Figure 6. Comparison of peritoneal fibrosis in EZH2-WT and EZH2-KO mice after high glucose PDF injury.

EZH2 knockout mice were generated by breeding our EZH2^loxP/loxP mice with tamoxifen-inducible Col1a2-Cre mice (Col1a2-CreER^+/−) and then backcrossing progeny to EZH2^loxP/loxP mice to obtain Col1a2-Cre⁺: EZH2^loxP/loxP mice (EZH2-KO) and Col1a2-Cre^-: EZH2^loxP/loxP mice (EZH2-WT). Both of EZH2-KO and EZH2-WT mice were received daily intraperitoneal injection of 100ml/kg peritoneal dialysis solution with 4.25% glucose for 28 days to establish peritoneal fibrosis model. (A) Photomicrographs (200X) show Masson trichrome staining of the peritoneum in each group. (B) The graph shows the positive area of the Masson-positive submesothelial area (blue) from ten random fields of six mice peritoneal samples. (C) The graph shows the thickness of the compact zone measured from ten random fields of six mice peritoneal samples. (D) Peritoneum tissue lysates were prepared and subjected to immunoblot analysis with antibodies against α-SMA, Collagen I, EZH2, H3K27me3, Histone H3 or GAPDH. (E) Expression levels of α-SMA, Collagen I and EZH2 were quantified by densitometry and normalized with GAPDH. (F) Expression level of H3K27me3 was quantified by densitometry and normalized with Histone H3. (G) Peritoneum tissue lysates were prepared and subjected to immunoblot analysis with antibodies against TIMP2, MMP2, MMP9, or GAPDH. (H) Expression levels of TIMP2, MMP2 and MMP9 were quantified by densitometry and normalized with GAPDH. Data are represented as the mean ± S.E.M (n = 6). Means with different superscript letters are significantly different from one another (P< 0.05). All scale bars = 20 μm.