肿瘤坏死因子-α在细菌脂多糖下调小鼠胎肝细胞色素P450 3A中的作用
发布时间：2019-06-14 14:00
【摘要】： 目的研究肿瘤坏死因子-α(TNF-α)在细菌脂多糖(LPS)下调小鼠胎肝细胞色素P450 3A(CYP3A)基因表达中的作用,并探讨低剂量LPS预处理对CYP3A下调的抑制效应。 方法本研究由5个实验组成:实验一、不给予妊娠小鼠任何处理,在妊娠第14、16、18 d剖杀孕鼠和出生后第4、21、70 d剖杀子鼠;实验二、经腹腔注射给予妊娠第17 d小鼠LPS(500μg/kg),LPS处理后不同时点(2、12 h)剖杀孕鼠;实验三、经腹腔注射给予妊娠第17 d小鼠不同剂量LPS(200、500μg/kg),LPS处理后12 h剖杀孕鼠;实验四、经腹腔注射给予妊娠第17 d小鼠LPS(500μg/kg),部分动物于LPS处理前30 min经腹腔注射给予TNF-α合成抑制剂己酮可可碱(PTX,100 mg/kg)进行预处理,LPS处理后12 h剖杀孕鼠;实验五、经腹腔注射给予妊娠第17 d小鼠LPS(500μg/kg),部分动物于LPS处理前24 h经腹腔注射给予低剂量LPS(10μg/kg),高剂量组于LPS处理后1.5 h和12 h剖杀孕鼠。采用RT-PCR技术分析母鼠肝脏、胎盘和胎肝组织Cyp3a11、TNF-α和PXR mRNA表达水平,Western blot技术检测胎肝组织CYP3A蛋白表达水平,ELISA法测定母鼠血清、羊水和胎肝组织TNF-α、IL-1β和IL-6蛋白含量。 结果妊娠第14 d后胎肝组织表达高水平的CYP3A,随着胎儿发育CYP3A表达水平逐步增加,出生后第4d达到成年小鼠表达水平的80%。孕期母鼠接触LPS显著下调胎肝组织Cyp3a11 mRNA和CYP3A蛋白表达水平,并呈明显的时间和剂量-效应关系;PTX预处理显著减弱LPS对胎肝CYP3A表达的下调。另一项研究结果显示,LPS处理(500μg/kg)前30 min经腹腔注射低剂量LPS(10μg/kg)显著抑制高剂量LPS对母鼠血清、羊水和胎肝组织TNF-α的诱导作用;低剂量LPS预处理也显著减弱LPS对母鼠肝脏和胎盘TNF-αmRNA的上调作用,而对胎肝TNF-αmRNA表达水平无显著影响;与其对高剂量LPS诱导TNF-α表达的抑制效应相一致,低剂量LPS预处理明显减弱LPS对小鼠胎肝组织CYP3A表达的下调作用。 结论母鼠孕期接触LPS显著下调胎肝CYP3A表达;源自母鼠血清和羊水的TNF-α至少部分参与了LPS对胎肝CYP3A的下调作用;低剂量LPS预处理通过抑制母鼠TNF-α的产生减弱LPS对胎肝CYP3A的下调作用。
[Abstract]:Objective To study the role of tumor necrosis factor-1 (TNF-1) in the down-regulation of the expression of cytochrome P450 3A (CYP3A) gene in mouse fetal hepatocyte cytochrome P450 3A (CYP3A), and to investigate the effect of low-dose LPS pretreatment on the down-regulation of CYP3A. Methods The study consisted of 5 experimental groups:1. No treatment was given to pregnant mice, and the rats were killed at the 14th, 16th and 18th day of gestation and the 4th, 21st and 70th day after birth. Kg), after LPS treatment, the pregnant rats were not killed at the same time (2,12 h); in the experiment, the rats were given different doses of LPS (200,500. mu.g/ kg) in the 17th day of gestation by intraperitoneal injection, and the pregnant rats were killed at 12 h after LPS treatment; and 4, LPS (500. mu.g/ kg) was given to the 17th day of pregnancy by intraperitoneal injection. Kg), in the first 30 minutes of LPS treatment, the mice were treated with the TNF-1 synthesis inhibitor hexanone (PTX,100 mg/ kg) for pretreatment, and the pregnant rats were killed at 12 h after LPS treatment; and 5, LPS (500. mu.g/ kg) in the 17th day of pregnancy was given by intraperitoneal injection. Kg), partial animals were given low dose of LPS (10. mu.g/ kg) by intraperitoneal injection 24 hours before LPS treatment and 1.5 h and 12 h after LPS treatment in high dose group. Methods: The expression of cyp3a11, TNF-1 and PXR mRNA in the liver, placenta and fetal liver tissues of female rats were analyzed by RT-PCR. The expression level of CYP3A protein in fetal liver was detected by Western blot, and the levels of TNF-1, IL-1 and IL-6 in the serum, amniotic fluid and fetal liver of the fetal liver were determined by Western blot. The results showed that the expression of CYP3A in the fetal liver after the 14th day of pregnancy increased gradually with the increase of the expression of CYP3A in the fetus, and the fourth day after birth reached the adult mouse. The levels of cyp3a11 mRNA and CYP3A protein in the fetal liver tissues were significantly reduced by exposure to LPS during pregnancy, and the time and dose-effect relationship was observed. The pretreatment of PTX significantly reduced LPS-to-fetal liver CYP. A further study showed that LPS treatment (500. mu.g/ kg) was injected intraperitoneally at a low dose of LPS (10. mu.g/ kg) to significantly inhibit the high dose of LPS in the serum, amniotic fluid and fetal liver tissue TNF of the maternal serum, amniotic fluid and fetal liver The effect of LPS on the expression of TNF-1 mRNA in fetal liver and placenta was not significantly affected by low-dose LPS pretreatment, and the expression of TNF-VEGF in fetal liver was not significantly affected by LPS pretreatment. The inhibitory effect was consistent, and the low-dose LPS pretreatment significantly reduced the LPS-induced CYP3A in the mouse fetal liver tissue. The effect of LPS on the regulation of CYP3A in the fetal liver was significantly reduced during the pregnancy. The low dose of LPS pretreatment was used to reduce the production of TNF-1 in the female mice and to decrease the impact of LPS on the fetal liver.
