补肾醒脑法对2型糖尿病认知功能障碍小鼠性激素信号通路相关激素水平、海马Erα和ErβmRNA的影响研究

发布时间：2018-01-17 17:03

本文关键词：补肾醒脑法对2型糖尿病认知功能障碍小鼠性激素信号通路相关激素水平、海马Erα和ErβmRNA的影响研究　出处：《成都中医药大学》2016年博士论文　论文类型：学位论文

【摘要】：本论文分为2型糖尿病(Type 2 Diabetes Mellitus T2DM)认知功能障碍与性激素信号通路相关性的理论探讨和科学研究两部分。第一部分理论探讨从祖国医学的角度出发,结合导师观点和自身研究,对消渴呆病(糖尿病认知功能障碍)的定义、病因病机、辨证论治和现代医学研究进行简要阐述。第二部分科学研究目的：研究2型糖尿病小鼠性激素信号通路中下丘脑-垂体-性腺轴(Hypothalamic-pituitary-gonadal axis HPG)中相关激素水平、海马中Er α和Er β mRNA水平的表达,进而明确补肾醒脑法改善2型糖尿病认知功能障碍小鼠的认知能力与其对性激素信号通路某些环节的调控的相关性。方法：健康清洁级雌性8周龄昆明小鼠,为正常对照组；健康SPF级(specific pathogen free无特定病原体级)雌性8周龄KKay小鼠为实验组。KKay小鼠,经去势手术和糖尿病认知功能障碍成模检测成功后随机分为模型对照组、阳性对照组、中药组和中药去势组(中药去势组先行去卵巢手术)。阳性对照组予吡拉西坦；中药组、中药去势组予醒脑益智合剂；KKay小鼠糖尿病造模成功后均予罗格列酮常规控制血糖。给药周期共计8周。在糖尿病成模后和给药结束后分别各进行一次Morris水迷宫实验。第二次水迷宫实验结束后,及时于冰上进行实验动物解剖及样本制备。酶联免疫分析法(Enzyme linked immunosorbent assay ELISA)检测各组小鼠相关性激素水平；实时荧光定量聚合酶链式反应(Quantitative Real-time Polymerase Chain Reaction QRT-PCR)测定海马中雌激素受体1 (Estrogen receptor Er a)和雌激素受体2 (Estrogen receptor Er β)mRNA的表达。结果：(1)第一次水迷宫实验结果中正常对照组和各组糖尿病小鼠平台潜伏期比较,各组均明显偏高(P0.05),可见T2DM小鼠认知能力显著低于正常小鼠。去卵巢后小鼠和其他各组小鼠比较P0.05,说明去势后急速衰老的T2DM小鼠认知能力显著低于正常小鼠和壮年的T2DM小鼠。第二次水迷宫实验结果中正常对照组和模型对照组、中药去势组的小鼠比较P0.05,说明这两组的认知能力显著低于正常对照组。给药前后比较：中药去势组、阳性对照组和中药组水迷宫成绩第二次都明显好于第一次(P0.05),即经吡拉西坦治疗后T2DM小鼠和醒脑益智合剂治疗后的T2DM小鼠、去势的T2DM小鼠认知能力均明显提高。(2)在HPG轴性激素水平研究中发现：模型对照组E2、P水平显著低于正常组(P0.05)。模型对照组、阳性对照组、中药去势组的LH、FSH、PRL水平显著高于正常组(P0.05),说明T2DM KKay小鼠、阳性对照组和急速衰老T2DM KKay小鼠的LH、FSH、PRL出现紊乱。而T的变化不大,没有明显统计学差异。在经过醒脑益智合剂治疗后中药组E2、P显著高于模型对照组(P0.05)。中药组和正常对照组,中药去势组与模型组E2、P、LH、FSH比较时P0.05,没有明显统计学差异,这说明使用醒脑益智合剂治疗2型糖尿病认知功能障碍KKay小鼠和其急速衰老小鼠模型是有效的,能调节HPG轴性激素的紊乱,纠正E2、P的低水平,降低LH、FSH的过高水平。(3)在小鼠海马Er α和Er β mRNA的表达研究中：T2DM KKay小鼠和去势的T2DM KKay小鼠Er α和Er β mRNA表达显著低于正常水平(P0.05)；模型对照组和中药组比较P0.05,有明显统计学差异,模型对照组和中药去势组比较P0.05,没有明显统计学差异,说明经醒脑益智合剂治疗T2DM KKay小鼠和去势T2DMKKay小鼠Er α和Er β mRNA表达能有所提高。中药治疗后T2DM KKay小鼠的Er α和Er β mRNA表达显著高于模型对照组；治疗后去势T2DM KKay小鼠Er α和Er βmRNA表达升高接近模型对照组。阳性对照组和中药去势组、模型对照组Er α mRNA表达比较P0.05,有明显统计学差异,说明吡拉西坦对小鼠Er α mRNA表达有一定影响,使Er α mRNA表达在一定程度上升高。模型对照组和阳性对照组Er β mRNA比较P0.05,没有明显统计学差异,说明吡拉西坦对T2DM小鼠Er β mRNA表达影响不大。结论：在常规控制血糖的基础上,醒脑益智合剂能有效提高T2DM认知功能障碍小鼠和去势后急速衰老的T2DM认知功能障碍小鼠的学习、记忆能力,其可能的作用机制可能是通过调整HPG性激素的水平,升高E2、P,降低LH、FSH;增强大脑海马Er α和Er β mRNA水平的表达而达到的。
[Abstract]:This paper is divided into type 2 diabetes (Type 2 Diabetes Mellitus T2DM) cognitive theory and scientific research related dysfunction and sexual hormone signaling pathways of two parts. Starting from the perspective of traditional Chinese medicine combined with the first part of the theory, research ideas and their mentor cancellation, stay thirsty disease (diabetic cognitive dysfunction) definition, etiology machine, TCM and modern medicine research were briefly described. The second part of the scientific research objective: To study the type 2 diabetic mice sex hormone signaling in the hypothalamic pituitary gonadal axis (Hypothalamic-pituitary-gonadal axis HPG) in hormone levels, expression of hippocampal Er alpha and Er beta mRNA level, then clearly improve Bushenxingnao method the cognitive ability of type 2 diabetic mice with cognitive impairment on sex hormone signaling pathway in some segments of the control. Methods: healthy female 8 week old Kunming mice as normal control group; SPF (specific pathogen free healthy specific pathogen free) female 8 week old KKay mice to.KKay mice in the experimental group, after castration and diabetic cognitive dysfunction model after successful detection were randomly divided into model group, positive control group, Chinese medicine group and traditional Chinese medicine the ovariectomized group (Chinese medicine group first castrated ovariectomy). Positive control group treated with piracetam; traditional Chinese medicine group, Chinese medicine group received castration xinnaoyizhi Decoction; KKay mice rats were treated with rosiglitazone. The conventional control of blood glucose administration period a total of 8 weeks. After diabetic model and after medication respectively. The first Morris water maze test. The end of the second after the water maze test, in experimental animal anatomy and sample preparation on ice. Enzyme linked immunosorbent assay (Enzyme linked immunosorbent assay ELISA) to detect the small Correlation between hormone levels in rats; real-time fluorescence quantitative