This study aims to construct the plasmid of human Runx1 and observe its possible effects on Runx1 gene expression in human pulmonary adenocarcinoma cells (A549). The shRNA sequence targeting human Runx1 was designed and synthesized, then inserted into pSuper plasmid by DNA recombination technology. The recombinant plasmid was confirmed by bacterial colonies PCR, enzyme digestion analysis and DNA sequencing. A549 cells were transfected by Runx1 shRNA plasmid. The inhibition efficiency of pSuper-Runx1-shRNA plasmid on Runx1 at mRNA level and protein level were measured with real-time PCR and Western blot. The results of real-time PCR and Western blot indicated that the mRNA and protein levels of Runx1 in A549 cells were inhibited by the pSuper-Runx1-shRNA expression plasmid, and the inhibition rate were 33% and 50%, respectively.
The aim of this study was to observe whether necroptosis is involved in the process of cardiac hypertrophy induced by pressure overload. SD rats underwent transverse abdominal aortic constriction (TAC) operation for establishing cardiac hypertrophy model. The structure and function of the left ventricle of rats were evaluated via echocardiography, left ventricular mass index, the expression of markers of cardiac hypertrophy and histological detection. Real-time PCR and Western blot were used to measure the gene and protein expression of receptor interacting protein kinase 1 and 3 (RIPK1 and RIPK3, the necroptosis markers) respectively. Four weeks after TAC operation, rat model for cardiac hypertrophy was established. The experimental data showed that the gene and protein expressions of RIPK1 and RIPK3 in the rat heart hypertrophic tissues after TAC for 4 weeks were increased significantly compared with those in the sham group. HE staining showed cardiomyocytes injury and hypertrophy in the hearts of TAC rat models. By transmission electron microscope, we observed that mitochondria of cardiomyocytes were damaged seriously in the TAC models. Treatment with losartan used, the selective antagonist of angiotensinⅡtypeⅠreceptor could improve the cardiac function of TAC rats. Moreover, losartan treatment decreased the expression of RIPK1 and RIPK3 in heart tissues of TAC rats. The results suggest that necroptosis occurrs in the process of cardiac hypertrophy with pressure overload, and losartan could alleviate the cardiac hypertrophy and inhibit necroptosis.