Influence of silencing toll-like receptor 4 with RNA interference on cardiomyocytes apoptosis induced by isoproterenol

In this study, we investigated the influence of silencing TLR4 with RNA interference (RNAi) on cardiomyocyte apoptosis induced by isoproterenol (ISO). Two plasmids containing specific to TLR4 siRNA sequences were constructed and transfected into primary cultured mice cardiomyocytes. The mRNA and protein expressions of TLR4 were analyzed by real-time polymerase chain reaction (PCR) and Western blotting respectively, and the plasmids that silenced TLR4 most significantly were selected, and renamed TL-R. The plasmids carrying a non-specific siRNA coding sequence (PCN) served as the negative control. Apoptosis of cardiomyocyte was induced by ISO at a concentration of 1 umol/L. Rates of cardiomyocyte apoptosis were evaluated by flow cytometery. The protein expression levels of Bcl-2 and Bax were detected by Western blotting. The plasmids contained siRNA sequences specific of TLR4 were constructed and transfected into the primary cultured cardiomyocytes successfully. Rates of cardiomyocyte apoptosis was obviously increased in three ISO-treat group. The results demonstrated that silencing TLR4 with RNA interference can significantly decreased cardiomyocyte apoptosis, accompanied by an increased expression of the antiapoptotic protein Bcl-2 and a decreased expression of the proapoptotic protein Bax. It was concluded that the recombinant plasmids could be successfully constructed, and transfected into the primary cultured cardiomyocytes. They could ameliorate the cardiomyocyte apoptosis induced by ISO efficiently.