透明质酸对游离脂肪酸引起的HA-VSMC表达IL-6、IL-8和TNF-α的影响

发布时间：2018-07-03 08:56

本文选题：游离脂肪酸 + 高分子量透明质酸　；参考：《宁夏医科大学》2017年硕士论文

【摘要】：目的探讨透明质酸(Hyaluronic acid,HA)对游离脂肪酸(Free fatty acid,FFA)引起的人主动脉血管平滑肌细胞(Human aortic vascular smooth muscle cells,HA-VSMC)表达白介素-6(Interleukin-6,IL-6)、白细胞介素-8(Interleukin-8,IL-8)和肿瘤坏死因子-α(Tumor necrosis factorα,TNF-α)的影响,为对血管病变的体内研究和临床治疗提供理论和实验依据。方法常规培养人HA-VSMC细胞系,采用不同浓度的FFA(0、100、200、400μmol/L)、不同浓度FFA(0、100、200、400μmol/L)+高分子量透明质酸(High molecular weight hyaluronic acid,HMW-HA,1mg/ml)和不同浓度FFA(0、100、200、400μmol/L)+低分子量透明质酸(Low molecular weight hyaluronic acid,LMW-HA,1mg/ml)处理细胞24、48h;采用MTT法检测FFA、FFA+LMW-HA和FFA+HMW-HA对HA-VSMC细胞存活率的影响;采用Ed U染色检测不同处理后细胞增殖活性;采用ELISA试剂盒检测培养上清IL-6、IL-8和TNF-α含量;采用实时荧光定量PCR法检测IL-6、IL-8和TNF-αm RNA表达水平;蛋白印记法(Western blot)检测IL-6、IL-8和TNF-α蛋白水平的表达。结果1、FFA在400μmol/L内可诱导HA-VSMC增殖且有时间和浓度依赖性;2、HMW-HA、LMW-HA均对FFA诱导的HA-VSMC增殖有抑制作用,且HMW-HA抑制作用强于LMW-HA;3、FFA单独和联合高低分子量HA处理细胞6、12、24、48h后细胞培养上清IL-6、IL-8和TNF-α随培养浓度和时间的增加而增加,HA可以抑制上述作用;4、荧光定量PCR和Western blot结果显示FFA+HMW-HA组和FFA+LMW-HA组IL-6、IL-8和TNF-αm RNA及其蛋白表达水平均低于FFA组,其中FFA+HMW-HA组又低于FFA+LMW-HA组,差异均有统计学意义(均P0.01)。结论(1)游离脂肪酸在400μmol/L内可诱导HA-VSMC增殖且有浓度依赖性;(2)HA能抑制FFA引起的HA-VSMC增殖,且HMW-HA抑制作用强于LMW-HA;(3)FFA显著促进HA-VSMC IL-6、IL-8和TNF-α的分泌,且有明显的时间和浓度依耐性;(4)FFA诱导HA-VSMC细胞IL-6、IL-8和TNF-αm RNA和蛋白表达升高;(5)HA对FFA引起的HA-VSMC细胞表达IL-6、IL-8和TNF-α有抑制作用,下调IL-6、IL-8和TNF-αm RNA和蛋白表达,且HMW-HA抑制效果强于LMW-HA。
[Abstract]:Objective to investigate the expression of Hyaluronic acid (HA) in human aortic vascular smooth muscle cells induced by Free fatty acid (FFA) and to express interleukin and tumor necrosis factor alpha. The effect of TNF- alpha, providing theoretical and experimental basis for the study and clinical treatment of vascular lesions. Methods routine culture of human HA-VSMC cell lines, using different concentrations of FFA (0100200400 mu mol/L), different concentrations of FFA (0100200400 mol/L) + high molecular weight hyaluronic acid (High molecular weight hyaluronic acid, HMW-HA, etc.) Different concentrations of FFA (0100200400 mu mol/L) + low molecular weight hyaluronic acid (Low molecular weight hyaluronic acid, LMW-HA, 1mg/ml) were used to treat cell 24,48h, and MTT method was used to detect the survival of the cells. The content of IL-6, IL-8 and TNF- alpha was raised and the RNA expression level of IL-6, IL-8 and TNF- alpha m was detected by real time fluorescence quantitative PCR method, and protein imprinting method (Western blot) was used to detect IL-6, expression and expression of alpha protein. Results 1 The proliferation of MC has a inhibitory effect, and the inhibitory effect of HMW-HA is stronger than that of LMW-HA; 3, FFA alone and combined high and low molecular weight HA cells 6,12,24,48h cell culture supernatant IL-6, IL-8 and TNF- alpha increase with the increase of culture concentration and time, HA can inhibit the effect. 4, fluorescent quantitative PCR and Western results show the group and the Western The expressions of IL-6, IL-8 and TNF- alpha m RNA and their protein expressions were lower than those in the FFA group, and the FFA+HMW-HA group was lower than the FFA+LMW-HA group. The difference was statistically significant (all P0.01). Conclusion (1) the free fatty acids can induce HA-VSMC proliferation and concentration dependence in 400 mu mol/L. (2) HA can inhibit the proliferation and the inhibitory effect is stronger than that of the FFA+LMW-HA. (3) (3) FFA significantly promoted the secretion of HA-VSMC IL-6, IL-8 and TNF- alpha, and had obvious time and concentration dependent tolerance; (4) FFA induced IL-6 in HA-VSMC cells, IL-8 and TNF- alpha m and protein expression increased; (5) The effect of the system is better than LMW-HA.
【学位授予单位】：宁夏医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R587.2;R54

【参考文献】