T1AM Attenuates the Hypoxia/Reoxygenation-Induced Necroptosis of H9C2 Cardiomyocytes via RIPK1/RIPK3 Pathway

Abstract

Purpose. To investigate the detailed mechanism of 3-iodothyronamine (T1AM) in cell apoptosis and programmed necrosis of hypoxia/reoxygenation- (H/R-) induced H9C2 injury. Materials and Methods. Cardiomyocyte H9C2 cells were cultured in vitro for the establishment of cardiomyocyte H/R models. Cells were randomly divided into four groups: the control group, H/R group, T1AM pretreatment group, T1AM pretreatment and H/R (6 μm T1AM+H/R) group. The degree of myocardial injury was determined by the detection of the cardiomyocyte inhibition rate by CCK8 and the detection of lactic dehydrogenase (LDH) activity. Cell apoptosis was assessed through TUNEL assay and flow cytometry analysis. The protein level and mRNA level of RIPK1, RIPK3, and CAMKII were detected by western blotting and qRT-PCR. Results. Compared with the control group, the cell inhibition rate was dramatically elevated in the H/R group. LDH release of cardiomyocytes was significantly increased. Protein and mRNA expressions of RIPK1, RIPK3, and CAMKII were significantly enhanced. Compared with the H/R group, the cell inhibition rate, LDH release, cardiomyocyte necroptosis rate, and protein and mRNA levels of RIPK1, RIPK3, and CAMKII of the T1AM+H/R group were significantly decreased. Conclusion. Pretreatment with T1AM could alleviate cardiomyocytes’ H/R injury and inhibit necroptosis of cardiomyocytes, which might exert a protective function upon activation of the RIPK1/RIPK3 pathway.