Experimental studies on the role of GATA4 in the endocardial cushions development / 中华心血管病杂志

ABSTRACT

<p><b>OBJECTIVE</b>To investigate the role of GATA4 gene in the endocardial cushions development.</p><p><b>METHODS</b>Target gene eukaryote expression vectors were constructed by pcDNA3.1(-) vector plasmid, and were identified by DNA sequence analysis. Recombinant plasmids were transfected into Hela cells with lipofectamine 2000, meanwhile Hela cells transfected with empty vector or those without transfection served as transfection control group and blank control group, respectively. Real-time PCR and Western blot were performed to detect the relative expression of mRNA and protein of transcription factors GATA4, Sox9, Scleraxis and ECM proteins Aggrecan, Tenascin in each group.</p><p><b>RESULTS</b>The relative mRNA expression of GATA4 in experimental group was significantly higher than in transfection control group and blank control group. GATA4 mRNA expression in Hela(GATA4), Hela(H436Y), Hela(Null) and Hela group was 310.83 ± 2.39, 146.35 ± 1.74, 0.94 ± 0.32, 1.00 ± 0.28, respectively (F = 72.508, P < 0.05). Western blot results were consistent with the results obtained by qRT-PCR. The relative mRNA and protein expressions of Sox9, Scleraxis, Aggrecan and Tenascin in both experimental groups were significantly higher than that in transfection control group and blank control group (P < 0.05), and above gene expressions were significantly downregulated in GATA4(H436Y) group, while they were similar between transfection control group and blank control group (all P > 0.05).</p><p><b>CONCLUSIONS</b>GATA4 H436Y mutation reduces it's transcriptional activation, which might serve as a theoretical framework to demonstrate the roles of GATA4 gene in endocardial cushion development.</p>