高血压病气虚血瘀证患者血清对CRL-1730凋亡相关蛋白及细胞活性的影响

发布时间：2018-04-18 17:29

本文选题：高血压病 + 气虚血瘀证　；参考：《广西中医药大学》2017年硕士论文

【摘要】：目的:为进一步明确高血压病气虚血瘀证患者血清诱导人脐静脉内皮细胞株(CRL-1730)凋亡的效应,并对其机制进行探讨,同时观察补阳还五汤对凋亡细胞的干预效应,为高血压病气虚血瘀证的临床防治提供实验依据。方法:分别收集高血压病气虚血瘀证患者、高血压病非气虚血瘀证、健康体检者的血清及补阳还五汤高、中、低剂量组含药血清,分别作用于体外培养的人脐静脉内皮细胞株,用ELISA检测细胞培养上清液凋亡相关蛋白含量,WB检测细胞凋亡相关蛋白表达,倒置相差显微镜观察细胞形态学,MTT法检测细胞增殖活性,台盼蓝法检测活细胞率,并观察补阳还五汤的保护作用。结果:1、ELISA检测结果:与健康组相比,高血压病气虚血瘀证组、高血压病非气虚血瘀证组细胞上清液中的β-微管蛋白、Caspase-3、Fas、FasL、Bax、Bax/Bcl-2表达量升高,差异有统计学意义(P0.05或P0.01),高血压病气虚血瘀证组、高血压病非气虚血瘀证组细胞上清液中的Bcl-2表达量降低,差异有统计学意义(P0.05或P0.01)。另外,与高血压病气虚血瘀证组相比,补阳还五汤高、中、低剂量组细胞上清液中的β-微管蛋白、Caspase-3、Fas、FasL、Bax、Bax/Bcl-2表达量降低,差异有统计学意义(P0.05或P0.01),Bcl-2表达量升高,差异有统计学意义(P0.05)。2、WB检测结果:与健康组相比,高血压病气虚血瘀证组、高血压病非气虚血瘀证组细胞中的β-微管蛋白、Caspase-3、Fas、FasL、Bax、Bcl-2、Bax/Bcl-2表达量无统计学差异(P0.05);与高血压病气虚血瘀证组相比,补阳还五汤高、中、低剂量组蛋白表达量也无统计学差异(P0.05)。3、由倒置相差显微镜观察发现,高血压病气虚血瘀证组存在细胞形态学改变。4、MTT法检测结果:与健康组相比,高血压病气虚血瘀证组、高血压病非气虚血瘀证组OD值减少,差异有统计学意义(P0.05);与高血压病气虚血瘀证组相比,补阳还五汤各剂量组OD值逐渐增加,差异有统计学差异(P0.05)。5、台盼蓝染色法检测结果:与健康组相比,高血压病气虚血瘀证组和高血压病非气虚血瘀证组均有不同程度降低;与高血压病气虚血瘀证组相比,补阳还五汤各剂量组活细胞计数百分比均升高。在光学显微镜下观察台盼蓝各组染色细胞分析,高血压病气虚血瘀证组、高血压病非气虚血瘀证组染色最多,补阳还五汤各剂量组次之,健康组染色最少。结论:高血压病气虚血瘀证患者血清可以诱导体外培养的人脐静脉血管内皮细胞株的凋亡,补阳还五汤对其凋亡有一定的保护作用,这种保护作用可能与降低β-微管蛋白和凋亡相关蛋白Bax、Fas、FasL、Caspase-3的表达量有关。
[Abstract]:Objective: to further clarify the effect of serum induced apoptosis of human umbilical vein endothelial cell line CRL-1730 in patients with qi deficiency and blood stasis syndrome of hypertension, and to explore its mechanism, and to observe the intervention effect of Buyang Huanwu decoction on apoptotic cells.To provide experimental basis for the clinical prevention and treatment of qi deficiency and blood stasis syndrome in hypertension.Methods: the serum of patients with qi deficiency and blood stasis syndrome of hypertension, non-qi deficiency and blood stasis syndrome of hypertension, the serum of healthy persons and the high, middle and low dosage groups of Buyang Huanwu decoction were collected respectively and acted on human umbilical vein endothelial cell lines cultured in vitro.The expression of apoptosis-related protein in supernatant of cell culture was detected by ELISA and WB. The cell proliferation activity was detected by inverted phase contrast microscope and the rate of living cells was detected by trypan blue method.The protective effect of Buyang Huanwu decoction was observed.Results: compared with the healthy group, the expression of 尾 -tubulin (尾 -tubulin) in the supernatant of the cell supernatant of hypertensive patients with qi deficiency and blood stasis syndrome, and the expression of 尾 -tubulin caspase-3, FasLFasLFasLBax-Bax-Bax-Bax-Bax / Bcl-2 was higher than that in the healthy group.The difference was statistically significant (P 0.05 or P 0.01). The expression of Bcl-2 in the supernatant of hypertension non-qi deficiency and blood stasis group was significantly lower than that of hypertension group (P 0.05 or P 0.01).In addition, compared with the group of qi deficiency and blood stasis syndrome of hypertension, the expression of 