1. Please check and comment entries here.
Table of Contents

    Topic review

    Mesenchymal Stem Cell-Derived Exosomes in Cartilage Regeneration

    View times: 26
    Submitted by:


    Exosomes are the small extracellular vesicles secreted by cells for intercellular communication. Exosomes are rich in therapeutic cargos such as microRNA (miRNA), long non-coding RNA (lncRNA), small interfering RNA (siRNA), DNA, protein, and lipids. Mesenchymal stem cell (MSC)-derived exosomes have been found contain miRNAs that modulate cartilage regeneration.

    1. Introduction

    Cartilage damage is very common and is one of the hallmarks of degenerative joint disorders such as osteoarthritis (OA) [1]. Age, gender (female), obesity, history of knee injury, joint overuse, joint abnormality, and genetics are known to increase the risk of OA [2][3]. Relying on cartilage self-repair is insufficient to restore the structure and function of the degenerated tissue as cartilage tissue has very poor regeneration potential [4]. Unlike most tissues, cartilage is avascular, alymphatic, and anueral and has low cellularity, with sparsely distributed chondrocytes embedded within the dense extracellular matrix (ECM) [5]. Thus, early intervention is critical for the management of OA. Generally, OA management can be categorized into non-pharmacological and pharmacological management modalities. Regardless of the treatment approaches, current interventions aim to prevent further damage and achieve symptoms control. Joint replacement surgery is typically prescribed when the damage is very extensive and severely affects the patient’s quality of life [6].
    Regenerative medicine is a new approach introduced in the last two decades to promote the regeneration of damaged cartilage. This approach is unique as it can stimulate cartilage regeneration, which cannot be achieved with conventional treatments. In recent years, instead of cells, more attention has been given to the paracrine factors secreted by cells. In particular, exosomes, a type of small extracellular vesicle (EV) secreted by cells for intercellular communication, have become the focus of recent studies. Exosomes are rich in proteins, lipids, and nucleic acids and have been documented to promote cartilage regeneration [7][8]. Previously, a battery of miRNAs was found to be involved in cartilage regeneration [9][10][11]

    2. Exosomes

    2.1. Categories

    Exosomes and ectosomes, which are derived from endosomes and the plasma membrane, respectively, are the two subtypes of lipid-bound EVs secreted into the extracellular environment. These EVs do not possess a nucleus structure; hence, they cannot replicate. According to the latest position statement indicated in the Minimal Information for Studies of Extracellular Vesicles 2018 (MISEV2018) report, exosome and ectosome terminologies should only be used if an experimental research design can establish the subcellular origin of the EV subtype being studied. Otherwise, operational terminologies for the subtypes of EVs should be adopted, which are based on (i) particle size (small EVs for particles smaller than 200 nm, and medium and large EVs for particles larger than 200 nm), (ii) surface markers expression or biochemical recognition (such as CD81+, CD63+ or annexin A5-stained), and (iii) the origin or condition of cells where the EVs are isolated (such as hypoxic EVs, apoptotic bodies, and podocyte EVs) [12].
    Exosomes can be either native or bioengineered, depending on whether they have been artificially manipulated or not. Both animals and plants can produce native exosomes, hence giving rise to animal-based exosomes or plant-based exosomes. Among the animal-based exosomes, they can be further classified depending on the type of cells that are producing the exosomes—either exosomes produced by normal and healthy cells or those that are synthesized from tumor cells. Amongst the variety of normal cells that can produce exosomes, MSC-derived exosomes have been given considerable attention due to the capability of MSCs to self-renew and undergo multilineage differentiation. Therefore, MSC-derived exosomes are being actively investigated for their tissue regeneration potential [13].

    2.2. Biogenesis

    Exosomes, the smallest form of EVs, with diameters ranging from 30–150 nm [14], are usually synthesized through an endosomal-sorting complex required for transport (ESCRT)-dependent or ESCRT-independent pathways [15][16][17][18]. The ESCRT-dependent pathway for exosome biogenesis usually involves a series of budding processes that are facilitated by complex protein machinery, which is primarily made up of four distinct ESCRT proteins (0 to III). This pathway is triggered when ubiquitinated proteins are recognized by ESCRT-0 and are sequestered into specific endosomal membrane domains. After further interacting with ESCRTs I/II and forming a total complex with ESCRT III, plasma membrane infolding begins, resulting in the formation of a cup-shaped early endosome that contains cell-surface proteins and soluble bioactive substances that are accumulated from the extracellular environment. As this early endosome buds inward, it matures into a late endosome that subsequently undergoes a secondary endosomal membrane invagination to form a multivesicular body containing intraluminal vesicles [16][19][20][21][22]. These intraluminal vesicles have similar membrane positioning to the cell surface, where extracellular domains of transmembrane protein face the extracellular environment while enclosing the cytosolic entities [23]. As the multivesicular body moves to the cell surface and fuses with it, the intraluminal vesicles are released into the extracellular space, giving rise to exosomes [13]. Other than ESCRT proteins, numerous other accessory proteins have been shown to be involved in the production of exosomes through the ESCRT-dependent pathway, including programmed cell death 6-interacting protein (ALIX); tumor susceptibility gene 101 protein (TSG101); heat shock cognate protein 70 (HSC70); vacuolar protein sorting-associated protein 4 (VPS4); heat shock protein 90 (HSP90); soluble N-ethylmaleimide-sensitive fusion attachment protein receptor (SNAREs); cluster of differentiation proteins 9, 81 and 63; and syndecan-1, synthenin-1, and tetraspanins [15][21][23][24]. In the alternative ESCRT-independent pathway, ceramides play an important role as their cone-shaped feature is speculated to promote microdomain-induced budding of the endosomal membrane. These ceramides are usually produced through the removal of the phosphocholine moiety by sphingomyelinases that are abundantly present in those microdomains. The presence of ceramides will also induce the lateral isolation of cargo within the endosomal membrane that leads to the formation of intraluminal vesicles within the multivesicular body, releasing exosomes once it fuses with the plasma membrane [25][26].

