芝麻素对兔主动脉内皮细胞JAK1表达的影响

发布时间：2018-04-08 21:32

  本文选题：芝麻素　切入点：JAK1　出处：《延边大学》2017年硕士论文

【摘要】：目的:本文通过研究及比较芝麻素与AG490在动脉粥样硬化形成过程中对兔主动脉内皮细胞JAK1表达的影响,探讨芝麻素在抗动脉粥样硬化机制中的作用。方法:(1)兔主动脉内皮细胞的分离、培养及传代。用传代成功后的对数生长期的主动脉内皮细胞进行实验。(2)细胞分组及处理:①正常对照组:在无菌条件下,将实验细胞于无血清的DMEM培养基中培养24-48小时;②ox-LDL组:将实验细胞在100mg/L浓度的ox-LDL培养基中培养24小时,然后在无血清的DMEM培养基中培养24小时;③AG490(JAK1抑制剂)组:将实验细胞培养于50 μ mol/L的AG490培养基中培养24小时,然后在ox-LDL培养基中培养24小时,最后在无血清的DMEM培养基中培养24小时;④芝麻素组:将实验细胞在芝麻素培养基(60μmol/L)中培养24小时,然后在100mg/L的ox-LDL培养基中培养24小时,最后在无血清的DEME培养基中培养24小时。(3)采用Real-time PCR方法检测兔主动脉内皮细胞增殖过程中芝麻素、AG490对JAK1表达的影响。结果:正常对照组:兔主动脉内皮细胞为梭形或多角形,胞浆丰富,边界清楚,可见典型的呈"铺路石状"镶嵌排列的细胞,单层分布。ox-LDL组:兔主动脉内皮细胞圆形细胞明显增多,边界不清,数目明显减少,凋亡小体出现,细胞碎片增多,细胞间隙变宽,偶有脱落。芝麻素组:兔主动脉内皮细胞呈多角形或梭形,边界清楚,排列接近正常对照组,但细胞数少于正常对照组细胞且多于ox-LDL组。PCR产物经1%琼脂糖凝胶电泳分析,在250bp处出现一条特异性条带。成功扩增出兔主动脉内皮细胞JAK1基因。以稀释的重组质粒作为PCR反应的模板,分别以各参数的上、下游引物为引物,经1%琼脂糖凝胶电泳分析表明,在2945bp处出现一条特异性条带。成功的扩增出兔主动脉内皮细胞JAK1基因的克隆质粒。ox-LDL组,芝麻素组,AG490组JAK1-mRNA表达明显高于正常对照组,有显著性差异P0.05;芝麻素组与AG490组JAK1-mRNA的表达明显低于ox-LDL组P0.05;芝麻素与AG490比较有显著性差异0.05。结论:芝麻素组与AG490组均可抑制JAK1-mRNA表达。芝麻素组抑制作用大于JAK1抑制剂AG490组,可作为防治心脑血管疾病新的靶点。
[Abstract]:Aim: to study and compare the effects of sesamin and AG490 on the expression of JAK1 in rabbit aortic endothelial cells during atherosclerosis, and to explore the role of sesamin in the mechanism of anti-atherosclerosis.Methods the rabbit aortic endothelial cells were isolated, cultured and subcultured.The aortic endothelial cells in logarithmic growth phase were divided into two groups and treated with 1: 1 normal control group: the experimental cells were cultured in serum-free DMEM medium for 24-48 hours under aseptic conditions.2ox-LDL group: the experimental cells were cultured in the ox-LDL medium of 100mg/L concentration for 24 hours and then in the serum-free DMEM medium for 24 hours. The experimental cells were cultured in 50 渭 mol/L AG490 medium for 24 hours.Then the cells were cultured in ox-LDL medium for 24 hours and in serum-free DMEM medium for 24 hours. The experimental cells were cultured in 60 渭 mol / L sesamin medium for 24 hours, and then in the ox-LDL medium of 100mg/L for 24 hours.Finally, cultured in serum-free DEME medium for 24 hours, Real-time PCR method was used to detect the effect of sesamin AG-490 on the expression of JAK1 during the proliferation of rabbit aortic endothelial cells.Results: in normal control group, the endothelial cells of rabbit aorta were fusiform or polygonal, with abundant cytoplasm and clear boundary.In the monolayer distribution of ox-LDL, the number of round cells in rabbit aortic endothelial cells was obviously increased, the boundary was not clear, the number of apoptotic bodies appeared, the number of apoptotic bodies appeared, the cell fragments increased, the cell gap became wider, and occasionally the cells fell off.In the sesamin group, the endothelial cells of rabbit aorta were polygonal or fusiform with clear boundary and close to the normal control group, but the number of cells was less than that of the normal control group and more than that of the ox-LDL group by 1% agarose gel electrophoresis.A specific band appeared at 250bp.The JAK1 gene of rabbit aortic endothelial cells was successfully amplified.The diluted recombinant plasmid was used as the template of PCR reaction. The primers of upstream and downstream of each parameter were used as primers respectively. The results of 1% agarose gel electrophoresis showed that there was a specific band in 2945bp.The cloned plasmid of JAK1 gene of rabbit aortic endothelial cells, ox-LDL group and sesamin group, were successfully amplified. The expression of JAK1-mRNA in AG490 group was significantly higher than that in normal control group.The expression of JAK1-mRNA in sesamin group and AG490 group was significantly lower than that in ox-LDL group (P 0.05), and there was a significant difference between sesamin group and AG490 group (P 0.05).Conclusion: both sesamin group and AG490 group can inhibit JAK1-mRNA expression.The inhibitory effect of sesamin group was greater than that of JAK1 inhibitor AG490 group, and could be used as a new target for prevention and treatment of cardiovascular and cerebrovascular diseases.
【学位授予单位】：延边大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R543.5
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本文编号：1723420