methods, a detailed comparative analysis is presented in Table 1.

Table 1.

The Comparisons of the Different Isolation Methods of Exosomes

Method	Principle	Processing Time	Cost	Purity	Exosome Integrity	Scalability	Main Contaminants	GMP Applicability	Disadvantages
Ultracentrifugation	Utilize the differences in sedimentation rate due to variations in particle size and density	>4 h	Middle	Low	Possible damage to exosomes due to high centrifugal force	Large volume (<25 mL)	Lipoproteins, vesicles, and viruses	Low	High costs of equipment, time-consuming, easy to mix impurities
Size exclusion chromatography	Utilize molecular sieves to achieve separation based on particle size differences	0.5–1 h	Middle	High	No damage to exosomes	Small volume (<1 mL)	Lipoproteins and viruses	High	High costs of chromatography columns; the sample may be diluted and the sample size is limited
Ultrafiltration	Selective separation using microporous membrane based on molecular size	1-3h	Middle	Medium	Possible damage to exosomes	Small volume (<15 mL)	Lipoproteins and viruses	High	Membrane pollution and flux decrease
Polymer precipitation	Inducing polymer phase separation by changing solution conditions	0.5–12 h	Low	Low	Possible damage to exosomes	High-throughput	Lipoproteins, viruses	Low	Introducing polymer interference
Immunoaffinity capture	Antigen- antibody binding	4–20 h	High	High	Elution may bring to damage to exosomes	Small volume (<3 mL)	Non-specific binding compound	Middle	High costs of specific antibodies and coupled magnetic beads; easy ligand detachment
Microfluidic technology	Microfluidic	<1 h	High	Variable	No damage to exosomes	Small volume	/	Low	High costs of equipment and chips; lack unified standards to compare and replicate the results

Notes: “cost” refers to the approximate expense of reagents and consumables required to process a single experimental sample. Based on the prevailing price lists from local suppliers and core facilities, we established the following criteria: Low cost: < ¥100 per sample; medium cost: ¥100- ¥500 per sample; High cost: > ¥500 per sample.

The application value of exosomes in neurodegenerative diseases highly depends on the quality of their isolation and purification. Currently, the predominant methodologies employed for exosome isolation in neurodegenerative disease research include ultracentrifugation, size-exclusion chromatography and ultrafiltration. Each of these techniques presents distinct advantages and limitations with respect to their application in diagnosis, therapeutic intervention, and mechanistic studies of neurodegenerative diseases. Specifically, size-exclusion chromatography is particularly advantageous for the identification of novel biomarkers, especially when handling precious biological specimens such as cerebrospinal fluid, owing to its gentle operational conditions that preserve exosomal integrity. Ultracentrifugation remains a conventional and widely utilized approach for biomarker discovery, therapeutic investigations, and elucidation of disease mechanisms, attributed to its relative convenience and rapid processing capabilities. Additionally, ultrafiltration serves a complementary role by enabling sample concentration and is often employed in conjunction with ultracentrifugation to facilitate expedited sample concentration.

Comparisons among these methods should be approached with caution due to inherent differences in recovery efficiency, purity, particle aggregation, co-isolation of lipoproteins, and preservation of biological activity can alter the apparent exosome cargo and experimental outcomes. Consequently, biomarkers or therapeutic effects identified using one isolation method may not be reproducible when alternative methods are employed. For clinical applications, the choice of isolation technique should therefore consider not only speed and yield but also factors such as purity, reproducibility, scalability, cost-effectiveness, and compliance with Good Manufacturing Practice (GMP) standards.

The identification of exosomes requires validation through multiple technical approaches and the integration of findings from diverse analytical perspectives. The identification of exosomes is currently dependent on three principal criteria according to the “Minimum Information for Studies of Extracellular Vesicles” published by the International Society of Extracellular Vesicles (ISEV): morphological assessment, morphological evaluation, and detection of specific marker proteins.33 The measurement of particle size mainly confirms whether the obtained particles belong to the size range of exosomes, which typically have a diameter size ranging from 30 to 150 nm.34 Morphological evaluation typically employs transmission electron microscopy (TEM) to ascertain whether vesicles exhibit the canonical “cup-shaped” or “double concave disc-shaped” structures.35 Detection of specific surface markers proteins of exosomes mainly involves the employment of protein immunoblotting, flow cytometry, and enzyme-linked immunosorbent assay (ELISA) to detect targeting markers, including CD9, CD63, CD81, HSP70, HSP90, Alix, and TSG101.36,37 However, reliance solely on particle size, morphology, and positive marker expression is insufficient to determine the purity, cellular origin, or functional equivalence of exosome preparations. Additional assessments, including evaluation of negative markers, contaminant profiling, particle-to-protein ratios, and functional potency assays, are essential, particularly for preparations intended for clinical use. Comprehensive validation is critical to ensure the reliability and reproducibility of subsequent research findings (Figure 1).

Figure 1.

The summary of the biogenesis and the identification of exosomes. The figure illustrates the formation process of exosomes, including endocytosis process, the process of early endosomes, intracellular multivesicular bodies, and exocytosis process. The primary biomarkers and detection technology of exosomes are also summarized. The blue arrow represents the process of exosome formation. The black arrow represents the structure of nucleic acids, while the blue lines represents the enlarged structure of exosomes.

Abbreviations: TEM, transmission electron microscope; NTA, nanoparticle tracking analysis; WB, Western blotting; FCM, flow cytometry; ELISA, enzyme linked immunosorbent assay; MHC, major histocompatibility complex; TSG101, tumor susceptibility 101; CD9, cluster of differentiation 9; CD63, cluster of differentiation 63; CD81, cluster of differentiation 81.