    2.3. Therapeutic Cargos of Exosomes

    Upon secretion into the extracellular space, exosomes will migrate and deliver their contents to their target cells, resulting in the alteration of gene expression as well as the modification of physiological and biological functions. The functional modification relies on the type of exosomal contents present within the exosomes [27]. Invariably, most exosomes are rich in proteins and lipids. To date, about 8000 proteins and 194 lipids have been found to be related to exosomes [28]. Along with these, exosomes are known to carry nucleic acid cargos, including mRNAs and miRNAs, for intercellular communication [29]. Since exosomes are secreted by numerous cell types and the exosomal content is heavily associated with the cell of origin, each exosome may have different roles in intercellular communication for varying physiological effects. For example, platelets secrete exosomes containing prostaglandins that modulate inflammatory activities [30]. Meanwhile, antigen-presenting cells, such as dendritic cells, release exosomes that carry functional major histocompatibility complexes class I and II (MHC I and MHC II) to attenuate or stimulate antigen-specific B-cell or T-cell responses [31]. Similarly, many other studies have also demonstrated the involvement of exosomes in various biological processes, including coagulation, blood vessel formation, red blood cell maturation, and removal of unwanted RNAs and proteins [32].
    In view of exosomes’ capability to carry their cargo to the target cells for participation in both normal and pathobiological mechanisms through intercellular communication, extensive investigations have been performed to exploit their therapeutic purposes to restore the diseased tissue to its normal phenotype. One of the approaches involves the isolation of these endogenous exosomes from body fluids so that they can be artificially enriched with therapeutic miRNAs before these bioengineered exosomes are reintroduced back into the patient’s body [29][32]. Exosomes have increasing popularity as a preferred drug delivery vehicle as they are non-immunogenic. In addition to that, other advantages of exosomes over MSCs are their homing ability to the site of injury and passing through tight junctions such as the blood–brain barrier. Since exosomes are very heterogeneous, they can also carry different proteins on their surface that can be delivered into the cell through receptor-mediated endocytosis upon interacting with the target cells [13]. Therefore, the exosome-assisted delivery approach has been utilized by various research groups to deliver a variety of therapeutic compounds to their target cells, including short interfering-RNA (siRNA), recombinant proteins, miRNA, and antagomirs, as well as anti-inflammatory and chemotherapeutic drugs [25]. For instance, exosomes equipped with siRNAs against the mRNA of Huntington’s disease have been successfully introduced into the cortical neurons of mice, resulting in an improvement of the murine Huntington’s disease condition [33]. Through the use of exogenous siRNAs, the MAPK gene in target lymphocytes and monocytes could be silenced. Hence, this proves the feasibility of bio-engineering isolated exosomes for the targeted delivery of therapeutic materials to a specific tissue to exert its function [34]. Alternatively, donor cells can be bio-engineered to contain the therapeutic materials, which will then be synthesized and encapsulated in the exosomes produced. These enriched exosomes can then be used for the treatment of specific diseases. An example would be a study done by Shimbo et al. that introduced miR-143 into MSCs, where this miRNA was then secreted and packaged into the exosomes. Upon delivery to osteosarcoma cells, the miR-143 present within the exosomes were able to significantly inhibit the migration of the cancerous cells [35]. In view of these and many other successful examples in utilizing exosomes as a delivery vehicle for therapeutic purposes, exosomes have been accepted as drug carriers for the treatment of various diseases in clinical trials [15].

