The number of capillaries selleck chemicals MG132 in each muscle fiber increased in the mice treated with either MSCs or simvastatin alone in comparison with the control group. The combined administration of simvastatin and MSCs resulted in the highest capillary density. Combination therapy enhances the differention of MSCs into endothelial cells in ischemic muscles To determine whether improved limb reperfusion by simvastatin and MSCs co therapy was associated with differentiation into endothelial cells, the number of incorporated DiI labeled MSCs into the mouse microvascular was detected by fluorescent stain ing against vWF. Histological and quantitative analyses showed that the number of incorporated MSCs was significantly greater in the com bination group relative to MSCs alone.
Combination therapy decreases cell apoptosis in vivo To determine whether improved limb reperfusion by the simvastatin/MSCs treatment was associated with increased ischemic muscle cells survival in Inhibitors,Modulators,Libraries vivo, the cell apoptosis was assessed in the ischemic muscle at days 21 after the treatment by TUNEL assay. Apoptosis as measured by TUNEL positive nuclei was signifi cantly decreased in ischemic muscle of simvastatin and MSCs treated mice versus vehicle treated mice. The co treatment of simvastatin and MSCs resulted in a further decrease of cell apoptosis. Combination therapy enhances the expression of VEGF protein in ischemic tissue To examine whether high dose simvastatin and MSCs co therapy improved postischemic neovascularization, the expression of VEGF protein was detected by western blot assay.
As can be seen in figure 6, the expression of VEGF significantly increased in the simvastatin group than in the control group. Moreover, the expression of VEGF was higher in MSCs group com pared with that in the simvastatin group, but was lower than that in the combination group. Effect of simvastatin on the cell viability of bone marrow derived Inhibitors,Modulators,Libraries MSCs in vitro In vitro, serum starvation induced bone marrow derived MSCs apoptosis, as indicated Inhibitors,Modulators,Libraries by flow cytometry and MTT assay. When incubated with 0. 01 umol/L of simvastatin, Inhibitors,Modulators,Libraries the percentage of apoptotic Inhibitors,Modulators,Libraries cells decreased and the viabi lity was visibly upregulated. However, pretreatment with 50 n M wortmannin, a PI3 K inhibitor, diminished the anti apoptotic effect of simvastatin. The cell viabi lity detected by MTT assay was significantly higher in sim vastatin treated group than that in the control group. Although the cell viabilities were higher in simvastatin wortmannin group than those in the control group, there was no significant difference between the two groups. These results indicated that PI3 K pathway was of impor tance for the anti ref 3 apoptotic role of simvastatin.