基于菊苣菊粉调节JNK和p38 MAPK信号通路对Ⅱ型糖尿病血糖作用机制的研究

发布时间：2018-01-07 05:11

本文关键词：基于菊苣菊粉调节JNK和p38 MAPK信号通路对Ⅱ型糖尿病血糖作用机制的研究　出处：《沈阳农业大学》2017年博士论文　论文类型：学位论文

【摘要】：菊苣菊粉(Chicory inulin,CI)是一种水溶性食物纤维,不能被人体消化,也是一种天然存在的多聚果糖,这种多聚果糖具有降血糖、降血脂以及提高免疫力等作用。以往的研究表明,菊苣菊粉能够促进胰岛分泌胰岛素,并通过影响糖代谢酶的活性抑制糖异生,从而产生降血糖的作用,但其干预Ⅱ型糖尿病的具体作用机制尚未完全阐明,因此本研究通过对Ⅱ型糖尿病模型大鼠口服菊苣菊粉,探讨其对Ⅱ型糖尿病模型大鼠的血糖、体重及糖脂代谢的影响,并通过体内和体外试验探究菊苣菊粉通过调节JNK和p38 MAPK信号通路对Ⅱ型糖尿病大鼠血糖的改善作用,揭示菊苣菊粉对Ⅱ型糖尿病大鼠降血糖作用的机制,为应用菊苣菊粉适合Ⅱ型糖尿病人群的食品开发提供相关的理论依据。本论文具体主要研究内容及结论如下:腹腔注射45 mg/kg链脲佐菌素(Streptozotocin,STZ),结合高脂饮食喂养构建T2DM大鼠模型,正常对照组大鼠静脉注射等体积生理盐水,常规饲料喂养。造模成功后随机分为6组:正常对照组(Control组)、模型组(T2DM组)、菊苣菊粉低剂量组(2.5g/kg CI组)、菊苣菊粉中剂量组(5g/kgCI组)、菊苣菊粉高剂量组(10g/kgCI组)、二甲双胍对照组(Metformin组)。连续灌胃给药8周,除正常对照组,剩余组大鼠采用高脂饮食喂养。8周给药期结束后,检测各组大鼠血糖浓度,测量记录大鼠体重。结果显示,与正常组大鼠相比,Ⅱ型糖尿病模型大鼠血糖浓度显著升高(p0.01),且Ⅱ型糖尿病模型大鼠体重均小于正常组大鼠(p0.01);说明菊苣菊粉可以降低Ⅱ型糖尿病模型大鼠血糖浓度,同时有效增加Ⅱ型糖尿病模型大鼠体重。研究菊苣菊粉对Ⅱ型糖尿病大鼠血脂代谢能力的影响,检测各组大鼠血红蛋白(Hemoglobin,HB)、糖化血红蛋白(Hemoglobin A1c,HbA1c)、胰岛素(Insulin,INS)、甘油三酯(Triglyceride,TG)、总胆固醇(Total cholesterol,TC)、游离脂肪酸(Nonesterified fatty acid,NEFA)等血脂代谢指标含量。结果显示,Ⅱ型糖尿病模型大鼠高脂喂养8周后血清中HB、HbA1c、INS、TG、TC、NEFA含量均显著性上升(p0.05),给药8周后,药物干预组HB、HbA1c、INS、TG、TC、NEFA含量均表现不同程度下降。说明菊苣菊粉能够改善Ⅱ型糖尿病大鼠血脂代谢紊乱。为了阐明菊苣菊粉改善糖尿病的分子机制,本研究通过Western blot法检测与糖脂代谢有关的JNK和p38 MAPK信号通路相关蛋白IRS、JNK及p38在肝脏和肌肉组织中的活化情况。结果显示,菊苣菊粉可以有效活化骨骼肌和肝脏组织中IRS信号,抑制JNK和p38 MAPK信号通路,并且菊苣菊粉这种干预作用具有一定的浓度依赖性。为进一步验证菊苣菊粉可以在体外通过调节JNK和p38 MAPK通路改善Ⅱ型糖尿病大鼠的血糖,采用MTT法检测不同浓度菊苣菊粉对小鼠成肌细胞C2Cl2和人肝癌细胞HepG2生长活性的影响,结果显示,在0～2.0mg/mL范围内,菊苣菊粉对C2C12和HepG2细胞生长均无显著性影响。通过荧光标记葡萄糖类似物法检测菊苣菊粉对C2C12和HepG2葡萄糖摄取能力的影响。结果显示,随着菊苣菊粉浓度的升高,C2C12和HepG2葡萄糖摄取能力也随之增强。同时Western blot结果显示,菊苣菊粉在体外亦能通过下调C2C12和HepG2细胞的JNK和p38 MAPK信号活性发挥作用。此外,为了进一步阐明JNK和p38 MAPK信号与菊苣菊粉在调控葡萄糖代谢中的作用,通过分别使用JNK信号抑制剂SP600125和p38 MAPK信号抑制剂SB203580处理,采用荧光标记葡萄糖类似物法检测细胞糖摄取能力的变化,结果显示,抑制JNK和p38 MAPK信号时可协同菊苣菊粉进一步增强细胞葡萄糖摄取能力。以上试验结果说明,菊苣菊粉可以降低Ⅱ型糖尿病模型大鼠的血糖、HB、HbA1c、INS、TG、TC、NEFA的浓度,改善Ⅱ型糖尿病大鼠血脂代谢功能,并有效增加Ⅱ型糖尿病模型大鼠的体重;同时菊苣菊粉能有效活化IRS信号,并通过抑制JNK和p38 MAPK信号通路增强细胞葡萄糖摄取能力,进而对Ⅱ型糖尿病大鼠的血糖起到改善作用。
[Abstract]:Chicory inulin (Chicory inulin CI) is a kind of water soluble dietary fiber can not be digested, but also polyfructose a natural existence, the polyfructose has hypoglycemic, hypolipidemic and improving immune function. Previous studies showed that chicory inulin can promote islet insulin secretion, and the effect of sugar metabolism enzyme activity inhibition of gluconeogenesis, resulting in hypoglycemic effect, but its specific mechanism of intervention of type II diabetes has not been fully elucidated, so this study of type II diabetes rat model of oral chicory inulin, explore the type II diabetic rats blood glucose, weight and lipid metabolism and, in vivo and in vitro study of chicory inulin by regulating the effects of JNK and p38 of MAPK signaling pathway on blood glucose in type II diabetic rats, to reveal the chicory inulin type II diabetes hypoglycemic rats The mechanism, to provide a theoretical basis for the development of food related application of chicory inulin for people with type 2 diabetes. In this paper the main research contents and conclusions are as follows: 45 mg/kg intraperitoneal injection of streptozotocin (Streptozotocin, STZ), to construct T2DM rat model with high fat diet, the rats in the normal control group vein injection of saline, conventional diet. The rats were randomly divided into 6 groups: normal control group (Control group), model group (T2DM group), low dose group of chicory inulin (2.5g/kg CI group), dose group of chicory inulin (5g/kgCI group), chicory inulin in high dose group (10g/kgCI group metformin), control group (group Metformin). Continuous gavage for 8 weeks, in addition to the normal control group, the remaining