肌肉肌醇、D-手性肌醇及二者合用对HepG2胰岛素抵抗细胞葡萄糖消耗量的改善作用

发布时间：2018-01-14 18:07

本文关键词：肌肉肌醇、D-手性肌醇及二者合用对HepG2胰岛素抵抗细胞葡萄糖消耗量的改善作用　出处：《青岛大学》2016年硕士论文　论文类型：学位论文

【摘要】：目的通过建立Hep G2胰岛素抵抗模型(Hep G2-IR),比较肌肉肌醇(myo-inositol,MI)、D-手性肌醇(D-chirol-inositol,DCI)及二者合用对Hep G2-IR细胞葡萄糖消耗量的影响,并探讨其作用的可能机制。方法1.CCK-8法检测不同浓度(0、25、35、45、55、65mmol/L)的葡萄糖对Hep G2细胞增殖活性的影响,不同浓度(0、0.25、0.5、1、2、4、8、16、32mmol/L)的MI、DCI及二者合用分别对Hep G2细胞增殖活性的影响,以确定其在实验中的作用剂量范围。2.浓度为55mmol/L的葡萄糖持续作用Hep G2细胞24h后,1nmol/L胰岛素刺激诱导Hep G2细胞15~25min,建立胰岛素抵抗细胞模型;通过葡萄糖氧化酶法(GOD-POD法)检测对照组与模型组葡萄糖消耗量,以鉴定模型是否成立。3.不同浓度(0.125、0.25、0.5mmol/L)的MI、DCI及二者合用分别作用Hep G2-IR细胞,GOD-POD法检测它们对葡萄糖消耗量的影响。Western-blot法检测MI、DCI及二者合用对Hep G2-IR细胞PI3K、p Akt、Akt蛋白表达的影响。结果1.与空白对照组相比,葡萄糖在35~55mmol/L浓度时,对Hep G2细胞增殖活性无影响,而葡萄糖在浓度为65mmol/L时,对Hep G2细胞增殖活性有显著的抑制作用(P0.05);0~0.5mmol/L浓度的MI、DCI对Hep G2细胞增殖活性无影响,而1~32mmol/L浓度的MI、DCI对Hep G2细胞增殖活性有显著的抑制作用(P0.01),且随MI、DCI浓度的增加,细胞增殖活性有逐渐降低的趋势;0~1mmol/L浓度的MI与DCI合用对Hep G2细胞增殖活性无影响,而2~32mmol/L浓度的MI与DCI合用对Hep G2细胞增殖活性有显著的抑制作用(P0.01),且随MI与DCI合用浓度的增加,细胞增殖活性有逐渐降低的趋势。2.GOD-POD法检测结果表明,与正常对照组相比,浓度为55mmol/L的葡萄糖持续作用Hep G2细胞24h,且1nmol/L胰岛素刺激15-25min后,被诱导的Hep G2细胞葡萄糖消耗量显著降低(P0.01),表明成功建立了Hep G2-IR细胞模型。3.GOD-POD法检测显示,与模型对照组相比,MI、DCI及二者合用组都可增加Hep G2-IR细胞葡萄糖消耗量(P0.05),且每个组随着干预浓度增加,葡萄糖消耗量有逐渐上升的趋势,在相同浓度下,合用组葡萄糖消耗量增加最显著(P0.05)。4.Western-blot分析显示,与模型对照组相比,MI、DCI及二者合用组可增加Hep G2-IR细胞PI3K蛋白表达及p Akt/Akt蛋白表达水平的比值,且每个组随着干预浓度增加,蛋白表达及蛋白表达水平的比值有逐渐上升的趋势,在相同浓度下,合用组提高的PI3K蛋白表达及p Akt/Akt蛋白表达水平的比值最显著(P0.05)。结论MI、DCI及二者合用可能通过提高Hep G2-IR细胞的PI3K蛋白表达及促进Akt的磷酸化,进而增加Hep G2-IR细胞葡萄糖消耗量,从而达到改善IR目的,其中合用作用效果最显著。
[Abstract]:Objective to compare the muscle myo-inositol (MIM) of Hep G2 insulin resistance model (Hep G2-IRI). Effects of D-chirol-inositol (DCI) and its combination on glucose consumption in Hep G2-IR cells. The possible mechanism of its action was also discussed. 1. CCK-8 method was used to detect the different concentrations of 25, 25, 35, 45 and 55. The effect of 65 mmol / L glucose on the proliferation activity of Hep G2 cells was observed. The effects of DCI and their combination on the proliferation of Hep G2 cells were studied. In order to determine the range of dose of glucose in the experiment. The concentration of glucose was 55 mmol / L after 24 hours of continuous treatment of Hep G2 cells. 1 nmol / L insulin stimulated Hep G2 cells were induced for 15 to 25 min to establish insulin resistance cell model. The glucose consumption of the control group and the model group was determined by glucose oxidase method (GOD-POD) to determine whether the model was established or not. Hep G2-IR cells were treated with 0.5 mmol / L MIDCI and their combination. GOD-POD assay was used to detect their effect on glucose consumption. Western-blot was used to detect PI3K in Hep G2-IR cells. Results 1.Compared with the blank control group, glucose at the concentration of 355mmol / L had no effect on the proliferation activity of Hep G2 cells. When the concentration of glucose was 65 mmol / L, the proliferative activity of Hep G2 cells was inhibited significantly (P 0.05). 0.5 mmol / L MI-DCI had no effect on the proliferation activity of Hep G2 cells, while the MI of 32 mmol / L concentration was not affected. The proliferation activity of Hep G2 cells was significantly inhibited by DCI, and the proliferation activity decreased gradually with the increase of MI-DCI concentration. The combination of 0 mmol / L MI and DCI had no effect on the proliferation activity of Hep G2 cells. The proliferation activity of Hep G2 cells was significantly inhibited by the combination of MI and DCI at the concentration of 32 mmol / L, and increased with the increase of the concentration of MI and DCI. The proliferation activity of the cells decreased gradually. 2. The results of GOD-POD assay showed that compared with the normal control group. Glucose at a concentration of 55 mmol / L continuously stimulated Hep G2 cells for 24 h, and 1 nmol / L insulin was stimulated for 15-25 min. The glucose consumption of induced Hep G2 cells decreased significantly, indicating that the Hep G2-IR cell model was successfully established. 3. Compared with the model control group, the glucose consumption of Hep G2-IR cells was increased in both groups, and each group increased with the intervention concentration. Glucose consumption increased gradually in the same concentration, the glucose consumption of the combined group increased most significantly (P0.05N. 4. Western-blot analysis showed. Compared with the model control group, the PI3K protein expression and the ratio of p Akt/Akt protein expression in Hep G2-IR cells were increased. With the increase of intervention concentration, the ratio of protein expression and protein expression level increased gradually in each group, at the same concentration. The ratio of PI3K protein expression and p Akt/Akt protein expression in combination group was significantly higher than that in control group (P 0.05). DCI and their combination may increase the glucose consumption of Hep G2-IR cells by increasing the expression of PI3K protein and the phosphorylation of Akt. In order to achieve the purpose of improving IR, the combined effect is the most significant.
【学位授予单位】：青岛大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：R587.1

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