Comparative analysis of adipose-, bone marrow-, and amniotic membrane-derived MSC secretomes and EVs reveals shared and source-specific therapeutic signatures for osteoarthritis

Abstract

Aim: Mesenchymal stromal cells (MSCs) exert their therapeutic effects in osteoarthritis (OA) primarily through paracrine signaling, including secreted proteins and extracellular vesicle (EV)-associated microRNAs (miRNAs). However, the contribution of tissue origin to the composition and function of these secretomes remains unclear. This study aimed to provide a comprehensive molecular and functional comparison of secretomes from adipose-derived (ASCs), bone marrow-derived MSCs (BMSCs) and human amniotic membrane-derived MSCs, with a specific focus on OA-relevant pathways.

Methods: MSCs were immunophenotyped by flow cytometry. Secretomes were profiled for 200 factors and 784 EV-miRNAs. Functional enrichment was performed using Gene Ontology and Reactome databases. In vitro, secretomes were tested on interleukin (IL)-1β-stimulated human chondrocytes to assess modulation of OA-related gene expression.

Results: All MSC secretomes shared a core of factors enriched in anti-inflammatory and matrix-regulatory functions. ASCs showed the differential expression of a few modulators, potentially shifting their chondroprotective phenotype. EV-miRNAs further distinguished the MSC types. ASCs and BMSCs clustered closely in both overall miRNA content and functional enrichment, which included pathways for extracellular matrix organization, angiogenesis and IL-6 signaling. BMSC- and ASC-EVs had a higher ratio of OA-protective to destructive miRNAs, including miR-24-3p, miR-125b-5p and miR-222-3p. Functional assays confirmed that all MSC secretomes were effective in suppressing key OA-related genes in inflamed chondrocytes, with ASCs and BMSCs having a stronger activity.

Conclusion: These findings support the development of MSC-derived cell-free therapies and emphasize the importance of molecular profiling in MSC source selection. Further studies are warranted to validate these observations and optimize MSC-based interventions for clinical translation in OA.

Keywords

Mesenchymal stromal cells / secretome / extracellular vesicles / miRNAs / osteoarthritis / chondroprotection / paracrine signaling / tissue regeneration

References

[1] Kloppenburg M,Cicuttini F.Osteoarthritis.Lancet2025;405:71-85

[2] Farinelli L,Gigante A.Pain management strategies in osteoarthritis.Biomedicines2024;12:805 PMCID:PMC11048725

[3] Kim EH,Park J.Progressing future osteoarthritis treatment toward precision medicine: integrating regenerative medicine, gene therapy and circadian biology.Exp Mol Med2025;57:1133-42 PMCID:PMC12229678

[4] Zhang H,Prantl L.Adipose tissue-derived therapies for osteoarthritis: multifaceted mechanisms and clinical prospects.Cells2025;14:669 PMCID:PMC12071781

[5] Kim GB,Park WT.Bone marrow aspirate concentrate: its uses in osteoarthritis.Int J Mol Sci2020;21:3224 PMCID:PMC7247342

[6] Forbes J,Knapik DM.The use of amniotic tissue-derived products in orthopedic surgery: a narrative review.Injury2024;55:111901

[7] Sanz-Nogués C.Current good manufacturing practice considerations for mesenchymal stromal cells as therapeutic agents.Biomater Biosyst2021;2:100018 PMCID:PMC9934414

[8] Di Matteo B,Vitale ND.Minimally manipulated mesenchymal stem cells for the treatment of knee osteoarthritis: a systematic review of clinical evidence.Stem Cells Int2019;2019:1735242 PMCID:PMC6710724

[9] Caplan AI.Mesenchymal stem cells: time to change the name!.Stem Cells Transl Med2017;6:1445-51 PMCID:PMC5689741

[10] Asgarpour K,Amiri F.Exosomal microRNAs derived from mesenchymal stem cells: cell-to-cell messages.Cell Commun Signal2020;18:149 PMCID:PMC7488404

[11] Shimomura K,Saseendar S.Exploring the potential of mesenchymal stem/stromal cell-derived extracellular vesicles as cell-free therapy for osteoarthritis: a narrative review.J Cartil Jt Preserv2024;4:100184

[12] Turlo AJ,Ramsbottom KA.Mesenchymal stromal cell secretome is affected by tissue source and donor age.Stem Cells2023;41:1047-59 PMCID:PMC10631804

[13] Dubey NK,Dubey R,Tsai FC.Revisiting the advances in isolation, characterization and secretome of adipose-derived stromal/stem cells.Int J Mol Sci2018;19:2200 PMCID:PMC6121360

[14] Trzyna A.Adipose-derived stem cells secretome and its potential application in “stem cell-free therapy”.Biomolecules2021;11:878 PMCID:PMC8231996

[15] Kehl D,Mallone A.Proteomic analysis of human mesenchymal stromal cell secretomes: a systematic comparison of the angiogenic potential.NPJ Regen Med2019;4:8 PMCID:PMC6467904

