Fig. 3

The differentiation of MSCs to α-smooth muscle actin (αSMA)⁺ cells in response to high mobility group box 1 (HMGB1) stimulation and receptor for advanced glycation end-product (RAGE) inhibition. The population of αSMA⁺ cells was determined by immunofluorescent staining (a) and flow cytometry (b). Both exogenous HMGB1 treatment and pLV-HMGB1 transfection inhibited platelet-derived growth factor (PDGF)-induced differentiation of MSCs to αSMA⁺ cells. The fraction of αSMA⁺ cells was decreased from 75% to 36% upon 100 ng/ml HMGB1 treatment, and from 53% to 18% after pLV-HMGB1 transfection. However, the inhibitory effect was blocked by RAGE knockdown. After the cells were transfected with pshRNA-RAGE1 and pshRNA-RAGE3, the percentages of αSMA⁺ cells increased from 38% to 55% and 61%, respectively. pshRNA-RAGE2 transfection had little effect. The experiments were repeated three times for each group. Group comparisons were made using the Mann-Whitney test. **P < 0.05. DAPI 4′,6-diamidino-2-phenylindole