The Pathogenesis and Risk Factor of Neurodegenerative Diseases

The pathogenesis of neurodegenerative diseases is an intricate and multifaceted process influenced by a combination of genetic, environmental, and age-related factors.38–40 These factors act synergistically, culminating the selective degeneration and functional impairment of specific neuronal populations.41 Protein misfolding and aggregation are considered the causes and central links of most neurodegenerative diseases.42 The biology of exosomes may provide a mechanistic bridge connecting these intracellular disturbances to the progressive propagation of pathological changes throughout neuronal and glial networks. Factors such as cellular stress, disruptions in autophagy, inflammatory responses, and mitochondrial dysfunction can modulate exosome secretion and the selection of their molecular cargo. Consequently, these exosomes may facilitate the intercellular transfer of pathological proteins, regulatory RNAs, lipids, or inflammatory mediators to recipient cells, thereby contributing to disease progression.

The pathogenesis of AD is initiated by the abnormal accumulation of amyloid-beta due to a combination of genetic and environmental factors. Soluble β-amyloid oligomers further trigger synaptic impairment and provoke neuroinflammatory responses.43,44 Subsequent neuroinflammation and cellular injury signals promote excessive phosphorylation of tau protein, ultimately resulting in the cortical dissemination and the formation of neurofibrillary tangles.45 Tau pathology is directly implicated in neuronal death and brain atrophy, which in turn results in irreversible and severe decline in cognitive function.46 In PD, the aggregation of alpha-synuclein into Lewy bodies predominantly affects dopaminergic neurons.47 PD is characterized as a progressive neurodegenerative disorder centered on alpha-synuclein pathology, which is driven by mitochondrial dysfunction, oxidative stress, and neuroinflammation, selectively targeting dopaminergic neuronal populations.48 Emerging evidence suggests that PD pathology may originate in peripheral sites, such as the enteric nervous system, and subsequently propagate to critical motor regions within the brain over time.49 ALS represents a multifactorial neurodegenerative condition influenced by genetic factors, dysregulation of RNA metabolism, impaired protein homeostasis, disrupted axonal transport, neuroinflammation, and mitochondrial dysfunction. The disease is pathologically characterized by the progressive degeneration of both upper and lower motor neurons, accompanied by abnormal cytoplasmic protein aggregates, notably including TDP-43.50,51 Evidence has demonstrated that approximately 5–10% of patients diagnosed with ALS have a family history of the condition.52 Furthermore, pathogenic genes including C9orf72, SOD1, TARDBP, and FUS have been identified as significant contributors to familial ALS, with pathogenic mutations constituting the primary etiological factors in disease onset.53 HD is an autosomal dominant neurodegenerative disorder marked by progressive neuronal loss within the basal ganglia and cerebral cortex. The etiology of HD is attributed to the abnormal expansion of a CAG trinucleotide repeat within the HTT gene.54 The principal pathogenic mechanism involves this single-gene mutation leading to the production of a mutant huntingtin protein (mHTT), which exerts toxic effects and induces multi-level cellular dysfunction.55

Mitochondria represent principal organelles responsible for cellular energy production, and their dysfunction leads to insufficient ATP generation. Neurons, characterized by their high energy demands, are particularly vulnerable to such deficits.56 Additionally, mitochondrial impairment can result in the overproduction of reactive oxygen species (ROS), which induce oxidative damage to lipids, proteins, and DNA, thereby compromising both mitochondrial integrity and neuronal structures.57 Chronic neuroinflammation can trigger hyperactivation of microglia and astrocytes, leading to the release of large amounts of pro-inflammatory cytokines and neurotoxic substances, exacerbating neuronal damage and synaptic loss.58 Furthermore, mitochondrial stress and neuroinflammation may also increase exosome secretion or modify exosomal content, thereby intensifying pathological intercellular communication. Disruptions in axonal transport and synaptic function also contribute to the dysregulation of nutrient delivery and signal transduction within the nervous system.59 Environmental factors and aging are associated with epigenetic modifications, such as DNA methylation and histone alterations, which modulate the expression of genes implicated in neuronal survival, inflammatory responses, and protein metabolism, thus playing a role in disease progression.60,61 Specifically, the failure of glutamate reuptake mechanisms leads to its excessive accumulation in the synaptic cleft.62 This glutamate excess, coupled with calcium overload and overactivation of N-methyl-D-aspartate (NMDA) receptors, results in a pronounced influx of calcium ions, triggering a cascade of enzymatic reactions that ultimately culminate in neuronal cell death.63 These convergent mechanisms may elucidate why exosomes are concurrently recognized as biomarkers of neuronal damage and proposed facilitators of disease propagation.