rats with high fat diet.8 weeks after the end of the period, the detection of blood glucose concentration in rats, measured body weights were recorded. The results showed that with the normal group Compared with rats, blood glucose concentration in rats model of type II diabetes significantly increased (P0.01), and the weight of type II diabetic rats were lower than normal rats (P0.01); that of chicory inulin can reduce the concentration of blood glucose in type II diabetic rat model, and effectively increase the body weight of model rats of type II diabetes sugar. Study of chicory inulin on lipid metabolism ability of type II diabetic rats, hemoglobin of rats (Hemoglobin, HB), glycosylated hemoglobin (Hemoglobin A1c, HbA1c), insulin (Insulin, INS), triglycerides (Triglyceride, TG), total cholesterol (Total, cholesterol, TC), free fatty acids (Nonesterified fatty acid NEFA, etc.) the blood lipid content. The results showed that high fat diet in type II diabetic rats after 8 weeks, serum HB, HbA1c, INS, TG, TC, NEFA were significantly increased (P0.05), 8 weeks after drug administration, drug intervention group HB, HbA1c, INS, TG, TC, NEFA content were decreased in different degrees. It can improve the chicory inulin dyslipidemia in type II diabetic rats. To elucidate the molecular mechanism of chicory inulin improve diabetes, the study by the Western blot method to detect the JNK and p38 with glucose and lipid metabolism of MAPK signaling pathway related proteins IRS, JNK and p38 activation in the liver and in the muscle tissue. The results showed that chicory inulin can effectively activate IRS signaling in skeletal muscle and liver tissue, inhibition of JNK and p38 MAPK signaling pathway, and the effect of this intervention with chicory inulin in a concentration dependent manner. In order to further verify the chicory inulin in vitro by regulating JNK and p38 MAPK pathway to improve type the blood sugar, effects of different concentrations of chicory inulin on detection of mouse myoblasts and C2Cl2 human hepatocarcinoma cell line HepG2 activity by MTT method. The results show that in 0 ~ 2. The range of 0mg/mL, chicory inulin had no significant effect on C2C12 and HepG2 cell growth. Detected by fluorescent labeling of chicory inulin glucose analogs of C2C12 and HepG2 glucose uptake. The results showed that with increasing concentration of C2C12 and HepG2 of chicory inulin, glucose uptake is enhanced. At the same time, Western blot showed that can also play the role of chicory inulin, by JNK and p38 MAPK signaling down-regulation of C2C12 and HepG2 cells in vitro. In addition, in order to further clarify the JNK and the p38 MAPK signal in the regulation of glucose metabolism and chicory inulin in effect, by respectively using JNK and p38 MAPK signal pathway inhibitor SP600125 SB203580 inhibitor, change by fluorescence the glucose analogue method to detect cell glucose uptake ability showed that inhibition of JNK and p38 MAPK signal can further enhance the synergy of Chicory Inulin Cell glucose uptake. These results indicate that the chicory inulin can reduce type II diabetic rats blood glucose, HB, HbA1c, INS, TG, TC, NEFA concentration, improve blood lipid metabolism in type II diabetic rats, and effectively increase the type II diabetic rats weight; chicory inulin can effectively at the same time activation of IRS signaling, and enhanced cellular glucose uptake through inhibition of JNK and p38 MAPK signaling pathway, then the type II diabetic rats blood sugar to improve.

【学位授予单位】：沈阳农业大学
【学位级别】：博士
【学位授予年份】：2017
【分类号】：TS201.4

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