Comparative proteomic analysis of rat adipose-derived mesenchymal stem cells and their exosomes

doi:10.12307/2025.046

Abstract

Abstract: BACKGROUND: Research on adipose-derived mesenchymal stem cells and their exosomes often uses proteomics to analyze their roles in tissue repair, but comparative proteomic analyses between the two are scarce.
OBJECTIVE: To analyze adipose-derived mesenchymal stem cells and their exosomes using proteomic analysis.
METHODS: Rat adipose-derived mesenchymal stem cells were isolated, cultured, and identified. Exosomes were then extracted from cell supernatant and identified. Differentially expressed proteins of adipose-derived mesenchymal stem cells and their exosomes were analyzed using DIA proteomics. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analyses were performed on these differentially expressed proteins.
RESULTS AND CONCLUSION: (1) Adipose-derived mesenchymal stem cells displayed a mainly spindle-shaped, fibroblast-like morphology. (2) The exosome suspension protein concentration was 7.66 mg/mL, as determined by the BCA method. (3) Exosomes exhibited a characteristic teacup shape with a visible double-layer membrane vesicle structure. The center presented a low electron density component. The exosomes showed a peak particle size distribution of 112.2 nm, a concentration of 7.5×1011 particles/mL, and an average diameter ranging from 70 to 140 nm. (4) Exosomes expressed high levels of surface marker proteins CD9 and TSG101. (5) Gene Ontology analysis of differentially expressed proteins revealed enrichment in the extracellular matrix and synapses, with functions related to ion binding, ribosome binding, and particularly cell adhesion and translation. (6) Kyoto Encyclopedia of Genes and Genomes pathway analysis indicated that the differentially expressed proteins were primarily involved in extracellular matrix receptor interaction, ribosome, and cytokine receptor interaction, and also associated with various metabolic diseases like cholesterol and thyroid disorders.

Key words: adipose mesenchymal stem cell, exosome, proteomics, comparative analysi, tissue repair

CLC Number:

Zhang Xinrui, Han Yue, Lei Lei, Liu Jianyu, Geng Chengkui. Comparative proteomic analysis of rat adipose-derived mesenchymal stem cells and their exosomes[J]. Chinese Journal of Tissue Engineering Research, 2025, 29(13): 2683-2689.