There are two categories of risk factor that affect the pathological process of neurodegenerative diseases: non-modifiable risk factors and modifiable risk factors.64,65 Non-modifiable risk factors mainly include age, sex, and genetic factors, whereas modifiable risk factors include cardiovascular and metabolic conditions, lifestyle, environmental exposures, comorbid diseases, and infections.15,66 Among these, age has been recognized as the most significant risk factor for neurodegenerative disorders.67 Aging is associated with a decline in cellular repair mechanisms and a diminished capacity to clear deleterious proteins, which adversely affects the proliferation and differentiation of neuronal cells.68 Current evidence indicates that the incidence of various neurodegenerative diseases increases exponentially with advancing age.69,70 Research data have demonstrated that the incidence of AD is significantly higher in women than in men,71 a phenomenon that may be attributable to factors that postmenopausal women experience a decrease in estrogen levels and the overall lifespan of women is generally greater than that of men.72 Conversely, PD exhibits a higher prevalence and incidence in males, a phenomenon potentially linked to the neuroprotective effects of sex hormones, genetic susceptibility differences, and variations in environmental and occupational exposures.73 Furthermore, evidence has demonstrated that several types of chronic diseases, including hypertension, diabetes mellitus, high cholesterol and obesity have been identified contributors to vascular dysfunction, cerebral hypoperfusion, blood-brain barrier disruption, and chronic inflammation. These pathophysiological changes significantly elevate the risk of AD and vascular dementia.74–76 In addition, sleep represents a critical period for the brain to clear metabolic waste such as the β-amyloid protein. It has been demonstrated that sleep disorders can accelerate the aggregation of β-amyloid protein.77 Additionally, behaviors such as smoking and excessive alcohol consumption induce oxidative stress and generate neurotoxic substances, thereby promoting neuronal damage and increasing susceptibility to neurodegenerative diseases.78 Further evidence suggests that traumatic brain injury, particularly repeated moderate to severe trauma, is a significant risk factor for AD and chronic traumatic encephalopathy.79 Similarly, Exposure to specific neurotoxins, including certain pesticides linked to elevated PD risk and heavy metals, can provoke neuroinflammation and neuronal injury, thereby facilitating the development of neurodegenerative conditions.80 Notably, psychiatric disorders such as depression and systemic inflammatory diseases also impact brain function. The pathogenesis of depression, characterized by neurotransmitter imbalances, neuroinflammation, and disrupted functional connectivity within brain regions, may contribute to neurodegeneration.81 Systemic inflammatory diseases have the potential to penetrate or influence the blood-brain barrier, activate central nervous system immune cells, exacerbate neuroinflammation, and consequently promote neurodegenerative processes82 (Figure 2).

Figure 2.

The summary of pathogenesis and risk factor of neurodegenerative diseases. The figure illustrates the primary pathogenesis of different types of neurodegenerative diseases, including AD, PD, HD, and ALS; It also summarizes the primary risk factors, including modified factors and non-modified factors. The blue and black arrows both represents the development process of the different type of diseases, while the dotted blue arrows represents the enlarged structure of microtube.

Abbreviations: sAPPβ, the soluble amyloid protein procurer β; sAPP, the soluble amyloid protein procurer; Aβ, amyloid β-protein; ALS, amyotrophiclateralsclerosis.

The Application of Exosomes in Neurodegenerative Diseases

The Diagnostic Application of Exosomes in Neurodegenerative Diseases

Exosomes are a type of nanoscale vesicles (30–150 nm) secreted by cells. They carry multiple types carrying a variety of specific, bioactive ingredients from the source cells, including proteins, RNAs, DNAs, and lipids. Exosome-based diagnostics represent a promising application for diagnosing neurodegenerative diseases. This is due to their ability to penetrate the blood-brain barrier, their ease of obtainability, their capacity for repeated sampling and their potential for dynamic monitoring. It provides a new approach to problem-solving, including early diagnosis and monitoring of disease progression. For example, Fan et al83 developed a unique micro-immobeads-based electromagnetic operating system. This system can automatically and accurately analyze the Aβ₄₂ protein derived from exosomes in blood samples of AD patients. Further statistical analysis demonstrated that a significant correlation between the exosome-derived Aβ₄₂ protein and the pathogenesis of AD. Similarly, Xu et al84 have developed a novel exosome isolation system including a novel drop-shaped porous microfluidic chip and an immunomagnetic bead based on rolling circle amplification (RCA). This special exosome isolation system can selectively screen for and obtain exosomes from serum samples, and accurately quantify the Aβ42 and p-Tau181 proteins in exosomes, which provide powerful tools for the early diagnosis and prevention of AD. Especially, Belcea et al85 developed an automated exosome isolation and diagnostic system for the early diagnosis of AD. This system is particularly useful for simultaneously isolating and screening of biomarkers derived from exosomes. This system can boast high purity and sensitivity, accurately detect and isolate biomarkers from various sources, making it a valuable asset for research and clinical applications. Furthermore, by combining various techniques, including tandem mass spectrometry proteomics technology, PCR technology, and colorimetric nano-enzyme assay, multiple types of exosome-derived proteins and mRNAs have been identified to be highly correlated with the pathogenesis of AD and identified as the biomarkers of AD.86–92 In addition, exosomes are also widely used as a diagnostic tool in PD. For example, Jang et al93 have developed a type of rapid and cost-effective exosome isolation technology based on the transferrin-conjugated magnetic nanoparticles. This technology can complete the isolation of exosomes within 35min. Following further analysis and statistics, it was identified that the following microRNAs were correlated with PD pathogenesis: miR-195-5p, miR-495-3p, miR-23b-3p, miR-30c-2-3p, miR-323a-3p and miR-27a-3p. Chen et al94 isolated the plasma exosomes from the PD patients and healthy individuals, and compared ferritin and transferrin receptor (TfR) levels in exosomes from the two different populations. The data demonstrated that PD patients had higher levels of ferritin and TfR than healthy individuals. Furthermore, statistical analysis revealed a significant correlation between ferritin and TfR levels and the progression of PD. These results suggest that ferritin and TfR derived from exosomes may serve as the diagnostic biomarkers for PD. For screening the early diagnostic biomarkers of ALS, Gautam et al95 isolated and analyzed the serum exosomes from the different members of one family with the SOD1D90A mutation. They identified a correlation between fibronectin 1 and the progression of ALS. Cheng et al96 analyzed the exosome-derived miRNAs from the peripheral blood of ALS patients and healthy individuals. Their findings revealed that five different exosomal miRNAs, including hsa-miR-199a-3p, hsa-miR-30b-5p, hsa-miR-501-3p, hsa-miR-103a-2-5p, and hsa-miR-181d-5p, were associated with ALS progression. This demonstrates that exosomal blood tests may serve as a potential diagnostic tool for ALS (Table 2 and Figure 3).

