Fig. 5
The miR-204/-211-Runx2 axis in regulation of NGF-Akt signaling and mesenchymal progenitor cell proliferation. a Western blot analysis shows activated Akt signaling in dKO CD45- BMSCs. The numbers below the blot indicate the relative levels of the denoted protein (except p-Akt normalized to total Akt, all proteins were normalized to β-actin) in each group compared to the control. n = 3. b IHC results of p-Akt in control and dKO mice. Blue arrowheads mark p-Akt-positive cells. Scale bar, 250 μm (left) or 50 μm (right). c qRT-PCR analysis of Ngf mRNA expression in CD45− BMSCs from control or dKO mice. **P < 0.01, unpaired Student’s t test. n = 3. d IHC results of NGF in control and dKO mice. Red arrowheads, NGF-positive cells. Scale bar, 50 μm. e qRT-PCR analysis of Ngf mRNAs shows that miR-204 or Runx2 siRNA decreases Ngf expression in CD45− BMSCs. ***P < 0.001, one-way ANOVA followed by the Tukey−Kramer test. n = 3. f Retroviral overexpression of two NGF isoforms in CD45− BMSCs from WT mice activates Akt signaling as shown by western blot. n = 3. g NGF overexpression increases proliferation of CD45− BMSCs determined by cell number counts. ***P < 0.001, one-way ANOVA. n = 3. h qRT-PCR analysis of CDK inhibitors in mouse CD45− BMSCs with GFP or NGF overexpression. **P < 0.01, ***P < 0.001, unpaired Student’s t test. n = 3. i Western blot analysis shows that Ngf knockdown downregulates Akt signaling in mouse CD45− BMSCs. n = 3. All data are shown as the mean ± s.d. Uncropped western blot scans are shown in Supplementary Data 1
