Protective effects of N-acetylcysteine on myocardial ischemia-reperfusion injury in the rat heart transplantation

Abstract

Objective To explore the protective effect of N-acetylcysteine (NAC) on graft ischemia-reperfusion injury (IRI) in the rat heart transplantation.Methods 100 healthv male Lewis rats and 20 healthy male BN rats were randomly divided into four groups.In control group,2 ml normal saline was infused via the inferior vena cave 30 min before donor harvest or implantation.In donor preconditioning group,NAC (300 mg/kg body weight,iv.) was infused via the inferior vena cave 30 min before donor harvest,and the recipient was not processed.In recipient preconditioning group,NAC (300 mg/kg body weight,iv.) was infused via the inferior vena cava 30 min before recipient implantation,and the donor was not processed; Grafts underwent cold storage in 4℃ conventional histidine-tryptophan-ketoglutarate(HTK)solution for 18 h prior to heterotopic transplantation (Lewis→Lewis).Another 20 male Lewis rats for recipients,and 20 male BN rats for donors were used for survival analysis.Graft function was monitored by daily palpation.Blood was drawn at 6 and 24 h post-reperfusion for analysis of creatine kinase isoenzvme MB (CK-MB),Troponin T (TnT),lactic acid dehydrogenase (LIDH) as markers of graft injury.Graft superoxide dismutase (SOD),and lipid hydroperoxide (LPO) activity were detected at 24 h after reperfusion.Tissue samples were taken 24 h after reperfusion for histological and ultrastructural changes.Inducible nitric oxide svnthase (iNOS) and endothelial nitric oxide syuthase (eNOS) protein expression was detected by using immunohistochemistrv.The expression of iNOS and eNOS mRNA was detected using by real-time quantitative polymerase chain reaction (Real-time PCR).Results Suaival rate of rats in the NAC pretreatment groups was significantly higher than in control group.The serum levels of CK-MB,TnT,LIDH at 6/24 h after reperfusion were reduced significantly in donor preconditioning group or recipient preconditioning group as compared with control group [for CK-MB,(10 681.11 ± 2052.01)/(12 903.29 ± 1789.46)/(31093.11 ± 6822.67) U/L at 6 h,and (1797.89 ± 383.96)/(3408.0 ± 1028.48)/(2230.40 ±450.03) U/Lat24h; forTnT,(45.76±7.46)/(17.90±2.87)/(49.51 ±6.41) μg/L at 6 h,and(11.08±1.53)/(7.06 ±0.71)/(20.28 ±2.90) μg/L at 24 h; for LDH,(4019.00 ±678.03)/(4235.17 ±585.88)/(8197.00 ±807.20) U/L at 6 h,and (1005.67 ± 164.71)/(774.00 ± 113.13)/(1619.50 ± 289.11) U/L at 24 h],respectively.LPO activity at 24 h after reperfusion was lower (9.48 ± 1.05)/(7.82 ± 1.56) vs.(10.90 ±2.28) μmol/L,and SOD activity was significautly higher in pretreatment groups (4.71 ± 1.09)/(4.57 ± 1.09) vs.(2.38 ±0.39) U/ml than in control group (P ＜0.05).Histological examinations (HE staining and electron microscopy) also proved that the injury in pretreatment group was alleviated as compared with control group.Immunohistochemical analysis and Real-time PCR revealed the decreases of iNOS (1.50 ± 0.22)/(1.63 ± 0.26) vs.(3.00 ± 0.15) and (0.95 ± 0.41) / (0.43 ± 0.17) vs.(1.00 ± 0.41),and the increase of eNOS (3.60 ± 0.16) / (3.40 ± 0.26) vs.(2.00 ± 0.21) and (3.06 ± 1.47)/(1.48 ± 0.53) vs.(1.00 ± 0.65) in pretreatment group as compared with control group.Conclusion NAC reduces IRI after heart transplantation and improves the outcome of the transplanted heart after prolonged cold preservation,which is most likely through anti-oxidation and enhancing the oxidase activity to reduce myocardial cell injury,thereby reducing mvocardial IRI. Key words: N-acetylcysteine; Heart transplation; Ischemia reperfusion injury