    3. Promoting Cartilage Repair Using Exosomes

    Plastic and reconstructive surgery can be used to restore the structure and function of cartilage defects. However, these treatments are more successful for minor defects. When it concerns a sizable cartilage defect, such treatment options still pose some limitations. Therefore, autologous or matrix-assisted implantation of chondrocytes could be a more effective treatment. Unfortunately, the use of these therapeutic approaches is also restricted by the lack of donor sites as a result of possible post-donation adverse effects as well as risks associated with the deterioration of graft tissue [36]. Recently, researchers have resorted to utilizing exosomes derived from stem cells as an alternative therapy due to the ease of accessibility, unlimited supply, and better stability. Besides that, exosomes are secretory products enriched with active molecules that could result in the desired therapeutic outcomes [37]. The following paragraphs describe the composition of exosomes that are desirable for therapeutics.
    Firstly, exosomes contain various types of growth factors and microRNAs that can stimulate cartilage regeneration. This notion arose through a study that demonstrated the successful repair of injured cartilage tissue when it was co-cultured with MSCs. It was eventually discovered that the cartilage tissue regeneration was achieved due to the presence of different growth factors secreted by MSCs, including fibroblast growth factor (FGF) 2, interleukin (IL)-6, and insulin-like growth factor (IGF). These biological factors possess the capability to induce the proliferation and matrix synthesis of chondrocytes, thus assisting in the healing of the injured cartilage [38]. In agreement with this, Jiang et al. observed enhanced osteochondral regenerative activity in the presence of exosomes derived from human umbilical cords. These exosomes were able to induce the proliferation and migration of chondrocytes, hence repairing the osteochondral knee injury in a rabbit model. The regenerative potential of the exosomes in this study was attributed to the presence of about 20 miRNA types that may exert positive effects in the regulation of the joint microenvironment [39]. Similar findings were observed in another study in which complete recovery of osteochondral defects in a rat model was achieved after treatment with MSC-derived exosomes. The restored cartilage and osteochondral bone displayed normal structural characteristics that resembled their normal counterparts, such as hyalinized cartilage with ideal surface regularity and good attachment to the adjoining cartilage, with well deposited ECM [40].
    Other than their regenerative capacity, exosomes were also found to possess chondroprotective effects. For instance, treatment of synovial explants with MSC-conditioned media was found to inhibit the expression of matrix degradative enzymes, including matrix metalloproteinase (MMP)-1, MMP-12, and IL-1b, thus facilitating the repair of cartilage tissue [41]. A separate in vitro study also pointed towards the aptitude of MSCs to protect chondrocytes in cartilage via the upregulation of type II collagen production [42] to resynthesize the matrix. Apart from restoring the cartilage matrix, enhanced expression of type II collagen could also prevent the hypertrophy of chondrocytes, thus avoiding the progression of cartilage degeneration [43]. Moreover, this chondrogenic protective effect has been demonstrated by Cosenza et al., who showed the ability of MSC-derived exosomes to exert protective effects on chondrocytes in a murine model with induced joint disease. This chondroprotective outcome was achieved through an increased expression of chondrocyte markers (such as aggrecan and type II collagen) while suppressing catabolic genes (including ADAMTS5 and MMP-13) to prolong the survival of OA-like chondrocytes induced with IL-1β [44]. Additionally, MSC-derived exosomes potentially assisted in cartilage repair by increasing the number of chondrocytes by preventing chondrocyte apoptosis through the upregulation of anti-apoptotic proteins, including Bcl-2 and survivin [45].
    Post-traumatic inflammation almost always occurs alongside cartilage injury, and it is a huge obstacle for cartilage repair. This is because cartilage is an avascular tissue; hence, they do not have the capability to resolve an inflammatory response, resulting in the cartilage being easily assaulted by the proinflammatory mediators [46]. Thus, it becomes a critical therapeutic obstacle to suppress inflammation prior to the repair of injured cartilage via regeneration. In this aspect, MSC-derived exosomes have proven to be valuable as they can exert an anti-inflammatory response. For example, adipose MSC-derived exosomes could decrease levels of pro-inflammatory mediators, reducing stimulation of NF-κB and activator protein 1 while increasing expression of anti-inflammatory cytokines and the expression of miR-100-5p, which binds to 3′UTR of mTOR to augment autophagy activity [8][47]. With similar mechanisms, exosomes isolated from bone marrow-derived MSCs [48] and umbilical-cord-derived MSCs [49] were able to relieve the pathological characteristics in diseased joints through decreasing the infiltration of inflammatory cells as well as repressing levels of inflammatory factors [50].