Figures/Tables 6

References

[1] MISHRA VK, SHIH HH, PARVEEN F, et al. Identifying the Therapeutic Significance of Mesenchymal Stem Cells. Cells. 2020;9(5):1145.
[2] LIAU LL, LOOI QH, CHIA WC, et al. Treatment of spinal cord injury with mesenchymal stem cells. Cell Biosci. 2020;10:112.
[3] RHODE SC, BEIER JP, RUHL T. Adipose tissue stem cells in peripheral nerve regeneration-In vitro and in vivo. J Neurosci Res. 2021;99(2): 545-560.
[4] ZHOU LN, WANG JC, ZILUNDU PLM, et al. A comparison of the use of adipose-derived and bone marrow-derived stem cells for peripheral nerve regeneration in vitro and in vivo. Stem Cell Res Ther. 2020; 11(1):153.
[5] AL-GHADBAN S, BUNNELL BA. Adipose Tissue-Derived Stem Cells: Immunomodulatory Effects and Therapeutic Potential. Physiology (Bethesda). 2020;35(2):125-133.
[6] WANG Y, FANG J, LIU B, et al. Reciprocal regulation of mesenchymal stem cells and immune responses. Cell Stem Cell. 2022;29(11): 1515-1530.
[7] WANG LT, LIU KJ, SYTWU HK, et al. Advances in mesenchymal stem cell therapy for immune and inflammatory diseases: Use of cell-free products and human pluripotent stem cell-derived mesenchymal stem cells. Stem Cells Transl Med. 2021;10(9):1288-1303.
[8] WILLMS E, JOHANSSON HJ, MÄGER I, et al. Cells release subpopulations of exosomes with distinct molecular and biological properties. Sci Rep. 2016;6:22519.
[9] SAILLIET N, ULLAH M, DUPUY A, et al. Extracellular Vesicles in Transplantation. Front Immunol. 2022;13:800018.
[10] HADE MD, SUIRE CN, MOSSELL J, et al. Extracellular vesicles: Emerging frontiers in wound healing. Med Res Rev. 2022;42(6):2102-2125.
[11] AN Y, LIN S, TAN X, et al. Exosomes from adipose-derived stem cells and application to skin wound healing. Cell Prolif. 2021;54(3):e12993.
[12] RECORD M, SILVENTE-POIROT S, POIROT M, et al. Extracellular vesicles: lipids as key components of their biogenesis and functions. J Lipid Res. 2018;59(8):1316-1324.
[13] HE C, ZHENG S, LUO Y, et al. Exosome Theranostics: Biology and Translational Medicine. Theranostics. 2018;8(1):237-255.
[14] HADE MD, SUIRE CN, SUO Z. Mesenchymal Stem Cell-Derived Exosomes: Applications in Regenerative Medicine. Cells. 2021;10(8): 1959.
[15] BU H, HE D, HE X, et al. Exosomes: Isolation, Analysis, and Applications in Cancer Detection and Therapy. Chembiochem. 2019;20(4):451-461.
[16] KALLURI R, LEBLEU VS. The biology, function, and biomedical applications of exosomes. Science. 2020;367(6478):eaau6977.
[17] WORTZEL I, DROR S, KENIFIC CM, et al. Exosome-Mediated Metastasis: Communication from a Distance. Dev Cell. 2019;49(3):347-360.
[18] WANG C, LI Z, LIU Y, et al. Exosomes in atherosclerosis: performers, bystanders, biomarkers, and therapeutic targets. Theranostics. 2021; 11(8):3996-4010.
[19] WATANABE Y, TSUCHIYA A, TERAI S. The development of mesenchymal stem cell therapy in the present, and the perspective of cell-free therapy in the future. Clin Mol Hepatol. 2021;27(1):70-80.
[20] FAMILTSEVA A, JEREMIC N, TYAGI SC. Exosomes: cell-created drug delivery systems. Mol Cell Biochem. 2019;459(1-2):1-6.
[21] ZHANG Y, BI J, HUANG J, et al. Exosome: A Review of Its Classification, Isolation Techniques, Storage, Diagnostic and Targeted Therapy Applications. Int J Nanomedicine. 2020;15:6917-6934.
[22] MONDAL J, PILLARISETTI S, JUNNUTHULA V, et al. Hybrid exosomes, exosome-like nanovesicles and engineered exosomes for therapeutic applications. J Control Release. 2023;353:1127-1149.
[23] 刘海琴,马华根,唐元瑜.原代大鼠脂肪间充质干细胞的体外培养扩增及鉴定[J].中国组织工程研究,2022,26(19):2953-2957.
[24] DOYLE LM, WANG MZ. Overview of Extracellular Vesicles, Their Origin, Composition, Purpose, and Methods for Exosome Isolation and Analysis. Cells. 2019;8(7):727.
[25] ZHAO R, ZHAO T, HE Z, et al. Composition, isolation, identification and function of adipose tissue-derived exosomes. Adipocyte. 2021;10(1): 587-604.
