铝和ApoE ε4联合作用对SH-SY5Y细胞tau蛋白磷酸化和Aβ表达的影响
发布时间：2018-11-03 15:49
【摘要】：目的 采用SH-SY5Y细胞研究环境因素铝和遗传因素ApoE S4基因分别及联合作用的情况下,对tau蛋白不同位点磷酸化水平和Aβ表达的影响,探讨铝和ApoEε4是否存在交互作用。 方法 培养并选取SH-SY5Y细胞,分别进行AICl3染毒和ApoE ε4转染处理,实验分组包括空白对照组、200μM AlCl3的低剂量组、400μM AlCl3的中剂量组、800μM AlCl3的高剂量组、空质粒组、ApoE ε4组、400μM AlCl3+ApoEε4组。AICl3染毒或ApoE ε4转染48h后,采用CCK-8试剂盒测定细胞活力；采用ELISA试剂盒测定总tau蛋白、tau-181、tau-231、au-262、tau-396和Aβ-40含量；采用析因设计的方法分析AICl3和ApoE s4是否存在交互作用。 结果 1.随着AlCl3染毒剂量的增高,SH-SY5Y细胞数量逐渐减少,突触变短消失；CCK-8检测结果显示细胞活力逐渐下降,200μM AlCl3的低剂量组、400μM AlCl3的中剂量组、800μM AICl3的高剂量组活力显著低于空白对照组(P0.05); ELISA检测结果显示,200μM AlCl3的低剂量组、400μM AlCl3的中剂量组、800μM AlCl3的高剂量组相较于空白对照组,总tau蛋白、tau-181、tau-231、tau-262、tau-396和Aβ-40含量均有显著升高(P0.05)。 2.经ApoE ε4转染后,SH-SY5Y细胞形态趋于圆形,易崩解死亡；CCK-8检测结果显示细胞活力明显下降,并显著低于空白对照组(P0.05); ELISA检测结果显示,ApoE ε4组细胞的tau蛋白和Aβ-40含量均显著高于空白对照组(P0.05)。 3.经AICl3和ApoE ε4独立和联合染毒后可观察到,两种因素联合作用下细胞丧失固有形态,其数量较空白对照和两因素独立作用时更为稀少；CCK-8检测结果显示细胞活力低下,与空白对照组有显著差异(P0.05); ELISA检测结果显示,ApoE s4组细胞的总tau蛋白、tau-181、tau-231、tau-262、tau-396和Aβ-40含量均显著高于空白对照组(P0.05)；根据以上结果经析因设计分析得出AlC13和ApoE ε4存在明显的交互作用(P0.05),并表现为协同效应。 结论 1.铝和ApoE ε4基因均可导致SH-SY5Y细胞形态改变,降低细胞活力,具有明显的细胞毒性作用； 2.铝和ApoE ε4基因均促使总tau、tau-181、tau-231、tau-262和tau-396磷酸化水平升高,Ap沉积增多； 3.铝和ApoE ε4基因同时存在的情况下两种因素具有交互作用,且表现为协同效应。
[Abstract]:Objective to study the effects of environmental factor aluminum and genetic factor ApoE S4 gene on the phosphorylation level of tau protein and the expression of A 尾 in SH-SY5Y cells, and to explore the interaction between aluminum and ApoE 蔚 4. Methods SH-SY5Y cells were cultured and treated with AICl3 and ApoE 蔚 4 transfection respectively. The experimental groups included blank control group, low dose group of 200 渭 M AlCl3, middle dose group of 400 渭 M AlCl3, high dose group of 800 渭 M AlCl3, empty plasmid group and ApoE 蔚 4 group. The cells were treated with AICl3 or ApoE 蔚 4 for 48 h, and the cell viability was measured by CCK-8 kit. The contents of total tau protein, tau-181,tau-231,au-262,tau-396 and A 尾 -40 were determined by ELISA kit, and the interaction between AICl3 and ApoE s4 was analyzed by factorial design. Result 1. With the increase of the dose of AlCl3, the number of SH-SY5Y cells gradually decreased, and the synapse became shorter and disappeared. The results of CCK-8 showed that the cell viability was decreased gradually. The activity of the low dose group of 200 渭 M AlCl3, the middle dose group of 400 渭 M AlCl3 group and the high dose group of 800 渭 M AICl3 group were significantly lower than that of the blank control group (P0.05). The results of ELISA showed that the low dose group of 200 渭 M AlCl3, the middle dose group of 400 渭 M AlCl3, and the high dose group of 800 渭 M AlCl3 compared with the control group, total tau protein, tau-181,tau-231,tau-262, The contents of tau-396 and A 尾 -40 were significantly increased (P0.05). 2. After transfection with ApoE 蔚 4, SH-SY5Y cells tended to be round in shape and prone to disintegration and death. The results of CCK-8 showed that the cell viability was significantly decreased and significantly lower than that of the blank control group (P0.05); ELISA results showed that the ApoE 蔚 4 cells tau protein and A 尾 -40 content were significantly higher than the blank control group (P0.05). 3. After AICl3 and ApoE 蔚 4 were treated independently and jointly, it was observed that the cells lost their inherent morphology under the combined treatment of two factors, and the number of the cells was less than that of the blank control and the two independent factors. The results of CCK-8 showed that the cell activity was low and there was significant difference between the control group and the control group (P0.05). ELISA results showed that the total tau protein, tau-181,tau-231,tau-262,tau-396 and A 尾 -40 contents in ApoE s4 cells were significantly higher than those in the blank control group (P0.05). According to the analysis of factorial design, the interaction between AlC13 and ApoE 蔚 4 was obvious (P0.05), and showed synergistic effect. Conclusion 1. Both aluminum and ApoE 蔚 4 gene can cause morphological changes and decrease cell viability of SH-SY5Y cells. 2. Both Al and ApoE 蔚 4 genes increased the phosphorylation level of total tau,tau-181,tau-231,tau-262 and tau-396, and increased Ap deposition. In the presence of both aluminum and ApoE 蔚 4 genes, the two factors have interaction and show synergistic effect.
