Fig. 2

Mitochondrial ROS production in control and patient fibroblasts after co-culturing with mesenchymal stem cells (MSC) or treatment with MSC supernatant. A ROS production as percentage (%) fluorescence intensity of control under steady-state conditions in control fibroblasts (Control) or patient fibroblasts carrying variants in either MT-ND6 (MT-ND6) or in MT-ND3 (m.10191T>C—MT-ND3a; m.10197G>A MT-ND3b) and after 72 h of co-cultivation with MSCs (+). ROS level elevation in patient cells is mitigated by co-culturing with MSCs. B ROS production under steady-state conditions and 10 days after removal of MSCs from the co-culturing system by blasticidin treatment. The mitigation of ROS level elevation persists even after MSC removal. C ROS production under steady-state conditions and after 72 h treatment with MSC supernatant (+SN). Supernatant treatment reduces ROS production in patient fibroblasts. D ROS production under steady-state conditions and after 72 h treatment with MSC supernatant followed by subsequent removal of MSC supernatant (SN) and culturing with regular medium (after SN) for 8 days. After removal of MSC supernatant, ROS production is similar to that under steady-state conditions. E ROS production under steady-state conditions compared to 72 h treatment with MSCs in a transwell (+TW) system. Paracrine effects of MSCs lead to reduction of ROS levels in patient fibroblasts. F ROS production under steady-state conditions and after 72 h treatment with MSCs in transwell system followed by subsequent removal of insert plate and culturing with regular medium (after TW) for 8 days. After removal of MSC, ROS production reduced to a lesser extent in patient MT-ND3b and MT-ND6. MT-ND3a levels are similar to untreated levels. Date are shown as mean ± SEM. **p < 0.01, ***p < 0.001. G Representative Western Blot results from cytosolic fractions analysed for SOD2 and SDHA protein levels in cells with (+) and without MSC treatment. H Quantification of SOD2 levels using the original blots from five independent experiments. Data are expressed as mean ± SD. *p < 0.05, ***p < 0.001. I Representative Western Blot results from cytosolic fractions analysed for HO-1, NQO1, GAPDH and α-Tubulin protein levels in cells with (+) and without MSC treatment. J Quantification of NQO1 levels using the original blots from four independent experiments. Data are expressed as mean ± SD. *p < 0.05, **p < 0.01, ***p < 0.001. K Quantification of HO-1 levels using the original blots from three independent experiments. Data are expressed as mean ± SD. *p < 0.05, **p < 0.01