【学位授予单位】：安徽医科大学
【学位级别】：硕士
【学位授予年份】：2009
【分类号】：R363
本文编号：2499441
[Abstract]:Objective To study the role of tumor necrosis factor-1 (TNF-1) in the down-regulation of the expression of cytochrome P450 3A (CYP3A) gene in mouse fetal hepatocyte cytochrome P450 3A (CYP3A), and to investigate the effect of low-dose LPS pretreatment on the down-regulation of CYP3A. Methods The study consisted of 5 experimental groups:1. No treatment was given to pregnant mice, and the rats were killed at the 14th, 16th and 18th day of gestation and the 4th, 21st and 70th day after birth. Kg), after LPS treatment, the pregnant rats were not killed at the same time (2,12 h); in the experiment, the rats were given different doses of LPS (200,500. mu.g/ kg) in the 17th day of gestation by intraperitoneal injection, and the pregnant rats were killed at 12 h after LPS treatment; and 4, LPS (500. mu.g/ kg) was given to the 17th day of pregnancy by intraperitoneal injection. Kg), in the first 30 minutes of LPS treatment, the mice were treated with the TNF-1 synthesis inhibitor hexanone (PTX,100 mg/ kg) for pretreatment, and the pregnant rats were killed at 12 h after LPS treatment; and 5, LPS (500. mu.g/ kg) in the 17th day of pregnancy was given by intraperitoneal injection. Kg), partial animals were given low dose of LPS (10. mu.g/ kg) by intraperitoneal injection 24 hours before LPS treatment and 1.5 h and 12 h after LPS treatment in high dose group. Methods: The expression of cyp3a11, TNF-1 and PXR mRNA in the liver, placenta and fetal liver tissues of female rats were analyzed by RT-PCR. The expression level of CYP3A protein in fetal liver was detected by Western blot, and the levels of TNF-1, IL-1 and IL-6 in the serum, amniotic fluid and fetal liver of the fetal liver were determined by Western blot. The results showed that the expression of CYP3A in the fetal liver after the 14th day of pregnancy increased gradually with the increase of the expression of CYP3A in the fetus, and the fourth day after birth reached the adult mouse. The levels of cyp3a11 mRNA and CYP3A protein in the fetal liver tissues were significantly reduced by exposure to LPS during pregnancy, and the time and dose-effect relationship was observed. The pretreatment of PTX significantly reduced LPS-to-fetal liver CYP. A further study showed that LPS treatment (500. mu.g/ kg) was injected intraperitoneally at a low dose of LPS (10. mu.g/ kg) to significantly inhibit the high dose of LPS in the serum, amniotic fluid and fetal liver tissue TNF of the maternal serum, amniotic fluid and fetal liver The effect of LPS on the expression of TNF-1 mRNA in fetal liver and placenta was not significantly affected by low-dose LPS pretreatment, and the expression of TNF-VEGF in fetal liver was not significantly affected by LPS pretreatment. The inhibitory effect was consistent, and the low-dose LPS pretreatment significantly reduced the LPS-induced CYP3A in the mouse fetal liver tissue. The effect of LPS on the regulation of CYP3A in the fetal liver was significantly reduced during the pregnancy. The low dose of LPS pretreatment was used to reduce the production of TNF-1 in the female mice and to decrease the impact of LPS on the fetal liver.
【学位授予单位】：安徽医科大学
【学位级别】：硕士
【学位授予年份】：2009
【分类号】：R363
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