Figure 4

Endocytosis inhibitors disrupt the actin cytoskeleton of fMSC. The morphology of the actin cytoskeleton was visualized with Alexa 568-conjugated phalloidin after incubation for 1 hr in serum-free (treatment) media, or treatment media + vehicle, blebbistatin or dynasore. A) Cells at high confluence are less prone to cytoskeletal change, whereas cells at lower confluence are impacted more by inhibitors (B and C shows enlargement of key regions of B). D) A schematic illustration of C, showing actin filaments as white lines, the outline of each cell as a red line, and membrane ruffles as blue lines or spots. A normal, filamentous actin cytoskeleton is seen in fMSC with treatment media. Treatment with vehicle (DMSO) has some impact on cytoskeletal integrity, with the partial loss of long actin filaments in most cells. Blebbistatin and dynasore treatment dramatically disrupts the actin cytoskeleton in most cells. The cells develop membrane ruffles, neural-like projections and begin detaching from the culture dish (x40 magnification).