polymerase chain reaction (Quantitative Real-time Polymerase Chain Reaction QRT-PCR) determination of estrogen receptor in hippocampus (Estrogen 1 receptor Er a) and estrogen receptor 2 (Estrogen receptor Er) on the expression of mRNA. Results: (1) the first water maze test results in the normal control group and diabetic group platform mice incubation period between groups were significantly higher (P0.05), the cognitive ability of visible T2DM mice was significantly lower than that of normal mice. The ovariectomized mice and the mice in other groups compared with P0.05, said T2DM cognitive ability rapid aging Ming after castration was significantly lower than that of normal mice and T2DM mice. In the second water maze experiment results in normal control group and model control group, Chinese medicine group were castrated P0.05, illustrate the cognitive ability of the two groups were significantly lower than the control group. Before administration After comparison: Chinese medicine castration group, positive control group and Chinese medicine group water maze test second times better than the first time (P0.05), by Laci Staw after the treatment of T2DM mice and xingnaoyizhi mixture in the treatment of T2DM after castration mice, T2DM mice cognitive ability were significantly improved. (2) found in the study of sex hormones the HPG axis: the model control group E2, the level of P was significantly lower than that of normal group (P0.05). The model control group, positive control group, Chinese medicine LH FSH, castration group, PRL level was significantly higher than that of normal group (P0.05), T2DM KKay mice, positive control group and the rapid aging T2DM mice with KKay LH FSH. PRL and T disorders. Little change, no significant statistical difference. After xingnaoyizhi mixture after treatment in Chinese medicine group E2, P were significantly higher than the model group (P0.05). The Chinese medicine group and normal control group, Chinese medicine group and the ovariectomized model group E2, P, LH, FSH is P0.05, there is no clear Significant statistical difference, which shows that the use of xingnaoyizhi mixture in the treatment of type 2 diabetic cognitive dysfunction in KKay mice and the rapid aging mice model is effective and can regulate the HPG axis hormone disorder, correct E2, lower level, P LH, the high level of FSH. (3) the expression of alpha and Er in hippocampus of mice Er beta mRNA: T2DM KKay T2DM KKay mice and ovariectomized mice Er alpha and Er beta mRNA expression was significantly lower than the normal level (P0.05); model group and Chinese medicine group P0.05, there was significant difference in model control group and Chinese medicine castration group P0.05, no statistically significant difference, which shows that the effect of Xingnao Yizhi mixture the treatment of T2DM KKay mice and ovariectomized T2DMKKay mice Er alpha and Er beta mRNA expression can be increased. The expression of T2DM in mice after treatment with Chinese medicine KKay Er alpha and Er beta mRNA was significantly higher than that in the model control group; after treatment of ovariectomized female T2DM KKay mice Expression of Er alpha and Er beta mRNA increased to the model group. The positive control group and Chinese medicine castration group, model control group, Er mRNA expression of P0.05, there was significant difference that piracetam has a certain effect on the mice Er alpha mRNA expression, the Er expression of mRNA alpha high rise in a certain extent. The model control group and the positive control group Er beta mRNA P0.05, no significant difference, indicating piracetam has little effect on the T2DM expression of mouse Er beta mRNA. Conclusion: Based on conventional glucose control, Xingnao Yizhi mixture can effectively improve the T2DM cognitive dysfunction in mice after castration and rapidly aging T2DM cognitive dysfunction in mice's learning and memory ability. And the possible mechanism may be HPG by adjusting the level of sex hormones, increased E2, P, lower LH, FSH; enhance the expression of hippocampal Er alpha and Er beta mRNA level and to achieve.

【学位授予单位】：成都中医药大学
【学位级别】：博士
【学位授予年份】：2016
【分类号】：R259;R277.7

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