Fig. 4

MSC-secreted Wnt4 and Wnt5a were both involved in the response of MS-1 cells to oxidative stress but may have opposing effects. a, b Confirmation of the knockdown efficiency of Wnt4-siRNA and Wnt5a-siRNA using qPCR and western blotting analysis. c MTT cell viability assay suggested that knockdown of Wnt5a further enhanced the beneficial effects of MSC-CM, whereas knockdown of Wnt4 did the opposite. d, e Transwell culturing of MSCs with MS-1 showed a similar trend; knockdown of Wnt5a in the MSCs improved their anti-apoptosis properties, and knockdown of Wnt4 hampered those effects (d shows the results of double staining of annexin V/PI flow cytometry and e shows the results of TUNEL staining). f Confirmation of changes in the apoptosis of MS-1 after Wnt5a/Wnt4 knockdown in the MSCs by cleaved caspase 3 western blotting analysis, together with the western blotting results of changes in the endothelium function indicated by p-eNOS, ICAM, and VCAM. Quantification of bands performed using ImageJ software. *p < 0.05, **p < 0.001. CM conditioned medium, NC negative control, siRNA silencing RNA, t-eNOS total endothelial nitric oxide synthase, p-eNOS phosphorylated-endothelial nitric oxide synthase, ICAM intercellular cell adhesion molecule, MSC, PI propidium iodide, VCAM vascular cell adhesion molecule