尾 -tubulin in the supernatant of cell supernatant of Buyang Huanwu decoction was higher than that of the control group, and the expression of 尾 -tubulin in the supernatant of Caspase-3 / Fas-Fas-Fas-Fas-Fas-Fas-Fas-FasLBax-Bax-Bax-Bax / Bcl-2 was significantly lower than that in the group of qi deficiency and blood stasis syndrome of hypertension.There was no significant difference in the expression of 尾 -tubulin caspase-3Fas-Fas-Fas-Fas-Fas-Bax-Bcl-2Bax-Bax / Bcl-2 in the cells of the hypertension group with qi deficiency and blood stasis syndrome, compared with that of the hypertension group, and the expression of 尾 -tubulin in the cells of the hypertension group was not significantly different from that of the hypertension group, and the expression of 尾 -tubulin protein in the cells of the hypertension group was not significantly different from that of the hypertension group, and the expression of 尾 -tubulin protein in the cells of the hypertension group was similar to that of the hypertension group.Buyang Huanwu decoction was high, medium and low dose histone expression had no statistical difference (P0.05N. 3). Observed by inverted phase contrast microscope, it was found that there were cell morphological changes and MTT assay in hypertension Qi deficiency and blood stasis syndrome group: compared with the healthy group,The OD value of hypertension non-qi deficiency and blood stasis syndrome group decreased, the difference was statistically significant (P 0.05), compared with hypertension qi deficiency and blood stasis syndrome group, the OD value of Buyang Huanwu decoction group increased gradually.The results of trypan blue staining: compared with the healthy group, the blood stasis syndrome group and the non-qi deficiency and blood stasis syndrome group were lower than those of the control group, and compared with the hypertension qi deficiency and blood stasis syndrome group, the blood stasis syndrome group was lower than that of the healthy control group, and compared with the healthy group, the blood stasis syndrome group and the blood stasis syndrome group were lower than those in the healthy control group.The percentage of living cells in each dose group of Buyang Huanwu decoction increased.The staining cells of trypan blue groups were observed under optical microscope. The staining was the most in the hypertension group with Qi deficiency and blood stasis syndrome, the most in the hypertension non-qi deficiency and blood stasis group, the second in the Buyang Huanwu decoction group, and the least in the healthy group.Conclusion: the serum of patients with qi deficiency and blood stasis syndrome of hypertension can induce apoptosis of human umbilical vein endothelial cell line cultured in vitro, and Buyang Huanwu decoction has certain protective effect on apoptosis.This protective effect may be related to the reduction of the expression of 尾 -tubulin and apoptosis-related protein Bax-Fas-Fas-FasL- Caspase-3.
【学位授予单位】：广西中医药大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R259

【参考文献】