论文部分内容阅读
[目的]观察饮用水有机提取物对大鼠肝脏谷胱甘肽-S-转移酶(glutathione-S-transferase,GSTs)活性及谷胱甘肽-S-转移酶A1(GSTA1)基因m RNA和蛋白表达的影响,探讨其在饮用水有机提取物肝脏损伤中作用。[方法]采用固相萃取法提取水样中有机污染物,50只SD大鼠随机分成5组,分别为空白对照组、溶剂对照组(玉米油)和低、中、高3个染毒组(剂量分别为每天5、20、80 L/kg·bw),进行经口灌胃染毒12周。分光光度法检测GSTs的活性,实时荧光定量聚合酶链反应法和Western blot法分别检测GSTA1基因的m RNA和蛋白质表达水平,同时检测肝功能各指标。[结果](1)大鼠肝脏GSTs酶活性:与空白对照、溶剂对照及低剂量组相比,中剂量[(50.66±5.62)U/mg蛋白]和高剂量组[(39.80±12.95)U/mg蛋白]的GSTs酶活性升高(P<0.05);与中剂量组相比,高剂量组GSTs的酶活性则明显降低(P<0.05)。(2)GSTA1的m RNA及蛋白表达水平:中、高剂量组高于空白对照组、溶剂对照及低剂量组(P<0.05);而与中剂量组相比,高剂量组GSTA1的m RNA表达水平下降(P<0.05)。Western blot检测结果显示,随染毒剂量的增加,GSTA1蛋白表达呈先升高后降低的趋势,与空白对照、溶剂对照及低剂量组比较,中、高剂量组的升高,差异具有统计学意义(P<0.05);而与中剂量组比较,高剂量GSTA1的蛋白表达则下降(P<0.05)。(3)肝功能指标:与对照组比较,血清胆碱酯酶(CHE)在中、高剂量染毒组升高,差异均具有统计学意义(P<0.05);丙氨酸氨基转移酶(ALT)、天冬氨酸氨基转换酶(AST)则仅在高剂量组升高(P<0.05)。总蛋白(TP)和白蛋白(ALB)在高剂量组下降(P<0.05)。GSTA1蛋白表达、GSTs活性与染毒大鼠肝脏CHE水平均呈正相关关系(r=0.490 5,r=0.685 2;P<0.05)。[结论]在本实验条件下,饮用水有机提取物较高剂量染毒可上调大鼠肝脏GSTA1的m RNA及蛋白质表达水平,调控GSTs活性改变,从而导致大鼠肝细胞对毒物的易感性增强,肝损伤加重。
[Objective] To observe the effect of drinking water organic extract on hepatic glutathione-S-transferase (GSTs) activity and glutathione-S-transferase A1 (GSTA1) gene m RNA and Protein expression, to explore its role in the liver injury of drinking water organic extract. [Method] Fifty SD rats were randomly divided into five groups, which were blank control group, solvent control group (corn oil) and low, medium and high concentration of three pollutants (Doses of 5, 20, 80 L / kg · bw, respectively) for oral administration for 12 weeks. The activity of GSTs was detected by spectrophotometry. The mRNA and protein expression levels of GSTA1 gene were detected by real-time fluorescence quantitative polymerase chain reaction and Western blot respectively. The indexes of liver function were also detected. [Results] (1) The hepatic GSTs activity in rats: Compared with the blank control group, the solvent control group and the low dose group, the middle dose [(50.66 ± 5.62) U / mg protein] and the high dose group [(39.80 ± 12.95) U / mg protein] (P <0.05). Compared with the middle dose group, the GSTs activity of high dose group was significantly decreased (P <0.05). (2) The mRNA and protein expression levels of GSTA1 in middle and high dose groups were higher than those in blank control group, solvent control group and low dose group (P <0.05). Compared with middle dose group, mST The expression level decreased (P <0.05). The result of Western blot showed that the expression of GSTA1 protein increased first and then decreased with the increase of exposure dose. Compared with the blank control group, solvent control group and low dose group, the expression of GSTA1 protein increased in middle and high dose group with statistically significant difference (P <0.05). Compared with the middle dose group, the protein expression of high dose GSTA1 decreased (P <0.05). (3) Index of liver function: Serum cholinesterase (CHE) levels were higher in middle-dose and high-dose groups than those in control group, with statistical significance (P <0.05); alanine aminotransferase ALT), aspartate aminotransferase (AST) increased only in high dose group (P <0.05). Total protein (TP) and albumin (ALB) decreased in high dose group (P <0.05). There was a positive correlation between GSTA1 protein expression and GSTs activity and liver CHE level (r = 0.490 5, r = 0.685 2; P <0.05). [Conclusion] Under this experimental condition, higher doses of organic extract of drinking water could up-regulate m RNA and protein expression of GSTA1 in rat liver and regulate the activity of GSTs, leading to increased susceptibility to hepatotoxicity in rat hepatocytes , Aggravating liver injury.