Fig. 4
Activation of BMPR2 signaling by BMP9 partially rescues RAD51 and reduces sensitivity to DNA damage agent. a Pulmonary microvascular endothelial cells (PMVECs) were treated with vehicle (Con, H2O) or mitomycin C (MMC) with or without BMP9 (10 ng/mL) for 14 h. RAD51 and BMPR2 protein relative to GAPDH were examined by immunoblot. γH2AX was studied to measure the amount of double-strand breaks. Representative image and the quantitation of six independent experiments (n = 6) are shown. b RAD51 and BMPR2 protein amount in PMVECs treated with vehicle (H2O, Con) or MMC with or without bone morphogenetic protein receptor kinase inhibitor LDN193189 (LDN; 100 nM) in combination with BMP9 for 14 h were examined by immunoblot and the quantitation of three independent experiments are shown (n = 3). γH2AX represents the amount of double-strand breaks. GAPDH was used for normalization. c RAD51 and GAPDH (loading control) protein amount in PMVECs treated with or without LDN193189 (100 nM) for 72 h. Representative image and the quantitation of three independent experiments are shown (n = 3). d PMVECs were treated with or without LDN193189 (100 nM) for 72 h, and the amount of DNA damage was measured by single-cell gel electrophoresis (alkaline comet assay). The percentage of cells with DNA damage was analyzed using ImageJ software. Representative images of alkaline comet assay and the quantitation of 136–162 cells are shown. Bars represent mean ± SEM from six different experiments per conditions in (a) and from three different experiments per conditions in (b–d). *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001 versus respective control. One-way ANOVA followed by Tukey’s multiple comparisons test was used in (a, b). Unpaired two-tail t-test was used in (c, d)
