Fig. 8: Modulation of bone morphogenetic receptor 2 (BMPR2) signaling pathway by Axl in human pulmonary arterial endothelial cells (hPAECs).

a, b Western blots and subsequent densitometry quantification of phospho-Axl, BMPR2, and ID1 after R428 treatment followed by Gas6 or BMP9 (2 h). Data from n = 3 biological independent experiments are presented as the n-fold change compared with DMSO-treated control (co) cells, defined with opened circles. Green and purple circles define Gas6 and BMP9. Light gray and dark gray circles define R428 + BMP9- and R428 + Gas6-treated conditions, respectively. c, d Representative western blots of Axl, BMPR2, and pSMAD1/5/8 and quantitative analyses of pSMAD1/5/8 expression upon the small interfering RNA (siRNA) treatment of Axl and BMPR2 in hPAECs. Opened circles define si scrambled (si co) cells, green and purple circles define BMP9 and Gas6 cells, respectively. Light gray and dark gray circles define siAxl and siBMPR2 cells, respectively. Data from n = 3 biological independent experiments are presented as the n-fold change compared with untreated control (co) cells. e Quantitative real-time PCR analyses of ID1 and ID2 after siRNA-mediated knock-down of Axl and BMPR2 and followed by BMP9 and Gas6 stimulations. Data from n = 3 biological independent experiments are presented as the n-fold change (2^−∆∆Ct) compared with scrambled siRNA control (si co). Statistical analyses were performed using one-way ANOVA with Tukey’s or Newman-Keuls post hoc test for multiple comparisons. a–e ^***P < 0.001, ^****P < 0.0001 versus untreated scrambled siRNA controls (si co) or DMSO-treated control (co) cells; a, b ^#P < 0.05, ^##P < 0.01 R428 + BMP9 and R428 + Gas6 versus R428; c–e ^#P < 0.05, ^##P < 0.01, ^###P < 0.001, ^####P < 0.0001 for si co + BMP9, si co + Gas6 versus si Axl + Gas6/BMP9, and siBMPR2 + BMP9/Gas6. In a Pan-actin, in c Vinculin served as a loading control. All the data represent the mean ± SEM.