[Mechanism of cleft palate in C57BL/6N mice induced by 2, 3, 7, 8-tetrachlorodibenzo-p-dioxin].

To explore the molecular mechanism of cleft palate in mice induced by 2, 3, 7, 8-tetrachlorodibenzo-p-dioxin (TCDD). The pregnant mice were randomly divided into TCDD-treated group (=42) and control group (=42). TCDD-treated group was given by gavage a single dose of TCDD (64 μg/kg) at 8: 00 AM on gestation day 10 (GD10) and the control group was given by gavage the isopyknic corn oil. At GD13-GD15, the fetal mice palate development was observed by HE staining. The mouse embryonic palatal mesenchymal cell proliferation was detected by 5-bromo-2-deoxyuridine (BrdU) immunofluorescence. The localization and expression of maternally expressed gene3 (MEG3) in mouse embryonic palatal mesenchymal cells was detected by situ hybridization and real-time PCR (RT-PCR). The key protein expressions of transforming growth factor-β (TGF-β)/Smad signaling pathway in mouse embryonic palatal mesenchyme were analyzed by Western blotting. The interaction of MEG3 and TGF-β receptor Ⅰ (TGF-βRⅠ) was examined by RNA binding protein immunoprecipitation (RIP). At GD13 and GD14, compared with the control group, the ratio of BrdU-positive cells in the palatal mesenchyme of TCDD-treated fetuses decreased significantly (GD13, 6.66, =0.003; GD14, 6.56, =0.003). However, at GD15, the ratio of BrdU-positive cells was significantly increased (5.98, =0.004). MEG3 was mainly expressed in the nuclei of fetal mouse palatal mesenchymal cells, and the expression of MEG3 in TCDD group was significantly increased at GD13, GD14 and GD15(GD13, 39.28, =0.012; GD14, 18.75, =0.042; GD15, 28.36, =0.045). At GD14, TCDD decreased the levels of p-Smad2 and Smad4 in embryonic palate mesenchymal cells (p-Smad2, 9.48, =0.001;Smad4, 63.10, =0.001), whereas the expression of Smad7 was significantly increased at GD14 (30.77, <0.001). The results of the RIP experiment showed that the amount of TGF-βRⅠ-bound MEG3 in mouse embryonic palatal mesenchymal cells in the TCDD group (23.940±1.301) was higher than that in the control group (8.537±1.523)(24.55, <0.001). MEG3 is involved in the suppression of mouse embryonic palatal mesenchymal cell proliferation, functioning at least in part via interacting with the TGF-βRⅠ protein and thereby suppressing Smad signaling in the context of TCDD induced cleft palate.