铝通过HPA轴激活NF-κB调控大鼠脾脏淋巴细胞凋亡机制的研究

发布时间：2018-05-21 01:17

本文选题：铝 + 大鼠　；参考：《东北农业大学》2017年硕士论文

【摘要】：本试验以80只5周龄体重约为115 g的Wistar大鼠为实验动物,随机均分为4组,分别为对照组(CG,0 mg/kg B.W.Al Cl3),低剂量组(LG,64 mg/kg B.W.Al Cl3),中剂量组(MG,128 mg/kg B.W.Al Cl3)和高剂量组(HG,256 mg/kg B.W.Al Cl3)。通过向饮水中添加六水三氯化铝(Al Cl3·6H2O)方式,复制铝中毒模型,实验期120 d。采用ELISA法检测大鼠血清促肾上腺皮质激素释放激素(CRH)、促肾上腺皮质激素(ACTH)和糖皮质激素(GCs)含量和大鼠脾脏淋巴细胞内caspase-9、caspase-3的活性;流氏细胞术检测脾脏淋巴细胞凋亡率;q RT-PCR法检测大鼠脾脏淋巴细胞细胞中糖皮质激素受体(GCR)、NF-κB、Bcl-2、Bax、caspase-9和caspase-3 m RNA表达量;western blot法检测GCR和NF-κB大鼠脾脏淋巴细胞核蛋白的表达量。目的在于探讨NF-κB在铝通过下丘脑-垂体-肾上腺(HPA)轴诱导大鼠脾脏淋巴细胞凋亡中的作用及机制,结果发现:(1)染铝组大鼠血清CRH、ACTH和GCs的含量均显著高于对照组(p0.05,p0.01);脾脏淋巴细胞内GCR m RNA和蛋白表达量均显著高于对照组(p0.05,p0.01),说明铝激活下丘脑-垂体-肾上腺(HPA)轴。(2)染铝组大鼠脾脏淋巴细胞早期和晚期凋亡率均显著高于对照组(p0.01);Bcl-2/Bax mRNA表达比率均显著低于对照组(p0.05,p0.01);caspase-9和caspase-3 m RNA表达量和活性均显著高于对照组(p0.05,p0.01),说明铝可诱导大鼠脾脏淋巴细胞凋亡。(3)染铝组大鼠脾脏淋巴细胞中NF-κB mRNA和核蛋白表达量均显著低于对照组(p0.01);淋巴细胞内GCR的核蛋白表达量与NF-κB的核蛋白表达量呈负相关;NF-κB mRNA表达量与Bcl-2 mRNA表达量呈正相关,而与Bax mRNA表达量呈负相关,说明NF-κB是铝通过HPA轴诱导大鼠脾脏淋巴细胞凋亡的关键因子。上述结果说明,铝通过HPA轴激活NF-κB调控的大鼠脾脏淋巴细胞凋亡。研究结果可为缓解铝免疫毒性的研究奠定基础。
[Abstract]:Eighty 5-week-old Wistar rats weighing about 115g were randomly divided into 4 groups: control group (CG 0 mg/kg B.W.Al Cl 3), low dose group (GG + 64 mg/kg B.W.Al Cl 3), middle dose group (MGN 128 mg/kg B.W.Al Cl 3) and high dose group (HGG 256 mg/kg B.W.Al Cl 3). The experimental animals were divided into three groups: the control group, the low dose group and the middle dose group, respectively. The model of aluminum poisoning was established by adding aluminum chloride (Al Cl3 6H 2O) into drinking water for 120 days. The contents of serum corticotropin releasing hormone (CRH), adrenocorticotropic hormone (ACTH) and glucocorticoid (GCs) and the activity of caspase-9 and caspase-3 in rat spleen lymphocytes were measured by ELISA method. Apoptosis rate of splenic lymphocytes was detected by flow cytometry and Q RT-PCR method was used to detect the expression of caspase-9 and caspase-3 m RNA in rat splenic lymphocytes. Western blot method was used to detect the expression of nuclear protein in spleen lymphocytes of GCR and NF- 魏 B rats. Objective to investigate the role and mechanism of NF- 魏 B in apoptosis of spleen lymphocytes in rats induced by aluminum via hypothalamus-pituitary-adrenal (HPA) axis. The results showed that the levels of serum CRHH-ACTH and GCs in the control group were significantly higher than those in the control group, and the expression of GCR m RNA and protein in spleen lymphocytes were significantly higher than those in the control group, indicating that aluminum activated hypothalamus-pituitary-adrenal (HPA) axis. The early and late apoptosis rate of spleen lymphocytes in aluminum exposed group was significantly higher than that in control group, which was significantly lower than that in control group. The expression and activity of caspase-9 and caspase-3 m RNA in control group were significantly higher than those in control group, which indicated that aluminum could induce spleen of rats. The expression of NF- 魏 B mRNA and nuclear protein in spleen lymphocytes of rats exposed to aluminum was significantly lower than that of control group (p0.01), and the expression of GCR in lymphocytes was negatively correlated with the expression of nuclear protein of NF- 魏 B. The expression of Bcl-2 mRNA was positively correlated. The expression of NF- 魏 B was negatively correlated with the expression of Bax mRNA, indicating that NF- 魏 B was the key factor of apoptosis of spleen lymphocytes induced by aluminum through the HPA axis in rats. These results suggest that aluminum activates NF- 魏 B induced apoptosis of spleen lymphocytes through HPA axis. The results may lay a foundation for the study of alleviating the immune toxicity of aluminum.
【学位授予单位】：东北农业大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R114

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