河豚毒素对缺血再灌注大鼠心肌细胞凋亡及相关蛋白表达的影响

Effects of Tetrodotoxin Cardioplegia on Cardiomyocyte Apoptosis and Related Protein Expression in Rat Hearts

目的研究河豚毒素(TTX)心脏停搏液对缺血再灌注大鼠心肌细胞凋亡及凋亡相关蛋白表达的影响。方法取Wistar大鼠24只,建立Langendorff-Neely离体心脏灌注模型,随机分为3组(n=8):基础组、STH-2组、TTX组。用TUNEL法检测心肌细胞凋亡指数,免疫组化测定凋亡相关蛋白Bcl-2、Bax、p53的表达,Westernblot检测Bcl-2蛋白表达。结果STH-2组凋亡指数(AI)(11.20±0.79)%高于TTX组(6.92±1.10)%和基础组(1.34±0.23)%(P<0.01)。TTX组Bcl-2的PEI为(22.63±1.10)%高于STH-2组(13.51±0.67)%和基础组(2.52±0.58)%(P<0.01);STH-2组Bax的PEI为(8.04±0.57)%高于TTX组(6.05±0.46)%和基础组(2.14±0.57)%(P<0.01)。TTX组Bcl-2/Bax为3.78±0.41高于STH-2组1.75±0.15(P<0.01)。TTX组Bcl-2的相对蛋白含量为(86.1±0.98)%高于STH-2组(53.2±1.76)%和基础组(23.7±2.61)%(P<0.01)。p53蛋白在各组均未见明显阳性表达。结论与STH-2比较,TTX能减少缺血再灌注大鼠心肌细胞凋亡数,其抗凋亡的机制可能与上调Bcl-2表达,下调Bax表达,提高Bcl-2/Bax比值有关。

OBJECTIVE To explore the effects of tetrodotoxin（TFX） cardioplegia on cardiomyocyte apoptosis and related protein expressions in rat hearts. METHODS Twenty four wistar rats were randomly divided into 3 groups（n = 8） ： Base group,STH-2 group and TTX group. Isolated rat Langendorff and Neely working heart models were estabished. The changes in cardiomyocyte apoptosis were detected by TUNEL, the expressions of Bcl-2 ,Bax protein were detected by immunohistochemieal assay in cardiomyocyte. Additionlly the expression of Bcl-2 protein in cardiomyocyte was detected by Western blot. RESULTS Apoptosis index（AI） in STH-2 group （ 11.20 ±0. 79） % was significantly higher than that in TTX group （ 6.92 ± 1.10） % and Base group （ 1.34 ± 0. 23 ） % （ P 〈 0. 01 ）. Bcl-2 protein expression index（PEI） in TTX group（22.63 ± 1.10）% increased as compared with STH-2 group （13.51 ±0.67 ）% and Base group（2.52 ±0.58 ） % （ P 〈 0.01 ） by immunohistochemical assay. Bax PEI in STH-2 group （ 8.04 ± 0.57 ） % increased as compared with TTX group（ 6.05 ± 0.46 ） % and Base group （ 2. 14± 0.57 ） % （ P 〈 0.01 ） by immunohistochemical assay. Bcl-2/Bax was 3.78±0.41 in TTX group, 1.75 ± 0. 15 in STH-2 group. Bcl-2 protein expression in TTX group （ 86. 1 ± 0.98 ） % increased as compared with STH-2 group（53.2 ± 1.76） % and Base group（23.7 ± 2.61 ） % （ P 〈 0.01 ） by Western blot. There was no sign of p53 protein expression in all groups by immunohistochemical assay in our experiment. CONCLUSION Compared with STH-2, TTX cardioplegia were all eviating the apoptosis of rat cardiomyocytes. The mechanism was probably up-regulating expression of Bcl-2 protein,down-regulating expression of bax protein and increasing Bcl-2/Bax.