【学位授予单位】：山西医科大学
【学位级别】：硕士
【学位授予年份】：2013
【分类号】：R749.16
本文编号：2308233
[Abstract]:Objective to study the effects of environmental factor aluminum and genetic factor ApoE S4 gene on the phosphorylation level of tau protein and the expression of A 尾 in SH-SY5Y cells, and to explore the interaction between aluminum and ApoE 蔚 4. Methods SH-SY5Y cells were cultured and treated with AICl3 and ApoE 蔚 4 transfection respectively. The experimental groups included blank control group, low dose group of 200 渭 M AlCl3, middle dose group of 400 渭 M AlCl3, high dose group of 800 渭 M AlCl3, empty plasmid group and ApoE 蔚 4 group. The cells were treated with AICl3 or ApoE 蔚 4 for 48 h, and the cell viability was measured by CCK-8 kit. The contents of total tau protein, tau-181,tau-231,au-262,tau-396 and A 尾 -40 were determined by ELISA kit, and the interaction between AICl3 and ApoE s4 was analyzed by factorial design. Result 1. With the increase of the dose of AlCl3, the number of SH-SY5Y cells gradually decreased, and the synapse became shorter and disappeared. The results of CCK-8 showed that the cell viability was decreased gradually. The activity of the low dose group of 200 渭 M AlCl3, the middle dose group of 400 渭 M AlCl3 group and the high dose group of 800 渭 M AICl3 group were significantly lower than that of the blank control group (P0.05). The results of ELISA showed that the low dose group of 200 渭 M AlCl3, the middle dose group of 400 渭 M AlCl3, and the high dose group of 800 渭 M AlCl3 compared with the control group, total tau protein, tau-181,tau-231,tau-262, The contents of tau-396 and A 尾 -40 were significantly increased (P0.05). 2. After transfection with ApoE 蔚 4, SH-SY5Y cells tended to be round in shape and prone to disintegration and death. The results of CCK-8 showed that the cell viability was significantly decreased and significantly lower than that of the blank control group (P0.05); ELISA results showed that the ApoE 蔚 4 cells tau protein and A 尾 -40 content were significantly higher than the blank control group (P0.05). 3. After AICl3 and ApoE 蔚 4 were treated independently and jointly, it was observed that the cells lost their inherent morphology under the combined treatment of two factors, and the number of the cells was less than that of the blank control and the two independent factors. The results of CCK-8 showed that the cell activity was low and there was significant difference between the control group and the control group (P0.05). ELISA results showed that the total tau protein, tau-181,tau-231,tau-262,tau-396 and A 尾 -40 contents in ApoE s4 cells were significantly higher than those in the blank control group (P0.05). According to the analysis of factorial design, the interaction between AlC13 and ApoE 蔚 4 was obvious (P0.05), and showed synergistic effect. Conclusion 1. Both aluminum and ApoE 蔚 4 gene can cause morphological changes and decrease cell viability of SH-SY5Y cells. 2. Both Al and ApoE 蔚 4 genes increased the phosphorylation level of total tau,tau-181,tau-231,tau-262 and tau-396, and increased Ap deposition. In the presence of both aluminum and ApoE 蔚 4 genes, the two factors have interaction and show synergistic effect.
【学位授予单位】：山西医科大学
【学位级别】：硕士
【学位授予年份】：2013
【分类号】：R749.16
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