| Objective: To establish a rat model of acute hepatic injury by exertional heat stroke,and to detect the expression of proteins in the liver tissue related to the mitochondrial apoptosis pathway,and to investigate the role of apoptosis in acute hepatic injury in rats with exertional heat stroke.Methods:(1)establish a rat model of acute hepatic injury by exertional heat stroke: after running training,20 rats which exercised correctly on the treadmill were selected.They were randomly divided into the experimental group and the control group,with 10 rats in each group.In the experimental temperature of 35±2℃ and the humidity of 80±10%,rats in the experimental group ran at a speed of 28 meters per second.Rats in the control group was fed as usual without running.(2)The serum ALT and AST levels of the rats were detected by automatic biochemical analysis detector.(3)The apoptosis of rat livers was observed by TUNEL assay.(4)The expression of Bax,bcl-2,cyt-c,caspase-3 and caspase-9 in the liver tissues was observed by immunohistochemical staining.The pathology were analyzed by a semi-quantitative score system.(5)The expression of Bax,bcl-2,cyt-c,caspase-3 and caspase-9 in the liver tissues was determined by western blot.The relative expression levels of the target proteins were presented as the ratio of the gray value of the target protein bands to the gray value of the β-actin bands.Results:(1)All rats in the experimental group experienced exertional heat stroke.The anal temperature of all rats in the experimental group was over 42.5 ℃.Their anal temperature was 42.6(42.5,42.7)℃,running time was 48.5±18.3 minutes.(2)The ALT level of the rats in the experimental group was 58.6(56.3,74.8)U/L,which was significantly increased compared with the control group(45.6±8.0 U/L;Z=3.024,P =0.002).The difference was statistically significant.The AST level of the rats in experimental group was 231.6(215.3-376.4)U/L,which was significantly increased compared with the control group(127.5±16.8 U/L;Z=3.780,P<0.001).The difference was statistically significant.(3)The AI of the rat hepatocytes in the experimental group was 1.40(0.98,2.55)%,which was significantly increased compared with the control group(0.05(0.00,0.33)%;Z=3.810,P<0.001).The difference was statistically significant.(4)Immunohistochemical staining of the proteins related to mitochondrial apoptosis pathway: The expression of Bax in the liver tissue in the experimental group was 2.00(2.00,2.50),which was significantly increased compared with the control group(0.50(0.00,1.00);Z=3.445,P<0.01).The difference was statistically significant.Expression of Bcl-2 in the liver tissue in the experimental group was 0.00(0.00,0.25),which was significantly decreased compared with the control group(1.00(0.00,1.25);Z=2.294,P=0.043).The difference was statistically significant.The expression of Cyt-c in the liver tissue in the experimental group was 2.00(1.00,2.50),which was significantly increased compared with the control group(1.00(0.00,1.00);Z=3.292,P<0.01).The difference was statistically significant.Expression of Caspase-9 in liver tissue of rats in experimental group was 1.00(0.75,1.00),which was significantly increased compared with the control group(0.00(0.00,0.25);Z=2.615,P =0.023).The difference was statistically significant.Expression of Caspase-3 in liver tissue of rats in experimental group was 2.00(1.00,2.50),which was significantly increased compared with the control group(0.00(0.00,0.00);Z=4.082,P<0.01).The difference was statistically significant.(5)Western blot of proteins related to the mitochondrial apoptosis pathway: Relative expression of Bax was 0.38(0.33,0.67),which was significantly increased compared with the control group(0.34(0.24,0.36);Z=2.395,P =0.015).The difference was statistically significant.Relative expression of Bcl-2 was 0.35(0.28,0.37),which was significantly decreased compared with the control group(0.39(0.36,0.66);Z=2.355,P =0.019).The difference was statistically significant.Relative expression of Cyt-c was 0.36(0.32,0.65),which was significantly increased compared with the control group(0.30±0.03;Z=2.318,P =0.019),.The difference was statistically significant.Relative expression of Caspase-9 was 0.36(0.32,0.60),which was significantly increased compared with the control group(0.29±0.05;Z=2.501,P =0.011),.The difference was statistically significant.Relative expression of Caspase-3 was 0.35(0.31,0.59),which was significantly increased compared with the control group(0.25±0.05;Z=3.064,P =0.001),.The difference was statistically significant.Conclusions:(1)Apoptosis is one of the important pathogenesis of acute hepatic injury in rats with exertional heat stroke.(2)Among proteins related to mitochondrial apoptosis pathway,Bax,Cyt-c,Caspase-3 and Caspase-9 were highly expressed in the liver tissues of rats with exertional heat stroke.The expression of Bcl-2,an apoptotic inhibitory protein,was decreased.It suggests that mitochondrial apoptosis pathway plays an important role in acute hepatic injury in rats with exertional heat stroke. |
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