Table 2.

The Summary of Details of the Diagnostic Application of Exosomes in Neurodegenerative Diseases

Disease Type	Sample Size	Sample Type	Isolation Method	Biomarker	AUC	Sensitivity and Specificity	Disease Stage	Reference
AD	Healthy individuals (n = 30) AD patients (n=20) Mild cognitive impairment (n = 10)	Plasma	Exosome isolation reagent kit	Exo-Aβ42	>0.9	Sensitivity was 80% and specificity was 80%	Early stage	[83]
AD	Healthy individuals (n=8) AD patients (n=8)	Serum	Microfluidic chip	Aβ42 and p-Tau181	>0.9	0.72 pM for Aβ42 and 0.038 pM for p-Tau181	Early stage	[84]
AD	Healthy individuals (n=100) AD patients (n=100)	Plasma	Automated high-purity exosome isolationbased AD diagnostics system	p-S396-Tau, p-T181-Tau, and Aβ1-42	/	1 copy/µL for nucleic acids and 5 copies/µL for proteins	Early stage	[85]
AD	AD patients (n = 26) Unaffected controls (n = 10)	Cerebrospinal fluid, serum and cell culture media	Exosome isolation reagent kit	miR-501-3p and miR-502-3p	0.79–0.91	/	Early stage	[86]
AD	/	Plasma	Ultracentri fugation	Exo-Aβ42	/	4.2× 104 particles/ mL	Early stage	[87]
AD	AD patients normal individuals	Blood	Ultracentri fugation	Exo-Aβ43	>0.95	Sensitivity was 95.0% and specificity was 95.0%	Early stage	[88]
AD	AD patients (n = 71) Healthy individuals (n = 71)	Serum	ExoQuick™ exosome isolation kit	miR-125b and miR-451a	miR-125b-1-3p was 0.765, miR-451awas 0.728	Sensitivity was 82.1% (67.9%) and specificity was 67.7% (72.6%)	Early stage	[89]
AD	AD patients (n = 15) Healthy individuals (n = 15)	Blood	Total exosome isolation reagent kit	GABRB3, GGTLC1, CADM1, FOXJ3, and ACY1	0.875	Sensitivity was 87.5% and specificity was 75%	Early stage	[90]
PD	AD patients (n = 278) Healthy individuals (n = 257)	/	/	GNAS, TUBB2A, RPL22, RPL5, and WNT5A	/	/	Early stage	[91]
PD	GSE16658 (PBMC, n = 32), GSE269776 (serum exosomes 2021, n = 76), and GSE269775 (serum exosomes 2020, n = 100)	Serum	/	miR-92b, miR-133a, miR-326, miR-125b, miR-148a, and miR-30b	0.791	Sensitivity was 100% and specificity was 43–69%	Early stage	[92]
PD	33 patients with PD, multiple Sclerosis (MS), and dementia	Plasma	Transferrin- conjugated magnetic nanoparticles	miR-195-5p, miR-495-3p, miR-23b-3P, miR-30c-2-3p, miR-323a-3p, and miR-27a-3p	/	/	All stage	[93]
PD	PD patients (n =43) Healthy individuals (n = 34)	Plasma	Total plasma exosome isolation kit	Ferritin and transferrin receptor	0.808	Sensitivity was 86.0% and specificity were 73.5%	Early stage	[94]
ALS	ALS patients (n =3) Healthy individuals (n = 3)	Serum	exoEasy kit	Fibronectin 1	/	/	Early stage	[95]
ALS	ALS patients (n =65)	Plasma	exoRNeasy midi kit	SOD1/C9orf72	0.8	Accuracy was 78.67%	All stage	[96]

Note: “/” represents that data not reported in the original study.

Abbreviations: AD, Alzheimer’s disease; Aβ42, amyloid-beta protein (1–42); AUC, area under the curve; Exo, exosomes; PD, Parkinson’s disease; miR, microRNA; GABRB3, gamma-aminobutyric acid type A receptor β3 gene; GGTLC1, gamma-glutamyltransferase light chain 1; CADM1, cell adhesion molecule 1; FOXJ3, forkhead box J3; ACY1, aminoacylase 1; GNAS, guanine nucleotide binding protein, alpha stimulating; TUBB2A, tubulin beta 2A class IIa; RPL22, ribosomal protein L22; RPL5, ribosomal protein L5; WNT5A, wingless-type MMTV integration site family, member 5A gene; PBMC, peripheral blood monoculear cell; ALS, amyotrophic lateral sclerosis; SOD1, superoxide dismutase 1; C9orf72, chromosome 9 open reading frame 72.

Figure 3.

The summary of the diagnostic application of exosomes in neurodegenerative diseases. The figure representative illustrates the principles of different types of isolation methods, capture methods, and analyzing method for exosome diagnosis application in neurodegenerative diseases. Blue and black lines represent the names of the different components used in exosome detection methods. Blue arrow shows the specific process of the extracellular vesicle detection method, and black arrow shows the corresponding sample and biomarker.

Abbreviations: Aβ, amyloid β-protein; ALS, amyotrophiclateralsclerosis; AD, Alzheimer’s disease; CD63, cluster of differentiation 63; ELISA, enzyme linked immunosorbent assay; RNASE 6, ribonuclease a family member 6; PPP5D1, PPP5 tetratricopeptide repeat domain containing 1; GNAS, guanine nucleotide binding protein, alpha stimulating; TUBB2A, tubulin beta 2A class IIa; RPL22, ribosomal protein L22; RPL5, ribosomal protein L5; WNT5A, Wnt family member 5A.

