Proteomic analysis of MSC‐derived apoptotic vesicles identifies Fas inheritance to ameliorate haemophilia a via activating platelet functions

Abstract

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.

Apoptotic MSCs and MSC-Derived Apoptotic Bodies as New Therapeutic Tools

“…The preparation of ApoBDs from MSCs usually includes the induction of apoptosis with staurosporine doses of 250 nM–0.5 μM for 12 h, followed by centrifugation at 800–1000 g to remove cell debris and then at 16000 g to precipitate the ApoBDs [ 108 , 109 , 113 ]. 200 μM H 2 O 2 for 8 h or nutrient deprivation by culturing MSCs in PBS for 24 h are also used as inducers of cell death in MSCs [ 112 ]. Le et al [ 114 ] suggested using high hydrostatic pressure at 50 MPa for 36 h to induce MSC apoptosis ex vivo.…”

“…In recent work, Zhang et al [ 112 ] compared the proteomic profiles of ApoBDs and exosomes derived from three different types of MSCs, including human BM-MSCs, human AD-MSCs, and mouse BM-MSCs and showed that ApoBD protein patterns are 80% matched between BM-MSCs and AD-MSCs. More functional proteins were present in ApoBDs compared to exosomes obtained from the same cells.…”

“…More functional proteins were present in ApoBDs compared to exosomes obtained from the same cells. The authors found 13 proteins that were present in ApoBDs but not in exosomes, including Fas, Integrin alpha-5, Syntaxin-4, CD44, RhoA, Caveolin-1, Cavin1, Rab-5C, RPS25, Lamin B1, VDAC-2, Calnexin, and Calreticulin, as well as one protein present exclusively in exosomes, but not in ApoBDs, namely syntenin-1 [ 112 ]. The main characteristics of different types of EVs are summarized in Table 1 .…”

“…Although ApoBDs induced higher numbers of activated CD62P + platelets and CD62P + platelet-derived microparticles, they failed to elevate more TF + platelets and TF + platelet-derived microparticles. These data suggest that ApoBDs rebalance coagulation through upregulation of platelet activity without altering other hemostatic factors [ 112 ].…”

Extracellular Vesicles for Dental Pulp and Periodontal Regeneration

“…ApoVs are derived from various types of cells, such as precursor cells, endothelial cells, immunocytes, and mesenchymal stem cells (MSCs) [ 19 ]. The buoyant density of ApoVs ranges from 1.118 to 1.228 g/mL, making them denser than sEVs (1.05–1.15 g/mL) and MVs (1.03–1.10 g/mL) [ 20 , 21 ]. Until now, the classification of ApoVs has been a matter of debate.…”