A review of the roles of exosomes in salivary gland diseases with an emphasis on primary Sjögren's syndrome
The salivary glands play a crucial role in producing and secreting saliva, which is vital for maintaining both oral and overall health. Salivary gland diseases represent a diverse group of conditions, including salivary gland dysfunction, obstructive diseases, inflammatory conditions, and neoplastic lesions. Among these, salivary gland dysfunction is commonly caused by primary Sjögren's Syndrome (pSS), radiotherapy for head and neck cancer, diabetes, menopause, and the use of certain medications. Other etiologies, such as those underlying sialolithiasis and certain neoplasms, also contribute to significant diagnostic and therapeutic challenges. These disorders significantly increase the risk of oral infections, dental caries, and xerostomia, ultimately reducing the patients' quality of life. Consequently, there is a pressing need for the development of new diagnostic and therapeutic strategies. One promising approach to addressing these challenges is through the study of exosomes—small extracellular vesicles (EVs) that transport bioactive molecules including proteins, lipids, DNA, mRNA, microRNA (miRNA), and long non-coding RNA. , , , , , , The lipid bilayer of exosomes protects their cargo from proteolytic degradation, enhancing stability and detectability. , , Consequently, research has shown that measuring exosomal proteins and RNA in plasma or saliva provides a more accurate representation of the physiological and pathological states of the salivary glands, positioning exosomes as promising non-invasive biomarkers for the early detection of salivary gland dysfunction. Exosomes also play a crucial role in intercellular communication by facilitating the transfer of bioactive molecules between cells. , Exosomes also offer therapeutic potential by selectively delivering bioactive molecules to target cells through ligand–receptor interactions. , , , , Exosomes derived from mesenchymal stem cells (MSCs) have garnered particular interest due to their low immunogenicity, stability, and manipulability, emerging as powerful therapeutic tools across a range of diseases. , , , , , , Studies have demonstrated that exosomes can suppress inflammation and promote tissue repair in salivary gland diseases, further underscoring their therapeutic potential. Despite the progress in understanding exosomes, no comprehensive review has yet synthesized their diagnostic and therapeutic roles in salivary gland diseases. This article seeks to address that gap by providing an in-depth review of the current research on the applications of exosomes in these conditions, highlighting their potential as both diagnostic and therapeutic tools. Narrative methodology and characteristics of selected studies The search strategy encompassed PubMed, Ovid MEDLINE, Embase, Web of Science, and CENTRAL databases. Our search involved the utilization of various keywords including ‘exosome,’ ‘extracellular vesicle,’ ‘salivary gland,’ ‘parotid,’ ‘submandibular,’ ‘sublingual,’ ‘hyposalivation,’ and ‘xerostomia,’ both individually and in combination, restricted to English-language publications. The final search was conducted in August 2024. The review adhered to the Preferred Reporting Items for Systematic Reviews and Meta-analyses (PRISMA) 2020 guideline ( ). A thorough review was carried out, which included research on exosome and/or EV, covering in vivo and in vitro experiments, as well as clinical trials. Our focus was to explore the potential of exosomes in the diagnosis and treatment of salivary gland diseases. XC and LL conducted individual screenings of the records by examining titles, abstracts, and complete texts. Subsequently, XC, CD, and SM extracted the data. Any inconsistencies in evaluations were resolved through constructive discussions with YC. Data collection was conducted using predesigned data extraction tables, and statistical analysis was performed in R (version 4.1.0). The bibliometrix package (version 4.1.2) was used for bibliometric analysis, following its established protocols. We retrieved a total of 809 records. After removing duplicates, 772 unique records remained. The titles and abstracts of these 772 records were screened, and 723 were excluded. Full-text analysis was conducted on the remaining 49 records, of which 31 studies were selected for inclusion in the final review ( ). The bibliometric analysis of the 31 original research articles highlights key aspects of the research landscape on salivary gland diseases. The articles, published between 2005 and 2024, show increased output after 2017, driven by advancements in molecular techniques and exosome research ( A). Kapsogeorgou EK (2005) and Aqrawi LA (2017) are the most cited studies, providing essential insights into pSS and EVs, shaping subsequent research in the field ( B). Keyword analysis reveals a focus on “Sjögren's syndrome,” “salivary glands,” and “extracellular vesicles,” with growing interest in mesenchymal stem cell-derived exosomes for PSS treatment, aligning with the focus of this review ( C). This analysis provides a concise overview of key contributors and emerging trends, offering a foundation for future research. The biogenesis of exosomes Exosomes were first identified as membrane vesicles with 5′-nucleotidase activity. Unlike ectosomes, which bud from the plasma membrane, exosomes originate from the endosomal system through the inward invagination of the endosomal membrane, forming multivesicular bodies (MVBs). These MVBs contain intraluminal vesicles that can either undergo lysosomal degradation or fuse with the plasma membrane to release exosomes, typically ranging from 40 to 160 nm in diameter. The biogenesis of exosomes is regulated by several molecular pathways, with the ESCRT (endosomal sorting complexes required for transport) machinery playing a primary role. , , However, ESCRT-independent mechanisms, such as ceramide production by neutral type II sphingomyelinase and the involvement of tetraspanins, are also crucial in endosomal sorting and exosome release. Rab GTPases, such as Rab27a and Rab27 b, further facilitate this process by regulating the transport of MVBs to the plasma membrane, ensuring proper vesicle formation, cargo sorting, and secretion. The salivary gland serves both as a source of exosome secretion and as a recipient for exosomes from other tissues. Salivary epithelial and mesenchymal cells can secrete exosomes, which interact with receptor cell membranes via surface molecules and homologous ligands, enabling their entry and subsequent regulation of the recipient cells' biological functions. , The first detection of exosome secretion by non-tumor salivary epithelial cells occurred in 2005, where exosomes containing ribonucleoproteins were identified. Later studies using Multidimensional Protein Identification Technology identified numerous proteins in parotid gland-derived exosomes, involved in signal transduction, immune responses, cell adhesion, and antigen presentation. Furthermore, salivary gland mesenchymal cells secrete exosomes containing miRNA-133 b-3p, which are transferred to the salivary epithelium. This process downregulates DIP2B expression, influencing the proliferation of epithelial progenitor cells and contributing to salivary gland morphogenesis. Salivary epithelial cells can also act as recipient cells for exosomes from other tissues. For instance, breast cancer-derived exosome-like microvesicles have been shown to interact with salivary gland epithelial cells (SGECs), leading to changes in the protein and mRNA composition of the exosomes they secrete. The salivary gland serves a dual role, both secreting exosomes that regulate recipient cells and acting as a target for exosomes from other tissues, influencing its expression and metabolism. This bidirectional interaction highlights the vital role of exosomes in mediating cellular communication. Given their regulatory functions, exosomes present significant potential for understanding, diagnosing, and developing treatments for salivary gland diseases, offering promising clinical applications. 