The Therapeutic Application of Exosomes in Neurodegenerative Diseases

Exosomes are a type of naturally targeted and low-immunogenic particle with a diameter of 30–150nm. In the therapeutic management of neurodegenerative diseases, they can carry proteins and miRNAs to across the blood-brain barrier, regulate the immune microenvironment, and directly or indirectly degrade neuropathological proteins (such as β-amyloid, α-synuclein, and tau). Additionally, they are vesicles that carry hundreds of proteins and miRNAs, which have various physiological effects such as inhibiting apoptosis, regulating autophagy, reducing inflammation, and promoting synaptic remodeling. As a type of vesicle with a cell membrane structure, exosomes are considered a natural drug delivery carrier and can be functionally modified through multiple pathways. The unique characteristics of these particles suggest a significant therapeutic potential in the treatment of neurodegenerative diseases. For example, to address issues relating to the delivery of drugs to the brain and the instability of miR-124-3p in vitro, Ke et al97 isolated the exosomes from microglia and modified them with miR-124-3p. Animal experiments demonstrated that miR-124-3p modified exosomes could cross the mouse blood-brain barrier to reach the lesion site. Furthermore, these exosomes have been shown to alleviate neuroinflammation and improve cognitive function in AD mice by regulating the MEKK3/NF-κB pathway. To further the delivery of exosomes to the brain, Yan et al98 employed low-intensity ultrasound to assist in delivering hADSC-Exos to AD mice to observe the therapeutic effect of their combined action. The data demonstrated that low-intensity ultrasound promoted the absorption of exosomes by target cells. Further animal experiments demonstrated that low intensity ultrasound assisted hADSC-Exos can improve the learning and memory of AD mice. Especially, Han et al99 developed a type of protein loading and release system based on exosomes. This special exosome-based delivery system can conduct the CRISPR-Cas-based epigenome editing in the AD mice, promote CpG site methylation in the Bace1 promoter and improve the recognition memory while decreasing the amyloid accumulation in AD mice. Evidence has also demonstrated the therapeutic role of exosome in other neurodegenerative diseases, including PD, ALS, and HD. For example, Prodromos et al100 conducted a non-controlled, single-arm study to observe the effect of AlloEx Exosome@ treatment on the symptoms in 18 patients which diagnosed with ALS, Kennedy disease, congenital myasthenic syndrome, or Lewy body dementia. Eighteen patients received a total of 32 AlloEx Exosome@ treatments. The findings of the study demonstrated that AlloEx Exosome@ treatment resulted in improvements in the clinical and strength parameters of the patients, with no adverse events occurring. It has been demonstrated that the intranasal administration of AlloEx Exosomes is preliminary evidence of safety and possible efficacy requiring confirmation in controlled trials, including ALS, Kennedy disease, congenital myasthenic syndrome, and Lewy body dementia. To observe the effect of heterochronic parabiosis on HD, Lee et al101 surgically connected the R6/2 mice with both young wild-type mice and old wild-type mice. They then built an HD cell model by differentiating neural stem cells. The data demonstrated that the symptoms of HD including mutant Huntingtin aggregation and cognition were improved. This evidence suggests that the therapeutic effect of exosomes in HD can be achieved through the heterochronic parabiosis method. The recent therapeutic applications of exosomes in neurodegenerative diseases, including AD, PD, HD, and ALS were summarized in Table 3.

Table 3.

The Summary of the Recent Therapeutic Application of Exosomes in Neurodegenerative Diseases

