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. Author manuscript; available in PMC: 2017 Sep 1.
Published in final edited form as: Bone. 2016 Jun 4;90:80–89. doi: 10.1016/j.bone.2016.06.003

Figure 6. TNF-induced proliferation of Hes1-GFP+ MSCs is prevented by PDGFRβ inhibition.

Figure 6

6-month-old Hes1-GFP/TNF-Tg mice and Hes1-GFP littermate control mice were injected with BrdU and bone marrow cells were stained with anti-BrdU or Annexin V antibodies plus or minus Propidium iodide (PI). CD45−/GFP+ cells were gated and the % of BrdU+ (A) and AnnexinV+/PI− cells (B) for proliferation and apoptosis were assessed, respectively. (C) GFP+ cells expressing PDGFRβ were examined for the percentage and mean fluorescence intensity (MFI). Values are mean plus SD of 5 mice. *p < 0.05 vs data from Hes1-GFP mice. (D) Bone marrow cells from Hes1-GFP mice were cultured to form CFU-F colonies and incorporated with BrdU. Cells were treated with TNF (10 ng/ml), PDGFRβ inhibitor (su16f, 1mM), or TNF plus su16f for 24 hours. BrdU+ cells were identified by IHC with anti-BrdU antibody. The percentage of BrdU+ within GFP+ cells and their GFP intensity were measured. Values are mean plus SD of 3 wells. Experiments were repeated 2 times. *p < 0.05 vs. PBS, and #p < 0.05 vs. TNF. (E) C3H10T1/2 cells were treated with TNF and/or Thapsigargin (Thap) for 12 hours. Expression of Hes1, PDGFRβ and Cyclin D1 protein levels were examined by Western blot. Experiments were repeated 2 times. (F) A cartoon showing potential relationship among TNF, Hes1 (Notch), PDGFRβ, and cell proliferation.