Figure 4
Rejuvenation of elderly AT-MSCs with infant AT-MSC-derived extracellular vesicles by inhibiting the accumulation of ROS. (A) The analysis of EVs under the transmission electron miscroscope. Bars indicated 200 nm (n = 5). (B) The particle size of EVs examined by a fiberoptic dynamic light scattering photometer (FDLS-3000) (n = 5). (C) The marker expression of EVs examined by Western blotting (n = 5). (D) The incorporation of PKH26-labeled EVs into elderly AT-MSCs, magnification 4 ×. Bar indicates 200 μM (n = 5). (E) The incorporating efficiency examined by a FACS analysis (n = 5). (F) The growth curve of AT-MSCs (n = 5). (G) The cellular senescence of elderly AT-MSCs with EV incorporation (n = 5). (H) The ROS expression of elderly AT-MSCs with EV incorporation (n = 5). (I) The SOD1 and SOD3 protein expression of elderly AT-MSCs with EV incorporation (n = 5). (J) The mRNA expression of growth factors in elderly AT-MSCs with EV incorporation (n = 5). (K) The transplantation study of elderly AT-MSCs with EV incorporation in streptozotocin-induced diabetic mice (n = 5). (L) The survival rate of db/db mice transplanted with AT-MSCs (n = 5). (M) The necrotic area of db/db mice transplanted with AT-MSCs. (N) Quantification of the necrotic area of db/db mice on day 10 post-transplantation with AT-MSCs (n = 5). In all above experiments, infant AT-MSCs and elderly AT-MSCs were derived from 5 different donors, respectively (n = 5). AT-MSCs were at passage 3 to passage 8, and the comparison was conducted with infant AT-MSCs and elderly AT-MSCs at the same passage number. EVs were isolated from infant AT-MSCs or elderly AT-MSCs from passage 4 to passage 6. The data represent the mean ± SD. ***P < 0.001, **P < 0.01, *P < 0.05, ns no significance. The experiments were performed in triplicate. Full-length Western blots are presented in Supplementary Figure S5.