    4. Exosomal miRNAs

    The cargo of stem cell-derived exosomes has been widely studied since 2010 when the therapeutic potential of MSC-derived exosomes was first described [51]. Although exosomes are enriched in many bioactive molecules, such as lipids, proteins, RNA, and mtDNA [52], the therapeutic potential of MSC-derived exosomes is usually rationalized with the presence of biologically relevant miRNA and proteins [53]. To date, more than 1000 proteins have been identified and mapped in MSC-derived exosomes, suggesting that the proteome of MSC-derived exosomes play a vital role in various biological processes such as cellular communication, exosome biogenesis, and tissue repair [54][55][56]. Similar to the MSC-derived proteins, many studies have suggested that MSC-derived miRNAs have the potential to modulate cell–cell communication as well as influence the progression of various diseases by regulating the signaling pathways of the recipient cells. 
    The miRNAs are a group of small, non-coding single-stranded RNAs, averaging 19–24 nucleotides, that regulate post-transcriptional gene expression. miRNAs are essential for physiological development and are involved in a variety of biological processes [57]. Abnormal expression of miRNAs is associated with several human diseases [58]. In addition to intrinsic cellular functions, miRNAs are secreted into extracellular fluids via EVs as signaling molecules mediating intercellular communication [59][60][61][62]. Hypothetically, each functional miRNA can interact with up to 200 mRNAs [63]. Intercellular communication can occur by several means, including receptor-mediated chemical interaction, direct cell–cell communication, and cytosolic synapses. These intercellular gene communications may occur not only in the microenvironment but can also occur at a distance through the secretion of exosomes into systemic circulation. Indeed, exosomes may be a more effective intercellular communication option, compared to proteins or small biochemical molecules such as mRNAs and miRNAs, that can regulate recipient cell protein production and gene expression. The ability of exosomes to deliver genetic material into cells at a distance also makes them ideal candidates for cell-free therapy. Over the decades, many studies have been conducted to study the potential of MSC-derived exosomes for treating OA and to identify the miRNAs that play a vital role in maintaining a healthy joint (Table 1).
    Table 1. List of miRNAs in MSC-derived exosomes related to chondrocytes and cartilage regeneration.
    miRNA Target Physiological Role Ref.
    miR-23b PKA Induce chondrogenic differentiation of human MSCs by inhibiting PKA signaling [64]
    miR-92a Noggin3 Targets Noggin3 and activates the PI3K/Akt/mTOR pathway to positively regulate the proliferation and matrix synthesis of chondroprogenitors [65]
    miR-125b ADAMTS-4 miR-125b overexpression suppresses IL-1-induced upregulation of ADAMTS-4 in human OA chondrocytes [67]
    miR-320 MMP-13 Downregulates MMP-13 expression in both the ATDC5 cell model of chondrogenesis and IL-1-treated primary mouse chondrocytes [68]
    miR-145 Sox9 miR-145 inhibition upregulates Sox9 expression and promotes MSC chondrogenesis [69]
    miR-221 MDM2 Downregulates MDM2 to prevent slug protein degradation that, in turn, negatively regulates chondroprogenitor proliferation [70]
    miR-22 PPARA, BMP-7 miR-22 inhibition upregulates BMP-7 and PPARA expression, inhibits IL-1 expression, and suppresses MMP-13 expression in OA chondrocytes [71]
    miR-92a-3p Wnt5a Regulate cartilage development and homeostasis by targeting Wnt5a [72]
    miR-135b Sp1a Promote chondrocyte proliferation and cartilage repair in OA by downregulating Sp1a in chondrocytes [73]
    miR-100-5p mTOR Inhibit mTOR signaling pathway to enhance chondrocyte autophagy [74]
    miR-140-5p YAP Enhance ECM secretion and induce proliferation and migration of articular chondrocytes via activating YAP as well as prevent osteoarthritic joint damage [75]
    miR-26a-5p PTGS2 Promote the survival of synovial fibroblasts and reduce synovitis [76]
    miR-136-5p ELF3 Inhibit cartilage degeneration in traumatic osteoarthritis [77]
    miR-127-3p CDH11-mediated Wnt/β-catenin pathway Inhibit CDH11, thereby blocking the Wnt/β-catenin pathway in chondrocytes and reducing the chondrocyte damage in osteoarthritic joints [78]
    miR-9-5p Syndecan-1 Has anti-inflammatory and cartilage protective effects on osteoarthritis [79]

    5. Summary

    In summary, exosomes contain miRNAs that can modulate cartilage repair and regeneration by enhancing proliferation, attenuating apoptosis, promoting chondrogenesis, increasing cartilage matrix secretion, and subsiding inflammation. Thus, exosome therapy has great potential for treating cartilage diseases and promoting cartilage regeneration,