The search strategy encompassed PubMed, Ovid MEDLINE, Embase, Web of Science, and CENTRAL databases. Our search involved the utilization of various keywords including ‘exosome,’ ‘extracellular vesicle,’ ‘salivary gland,’ ‘parotid,’ ‘submandibular,’ ‘sublingual,’ ‘hyposalivation,’ and ‘xerostomia,’ both individually and in combination, restricted to English-language publications. The final search was conducted in August 2024. The review adhered to the Preferred Reporting Items for Systematic Reviews and Meta-analyses (PRISMA) 2020 guideline ( ). A thorough review was carried out, which included research on exosome and/or EV, covering in vivo and in vitro experiments, as well as clinical trials. Our focus was to explore the potential of exosomes in the diagnosis and treatment of salivary gland diseases. XC and LL conducted individual screenings of the records by examining titles, abstracts, and complete texts. Subsequently, XC, CD, and SM extracted the data. Any inconsistencies in evaluations were resolved through constructive discussions with YC. Data collection was conducted using predesigned data extraction tables, and statistical analysis was performed in R (version 4.1.0). The bibliometrix package (version 4.1.2) was used for bibliometric analysis, following its established protocols. We retrieved a total of 809 records. After removing duplicates, 772 unique records remained. The titles and abstracts of these 772 records were screened, and 723 were excluded. Full-text analysis was conducted on the remaining 49 records, of which 31 studies were selected for inclusion in the final review ( ). The bibliometric analysis of the 31 original research articles highlights key aspects of the research landscape on salivary gland diseases. The articles, published between 2005 and 2024, show increased output after 2017, driven by advancements in molecular techniques and exosome research ( A). Kapsogeorgou EK (2005) and Aqrawi LA (2017) are the most cited studies, providing essential insights into pSS and EVs, shaping subsequent research in the field ( B). Keyword analysis reveals a focus on “Sjögren's syndrome,” “salivary glands,” and “extracellular vesicles,” with growing interest in mesenchymal stem cell-derived exosomes for PSS treatment, aligning with the focus of this review ( C). This analysis provides a concise overview of key contributors and emerging trends, offering a foundation for future research. 

