Fig 1.
Characterization of iMSCs and ADMSCs by flow cytometry and differentiation potential.
(A) Histograms of flow cytometric analysis showing the expression of positive human MSC surface markers and the negative markers cocktail for iMSCs and ADMSCs. (B) The percentages of iMSCs expressing the human MSCs markers compared to ADMSCs. (C) Alizarin Red staining of calcium deposits in the differentiated osteogenic cells. (D) Oil Red O staining of fat vacuoles in the adipocyte differentiated cells. Scale bar: 20 μm.
Fig 2.
Characterization of iMSC-EVs and ADMSC-EVs.
(A&B) Flow cytometry histograms and graph analysis of tetraspanin markers CD9, CD81, and CD63 on iMSC-EV and ADMSC-EV. (C) Size distribution of iMSC-EVs and ADMSC-EVs measured by DLS. (D) Representative images of iMSC-EVs and ADMSC-EVs by transmission electron microscopy. Scale bar: 1 μm.
Fig 3.
iMSC-EVs and ADMSC-EVs are internalized by HDFs.
Representative images showing the uptake of DiI-labeled iMSC-EVs and ADMSC-EVs by human dermal fibroblasts (HDFs). DiI, a lipophilic dye, was used to stain EV membranes before incubation with HDFs. Scale bar: 50 μm.
Fig 4.
Effect of iMSC-EVs and ADMSC-EVs on cell viability.
(A) MTT results of HDF samples treated with iMSC-EVs, ADMSC-EVs and HDF-EVs after 48 and 72 hours of cell seeding. (*p-value ≤ 0.05, ** p-value ≤ 0.01, ***p-value ≤ 0.001). The effect of iMSC-EVs and ADMSC-EVs on cell apoptosis of (B) HDF cells and (C) ADMSCs as illustrated by the graphs showing the percentage of apoptotic cells after treatment with iMSC-EVs and ADMSC-EVs (**p-value ≤ 0.01). (D) Representative images of SA β-Gal staining for senescent HDF cells in the presence or absence of iMSC-EVs and ADMSC-EVs. Scale bar: 200 μm. (E) The percentage of senescent cells after EVs treatment calculated as the number of SA-β-Gal positive cells divided by the numver of total cells.
Fig 5.
The effect of iMSC-EVs and ADMSC-EVs on wound healing.
(A) Representative ADMSC images showing the scratched area at different time points 0, 24, and 48 hours (Evos, 4X magnification). Scale bar: 200 μm. (B) The quantification of scratched area μm2 in ADMSC cultures. (* p < 0.05, **** p-value <0.0001, ns-not significant). (C) Representative HDFs images showing the scratched area at 0, 24 and 48 hours (Evos, 4 X magnification) Scale bar: 200 μm. (D) The quantification of the scratched area in HDF cultures as a percentage.