TGFβ1-Induced Differentiation of Human Bone Marrow-Derived MSCs Is Mediated by Changes to the Actin Cytoskeleton

Figure 5

Inhibition of actin polymerization promotes adipogenic differentiation but inhibits osteogenic differentiation in MSCs. MSCs underwent osteogenic or adipogenic differentiation by culturing cells in the appropriate medium for 7 days. Cells also underwent the indicated treatments. (a) Mineralized calcium deposition, as determined by alizarin red S staining in MSCs that were osteoinduced (OS), osteoinduced with TGFβ1 treatment two days prior to induction (OS + TGFβ1), osteoinduced with CYD treatment at the onset of induction (OS + CYD), or osteoinduced with both TGFβ1 and CYD treatment at the time points described above (OS + TGFβ1 + CYD). Lower panel: micrograph of stained wells. Upper panel: quantification of mineralized matrix formation under the indicated treatment conditions. Data are shown as the mean ± SD of three independent experiments (; ). (b) Gene expression of the osteogenic markers ALPL, RUNX2, and OCN, normalized to GAPDH, as determined by qRT-PCR. Cells were either not induced (NI) or induced under the conditions described in (a). Data are shown as the mean ± SD of three independent experiments (; , ). (c) Adipogenic differentiation of MSCs that were adipoinduced (AD), adipoinduced with TGFβ1 treatment 2 days prior to induction (AD + TGFβ1), adipoinduced with CYD treatment, initiated at the onset of induction (AD + CYD), or adipoinduced with both TGFβ1 and CYD treatment at the time points described above (AD + TGFβ1 + CYD). Lower panel: Oil red O staining of the indicated cells. Upper panel: Nile red quantification of oil content under the indicated conditions. (d) Gene expression of the adipogenic marker genes PPARG and LPL, determined by qRT-PCR and normalized to GAPDH, under the indicated treatment regimens. Data are shown as the mean ± SD of three independent experiments (, ). All controls were treated with vehicle only.

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