[16] Zhuang J,Sun R.Multifunctional exosomes derived from bone marrow stem cells for fulfilled osseointegration.Front Chem2022;10:984131 PMCID:PMC9441814

[17] Ragni E,Perucca Orfei C.Amniotic membrane-mesenchymal stromal cells secreted factors and extracellular vesicle-miRNAs: anti-inflammatory and regenerative features for musculoskeletal tissues.Stem Cells Transl Med2021;10:1044-62 PMCID:PMC8235131

[18] Muntiu A,Vincenzoni F.Disclosing the molecular profile of the human amniotic mesenchymal stromal cell secretome by filter-aided sample preparation proteomic characterization.Stem Cell Res Ther2023;14:339 PMCID:PMC10683150

[19] Zuk PA,Mizuno H.Multilineage cells from human adipose tissue: implications for cell-based therapies.Tissue Eng2001;7:211-28

[20] Giordano R,Isalberti M.Autologous mesenchymal stem cell therapy for progressive supranuclear palsy: translation into a phase I controlled, randomized clinical study.J Transl Med2014;12:14 PMCID:PMC3912501

[21] Papait A,Cargnoni A.Comparison of EV-free fraction, EVs, and total secretome of amniotic mesenchymal stromal cells for their immunomodulatory potential: a translational perspective.Front Immunol2022;13:960909 PMCID:PMC9424831

[22] Szklarczyk D,Koutrouli M.The STRING database in 2023: protein-protein association networks and functional enrichment analyses for any sequenced genome of interest.Nucleic Acids Res2023;51:D638-46 PMCID:PMC9825434

[23] Cavalleri T,Favero C.Plasmatic extracellular vesicle microRNAs in malignant pleural mesothelioma and asbestos-exposed subjects suggest a 2-miRNA signature as potential biomarker of disease.PLoS One2017;12:e0176680 PMCID:PMC5417506

[24] D’haene B,Hellemans J.miRNA expression profiling: from reference genes to global mean normalization.Methods Mol Biol2012;822:261-72

[25] Aparicio-Puerta E,Schmartz GP,Fehlmann T.miEAA 2023: updates, new functional microRNA sets and improved enrichment visualizations.Nucleic Acids Res2023;51:W319-25 PMCID:PMC10320052

[26] Metsalu T.ClustVis: a web tool for visualizing clustering of multivariate data using Principal Component Analysis and heatmap.Nucleic Acids Res2015;43:W566-70 PMCID:PMC4489295

[27] Iulian Stanciugelu S,Selaru C.Osteoarthritis and microRNAs: do they provide novel insights into the pathophysiology of this degenerative disorder?.Life2022;12:1914 PMCID:PMC9692287

[28] Endisha H,Jurisica I.The complex landscape of microRNAs in articular cartilage: biology, pathology, and therapeutic targets.JCI Insight2018;3:e121630 PMCID:PMC6171796

[29] Dominici M,Mueller I.Minimal criteria for defining multipotent mesenchymal stromal cells. The International Society for Cellular Therapy position statement.Cytotherapy2006;8:315-7

[30] Tzavlaki K.TGF-β signaling.Biomolecules2020;10:487 PMCID:PMC7175140

[31] Wang W,Lyons KM.TGFβ signaling in cartilage development and maintenance.Birth Defects Res C Embryo Today2014;102:37-51 PMCID:PMC4267887

[32] Fan W,Wang Y.CD105 promotes chondrogenesis of synovium-derived mesenchymal stem cells through Smad2 signaling.Biochem Biophys Res Commun2016;474:338-44

[33] Chang CB,Kim EM,Seong SC.Chondrogenic potentials of human synovium-derived cells sorted by specific surface markers.Osteoarthritis Cartilage2013;21:190-9

[34] Wu CC,Sytwu HK,Chang DM.CD146+ mesenchymal stem cells display greater therapeutic potential than CD146- cells for treating collagen-induced arthritis in mice.Stem Cell Res Ther2016;7:23 PMCID:PMC4741021

[35] Li X,Zha K.Enrichment of CD146⁺ adipose-derived stem cells in combination with articular cartilage extracellular matrix scaffold promotes cartilage regeneration.Theranostics2019;9:5105-21 PMCID:PMC6691381

[36] Mukherjee A.The role of inflammatory mediators and matrix metalloproteinases (MMPs) in the progression of osteoarthritis.Biomater Biosyst2024;13:100090 PMCID:PMC10910010

[37] Wilkinson DJ.Serpins in cartilage and osteoarthritis: what do we know?.Biochem Soc Trans2021;49:1013-26 PMCID:PMC8106492

[38] Ripmeester EGJ,van den Akker GGH.BMP7 reduces the fibrocartilage chondrocyte phenotype.Sci Rep2021;11:19663 PMCID:PMC8490443