Exosomes were first identified as membrane vesicles with 5′-nucleotidase activity. Unlike ectosomes, which bud from the plasma membrane, exosomes originate from the endosomal system through the inward invagination of the endosomal membrane, forming multivesicular bodies (MVBs). These MVBs contain intraluminal vesicles that can either undergo lysosomal degradation or fuse with the plasma membrane to release exosomes, typically ranging from 40 to 160 nm in diameter. The biogenesis of exosomes is regulated by several molecular pathways, with the ESCRT (endosomal sorting complexes required for transport) machinery playing a primary role. , , However, ESCRT-independent mechanisms, such as ceramide production by neutral type II sphingomyelinase and the involvement of tetraspanins, are also crucial in endosomal sorting and exosome release. Rab GTPases, such as Rab27a and Rab27 b, further facilitate this process by regulating the transport of MVBs to the plasma membrane, ensuring proper vesicle formation, cargo sorting, and secretion. The salivary gland serves both as a source of exosome secretion and as a recipient for exosomes from other tissues. Salivary epithelial and mesenchymal cells can secrete exosomes, which interact with receptor cell membranes via surface molecules and homologous ligands, enabling their entry and subsequent regulation of the recipient cells' biological functions. , The first detection of exosome secretion by non-tumor salivary epithelial cells occurred in 2005, where exosomes containing ribonucleoproteins were identified. Later studies using Multidimensional Protein Identification Technology identified numerous proteins in parotid gland-derived exosomes, involved in signal transduction, immune responses, cell adhesion, and antigen presentation. Furthermore, salivary gland mesenchymal cells secrete exosomes containing miRNA-133 b-3p, which are transferred to the salivary epithelium. This process downregulates DIP2B expression, influencing the proliferation of epithelial progenitor cells and contributing to salivary gland morphogenesis. Salivary epithelial cells can also act as recipient cells for exosomes from other tissues. For instance, breast cancer-derived exosome-like microvesicles have been shown to interact with salivary gland epithelial cells (SGECs), leading to changes in the protein and mRNA composition of the exosomes they secrete. The salivary gland serves a dual role, both secreting exosomes that regulate recipient cells and acting as a target for exosomes from other tissues, influencing its expression and metabolism. This bidirectional interaction highlights the vital role of exosomes in mediating cellular communication. Given their regulatory functions, exosomes present significant potential for understanding, diagnosing, and developing treatments for salivary gland diseases, offering promising clinical applications. 

