Fig. 5
From: YTHDF2 reduction fuels inflammation and vascular abnormalization in hepatocellular carcinoma
YTHDF2 suppresses HCC by targeting IL11 and SERPINE2 mRNAs in a m6A-reader fashion. (a) An empty vector (EV) or vectors encoding wild-type (WT) or mutant YTHDF2 (W432A and W486A) were transduced into SMMC7721 cells with a YTHDF2-knockdown (KD) background. Proliferative activity of indicated SMMC7721 cells was assessed by CCK8 (left) or WST1 assay (right). n = 5 biological replicates. (b) Numbers of endothelial tubes formed by HUVEC cocultured with indicated SMMC7721 cells grown for 24 h under Nx or Hx. n = 3 biological replicates. (c) RT-qPCR analysis of the relative mRNA levels of IL11 (left) and SERPINE2 (right) in indicated SMMC7721 cells grown for 24 h under Nx or Hx. n = 3 biological replicates. (d) ELISA analysis of the quantitative protein levels of IL-11 in the culture supernatant in indicated SMMC7721 cells grown for 24 h under Nx or Hx. n = 3 biological replicates. (e) Immunoblots showing STAT3 phosphorylation and Serpin E2 expression in indicated SMMC7721 cells grown for 24 h under Nx or Hx. n = 2 independent experiments. (f) Indicated SMMC7721 cells were injected subcutaneously into NPG mice and tumor volumes were measured at indicated dates. n = 5 mice. (g) Control shRNA or shRNAs targeting IL11 and SERPINE2 were transduced into SMMC7721 cells with a YTHDF2-knockdown (KD) background. Proliferative activity of indicated SMMC7721 cells was assessed by CCK8 (upper) or WST1 assay (lower). n = 5 biological replicates. (h) Numbers of endothelial tubes formed by HUVEC cocultured with indicated SMMC7721 cells. n = 3 biological replicates. (i) Indicated SMMC7721 cells were injected subcutaneously into NPG mice and tumor volumes were measured at indicated time points. n = 5 mice. (j) Immunohistological staining of IL-11 and SerpinE2 in human HCC tissue arrays divided as YTHDF2Low and YTHDF2High categories according to median integrated optical density (IOD) value of YTHDF2. n = 143 patients. Scale bar, 200 μm. Error bars indicate means ± SEM *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001. P-values were determined by two tailed t-test