Fig. 4.

Effects of cholesterol and caveolin-1 (CAV-1) perturbation on caveolae content in MSCs. Sucrose density centrifugation was used to fractionate membrane preparations derived from the five MSC groups: untreated MSCs, cholesterol-depleted MSCs (MβCD), cholesterol-enriched MSCs (Chol), MSCs transfected with control, nonspecific siRNA (si Ctrl), and MSCs transfected with CAV-1 siRNA (si CAV-1). Upon ultracentrifugation, buoyant membrane rafts float to the upper fractions (4–7) of the 12-fraction sucrose gradient and are separated from intracellular fractions (9–12). Caveolae are identified on the basis of CAV-1 content. a CAV-1 immunoblotting of the fractions. This result is representative of three, pooled human MSC donor sources tested (Additional file 1: Table S1). b CAV-1 signal/total protein amount used for fractionation for each experimental group analyzed by densitometry. Data represent mean ± SD of all experimental replicates, analyzed using NIH ImageJ; n > 3. These results indicate that changes in membrane cholesterol levels significantly affect the caveolae content in MSCs. *p < 0.05, **p < 0.001, versus untreated group; ^#p < 0.001, versus si Ctrl group