    This entry is adapted from 10.3390/ph14111093


    1. Shane Anderson, A.; Loeser, R.F. Why is osteoarthritis an age-related disease? Best Pract. Res. Clin. Rheumatol. 2010, 24, 15–26.
    2. Pang, K.-L.; Chow, Y.Y.; Leong, L.M.; Law, J.X.; Ghafar, N.A.; Soelaiman, I.N.; Chin, K.-Y. Establishing SW1353 Chondrocytes as a Cellular Model of Chondrolysis. Life 2021, 11, 272.
    3. Liau, L.L.; Hassan, M.N.F.B.; Tang, Y.L.; Ng, M.H.; Law, J.X. Feasibility of Human Platelet Lysate as an Alternative to Foetal Bovine Serum for In Vitro Expansion of Chondrocytes. Int. J. Mol. Sci. 2021, 22, 1269.
    4. Armiento, A.R.; Alini, M.; Stoddart, M.J. Articular fibrocartilage—Why does hyaline cartilage fail to repair? Adv. Drug Deliv. Rev. 2019, 146, 289–305.
    5. Liu, Y.; Zhou, G.; Cao, Y. Recent Progress in Cartilage Tissue Engineering—Our Experience and Future Directions. Engineering 2017, 3, 28–35.
    6. Mora, J.C.; Przkora, R.; Cruz-Almeida, Y. Knee osteoarthritis: Pathophysiology and current treatment modalities. J. Pain Res. 2018, 11, 2189–2196.
    7. Liau, L.L.; Al-Masawa, M.E.; Koh, B.; Looi, Q.H.; Foo, J.B.; Lee, S.H.; Cheah, F.C.; Law, J.X. The Potential of Mesenchymal Stromal Cell as Therapy in Neonatal Diseases. Front. Pediatr. 2020, 8, 591693.
    8. Zhou, Q.; Cai, Y.; Jiang, Y.; Lin, X. Exosomes in osteoarthritis and cartilage injury: Advanced development and potential therapeutic strategies. Int. J. Biol. Sci. 2020, 16, 1811.
    9. Duan, L.; Liang, Y.; Xu, X.; Xiao, Y.; Wang, D. Recent progress on the role of miR-140 in cartilage matrix remodelling and its implications for osteoarthritis treatment. Arthrit. Res. Ther. 2020, 22, 194.
    10. Baek, D.; Lee, K.-M.; Park, K.W.; Suh, J.W.; Choi, S.M.; Park, K.H.; Lee, J.W.; Kim, S.-H. Inhibition of miR-449a Promotes Cartilage Regeneration and Prevents Progression of Osteoarthritis in In Vivo Rat Models. Mol. Ther. Nucleic Acids 2018, 13, 322–333.
    11. Lolli, A.; Sivasubramaniyan, K.; Vainieri, M.L.; Oieni, J.; Kops, N.; Yayon, A.; van Osch, G.J.V.M. Hydrogel-based delivery of antimiR-221 enhances cartilage regeneration by endogenous cells. J. Control. Release 2019, 309, 220–230.
    12. Thery, C.; Witwer, K.W.; Aikawa, E.; Alcaraz, M.J.; Anderson, J.D.; Andriantsitohaina, R.; Antoniou, A.; Arab, T.; Archer, F.; Atkin-Smith, G.K.; et al. Minimal information for studies of extracellular vesicles 2018 (MISEV2018): A position statement of the International Society for Extracellular Vesicles and update of the MISEV2014 guidelines. J. Extracell. Vesicles 2018, 7, 1535750.
    13. Zhang, Y.; Bi, J.; Huang, J.; Tang, Y.; Du, S.; Li, P. Exosome: A Review of Its Classification, Isolation Techniques, Storage, Diagnostic and Targeted Therapy Applications. Int. J. Nanomed. 2020, 15, 6917–6934.
    14. Tan, K.L.; Chia, W.C.; How, C.W.; Tor, Y.S.; Show, P.L.; Looi, Q.H.D.; Foo, J.B. Benchtop Isolation and Characterisation of Small Extracellular Vesicles from Human Mesenchymal Stem Cells. Mol. Biotechnol. 2021, 63, 780–791.
    15. Tschuschke, M.; Kocherova, I.; Bryja, A.; Mozdziak, P.; Angelova Volponi, A.; Janowicz, K.; Sibiak, R.; Piotrowska-Kempisty, H.; Iżycki, D.; Bukowska, D. Inclusion biogenesis, methods of isolation and clinical application of human cellular exosomes. J. Clin. Med. 2020, 9, 436.
    16. Lorenc, T.; Chrzanowski, J.; Olejarz, W. Current Perspectives on Clinical Use of Exosomes as a Personalized Contrast Media and Theranostics. Cancers 2020, 12, 3386.
    17. Katzmann, D.J.; Babst, M.; Emr, S.D. Ubiquitin-dependent sorting into the multivesicular body pathway requires the function of a conserved endosomal protein sorting complex, ESCRT-I. Cell 2001, 106, 145–155.
    18. Wei, D.; Zhan, W.; Gao, Y.; Huang, L.; Gong, R.; Wang, W.; Zhang, R.; Wu, Y.; Gao, S.; Kang, T. RAB31 marks and controls an ESCRT-independent exosome pathway. Cell Res. 2021, 31, 157–177.
    19. Hong, P.; Yang, H.; Wu, Y.; Li, K.; Tang, Z. The functions and clinical application potential of exosomes derived from adipose mesenchymal stem cells: A comprehensive review. Stem Cell Res. Ther. 2019, 10, 242.
    20. Avalos-Padilla, Y.; Georgiev, V.N.; Lantero, E.; Pujals, S.; Verhoef, R.N.; Borgheti-Cardoso, L.; Albertazzi, L.; Dimova, R.; Fernandez-Busquets, X. The ESCRT-III machinery participates in the production of extracellular vesicles and protein export during Plasmodium falciparum infection. PLoS Pathog. 2021, 17, e1009455.