[26] HAN Y, LI X, ZHANG Y, et al. Mesenchymal Stem Cells for Regenerative Medicine. Cells. 2019;8(8):886.
[27] PIXLEY JS. Mesenchymal stem cells to treat type 1 diabetes. Biochim Biophys Acta Mol Basis Dis. 2020;1866(4):165315.
[28] SONG Y, DU H, DAI C, et al. Human adipose-derived mesenchymal stem cells for osteoarthritis: a pilot study with long-term follow-up and repeated injections. Regen Med. 2018;13(3):295-307.
[29] SHERMAN LS, ROMAGANO MP, WILLIAMS SF, et al. Mesenchymal stem cell therapies in brain disease. Semin Cell Dev Biol. 2019;95:111-119.
[30] CHEN SY, YANG RL, WU XC, et al. Mesenchymal Stem Cell Transplantation: Neuroprotection and Nerve Regeneration After Spinal Cord Injury. J Inflamm Res. 2023;16:4763-4776.
[31] FATHOLLAHI A, GABALOU NB, ASLANI S. Mesenchymal stem cell transplantation in systemic lupus erythematous, a mesenchymal stem cell disorder. Lupus. 2018;27(7):1053-1064.
[32] REGMI S, PATHAK S, KIM JO, et al. Mesenchymal stem cell therapy for the treatment of inflammatory diseases: Challenges, opportunities, and future perspectives. Eur J Cell Biol. 2019;98(5-8):151041.
[33] MAZINI L, ROCHETTE L, ADMOU B, et al. Hopes and Limits of Adipose-Derived Stem Cells (ADSCs) and Mesenchymal Stem Cells (MSCs) in Wound Healing. Int J Mol Sci. 2020;21(4):1306.
[34] XUNIAN Z, KALLURI R. Biology and therapeutic potential of mesenchymal stem cell-derived exosomes. Cancer Sci. 2020;111(9): 3100-3110.
[35] MELDOLESI J. Exosomes and Ectosomes in Intercellular Communication. Curr Biol. 2018;28(8):R435-R444.
[36] TANG Y, ZHOU Y, LI HJ. Advances in mesenchymal stem cell exosomes: a review. Stem Cell Res Ther. 2021;12(1):71.
[37] AGUIAR KOGA BA, FERNANDES LA, FRATINI P, et al. Role of MSC-derived small extracellular vesicles in tissue repair and regeneration. Front Cell Dev Biol. 2023;10:1047094.
[38] SUN Y, LIU G, ZHANG K, et al. Mesenchymal stem cells-derived exosomes for drug delivery. Stem Cell Res Ther. 2021;12(1):561.
[39] HARRELL CR, VOLAREVIC V, DJONOV V, et al. Therapeutic Potential of Exosomes Derived from Adipose Tissue-Sourced Mesenchymal Stem Cells in the Treatment of Neural and Retinal Diseases. Int J Mol Sci. 2022;23(9):4487.
[40] FANG Y, ZHANG Y, ZHOU J, et al. Adipose-derived mesenchymal stem cell exosomes: a novel pathway for tissues repair. Cell Tissue Bank. 2019;20(2):153-161.
[41] GOODARZI P, ALAVI-MOGHADAM S, PAYAB M, et al. Metabolomics Analysis of Mesenchymal Stem Cells. Int J Mol Cell Med. 2019; 8(Suppl1):30-40.
[42] TOH WS, LAI RC, ZHANG B, et al. MSC exosome works through a protein-based mechanism of action. Biochem Soc Trans. 2018;46(4): 843-853.
[43] KIMIZ-GEBOLOGLU I, ONCEL SS. Exosomes: Large-scale production, isolation, drug loading efficiency, and biodistribution and uptake. J Control Release. 2022;347:533-543.
[44] PEGTEL DM, GOULD SJ. Exosomes. Annu Rev Biochem. 2019;88: 487-514.
[45] SALUNKHE S, DHEERAJ, BASAK M, et al. Surface functionalization of exosomes for target-specific delivery and in vivo imaging & tracking: Strategies and significance. J Control Release. 2020;326:599-614.
[46] YAMASHITA T, TAKAHASHI Y, TAKAKURA Y. Possibility of Exosome-Based Therapeutics and Challenges in Production of Exosomes Eligible for Therapeutic Application. Biol Pharm Bull. 2018;41(6):835-842.
[47] ALONSO-ALONSO ML, GARCÍA-POSADAS L, DIEBOLD Y. Extracellular Vesicles from Human Adipose-Derived Mesenchymal Stem Cells: A Review of Common Cargos. Stem Cell Rev Rep. 2022;18(3):854-901.
[48] ZHU A, LIU N, SHANG Y, et al. Signaling pathways of adipose stem cell-derived exosomes promoting muscle regeneration. Chin Med J (Engl). 2022;135(21):2525-2534.
[49] LEE M, BAN JJ, YANG S, et al. The exosome of adipose-derived stem cells reduces β-amyloid pathology and apoptosis of neuronal cells derived from the transgenic mouse model of Alzheimer’s disease. Brain Res. 2018;1691:87-93.