Primary Sjögren's syndrome pSS is a chronic autoimmune disease characterized by lymphocytic infiltration of the salivary and lachrymal glands, leading to xerostomia and xerophthalmia. Although the exact etiology of pSS remains unclear, recent studies focusing on exosomes and EVs have offered new insights into its underlying pathological mechanisms, particularly in the context of intercellular communication. These findings not only enhance our understanding of disease progression but also provide a foundation for potential advancements in diagnosis and therapeutic strategies ( ). Pathogenesis of primary Sjögren's syndrome Previous research suggests that environmental triggers, such as Epstein–Barr virus (EBV) infection, may promote the release of Ro/SSA and La/SSB ribonucleoproteins (RNPs), thereby activating the innate immune system and the production of interferons, which play a crucial role in the onset of the disease. Further studies have identified elevated levels of EBV-specific microRNA (EBV-miRNA-BART13-3p) in the B cells and SGECs of pSS patients. This miRNA can be transferred from B cells to SGECs via exosomes, where it inhibits the expression of crucial proteins like stromal interacting molecule 1 (STIM1) and aquaporin 5 (AQP5). This inhibition may result in the loss of store-operated Ca2+ entry (SOCE) and reduced activation of nuclear factor of activated T cells (NFAT), which ultimately contributes to impaired saliva secretion ( ). Additionally, exosomes secreted by activated T cells in pSS patients contain miRNA-142-3p, which can be transferred to glandular cells. This miRNA targets key intracellular signaling elements—such as sarco(endo)plasmic reticulum Ca2+ ATPase 2 b (SERCA2B), ryanodine receptor 2 (RyR2), and adenylate cyclase 9 (AC9)—disrupting Ca2+ signaling and cAMP production. The resulting decrease in protein production in SGECs may further contribute to epithelial dysfunction. These findings suggest that exosome-mediated transfer of miRNAs from immune cells to epithelial cells plays a pivotal role in pSS pathogenesis by impairing salivary gland function. Given the understanding of these mechanisms, recent research has turned towards the development of exosome-based biomarkers for the early diagnosis of pSS, driven by the need for more accurate and non-invasive. Diagnosis of primary Sjögren's syndrome Recent studies have highlighted the potential of exosome-derived biomarkers in the early diagnosis of pSS, addressing limitations in conventional diagnostic methods. Exosomes, with their protective lipid bilayer, ensure the stability of their contents, such as miRNAs and proteins, making them promising candidates for biomarkers. In 2005, Kapsogeorgou et al. discovered that exosomes secreted by SGECs in pSS patients contain autoantigenic Ro/SSA, La/SSB, and Sm RNPs, as well as epithelial-specific cytokeratins. This study identified intracellular RNPs in exosomes from SGECs, providing new insights into the potential role of exosomes in antigen presentation mechanisms within the immune system. Subsequent studies revealed the upregulation of proteins involved in innate immunity (LCN2), cell signaling (CALM), and wound repair (GRN and CALML5) in the saliva of pSS patients, suggesting a distinct exosomal protein signature associated with the disease. Additionally, elevated levels of exosomal circular RNAs (circ-IQGAP2 and circ-ZC3H6) have been found in both minor salivary glands and serum samples from pSS patients. Studies using the Non-Obese Diabetic (NOD) mouse model, a widely accepted model for pSS, also identified five upregulated miRNAs (miRNA-127-3p, miRNA-409-3p, miRNA-410-3p, miRNA-541-5p, and miRNA-540-5p) in serum exosomes. Furthermore, a comparison of the EV-enriched saliva sub-proteome of pSS patients with the whole saliva proteome revealed 121 differentially expressed proteins. This discovery underscores the potential of exosome-based biomarkers to provide a comprehensive molecular profile of pSS. Emerging studies have also linked exosome-derived proteins to novel pathological mechanisms in pSS. For instance, differentially expressed proteins related to ferroptosis, such as ceruloplasmin and transferrin, were found to be downregulated in exosomes from the plasma of pSS patients. This was the first study to establish a connection between ferroptosis and epithelial cell damage in pSS, offering new perspectives on disease mechanisms and potential therapeutic targets. In summary, exosome-based biomarkers, including proteins, circular RNAs, and miRNAs, show great promise for the early detection of pSS. These developments could lead to more timely and accurate diagnoses, ultimately improving patient outcomes. Treatment of primary Sjögren's syndrome The treatment of pSS remains a significant challenge, as current therapies are primarily focused on symptom management. Although B-cell-targeted therapies, such as rituximab and belimumab, have been introduced in recent years, their non-selective depletion of B cells offers limited improvement in managing the dry symptoms associated with pSS. At present, mesenchymal stem cell-derived exosomes (MSCs-exos) have shown promise in mimicking the immunomodulatory and tissue repair functions of MSCs. Recent research underscores the critical role of specific miRNAs encapsulated within exosomes or EVs in treating pSS. For example, exosomes derived from early-passage induced pluripotent stem cells-derived MSCs (iPSC-MSCs) are more effective in suppressing inflammatory responses and regulating immune function compared to late-passage exosomes. This effect is primarily mediated by molecules such as TGF-β1, miRNA-21, and miRNA-125 b. And exosomes derived from lip salivary gland MSCs (LGMSC-exos) contain miRNA-125 b, which targets PRDM1, a key regulator of plasma cell differentiation. Similarly, miRNA-10a-5p in exosomes from myeloid-derived suppressor cells, which effectively inhibits B cell proliferation by targeting Bcl-6. Further emphasizing the therapeutic potential of miRNA-loaded exosomes in modulating immune responses, miRNA-10a-5p within exosomes significantly impacts autoimmune conditions like pSS by altering critical immune cell functions. Additionally, miRNAs in olfactory ecto-MSCs-derived exosomes influence T follicular helper cell responses, adding another dimension to miRNA-mediated regulation in pSS. Exosomes derived from bone marrow MSCs and iPSCs have been shown to inhibit Th17 cell differentiation while promoting Treg cell expansion, resulting in reduced lymphocytic infiltration in salivary glands and decreased serum autoantibody levels. Similar effects were observed with exosomes from labial gland-derived MSCs, which play a key role in regulating the Treg/Th17 balance, crucial for controlling inflammatory responses in pSS. Further studies demonstrated that exosomes from human umbilical cord MSCs exhibit immunosuppressive properties, reducing inflammation by modulating the Treg/Th17 balance and improving gut microbiota composition. , These findings collectively highlight exosomes' capacity to alleviate pSS symptoms by targeting immune regulatory pathways. In addition to their immunomodulatory effects, exosomes have shown potential in enhancing tissue repair and cellular function. Exosomes from dental pulp stem cells (DPSC-exos) enhance salivary gland epithelial function through the GPER-mediated cAMP/PKA/CREB pathway, underscoring a signaling pathway-driven mechanism. Similarly, that exosomes from human exfoliated deciduous teeth reduce cellular apoptosis and enhance glandular function by suppressing p-ERK1/2-mediated apoptosis. Collectively, exosomes play a critical role in the pathogenesis, diagnosis, and treatment of pSS. Mechanistically, exosomes from EBV-infected B cells and activated T cells transfer key miRNAs, such as EBV-miRNA-BART13-3p and miRNA-142-3p, to salivary gland epithelial cells, disrupting calcium signaling pathways and reducing saliva secretion. These exosomes contribute to glandular dysfunction by downregulating key proteins like STIM1 and AQP5. Diagnostically, exosomes offer non-invasive