Disease Type	Source	Isolation Method		Modification		Reference
AD	Microglia	Ultracentrifugation		miR-124-3p modified		[97]
AD	hADSC	/		Low-intensity ultrasound		[98]
AD	Regulatory T cells	Ultracentrifugation		β-secretase responsive and β-amyloid targeted		[102]
AD	HEK-293T cells	Ultracentrifugation		Encapsulated VU0155069		[103]
AD	MSCs	ExoQuick-TC kit		Curcumin modified		[104]
AD	BMSCs	Ultracentrifugation		miR-214-3p and CD151 modified		[105]
AD	hUC-MSCs	Ultracentrifugation		BACE1 siRNA and berberine modified		[106]
AD	Blood	Ultracentrifugation		/		[107]
AD	HEK293T cells	Tangential flow filtration		mMaple3 modified		[99]
AD	SH-WT cells	Ultracentrifugation		miR-124-modified		[108]
AD	Dendritic cells	Polymer precipitation		Cholecystokinin and somatostatin modified		[109]
AD	Macrophage	Ultracentrifugation		Curcumin and methylene blue modified		[110]
AD	Microglia	Ultracentrifugation		/		[111]
PD	Microglia	ExoQuick-TC kit		IL-17A modified		[112]
PD	Neural stem cells	Ultrafiltration		/		[113]
PD	OM-MSCs	Ultracentrifugation		/		[114]
PD	BMMSCs	Ultracentrifugation		/		[115]
PD	hASCs	Ultracentrifugation		/		[116]
PD	hADSC	Ultracentrifugation		/		[117]
PD	Umbilical cord blood	Ultracentrifugation		/		[118]
PD	hUCMSCs	Exo-spin kit		Baicalein and oleuropein modified		[119]
PD	MSCs	Ultracentrifugation		/		[120]
PD	MSCs	Ultracentrifugation		FTO-targeted siRNAs modified		[121]
ALS	Cerebrospinal fluid	Total exosome isolation kit		/		[122]
ALS	hUCMSCs	/		/		[100]
HD	HD cells	Exo-quick exosomeprecipitation solution		/		[101]
Animal Model/Cell Model	Sample Size Per Group	Dose and Administration Schedule	Therapeutic Effect	Mechanism	Key Safety Observations	Evidence Level	Reference
AD mouse	n=10	Intravenous administration	Improved cognitive function	Regulating MEKK3/NF-κB	Not reported	Preclinical	[97]
APP/PS1 mice	n=6	Intranasal administration, every 3 days for 2 months	Improved cognition and neurogenesis	Regulating Fos/Kcnj13	Not reported	Preclinical	[98]
3xTg-AD mice	n=6	/	Decreased β-amyloid accumulation and inflammation	/	Not reported	Preclinical	[102]
CD-1 mice	n=3	Intravenous administration, at a dosage of 1mg/kg	Improved the cognitive function	/	Not reported	Preclinical	[103]
STZ-induced rat	n=8	Intraperitoneal injection, at a dosage of 30μg/body	Improved the memory and learning skill	/	Not reported	Preclinical	[104]
Pentobarbital sodium+ Aβ1–42 induced AD rat	n=8	Intravenous administration, at a dose of 100μL	Decreasing neuronal apoptosis	Regulating miR-214-3p/CD151	Not reported	Preclinical	[105]
5xFAD mice	n=6	Intranasal administration, at a dose of 1 mg/mL exosomes every 3 weeks for 4 weeks	Decreasing β-amyloid deposition and inflammation	/	Not reported	Preclinical	[106]
APP/PS1 mice	n=6	/	Improved cognitive function	Regulating Nsmaf, Gnai3, and Akt3	Not reported	Preclinical	[107]
5xFAD and 3xTg-AD mice	n=3	Intravenous injection	Improved recognition memory	/	Not reported	Preclinical	[99]
SH-WT/SH-SWE neuroblastoma cells, microglia, and IM-HA astrocytes	n=3	/	Improved neurodegeneration	/	Not reported	Preclinical	[108]
Human neuroblastoma SH-SY5Y cells and BALB/c mice	n=3	Intravenous injection, at a dosage of 2.5×1010/250 µL	Decreasing β-amyloid production	/	Not reported	Preclinical	[109]
Okadaic acid induced AD mice	n=3	Intraperitoneal injection, at a dosage of 150μL per day for 7 days	Improved cognitive dysfunction	Regulating AKT/GSK-3β	Not reported	Preclinical	[110]
Aβ1-42 induced mice	n=6	Intravenous injection, at a dosage of 1µg/µL per week for 2 months	Alleviated inflammation and oxidative stress	Regulating Wnt/β-catenin	Not reported	Preclinical	[111]
MPTP-induce mice	n=6	Intravenous injection, at a dosage of 200µg	Improved neurodegeneration and motor impairment	/	Not reported	Preclinical	[112]
MPTP-induce mice	n=6	Intravenous injection, at a dosage of 7.32×1010 particles/mL) on days 3, 17, and 31	Alleviated vascular injury	/	Not reported	Preclinical	[113]
MPTP-induce mice	n=6	Stereotactic injection, at a dosage of 1.4×108 nanoparticles per week for 6 weeks	Ameliorated oxidative stress	Regulating TP53INP1/IGF2BP1	Not reported	Preclinical	[114]
Rotenone-induced rat	n=10	Intravenous administration, at a dose of 1mL per week for 3 weeks	Improved motor function	/	Not reported	Preclinical	[115]
MitoPark mice	n=6	Intravenous administration, at a dose of 0.1–1μg per month for 3 months	Improved neuroinflammation	/	Not reported	Preclinical	[116]
Rotenone-induced rat	n=5	Intravenous administration, at a dose of 500µg/kg per three day for 12 days	Promoted neuronal function	Regulating MAPK	Not reported	Preclinical	[117]
MPTP-induce mice	n=5	Intravenous administration, at a dose of 20μg every other day for 14 days	Ameliorated motor dysfunction	Regulating MAPK p38 and ERK 1/2	Not reported	Preclinical	[118]
SHSY5Y cells and hCMEC/D3 cells	n=3	/	Reduced α-synuclein pathogenicity	/	Not reported	Preclinical	[119]
MPTP-induce mice	n=7	Intraperitoneal injection, at a dosage of (2.44 × 1010, 7.32 × 1010, 1.22 × 1011 particles/mL twice a week for 5 weeks	Ameliorated motor deficits	Regulating Nox4/ROS/Nrf2	Not reported	Preclinical	[120]
MPTP-induce mice	n=3	/	Alleviated dopaminergic neuronal death	/	/	/	[121]
G93A (A) and ALS111 mice	n=9	Stereotactic injection, at a dose of 7×107 and 7×108	Improved the activity of neuron	/	Not reported	Preclinical	[122]
/	n=1-12	Intranasal administration, received a single dose (1.25×) and a total dose of 2.25×(2.5×) of AlloEx Exosomes	Improved cognitive dysfunction	/	Not reported	Clinical case series	[100]
R6/2 mice and neural stem cells	n=6	/	Improved mitochondria dysfunction	/	/	/	[101]

Note: “/” represents that data not reported in the original study.

Abbreviations: AD, Alzheimer’s Disease; APP, Amyloid precursor protein; PS1, presenilin-1; MPTP, 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine; MEKK3, mitogen-activated protein kinase/extracellular signal-regulated kinase kinase kinase 3; NF-κB, Nuclear factor-κB; hADSCs, human adipose derived stem cells; Fos, proto-oncogene, AP-1 transcription factor subunit; Kcnj13, potassium inwardly rectifying channel subfamily J member 13; HEK-293T cells, human embryonic kidney 293T cells; CD151, cluster of differentiation 151; BMSCs, bone marrow mesenchymal stem cells; hUCMSCs, human umbilical cord mesenchymal stem cells; BACE1, Beta-site amyloid precursor protein cleaving enzyme 1; Nsmaf, neutral sphingomyelinase activation associated factor; Gnai3, G protein subunit alpha i3; AKT, serine/threonine kinase 3; IL-17A,interleukin-17A; OM-MSCs, olfactory mucosa MSCs; TP53INP1, tumor protein 53-induced nuclear protein 1; IGF2BP1, insulin like growth factor 2 mRNA binding protein 1; MAPK p38, P38 mitogen-activated protein kinase; ERK 1/2, Extracellular signal-regulated kinase 1/2; NOX4, NADPH oxidase 4; ROS, reactive oxygen species; Nrf2, nuclear factor erythroid 2-related factor 2.