    21. Morita, E.; Colf, L.A.; Karren, M.A.; Sandrin, V.; Rodesch, C.K.; Sundquist, W.I. Human ESCRT-III and VPS4 proteins are required for centrosome and spindle maintenance. Proc. Natl. Acad. Sci. USA 2010, 107, 12889–12894.
    22. Tang, S.; Buchkovich, N.J.; Henne, W.M.; Banjade, S.; Kim, Y.J.; Emr, S.D. ESCRT-III activation by parallel action of ESCRT-I/II and ESCRT-0/Bro1 during MVB biogenesis. eLife 2016, 5, e15507.
    23. Bryl, R.; Borowiec, B.; Siroma, R.S.; Pinto, N.; Melo, M.A.; Shibli, J.A.; Dyszkiewicz-Konwińska, M. Current application of exosomes in medicine. Med. J. Cell Biol. 2020, 8, 101–111.
    24. Lee, I.J.; Stokasimov, E.; Dempsey, N.; Varberg, J.M.; Jacob, E.; Jaspersen, S.L.; Pellman, D. Factors promoting nuclear envelope assembly independent of the canonical ESCRT pathway. J. Cell Biol. 2020, 219, e201908232.
    25. Zhang, Y.; Liu, Y.; Liu, H.; Tang, W.H. Exosomes: Biogenesis, biologic function and clinical potential. Cell Biosci. 2019, 9, 1–18.
    26. Trajkovic, K.; Hsu, C.; Chiantia, S.; Rajendran, L.; Wenzel, D.; Wieland, F.; Schwille, P.; Brugger, B.; Simons, M. Ceramide triggers budding of exosome vesicles into multivesicular endosomes. Science 2008, 319, 1244–1247.
    27. Wei, H.; Chen, Q.; Lin, L.; Sha, C.; Li, T.; Liu, Y.; Yin, X.; Xu, Y.; Chen, L.; Gao, W. Regulation of exosome production and cargo sorting. Int. J. Biol. Sci. 2021, 17, 163.
    28. Sorop, A.; Constantinescu, D.; Cojocaru, F.; Dinischiotu, A.; Cucu, D.; Dima, S.O. Exosomal microRNAs as Biomarkers and Therapeutic Targets for Hepatocellular Carcinoma. Int. J. Mol. Sci. 2021, 22, 4997.
    29. Munir, J.; Yoon, J.K.; Ryu, S. Therapeutic miRNA-Enriched Extracellular Vesicles: Current Approaches and Future Prospects. Cells 2020, 9, 2271.
    30. Heijnen, H.F.G.; Schiel, A.E.; Fijnheer, R.; Geuze, H.J.; Sixma, J.J. Activated Platelets Release Two Types of Membrane Vesicles: Microvesicles by Surface Shedding and Exosomes Derived From Exocytosis of Multivesicular Bodies and α-Granules. Blood 1999, 94, 3791–3799.
    31. Shenoda, B.B.; Ajit, S.K. Modulation of immune responses by exosomes derived from antigen-presenting cells. Clin. Med. Insights Pathol. 2016, 9, CPath-S39925.
    32. Ha, D.; Yang, N.; Nadithe, V. Exosomes as therapeutic drug carriers and delivery vehicles across biological membranes: Current perspectives and future challenges. Acta Pharm. Sin. B 2016, 6, 287–296.
    33. Didiot, M.-C.; Hall, L.M.; Coles, A.H.; Haraszti, R.A.; Godinho, B.M.; Chase, K.; Sapp, E.; Ly, S.; Alterman, J.F.; Hassler, M.R. Exosome-mediated delivery of hydrophobically modified siRNA for huntingtin mRNA silencing. Mol. Ther. 2016, 24, 1836–1847.
    34. Wahlgren, J.; Karlson, T.D.L.; Brisslert, M.; Vaziri Sani, F.; Telemo, E.; Sunnerhagen, P.; Valadi, H. Plasma exosomes can deliver exogenous short interfering RNA to monocytes and lymphocytes. Nucleic Acids Res. 2012, 40, e130.
    35. Shimbo, K.; Miyaki, S.; Ishitobi, H.; Kato, Y.; Kubo, T.; Shimose, S.; Ochi, M. Exosome-formed synthetic microRNA-143 is transferred to osteosarcoma cells and inhibits their migration. Biochem. Biophys. Res. Commun. 2014, 445, 381–387.
    36. Branam, G.M.; Saber, A.Y. Osteochondral Autograft Transplantation; StatPearls, 2021. Available online: https://www.ncbi.nlm.nih.gov/books/NBK560655/ (accessed on 25 October 2021).
    37. Chen, Y.; Xue, K.; Zhang, X.; Zheng, Z.; Liu, K. Exosomes derived from mature chondrocytes facilitate subcutaneous stable ectopic chondrogenesis of cartilage progenitor cells. Stem Cell Res. Ther. 2018, 9, 1–14.
    38. Liu, Y.; Ma, Y.; Zhang, J.; Yuan, Y.; Wang, J. Exosomes: A novel therapeutic agent for cartilage and bone tissue regeneration. Dose-Response 2019, 17, 1559325819892702.
    39. Jiang, S.; Tian, G.; Yang, Z.; Gao, X.; Wang, F.; Li, J.; Tian, Z.; Huang, B.; Wei, F.; Sang, X.; et al. Enhancement of acellular cartilage matrix scaffold by Wharton’s jelly mesenchymal stem cell-derived exosomes to promote osteochondral regeneration. Bioact. Mater. 2021, 6, 2711–2728.
    40. Zhang, S.; Chu, W.C.; Lai, R.C.; Lim, S.K.; Hui, J.H.; Toh, W.S. Exosomes derived from human embryonic mesenchymal stem cells promote osteochondral regeneration. Osteoarthr. Cartil. 2016, 24, 2135–2140.