biomarkers, with molecules such as Ro/SSA, La/SSB, and circular RNAs (e.g., circ-IQGAP2) found in serum, saliva, and tears, facilitating early detection and disease monitoring. In terms of treatment, MSC-exos demonstrate significant potential by modulating immune responses, balancing Treg and Th17 cells, reducing inflammation, and promoting tissue repair. However, challenges such as exosome heterogeneity, limited standardization in isolation methods, and the need for large-scale clinical trials remain in pSS. Addressing these issues will be crucial for translating exosome-based therapies into clinical practice. Future research should focus on refining exosome-based diagnostics and expanding the clinical use of MSC-exos to restore salivary gland function and improve pSS symptoms. Radiation-induced salivary gland dysfunction Exosome-based therapies have shown potential in addressing the underlying causes of salivary gland dysfunction. In particular, radiotherapy-induced xerostomia, a prevalent and severe side effect affecting head and neck cancer survivors, has been a target for innovative therapeutic approaches. For instance, exosomes derived from human dental pulp stem cells have been shown to reduce inflammatory cytokine production and reverse oxidative stress in submandibular gland cells, thereby preventing cellular senescence induced by radiation. Similarly, exosomes from hypoxia-pretreated human urine-derived stem cells have demonstrated the ability to repair radiation-induced salivary gland injuries by activating the Wnt3a/GSK3β pathway. Moreover, exosomes derived from salivary gland organoids assembled through magnetic 3D bioassembly have emerged as a promising strategy for mitigating epithelial damage in irradiated submandibular gland models. Diabetes-related salivary gland dysfunction Diabetes is one of the most common metabolic diseases affecting salivary function. More than half of diabetic patients suffer from xerostomia and salivary gland hypofunction. Research has suggested that circulating exosomal miRNAs may be related to insulin resistance. Further experiments demonstrated that bone marrow MSC-derived exosomes can inhibit diabetic sequelae in the salivary glands and its complications by inhibiting TGFβ and its related pathway via Smad 2 and Smad 3 in streptozotocin-induced diabetic rats. Additionally, research indicated that salivary exosomes reduced blood glucose levels and enhanced salivary gland function in diabetic rats. Menopause-related salivary gland dysfunction Menopause is often accompanied by physiological and functional changes in the oral cavity. Previous studies have indicated that hormone deficiency post-menopause may lead to reduced salivary gland secretion. Currently, there is no standard effective treatment for salivary gland dysfunction occurring after menopause. However, tonsil-MSC-derived EVs alleviated salivary gland dysfunction caused by menopause through anti-inflammatory and anti-fibrosis mechanisms in ovariectomized rats. Further research is needed to identify the specific miRNAs and proteins that play a critical role in post-menopausal salivary gland dysfunction. Sialolithiasis Sialolithiasis, or salivary calculi, is characterized by the mechanical obstruction of the salivary glands' excretory ducts due to the partial deposition of calcific materials. It accounts for one-third of all salivary gland disorders, with the majority of calculi occurring in the submandibular glands. The accumulation of salivary stones can lead to recurrent chronic sialadenitis, as the presence of these stones promotes bacterial aggregation, which can cause inflammation and infection. Although the exact mechanisms behind sialolith formation remain unclear, various theories have been proposed. Some suggests that changes in ion concentration in saliva, the presence of bacteria or foreign bodies in the ducts, or mucous plugs may initiate stone formation. , , The organic core theory posits that the organic components at the center of calcification, such as bacteria, foreign bodies, or desquamated epithelial cells, act as nucleating sites for calculus formation. , In contrast, some theories suggest that stone formation is secondary to sialadenitis, where the swelling during chronic inflammation leads to salivary stasis and the subsequent formation of a calcium-rich core. Some studies have found no organic components in the central area of the calculus core. , However, recent research has provided evidence that exosomes may play a role in the formation and development of sialolithiasis. Using liquid chromatography-mass spectrometry and microscopy, exosomes were confirmed to be present in salivary calculi. Additionally, numerous EVs and degraded cytoplasmic organelles were observed in the central nidus of the stones through transmission electron microscopy. Further examination also revealed the presence of exosomes and bi-layered bacterial cell membranes in the central nidus of a patient with recurrent sialolithiasis. These findings suggest that exosomes might contribute to the pathogenesis of sialolithiasis, although further research is needed to elucidate the exact mechanisms involved. Salivary adenoid cystic carcinoma SACC accounts for approximately 1 % of head and neck tumors and 10 % of salivary gland tumors. Due to its proximity to critical anatomical structures, achieving negative surgical margins during treatment is often difficult. This challenge contributes to a high recurrence rate, with nearly 40 % of patients experiencing local recurrence within five years. Additionally, SACC has a significant risk of distant metastasis, ranging from 8 % to 60 %, resulting in a generally poor long-term prognosis. SACC is primarily characterized by hematogenous metastasis, with the lungs being the most commonly affected site. Exosomes, small vesicles secreted by cancer cells, have been increasingly recognized as playing a crucial role in this metastatic process. , , PROMININ-1 (CD133) is significantly expressed in SACC and is released into saliva through exosomes or EVs. This suggests that exosomes could be involved in both the diagnosis and progression of SACC. Further supporting the role of exosomes in SACC, the upregulation of epithelial regulatory proteins in SACC cells induces epithelial–mesenchymal transition by modulating the GLI1/E-cadherin axis. This process not only increases the expression of pro-angiogenic factors like VEGFA, bFGF, and IL-8 but also facilitates their transfer via exosomes. These exosomes enhance angiogenesis within the tumor microenvironment and increase vascular permeability in the pre-metastatic lung microenvironment, thereby promoting metastasis. Exosomes from SACC-83 cells further enhance these invasive capabilities by increasing endothelial cell permeability through targeting tight junction proteins such as claudin-1, ZO-1, and β-catenin. Notably, SACC cell-derived exosomes contribute to angiogenesis and local vascular microleakage by transporting miRNA-23 b-3p, suggesting that this miRNA could serve as a potential biomarker for distant metastasis in SACC. Current challenges in clinical integration of exosomes for salivary gland diseases Despite their promising potential, several challenges need to be addressed before exosomes can be fully integrated into clinical practice for salivary gland diseases. One major challenge is improving techniques for exosome extraction and purification, particularly from oral fluids or serum, to develop standardized protocols and establish reliable diagnostic standards. Additionally, the heterogeneity of exosomes, due to their diverse origins and cargo, limits their application as biomarkers. Further exploration is needed to understand the mechanisms involved in salivary gland diseases and to identify novel exosome-based biomarkers. Finally, clinical trials involving exosomes in salivary gland diseases remain limited, often relying on small sample sizes. Expanding research to larger-scale population studies is essential for translating basic scientific discoveries into practical clinical applications. 