[39] Saraiva M,O’Garra A.Biology and therapeutic potential of interleukin-10.J Exp Med2020;217:e20190418 PMCID:PMC7037253

[40] Al-Qahtani AA,Al-Qahtani AA.Pro-inflammatory and anti-inflammatory interleukins in infectious diseases: a comprehensive review.Trop Med Infect Dis2024;9:13 PMCID:PMC10818686

[41] van Helvoort EM, van der Heijden E, van Roon JAG, Eijkelkamp N, Mastbergen SC. The role of interleukin-4 and interleukin-10 in osteoarthritic joint disease: a systematic narrative review.Cartilage2022;13:19476035221098167 PMCID:PMC9251827

[42] Arend WP.The balance between IL-1 and IL-1Ra in disease.Cytokine Growth Factor Rev2002;13:323-40

[43] Jayasuriya CT,Li J.Molecular characterization of mesenchymal stem cells in human osteoarthritis cartilage reveals contribution to the OA phenotype.Sci Rep2018;8:7044 PMCID:PMC5935742

[44] Zhang Y,Vithran DTA,Xiao W.CC chemokines and receptors in osteoarthritis: new insights and potential targets.Arthritis Res Ther2023;25:113 PMCID:PMC10316577

[45] Reboul P,Martel-Pelletier J.Hepatocyte growth factor in osteoarthritis: when bone and cartilage decide to have a chat.Arthritis Res Ther2003;5:162 PMCID:PMC2833953

[46] Bollmann M,Garbers C.Interleukin-11 - a new cytokine in osteoarthritis?.Osteoarthritis and Cartilage2022;30:S168-9

[47] Kohrs L,Lokau J.The biology of interleukin-6 family cytokines is regulated by glycosylation.Biochem J2025;482:535-51 PMCID:PMC12179393

[48] Yan D,Chen D,Im HJ.Bovine lactoferricin-induced anti-inflammation is, in part, via up-regulation of interleukin-11 by secondary activation of STAT3 in human articular cartilage.J Biol Chem2013;288:31655-69 PMCID:PMC3814761

[49] Yang T,Li Y.MicroRNA-146a-5p alleviates the pathogenesis of osteoarthritis by inhibiting SDF-1/CXCR4-induced chondrocyte autophagy.Int Immunopharmacol2023;117:109938

[50] Qin H,He Y.Silencing miR-146a-5p protects against injury-induced osteoarthritis in mice.Biomolecules2023;13:123 PMCID:PMC9856058

[51] Zhang X,Zhao J.miR-146a facilitates osteoarthritis by regulating cartilage homeostasis via targeting Camk2d and Ppp3r2.Cell Death Dis2017;8:e2734 PMCID:PMC5477577

[52] Zhang H,Li D.miR-146a-5p promotes chondrocyte apoptosis and inhibits autophagy of osteoarthritis by targeting NUMB.Cartilage2021;13:1467S-77 PMCID:PMC8804840

[53] Xu J,Ding R.MiR-24-3p attenuates IL-1β-induced chondrocyte injury associated with osteoarthritis by targeting BCL2L12.J Orthop Surg Res2021;16:371 PMCID:PMC8194242

[54] Philipot D,Platano D.p16INK4a and its regulator miR-24 link senescence and chondrocyte terminal differentiation-associated matrix remodeling in osteoarthritis.Arthritis Res Ther2014;16:R58 PMCID:PMC4060445

[55] Rasheed Z,Abdulmonem WA.MicroRNA-125b-5p regulates IL-1β induced inflammatory genes via targeting TRAF6-mediated MAPKs and NF-κB signaling in human osteoarthritic chondrocytes.Sci Rep2019;9:6882 PMCID:PMC6499837

[56] Li J,Chen K.Upregulation of miR-125b-5p relieves chondrocyte inflammation and apoptosis in osteoarthritis by repressing the YAP1/NF-κB pathway.aging2021;13:

[57] Song J,Kim D,Chun CH.MicroRNA-222 regulates MMP-13 via targeting HDAC-4 during osteoarthritis pathogenesis.BBA Clin2015;3:79-89 PMCID:PMC4661531

[58] Stadnik PS,Tarn J.Regulation of microRNA-221, -222, -21 and -27 in articular cartilage subjected to abnormal compressive forces.J Physiol2021;599:143-55 PMCID:PMC8132181

[59] Vincent TL.IL-1 in osteoarthritis: time for a critical review of the literature.F1000Res2019;8:934 PMCID:PMC6589928

[60] Alahdal M,Ouyang H.The role of indoleamine 2,3 dioxygenase 1 in the osteoarthritis.Am J Transl Res2020;12:2322-43 PMCID:PMC7344072

[61] Chen H,Hu S.Molecular mechanisms of chondrocyte proliferation and differentiation.Front Cell Dev Biol2021;9:664168 PMCID:PMC8194090