Fig. 3

5-aza-2′-deoxycytidine (5azadC) enhances FLT1 expression and induces sFLT1 secretion in choriocarcinoma cells. Three choriocarcinoma cell lines were incubated for 5 days in the presence of 0.1% DMSO (vehicle control) or 10 μM 5azadC. Culture media were changed daily and on the fifth day the conditioned media were collected for ELISA and Western blotting. a The mRNA expression levels of three FLT1 splice variants in the three choriocarcinoma cell lines. Results are expressed as a ratio relative to the expression of GAPDH mRNA. b Measurement of sFLT1 secreted from the three choriocarcinoma cell lines. The amount of sFLT1 in the conditioned media was measured by ELISA, and was then normalized to the protein content of each cell. c Immunoprecipitation of FLT1 isoforms from the cell lysates. Numbers represent the amount of protein subjected to immunoprecipitation. The indicated percentage of cell lysate was used as the input. d Western blotting of sFLT1 proteins secreted into the conditioned media. Numbers show the volume of media subjected to heparin-affinity pull-down. Uncropped images of Western blots are presented in Additional file 1: Figure S6. All values represent the mean ± SD (n = 3). Asterisks indicate a significant difference (P < 0.05). ND: not detected