pSS is a chronic autoimmune disease characterized by lymphocytic infiltration of the salivary and lachrymal glands, leading to xerostomia and xerophthalmia. Although the exact etiology of pSS remains unclear, recent studies focusing on exosomes and EVs have offered new insights into its underlying pathological mechanisms, particularly in the context of intercellular communication. These findings not only enhance our understanding of disease progression but also provide a foundation for potential advancements in diagnosis and therapeutic strategies ( ). 

Previous research suggests that environmental triggers, such as Epstein–Barr virus (EBV) infection, may promote the release of Ro/SSA and La/SSB ribonucleoproteins (RNPs), thereby activating the innate immune system and the production of interferons, which play a crucial role in the onset of the disease. Further studies have identified elevated levels of EBV-specific microRNA (EBV-miRNA-BART13-3p) in the B cells and SGECs of pSS patients. This miRNA can be transferred from B cells to SGECs via exosomes, where it inhibits the expression of crucial proteins like stromal interacting molecule 1 (STIM1) and aquaporin 5 (AQP5). This inhibition may result in the loss of store-operated Ca2+ entry (SOCE) and reduced activation of nuclear factor of activated T cells (NFAT), which ultimately contributes to impaired saliva secretion ( ). Additionally, exosomes secreted by activated T cells in pSS patients contain miRNA-142-3p, which can be transferred to glandular cells. This miRNA targets key intracellular signaling elements—such as sarco(endo)plasmic reticulum Ca2+ ATPase 2 b (SERCA2B), ryanodine receptor 2 (RyR2), and adenylate cyclase 9 (AC9)—disrupting Ca2+ signaling and cAMP production. The resulting decrease in protein production in SGECs may further contribute to epithelial dysfunction. These findings suggest that exosome-mediated transfer of miRNAs from immune cells to epithelial cells plays a pivotal role in pSS pathogenesis by impairing salivary gland function. Given the understanding of these mechanisms, recent research has turned towards the development of exosome-based biomarkers for the early diagnosis of pSS, driven by the need for more accurate and non-invasive. 

Recent studies have highlighted the potential of exosome-derived biomarkers in the early diagnosis of pSS, addressing limitations in conventional diagnostic methods. Exosomes, with their protective lipid bilayer, ensure the stability of their contents, such as miRNAs and proteins, making them promising candidates for biomarkers. In 2005, Kapsogeorgou et al. discovered that exosomes secreted by SGECs in pSS patients contain autoantigenic Ro/SSA, La/SSB, and Sm RNPs, as well as epithelial-specific cytokeratins. This study identified intracellular RNPs in exosomes from SGECs, providing new insights into the potential role of exosomes in antigen presentation mechanisms within the immune system. Subsequent studies revealed the upregulation of proteins involved in innate immunity (LCN2), cell signaling (CALM), and wound repair (GRN and CALML5) in the saliva of pSS patients, suggesting a distinct exosomal protein signature associated with the disease. Additionally, elevated levels of exosomal circular RNAs (circ-IQGAP2 and circ-ZC3H6) have been found in both minor salivary glands and serum samples from pSS patients. Studies using the Non-Obese Diabetic (NOD) mouse model, a widely accepted model for pSS, also identified five upregulated miRNAs (miRNA-127-3p, miRNA-409-3p, miRNA-410-3p, miRNA-541-5p, and miRNA-540-5p) in serum exosomes. Furthermore, a comparison of the EV-enriched saliva sub-proteome of pSS patients with the whole saliva proteome revealed 121 differentially expressed proteins. This discovery underscores the potential of exosome-based biomarkers to provide a comprehensive molecular profile of pSS. Emerging studies have also linked exosome-derived proteins to novel pathological mechanisms in pSS. For instance, differentially expressed proteins related to ferroptosis, such as ceruloplasmin and transferrin, were found to be downregulated in exosomes from the plasma of pSS patients. This was the first study to establish a connection between ferroptosis and epithelial cell damage in pSS, offering new perspectives on disease mechanisms and potential therapeutic targets. In summary, exosome-based biomarkers, including proteins, circular RNAs, and miRNAs, show great promise for the early detection of pSS. These developments could lead to more timely and accurate diagnoses, ultimately improving patient outcomes. 

