Fig. 3.

RASSF1A promotes proliferation and glycolytic shift in hypoxic human PASMCs. Human PASMCs (a–c) were transfected with a RASSF1 siRNA (si-RASSF1) and control siRNA (si-Control) or c si-RASSF1 and HIF1α siRNA (si-HIF1A) in combination or b RASSF1A-FLAG and empty vector (EV). 24 h after transfection, cells were exposed to normoxia or hypoxia for 48 h and proliferation was measured by BrdU incorporation assay. Human PASMCs were transfected with d, e si-RASSF1 and si-Control or f, g RASSF1A-FLAG and EV. 24 h after transfection, cells were exposed to normoxia or hypoxia for 24 h. d, f Real time PCRs for indicated genes were performed. e, g Cell lysates were subjected to (left) western blotting, followed by (right) densitometric quantification of relative PDK1, LDHA and HK2 to ACTB expression. h HEK293 cells were transfected with RASSF1A-FLAG or EV and 24 h after transfection, exposed to normoxia or hypoxia for 24 h and intracellular lactate production was measured. *P < 0.05, **P < 0.01, ***P < 0.001 compared to a, c–e si-Control (hypoxia) or b, f–h EV (hypoxia), one-way ANOVA followed by SNK multiple comparison test. c ^§P < 0.01 compared to si-RASSF1, two-way ANOVA. n = 3 independent experiments from 3 biological replicates each; data represent mean ± s.e.m.