    41. To, K.; Romain, K.; Mak, C.; Kamaraj, A.; Henson, F.; Khan, W. The treatment of cartilage damage using human mesenchymal stem cell-derived extracellular vesicles: A systematic review of in vivo studies. Front. Bioeng. Biotechnol. 2020, 8, 580.
    42. Vonk, L.A.; van Dooremalen, S.F.; Liv, N.; Klumperman, J.; Coffer, P.J.; Saris, D.B.; Lorenowicz, M.J. Mesenchymal stromal/stem cell-derived extracellular vesicles promote human cartilage regeneration in vitro. Theranostics 2018, 8, 906.
    43. Lian, C.; Wang, X.; Qiu, X.; Wu, Z.; Gao, B.; Liu, L.; Liang, G.; Zhou, H.; Yang, X.; Peng, Y.; et al. Collagen type II suppresses articular chondrocyte hypertrophy and osteoarthritis progression by promoting integrin beta1-SMAD1 interaction. Bone Res. 2019, 7, 8.
    44. Cosenza, S.; Ruiz, M.; Toupet, K.; Jorgensen, C.; Noël, D. Mesenchymal stem cells derived exosomes and microparticles protect cartilage and bone from degradation in osteoarthritis. Sci. Rep. 2017, 7, 1–12.
    45. Zhang, S.; Chuah, S.J.; Lai, R.C.; Hui, J.H.P.; Lim, S.K.; Toh, W.S. MSC exosomes mediate cartilage repair by enhancing proliferation, attenuating apoptosis and modulating immune reactivity. Biomaterials 2018, 156, 16–27.
    46. Zhang, Y.; Pizzute, T.; Pei, M. Anti-inflammatory strategies in cartilage repair. Tissue Eng. Part B Rev. 2014, 20, 655–668.
    47. Tofiño-Vian, M.; Guillén, M.I.; Del Caz, M.D.P.; Silvestre, A.; Alcaraz, M.J. Microvesicles from human adipose tissue-derived mesenchymal stem cells as a new protective strategy in osteoarthritic chondrocytes. Cell. Physiol. Biochem. 2018, 47, 11–25.
    48. Ni, Z.; Zhou, S.; Li, S.; Kuang, L.; Chen, H.; Luo, X.; Ouyang, J.; He, M.; Du, X.; Chen, L. Exosomes: Roles and therapeutic potential in osteoarthritis. Bone Res. 2020, 8, 1–18.
    49. Yan, L.; Wu, X. Exosomes produced from 3D cultures of umbilical cord mesenchymal stem cells in a hollow-fiber bioreactor show improved osteochondral regeneration activity. Cell Biol. Toxicol. 2020, 36, 165–178.
    50. Zhang, S.; Teo, K.Y.W.; Chuah, S.J.; Lai, R.C.; Lim, S.K.; Toh, W.S. MSC exosomes alleviate temporomandibular joint osteoarthritis by attenuating inflammation and restoring matrix homeostasis. Biomaterials 2019, 200, 35–47.
    51. Lai, R.C.; Arslan, F.; Lee, M.M.; Sze, N.S.K.; Choo, A.; Chen, T.S.; Salto-Tellez, M.; Timmers, L.; Lee, C.N.; El Oakley, R.M. Exosome secreted by MSC reduces myocardial ischemia/reperfusion injury. Stem Cell Res. 2010, 4, 214–222.
    52. Yin, K.; Wang, S.; Zhao, R.C. Exosomes from mesenchymal stem/stromal cells: A new therapeutic paradigm. Biomark. Res. 2019, 7, 1–8.
    53. Toh, W.S.; Lai, R.C.; Zhang, B.; Lim, S.K. MSC exosome works through a protein-based mechanism of action. Biochem. Soc. Trans. 2018, 46, 843–853.
    54. Lai, R.C.; Tan, S.S.; Teh, B.J.; Sze, S.K.; Arslan, F.; De Kleijn, D.P.; Choo, A.; Lim, S.K. Proteolytic potential of the MSC exosome proteome: Implications for an exosome-mediated delivery of therapeutic proteasome. Int. J. Proteom. 2012, 2012, 971907.
    55. Anderson, J.D.; Johansson, H.J.; Graham, C.S.; Vesterlund, M.; Pham, M.T.; Bramlett, C.S.; Montgomery, E.N.; Mellema, M.S.; Bardini, R.L.; Contreras, Z. Comprehensive proteomic analysis of mesenchymal stem cell exosomes reveals modulation of angiogenesis via nuclear factor-kappaB signaling. Stem Cells 2016, 34, 601–613.
    56. Collino, F.; Pomatto, M.; Bruno, S.; Lindoso, R.S.; Tapparo, M.; Sicheng, W.; Quesenberry, P.; Camussi, G. Exosome and microvesicle-enriched fractions isolated from mesenchymal stem cells by gradient separation showed different molecular signatures and functions on renal tubular epithelial cells. Stem Cell Rev. 2017, 13, 226.
    57. Fu, G.; Brkić, J.; Hayder, H.; Peng, C. MicroRNAs in Human Placental Development and Pregnancy Complications. Int. J. Mol. Sci. 2013, 14, 5519–5544.
    58. Paul, P.; Chakraborty, A.; Sarkar, D.; Langthasa, M.; Rahman, M.; Bari, M.; Singha, R.K.S.; Malakar, A.K.; Chakraborty, S. Interplay between miRNAs and human diseases. J. Cell. Physiol. 2018, 233, 2007–2018.
    59. Pitt, J.M.; Kroemer, G.; Zitvogel, L. Extracellular vesicles: Masters of intercellular communication and potential clinical interventions. J. Clin. Investig. 2016, 126, 1139–1143.