The treatment of pSS remains a significant challenge, as current therapies are primarily focused on symptom management. Although B-cell-targeted therapies, such as rituximab and belimumab, have been introduced in recent years, their non-selective depletion of B cells offers limited improvement in managing the dry symptoms associated with pSS. At present, mesenchymal stem cell-derived exosomes (MSCs-exos) have shown promise in mimicking the immunomodulatory and tissue repair functions of MSCs. Recent research underscores the critical role of specific miRNAs encapsulated within exosomes or EVs in treating pSS. For example, exosomes derived from early-passage induced pluripotent stem cells-derived MSCs (iPSC-MSCs) are more effective in suppressing inflammatory responses and regulating immune function compared to late-passage exosomes. This effect is primarily mediated by molecules such as TGF-β1, miRNA-21, and miRNA-125 b. And exosomes derived from lip salivary gland MSCs (LGMSC-exos) contain miRNA-125 b, which targets PRDM1, a key regulator of plasma cell differentiation. Similarly, miRNA-10a-5p in exosomes from myeloid-derived suppressor cells, which effectively inhibits B cell proliferation by targeting Bcl-6. Further emphasizing the therapeutic potential of miRNA-loaded exosomes in modulating immune responses, miRNA-10a-5p within exosomes significantly impacts autoimmune conditions like pSS by altering critical immune cell functions. Additionally, miRNAs in olfactory ecto-MSCs-derived exosomes influence T follicular helper cell responses, adding another dimension to miRNA-mediated regulation in pSS. Exosomes derived from bone marrow MSCs and iPSCs have been shown to inhibit Th17 cell differentiation while promoting Treg cell expansion, resulting in reduced lymphocytic infiltration in salivary glands and decreased serum autoantibody levels. Similar effects were observed with exosomes from labial gland-derived MSCs, which play a key role in regulating the Treg/Th17 balance, crucial for controlling inflammatory responses in pSS. Further studies demonstrated that exosomes from human umbilical cord MSCs exhibit immunosuppressive properties, reducing inflammation by modulating the Treg/Th17 balance and improving gut microbiota composition. , These findings collectively highlight exosomes' capacity to alleviate pSS symptoms by targeting immune regulatory pathways. In addition to their immunomodulatory effects, exosomes have shown potential in enhancing tissue repair and cellular function. Exosomes from dental pulp stem cells (DPSC-exos) enhance salivary gland epithelial function through the GPER-mediated cAMP/PKA/CREB pathway, underscoring a signaling pathway-driven mechanism. Similarly, that exosomes from human exfoliated deciduous teeth reduce cellular apoptosis and enhance glandular function by suppressing p-ERK1/2-mediated apoptosis. Collectively, exosomes play a critical role in the pathogenesis, diagnosis, and treatment of pSS. Mechanistically, exosomes from EBV-infected B cells and activated T cells transfer key miRNAs, such as EBV-miRNA-BART13-3p and miRNA-142-3p, to salivary gland epithelial cells, disrupting calcium signaling pathways and reducing saliva secretion. These exosomes contribute to glandular dysfunction by downregulating key proteins like STIM1 and AQP5. Diagnostically, exosomes offer non-invasive biomarkers, with molecules such as Ro/SSA, La/SSB, and circular RNAs (e.g., circ-IQGAP2) found in serum, saliva, and tears, facilitating early detection and disease monitoring. In terms of treatment, MSC-exos demonstrate significant potential by modulating immune responses, balancing Treg and Th17 cells, reducing inflammation, and promoting tissue repair. However, challenges such as exosome heterogeneity, limited standardization in isolation methods, and the need for large-scale clinical trials remain in pSS. Addressing these issues will be crucial for translating exosome-based therapies into clinical practice. Future research should focus on refining exosome-based diagnostics and expanding the clinical use of MSC-exos to restore salivary gland function and improve pSS symptoms. 

Exosome-based therapies have shown potential in addressing the underlying causes of salivary gland dysfunction. In particular, radiotherapy-induced xerostomia, a prevalent and severe side effect affecting head and neck cancer survivors, has been a target for innovative therapeutic approaches. For instance, exosomes derived from human dental pulp stem cells have been shown to reduce inflammatory cytokine production and reverse oxidative stress in submandibular gland cells, thereby preventing cellular senescence induced by radiation. Similarly, exosomes from hypoxia-pretreated human urine-derived stem cells have demonstrated the ability to repair radiation-induced salivary gland injuries by activating the Wnt3a/GSK3β pathway. Moreover, exosomes derived from salivary gland organoids assembled through magnetic 3D bioassembly have emerged as a promising strategy for mitigating epithelial damage in irradiated submandibular gland models. 

Diabetes is one of the most common metabolic diseases affecting salivary function. More than half of diabetic patients suffer from xerostomia and salivary gland hypofunction. Research has suggested that circulating exosomal miRNAs may be related to insulin resistance. Further experiments demonstrated that bone marrow MSC-derived exosomes can inhibit diabetic sequelae in the salivary glands and its complications by inhibiting TGFβ and its related pathway via Smad 2 and Smad 3 in streptozotocin-induced diabetic rats. Additionally, research indicated that salivary exosomes reduced blood glucose levels and enhanced salivary gland function in diabetic rats. 

Menopause is often accompanied by physiological and functional changes in the oral cavity. Previous studies have indicated that hormone deficiency post-menopause may lead to reduced salivary gland secretion. Currently, there is no standard effective treatment for salivary gland dysfunction occurring after menopause. However, tonsil-MSC-derived EVs alleviated salivary gland dysfunction caused by menopause through anti-inflammatory and anti-fibrosis mechanisms in ovariectomized rats. Further research is needed to identify the specific miRNAs and proteins that play a critical role in post-menopausal salivary gland dysfunction. 

