Fig. 1
Mitochondria of MSCs successfully transferred to H2O2-induced tenocytes alleviate in vitro apoptosis. A Representative fluorescence images showing MitoTracker Red CMXRos-labeled MSC mitochondria (red) in tenocytes (green). Nuclei are counterstained with Hoechst 33,342 (blue). Scale bars: 200 × , 25 µm; 400 × , 10 μm (magnified graphs). B Flow cytometry results displaying annexin V-/PI- (viable), annexin V + /PI- (early apoptotic), annexin V + /PI + (late apoptotic) , or annexin V-/PI + (necrotic) tenocytes (n = 3). C Tenocyte proliferation was determined using CellTrace™ Violet by flow cytometry (n = 6). D Gene expression of proliferation factor Ki67. The relative gene expression level (Gene/GAPDH relative to the control) was measured by RT-qPCR (n = 3). MSC treatment resulted in an increase of anti-apoptotic gene Bcl-2 (E), and decrease of pro-apoptotic genes, including Bax (F), caspase 3 (G), caspase 9 (H), and Cyt-c (I), in H2O2-injured tenocytes (E-I, n = 3). Mitochondrial transfer prevented H2O2-induced mitochondrial apoptosis gene expression, compared to the mitochondrial transfer interference by CB. Bcl-2 protein expression (protein/β-actin relative to control) (K, O) was shown by western blot analysis to be significantly upregulated in MSC group, whereas expression of AIF (J, M), Smac/DIABLO (J, M), Cyt-c (J, N), Bax (K, O), caspase 3 (L, Q), and caspase 9 (L, Q) were downregulated, compared to H2O2-treated tenocytes (J-L, n = 3). P Expression rate of Bcl-2 and Bax. Data represent mean ± SD. Statistical significance of the differences among groups was determined by ANOVA (Bonferroni or LSD method). *P < 0.05, **P < 0.01, ***P < 0.001