Several additional challenges must be overcome before exosome-based therapies can be clinically translated. These include the absence of a universally accepted product definition, variation between batches, incomplete characterization of active and unwanted cargo, insufficiently validated potency assays, uncertain biodistribution, limited exposure to the brain after systemic administration, and inadequate long-term safety data. Furthermore, surface engineering or therapeutic cargo loading may alter the biological behavior, immunogenicity, and regulatory classification of the final product.

The Application of Exosomes in the Pathogenesis of Neurodegenerative Diseases

Exosomes are a specific type of extracellular vesicle characterised by their unique structure and function. They have been shown to play a dual role in neurodegenerative diseases. On the one hand, they have great potential as diagnostic and therapeutic tools for neurodegenerative diseases. However, the transportation of pathological proteins by exosomes, including β-amyloid, α-synuclein, and hyperphosphorylated tau protein, can facilitate the propagation of neurodegenerative diseases by spreading between neurons in the brain and accelerating the pathological diffusion of neurodegenerative diseases, becoming an exacerbating factor in the progression of neurodegenerative diseases. AD is a neurodegenerative disease closely related to the presence of β-amyloid plaques and tau protein neurofibrillary tangles in the brain. Evidence suggests that exosome samples isolated from β-amyloid plaques have been found to express flotilin-1 and Alix. Furthermore, analysis has revealed that the β-amyloid peptide derived from exosome samples is implicated in the formation of amyloid plaques. Alix, a derivative of exosome, has been demonstrated to stimulate the secretion of exosomes by amyloid-β plaques.123 Further research has revealed the presence of exosome-associated β-amyloid in SH-SY5Y cells derived from cerebral fluid in PD patients. Moreover, amyloid-β can be transmitted through exosomes, inhibiting the generation of exosomes and reducing the pathogenesis of AD.124,125 To further elucidate the role of exosome release and autophagy in the neuroprotective effects of acacetin, Nilufer et al126 treated a BMMSC model induced by NH₄OH-soluble Aβ_1-42 peptide with acacetin. The results showed that the NH₄OH-soluble Aβ_1-42 peptide increased the expression of Alix protein in BMMSCs, and acacetin could decrease the expression of Alix protein in NH₄OH-soluble Aβ_1-42 peptide-induced BMMSCs. These data demonstrated that acacetin has an effect on the regulation of the exosome release in BMMSCs. This suggests that exosomes play a role in the regulation of the pathogenesis mechanism of AD. To understanding the effect of complement receptor 3 on the progression of PD, Ma et al127 firstly subjected microglia to lipopolysaccharide (LPS) treatment. The data found that LPS treatment increased the exosome secretion by microglia. The inhibition of the expression of complement receptor 3 resulted in a decrease in the effect of LPS on exosome release in microglia. Furthermore, they identified that the complement receptor 3-NOX2 axis participated in the process of the effect of LPS on exosome release in microglia. Similarly, Citron et al128 investigated the effect of treadmill exercise on PD rats. The data demonstrated that this type of treadmill exercise alleviated the neurodegeneration in PD rats, though no effect on Lewy body-like inclusion burden was observed. Further RNA sequencing analysis demonstrated that treadmill exercise could modify the exosome release and exosome content in PD rats. Following analysis, 27 RNA molecules were found to be expressed at higher levels in the exosome sample. Of these, three mRNAs were identified as having the potential to enhance neuroprotection in PD rats (Figure 4).

Figure 4.

The summary of the application of exosomes in the pathogenesis of neurodegenerative diseases. The figure representative illustrates exosomes participated in amyloid-β plaques-mediated pathogenesis of AD, the neuroprotective effects of acacetin in PD, and the treatment of treadmill exercise in PD. Blue arrow represents specific molecules, protein markers, or exosomes. Blue flat-headed arrow represents an enlarged exosome structure. Blue dotted line represents a relatively independent occurrence process.

Abbreviations: Alix, ALG-2-interacting protein; BMMSCs, bone marrow mesenchymal stem cells; Aβ, amyloid β-protein.