    60. Yang, T.; Gu, H.; Chen, X.; Fu, S.; Wang, C.; Xu, H.; Feng, Q.; Ni, Y. Cardiac hypertrophy and dysfunction induced by overexpression of miR-214 in vivo. J. Surg. Res. 2014, 192, 317–325.
    61. Wang, J.; Chen, J.; Sen, S. MicroRNA as Biomarkers and Diagnostics. J. Cell. Physiol. 2016, 231, 25–30.
    62. Alvarez-Garcia, I.; Miska, E. MicroRNA functions in animal development and human disease. Development 2005, 132, 4653–4662.
    63. Krek, A.; Grün, D.; Poy, M.; Wolf, R.; Rosenberg, L.; Epstein, E.; MacMenamin, P.; da Piedade, I.; Gunsalus, K.; Stoffel, M.; et al. Combinatorial microRNA target predictions. Nat. Genet. 2005, 37, 495–500.
    64. Ham, O.; Song, B.; Lee, S.; Choi, E.; Cha, M.; Lee, C.; Park, J.; Kim, L.; Chang, W.; Lim, S.; et al. The role of microRNA-23b in the differentiation of MSC into chondrocyte by targeting protein kinase A signaling. Biomaterials 2012, 33, 4500–4507.
    65. Ning, G.; Liu, X.; Dai, M.; Meng, A.; Wang, Q. MicroRNA-92a upholds Bmp signaling by targeting noggin3 during pharyngeal cartilage formation. Dev. Cell 2013, 24, 283–295.
    66. Hou, C.; Zhang, Z.; Zhang, Z.; Wu, P.; Zhao, X.; Fu, M.; Sheng, P.; Kang, Y.; Liao, W. Presence and function of microRNA-92a in chondrogenic ATDC5 and adipose-derived mesenchymal stem cells. Mol. Med. Rep. 2015, 12, 4877–4886.
    67. Matsukawa, T.; Sakai, T.; Yonezawa, T.; Hiraiwa, H.; Hamada, T.; Nakashima, M.; Ono, Y.; Ishizuka, S.; Nakahara, H.; Lotz, M.K.; et al. MicroRNA-125b regulates the expression of aggrecanase-1 (ADAMTS-4) in human osteoarthritic chondrocytes. Arthrit. Res. Ther. 2013, 15, R28.
    68. Meng, F.; Zhang, Z.; Chen, W.; Huang, G.; He, A.; Hou, C.; Long, Y.; Yang, Z.; Zhang, Z.; Liao, W. MicroRNA-320 regulates matrix metalloproteinase-13 expression in chondrogenesis and interleukin-1β-induced chondrocyte responses. Osteoarthr. Cartil. 2016, 24, 932–941.
    69. Yang, B.; Guo, H.; Zhang, Y.; Chen, L.; Ying, D.; Dong, S. MicroRNA-145 regulates chondrogenic differentiation of mesenchymal stem cells by targeting Sox9. PLoS ONE 2011, 6, e21679.
    70. Kim, D.; Song, J.; Jin, E. MicroRNA-221 regulates chondrogenic differentiation through promoting proteosomal degradation of slug by targeting Mdm2. J. Biol. Chem. 2010, 285, 26900–26907.
    71. Iliopoulos, D.; Malizos, K.; Oikonomou, P.; Tsezou, A. Integrative microRNA and proteomic approaches identify novel osteoarthritis genes and their collaborative metabolic and inflammatory networks. PLoS ONE 2008, 3, e3740.
    72. Mao, G.; Zhang, Z.; Hu, S.; Zhang, Z.; Chang, Z.; Huang, Z.; Liao, W.; Kang, Y. Exosomes derived from miR-92a-3p-overexpressing human mesenchymal stem cells enhance chondrogenesis and suppress cartilage degradation via targeting WNT5A. Stem Cell Res. Ther. 2018, 9, 247.
    73. Wang, R.; Xu, B.; Xu, H. TGF-β1 promoted chondrocyte proliferation by regulating Sp1 through MSC-exosomes derived miR-135b. Cell Cycle 2018, 17, 2756–2765.
    74. Wu, J.; Kuang, L.; Chen, C.; Yang, J.; Zeng, W.; Li, T.; Chen, H.; Huang, S.; Fu, Z.; Li, J.; et al. miR-100-5p-abundant exosomes derived from infrapatellar fat pad MSCs protect articular cartilage and ameliorate gait abnormalities via inhibition of mTOR in osteoarthritis. Biomaterials 2019, 206, 87–100.
    75. Tao, S.-C.; Yuan, T.; Zhang, Y.-L.; Yin, W.-J.; Guo, S.-C.; Zhang, C.-Q. Exosomes derived from miR-140-5p-overexpressing human synovial mesenchymal stem cells enhance cartilage tissue regeneration and prevent osteoarthritis of the knee in a rat model. Theranostics 2017, 7, 180.
    76. Jin, Z.; Ren, J.; Qi, S. Human bone mesenchymal stem cells-derived exosomes overexpressing microRNA-26a-5p alleviate osteoarthritis via down-regulation of PTGS2. Int. Immunopharmacol. 2020, 78, 105946.
    77. Chen, X.; Shi, Y.; Xue, P.; Ma, X.; Li, J.; Zhang, J. Mesenchymal stem cell-derived exosomal microRNA-136-5p inhibits chondrocyte degeneration in traumatic osteoarthritis by targeting ELF3. Arthrit. Res. Ther. 2020, 22, 1–13.
    78. Dong, J.; Li, L.; Fang, X.; Zang, M. Exosome-Encapsulated microRNA-127-3p Released from Bone Marrow-Derived Mesenchymal Stem Cells Alleviates Osteoarthritis Through Regulating CDH11-Mediated Wnt/β-Catenin Pathway. J. Pain Res. 2021, 14, 297–310.
    79. Jin, Z.; Ren, J.; Qi, S. Exosomal miR-9-5p secreted by bone marrow-derived mesenchymal stem cells alleviates osteoarthritis by inhibiting syndecan-1. Cell Tissue Res. 2020, 381, 99–114.