Fig. 3

PLCE1 promotes proliferation and angiogenesis of ESCC cells in vitro and induces aggressiveness in vivo. a Eca109 and EC9706 cells treated with U73122 at 0, 2, 5, and 10 concentrations for 24 and 48 h. ShR-PLCE1 were transfected at a MOI of 15 for 60 h. PLCE1 expression measured by Western blot. b Real-time PCR analysis demonstrating relationship between PLCE1 expression and apoptosis. Color represents intensity scale for vector of PLCE1 shRNA versus control, as calculated by log2 transformation. c Eca109 and EC9706 cells treated shR-PLCE1 or U73122 at the indicated concentration for 0, 24, 48, 72, and 96 h. Cell viability measured by MTT and presented as means ± SD from three separate experiments. d TUNEL staining of cells treated as indicated; e shR-PLCE1 on apoptosis-related proteins assayed by Western blot. β-Actin was a loading control. f Real-time PCR analysis demonstrated the positive relationship between PLCE1 expression and angiogenesis. Pseudo-color represents intensity scale for the vector or PLCE1 shRNA versus control, as calculated by log2 transformation. g Tube formation by indicated cells. h Effects of shR-PLCE1 on VEGF-C protein expression as detected by Western blot. i Xenograft model in nude mice; representative images of tumors from all mice in each group. Mean tumor weights j and tumor volume growth curves k for tumors formed by the indicated cells. l H&E and IHC staining demonstrated that PLCE1 induced the aggressive phenotype of ESCC cells in vivo. Scale bar, 100 μm. Microvascular density m show that PLCE1 promotes resistance to apoptosis and angiogenesis in vivo. All data are presented as mean ± SD. *P < 0.05, ***P < 0.001