Conclusion and Perspectives

Neurodegenerative diseases are chronic and progressive neurological disorders caused by the degeneration of neurons. Epidemiologically, these diseases exhibit a high incidence rate, rapid progression, and impose a substantial disease burden, resulting in significant economic costs for both patients and society. Exosomes possess distinctive properties-such as their inherent ability to traverse biological barriers, high biocompatibility, low immunogenicity, intrinsic immunoregulatory functions, and a natural capacity for drug delivery vehicles-that confer unique advantages for their application in neurodegenerative disease. However, several challenges hinder the clinical translation of exosome-based therapies for these conditions. (1) The production and purification of exosomes should be carried out in accordance with the established protocols to ensure consistency and quality control. Exosomes are a type of nanoscale cellular vesicles that contain DNAs, RNAs, and lipids. There are various surface markers for identifying exosomes. This results in heterogeneity in size, molecular content, and functional properties across different exosome batches. Currently, ultracentrifugation remains the predominant method for vesicle isolation; however, this technique poses difficulties in achieving large-scale, stable production. The inherent heterogeneity of exosomes, coupled with the limitations of existing isolation methods, restricts the scalability and compliance with Good Manufacturing Practice (GMP) standards necessary for clinical-grade exosome production. Emerging approaches, such as the use of microcarriers for three-dimensional cell culture and tangential flow filtration (TFF) technology, offer potential solutions by enabling real-time monitoring of critical steps in cell culture and product purification, thereby facilitating the amplification of bioactive exosome yields. (2) The natural low immunogenicity is the reason why exosomes are considered ideal carriers. However, this’ low immunogenicity still poses certain risks in clinical applications. Repeated administration of exosomes may trigger immune reactions or activate the complement system, which poses a significant safety risk. The immunogenicity of exosomes varies depending on the cell source. Pollutants (such as impurity proteins and endotoxins) introduced during the isolation and purification process may also trigger immune reactions. Specifically, data on the long-term safety of exosomes is still limited. Delayed neuroinflammation, ectopic gene regulation, unintended effects on tumor-related pathways, and accumulation in non-target organs are the important safety concerns. To solve these problems, cell sources with low immunogenicity should be prioritized, purification processes should be optimized, the surface of the exosomes should be modified and immune tolerance should be induced. (3) Precise delivery of exosomes to brain lesions is a key to the treatment of neurodegenerative diseases. In fact, natural exosomes lack specific targeting ability and are easily intercepted by organs such as the liver, spleen, and lungs after entering the body. This results in a very low number of exosomes reaching the brain. The presence of the blood-brain barrier (BBB) also significantly reduces the efficiency with which exosomes reach the site of brain lesions. One potential solution to this issue is the modification of ligands on the surface of exosomes that can specifically recognize brain targets and achieve active targeting. Simultaneously, optimizing delivery routes is a key priority. This can be achieved through non-invasive “nose-to-brain” direct delivery using nasal administration, or through intrathecal/intraventricular administration, which can directly inject extracellular vesicles into the cerebrospinal fluid, bypassing the BBB. This approach has the potential to increase the number of extracellular vesicles in the brain, leading to effective treatment of neurodegenerative diseases. (4) The limitations of the targeting of exosomes and the uncertainty of their distribution in vivo. Following natural exosomes enter the human body, the accurate location of the lesion site and the subsequent exertion of their effects by the exosomes is crucial for maximising their therapeutic effectiveness. Evidence has demonstrated that the majority of exosomes in the body are rapidly cleared by the reticuloendothelial system, and are unable to brain accurately. While engineering modifications can significantly enhance the targeting of exosomes, but targeted modifications will increase the complexity of the production procedures and product measurements. Genetic engineering and chemical modification may be ideal solutions to this problem. Specific genetic modifications carried out on the mother cell result in the secreted exosomes naturally carrying specific navigation molecules to reach the lesion site. And chemical modification can then be used to “post-process” the isolated and purified exosomes. This involves the use of chemical reactions to connect navigation molecules to the surface of the exosomes, achieving precise targeting of the target cells. (5) The limitations of drug loading and drug stability in exosomes. Exosomes are a type of nanoscale extracellular vesicles that are highly compatible and can be used to encapsulate various active ingredients for drug delivery. One of the key challenges in the effective application of drugs is the efficient loading of drugs into exosomes without compromising their integrity during storage and transportation. In addition, a technical challenge lies in the use of EXDOs for the loading of hydrophilic macromolecular drugs. Currently, a variety of innovative fusion technologies, including fusion, hybridization, and light control, can be used to load a range of drugs efficiently, from small molecules to large molecules. These technologies could be the ideal solution to the aforementioned problems. (6) The limitation of being lack of large-scale clinical validation and standardization. Currently, most clinical trials on the application of exosomes are small-scale, retrospective studies, making it difficult to verify their true clinical values. Different studies use different combinations of biomarkers and detection thresholds, lacking a unified clinical decision-making standard. Furthermore, regulatory classification may vary depending on the cell source, manufacturing process, therapeutic cargo, and intended use. Exosome products may be regulated as biological products, cell-derived products, gene therapy-related products, drug-delivery systems, or combination products. This regulatory uncertainty complicates clinical development and emphasizes the need for product-specific criteria regarding identity, purity, potency, sterility, safety, and release testing. Conducting rigorously designed prospective multicenter clinical studies, synchronously developing data interpretation standards and clinical decision thresholds, may help to promote the application of exosomes in the treatment of neurodegenerative diseases.

Future studies should therefore prioritize standardized reporting, qualified cell banks, GMP-compatible manufacturing, validated potency assays, quantitative pharmacokinetic and biodistribution studies, appropriate disease controls, prospective multicenter biomarker validation, randomized controlled trials, and long-term post-treatment surveillance. These requirements are essential for distinguishing promising experimental findings from clinically meaningful and reproducible benefits.

It should be noted that not all of the primary diagnostic studies included in the summary reported complete diagnostic accuracy metrics (eg, AUC, sensitivity, or specificity); where such data are unavailable, they are indicated as “/” in the table This reflects a limitation of the published literature and underscores the need for more standardized reporting of diagnostic performance parameters in future studies.

Exosomes have demonstrated significant potential for application in the field of neurodegenerative diseases, encompassing their role in pathological mechanisms, the identification of diagnostic biomarkers, and the development of therapeutic interventions. With the resolution of the following issues, including the production and purification of exosomes, the targeting of exosomes and the uncertainty of their distribution in vivo, drug loading and drug stability of exosomes, being lacking in large-scale clinical validation and standardization, will accelerate the application of exosomes in neurodegenerative diseases.

Data Sharing Statement

The original contributions presented in the study were included in the article, and further inquiries could be directed to the corresponding author.

Disclosure

The authors report no conflicts of interest in this work.