Objective: To explore the effect of N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide(W7) on the differentiation from human adipose-derived mesenchymal stem cells (hADSCs) to endothelial cells.
Methods: hADSCs were cultured with serum-free differential medium containing 40 ng/ml vascular endothelial growth factor (VEGF) and 10ng/ml basic fibroblast growth factor (bFGF). Cells were divided into control group (differential medium without W7), high-dose group (containing 30 μmol/L W7), medium-dose group (containing 20 μmol/L W7), and low-dose group ( containing 10 μmol/L W7). The hADSCs were cultured for 8 days, and then the changes in the phenotypes of von Willebrand factor (vWF) and vessel-selective cadherin (VE-Cadherin) were detected by flow cytometry (FCM). The intracellular Ca(2+) labeled with Fluo-3 was detected by laser confocal microscopy. After hADSCs planting on Matrigel, their angiogenic potentials were observed under the inverted phase contrast microscope, and the expression of extracellular regulated kinase (ERK) and phosphorylated extracellular regulated kinase (p-ERK) were evaluated by Western blot.
Results: After the hADSCs were cultured for 8 days, compared with the control group, the expressions of vWF and VE-Cadherin significantly increased along with the decrease of W7 level and the intracellular Ca(2+) also significantly increased (Pü0.01). Lumina-like vascular structure was formed in W7 treatment groups, but not in the blank control group. Compared with the blank control group, the expression of ERK showed no significant in W7 treatment groups (high-, medium-, and low-dose groups)(P>0.05); however, along with the decrease of W7 levels, the expression of p-ERK significantly increased(P<0.05).
Conclusion: W7 in proper levels can effectively induce the differentiation from hADSCs to endothelium by increasing the intracellular Ca(2+) level and thus activating the ERK/MAPK pathway.