Role of protein kinase C in reduction of hypoxia-reoxygenation injury by hypoxic preconditioning or norepinephrine preconditioning in cultured neonatal rat cardiomyocytes

Abstract

Objective To evaluate the role of protein kinase C （PKC） in reduction of hypoxia-reoxygenation （H/R） injury by hypoxic preconditioning or norepinephrine preconditioning in cultured neonatal rat cardiomyocytes. Methods Primary cultured neonatal rat cardiomyocytes were randomly divided into 6 groups （n = 25 each）： control group （group Ⅰ）, H/R group （group Ⅱ）, hypoxia preconditioning group （group Ⅲ）, norepinephrine preconditioning group （group Ⅳ）, H7 ＋ hypoxia preconditioning group （group Ⅴ） and H7 ＋ norepinephrine preconditioning group （group Ⅵ）. In group Ⅱ , the cardiomyocytes were exposed to 3 h of hypoxia followed by 1 h of reoxygenation. In group Ⅲ, the cells were subjected to 20 min of hypoxia followed by 20 min of reoxygenation before H/R. Norepinephrine was added to the culture medium with a final concentration of 10- 7 mol/L,and then the cells were cultured for 30 min before H/R in group Ⅳ. H7 was added to the culture medium with a final concentration of 5 × 10-5 mol/L, the cells were then cultured for 10 min, and the following procedures before H/R were the same as thase described in group Ⅴ . H7 was added to the culture medium with a final concentration of 5 × 10-5 mol/L, the cells were then cultured for 10 min, and the following procedures were the same as those described in group Ⅵ. The cell survival rate, the activities of LDH and CK in the supernatant, and the content of MDA and activity of SOD in cardiomyocytes were determined. Results The cell survival rate and activity of SOD were significantly lower, while the LDH and CK activities and MDA content higher in group Ⅱ than in group Ⅰ ,in group Ⅴ than in group Ⅲ, and in group Ⅵ than in group Ⅳ （P 〈 0.01）. The cell survival rate and activity of SOD were significantly increased, while the LDH and CK activities and MDA content decreased in group Ⅲ and Ⅳ compared with group Ⅱ （P〈0.01）.Conclusion The activiation of PKC is involved in the reduction of H/R injury by hypoxic preconditioning or norepinephrine preconditioning in cultured neonatal rat cardiomyocytes. Key words: Protein kinase C; Anoxia; Norepinephrine; Ischemic preconditioning; Myocytes,cardiac; Cell hypoxia; Oxygen