Apoptotic vesicles (apoVs) are apoptotic cell‐derived nanosized vesicles that play a crucial role in multiple pathophysiological settings. However, their detailed characteristics, specific surface markers, and biological properties are not fully elucidated. In this study, we compared mesenchymal stem cell (MSC)‐derived apoVs and exosomes from three different types of MSCs including human bone marrow MSCs (hBMSCs), human adipose MSCs (hASCs), and mouse bone marrow MSCs (mBMSCs). We established a unique protein map of MSC‐derived apoVs and identified the differences between apoVs and exosomes in terms of functional protein cargo and surface markers. Furthermore, we identified 13 proteins specifically enriched in apoVs compared to exosomes, which can be used as apoV‐specific biomarkers. In addition, we showed that apoVs inherited apoptotic imprints such as Fas to ameliorate haemophilia A in factor VIII knockout mice via binding to the platelets’ FasL to activate platelet functions, and therefore rescuing the blood clotting disorder. In summary, we systemically characterized MSC‐derived apoVs and identified their therapeutic role in haemophilia A treatment through a previously unknown Fas/FasL linkage mechanism.

Mesenchymal stem cells (MSCs) are easily isolated from tissues and have the ability to self-proliferate and to differentiate along different cell lineages, such that they are commonly used in tissue regeneration. [10][11][12][13][14] Several clinical studies have shown that MSCs are safe and effective when used in tissue repair and wound healing. [15][16][17][18][19][20][21] However, despite the therapeutic efficacy of MSCs, there are significant costs and challenges associated with their therapeutic use, because they require strict monitoring of manufacturing, processing, and storage to ensure optimal viability and potency following transplantation. [22,23] Some studies have shown that stem cells transplanted into the body initially undergo significant apoptosis [24][25][26][27][28][29] and that the direct transplantation of apoptotic stem cells can have lasting effects. [30] Apoptosis plays an important role in maintaining physiological homeostasis. [31][32][33] Apoptotic vesicles (apoVs) are extracellular vesicles rich in proteins, RNA, and lipids that are released during apoptosis. [34] Their role is to mediate the transfer of substances and signal exchange between cells and thus maintain homeostasis. In a previous study, we examined the characteristics and specific markers of MSC-derived apoVs and found that proteins on the surface of apoVs regulated platelet aggregation. [35] In another study, we demonstrated the therapeutic effect of MSC-derived apoVs, including in type 2 diabetes, through their interactions with hepatic macrophages. [36] We also found that apoVs activate the Fas pathway in multiple myeloma cells and thus induce apoptosis in these tumor cells. [37] Pluripotent stem cell (PSC)-derived apoVs (PSC-apoVs) can inherit pluripotent molecules from PSCs to stimulate adult stem cells and promote wound healing in mouse skin. [38] Exogenous apoVs (apoEVs) have been reported to promote wound healing and hair growth by activating the Wnt/β-catenin pathway. [39] The administration of apoVs can prevent Th17 differentiation and memory T cell formation to ameliorate inflammation and joint erosion in a mouse model of arthritis. [40] While mouse MSC-derived apoVs has been used to treat osteoporosis, [41] whether apoVs derived from human bone marrow mesenchymal stem cells (hBMMSCs) can impact bone metabolism is unknown. Thus, we investigated the regulatory role of apoVs in bone metabolism and therapeutic effects on bone defects and bone loss. Our results provide an important theoretical basis for novel clinical applications of hBMMSCs-derived apoVs. Mesenchymal stem cells (MSCs) are widely used in the treatment of diseases. After their in vivo application, MSCs undergo apoptosis and release apoptotic vesicles (apoVs). This study investigates the role of apoVs derived from human bone marrow mesenchymal stem cells (hBMMSCs) in bone metabolism and the molecular mechanism of the observed effects. The results show that apoVs can promote osteogenesis and inhibit osteoclast formation in vitro and in vivo. ApoVs may t...

Background In tissue engineering, mesenchymal stem cells (MSCs) are common seed cells because of abundant sources, strong proliferation ability and immunomodulatory function. Numerous researches have demonstrated that MSC-macrophage crosstalk played a key role in the tissue engineering. Macrophages could regulate the differentiation of MSCs via different molecular mechanisms, including extracellular vesicles. Apoptotic macrophages could generate large amounts of apoptotic vesicles (apoVs). ApoVs are rich in proteins, RNA (microRNAs, mRNAs, ncRNAs, etc.) and lipids, and are a key intercellular communication mediator that can exert different regulatory effects on recipient cells. MiRNAs account for about half of the total RNAs of extracellular vesicles, and play important roles in biological processes such as cell proliferation and differentiation, whereas the functions of macrophage-derived apoVs remain largely unknown. There was no research to clarify the role of macrophage-derived apoVs in MSC fate choices. In this study, we aimed to characterize macrophage-derived apoVs, and investigate the roles of macrophage-derived apoVs in the fate commitment of MSCs. Methods We characterized macrophage-derived apoVs, and investigated their role in MSC osteogenesis and adipogenesis in vitro and in vivo. Furthermore, we performed microRNA loss- and gain-of-function experiments and western blot to determine the molecular mechanism. Results Macrophages could produce a large number of apoVs after apoptosis. MSCs could uptake apoVs. Then, we found that macrophage-derived apoVs inhibited osteogenesis and promoted adipogenesis of MSCs in vitro and in vivo. In mechanism, apoVs were enriched for microRNA155 (miR155), and apoVs regulated osteogenesis and adipogenesis of MSCs by delivering miR155. Besides, miR155 regulated osteogenesis and adipogenesis of MSCs cultured with macrophage-derived apoVs via the SMAD2 signaling pathway. Conclusions Macrophage-derived apoVs could regulate the osteogenesis and adipogenesis of MSCs through delivering miR155, which provided novel insights for MSC-mediated tissue engineering.

hBMMSC-apoVs can promote mesenchymal stem cell (MSC) osteogenesis and inhibit osteoclast formation in vitro and in vivo. The mechanisms include the release of miR1324, which inhibits the expression of the target gene SNX14, thereby activating the SMAD1/5 pathway in target cells. In article number 2205813, Xiao Zhang, Yunsong Liu, and co-workers suggest a therapeutic strategy for the treatment of bone loss, including bone tissue engineering through a cell-free approach.