Sialolithiasis, or salivary calculi, is characterized by the mechanical obstruction of the salivary glands' excretory ducts due to the partial deposition of calcific materials. It accounts for one-third of all salivary gland disorders, with the majority of calculi occurring in the submandibular glands. The accumulation of salivary stones can lead to recurrent chronic sialadenitis, as the presence of these stones promotes bacterial aggregation, which can cause inflammation and infection. Although the exact mechanisms behind sialolith formation remain unclear, various theories have been proposed. Some suggests that changes in ion concentration in saliva, the presence of bacteria or foreign bodies in the ducts, or mucous plugs may initiate stone formation. , , The organic core theory posits that the organic components at the center of calcification, such as bacteria, foreign bodies, or desquamated epithelial cells, act as nucleating sites for calculus formation. , In contrast, some theories suggest that stone formation is secondary to sialadenitis, where the swelling during chronic inflammation leads to salivary stasis and the subsequent formation of a calcium-rich core. Some studies have found no organic components in the central area of the calculus core. , However, recent research has provided evidence that exosomes may play a role in the formation and development of sialolithiasis. Using liquid chromatography-mass spectrometry and microscopy, exosomes were confirmed to be present in salivary calculi. Additionally, numerous EVs and degraded cytoplasmic organelles were observed in the central nidus of the stones through transmission electron microscopy. Further examination also revealed the presence of exosomes and bi-layered bacterial cell membranes in the central nidus of a patient with recurrent sialolithiasis. These findings suggest that exosomes might contribute to the pathogenesis of sialolithiasis, although further research is needed to elucidate the exact mechanisms involved. 

SACC accounts for approximately 1 % of head and neck tumors and 10 % of salivary gland tumors. Due to its proximity to critical anatomical structures, achieving negative surgical margins during treatment is often difficult. This challenge contributes to a high recurrence rate, with nearly 40 % of patients experiencing local recurrence within five years. Additionally, SACC has a significant risk of distant metastasis, ranging from 8 % to 60 %, resulting in a generally poor long-term prognosis. SACC is primarily characterized by hematogenous metastasis, with the lungs being the most commonly affected site. Exosomes, small vesicles secreted by cancer cells, have been increasingly recognized as playing a crucial role in this metastatic process. , , PROMININ-1 (CD133) is significantly expressed in SACC and is released into saliva through exosomes or EVs. This suggests that exosomes could be involved in both the diagnosis and progression of SACC. Further supporting the role of exosomes in SACC, the upregulation of epithelial regulatory proteins in SACC cells induces epithelial–mesenchymal transition by modulating the GLI1/E-cadherin axis. This process not only increases the expression of pro-angiogenic factors like VEGFA, bFGF, and IL-8 but also facilitates their transfer via exosomes. These exosomes enhance angiogenesis within the tumor microenvironment and increase vascular permeability in the pre-metastatic lung microenvironment, thereby promoting metastasis. Exosomes from SACC-83 cells further enhance these invasive capabilities by increasing endothelial cell permeability through targeting tight junction proteins such as claudin-1, ZO-1, and β-catenin. Notably, SACC cell-derived exosomes contribute to angiogenesis and local vascular microleakage by transporting miRNA-23 b-3p, suggesting that this miRNA could serve as a potential biomarker for distant metastasis in SACC. 

Despite their promising potential, several challenges need to be addressed before exosomes can be fully integrated into clinical practice for salivary gland diseases. One major challenge is improving techniques for exosome extraction and purification, particularly from oral fluids or serum, to develop standardized protocols and establish reliable diagnostic standards. Additionally, the heterogeneity of exosomes, due to their diverse origins and cargo, limits their application as biomarkers. Further exploration is needed to understand the mechanisms involved in salivary gland diseases and to identify novel exosome-based biomarkers. Finally, clinical trials involving exosomes in salivary gland diseases remain limited, often relying on small sample sizes. Expanding research to larger-scale population studies is essential for translating basic scientific discoveries into practical clinical applications. 

Exosomes offer substantial potential to improve clinical outcomes in salivary gland diseases, particularly as non-invasive biomarkers and therapeutic tools. In conditions like pSS and sialolithiasis, exosomes can enable earlier and more precise diagnosis due to their molecular stability and specificity. Moreover, exosomes derived from MSCs show promise in modulating immune responses and promoting tissue repair, opening the door to novel therapeutic strategies. For these potentials to be fully realized, future research must prioritize large-scale clinical trials to validate exosome-based diagnostics and treatments. Further exploration of the mechanisms by which exosomes regulate immune responses and repair damaged tissues will be essential. Additionally, advancements in exosome engineering, such as customizing their cargo for specific therapeutic targets, could transform exosomes into a cornerstone of personalized medicine. Identifying disease-specific biomarkers will also be critical to refining diagnostic precision and enhancing therapeutic interventions. 

Exosomes and their cargo play a critical role in intercellular communication, positioning them as promising biomarkers, therapeutic targets, and drug delivery vehicles for salivary gland diseases. However, continued research is necessary to fully realize these possibilities and to integrate exosome-based diagnostics and therapies into routine clinical practice. 

The authors have no conflicts of interest relevant to this article. 

