【摘要】：研究背景2型糖尿病是一種年齡相關(guān)性疾病,隨著病程進(jìn)展,胰島β細(xì)胞功能及含量逐漸衰減。研究發(fā)現(xiàn)生長(zhǎng)分化因子]1(GDF11)可以改善多種器官衰老表型及正向調(diào)控胰島β細(xì)胞的分化成熟。但是,尚無(wú)關(guān)于GDF11對(duì)成年胰島β細(xì)胞作用的文獻(xiàn)報(bào)道。研究目的本研究旨在明確GDF11對(duì)糖尿病小鼠胰島β細(xì)胞功能及含量的影響及可能機(jī)制。方法購(gòu)買(mǎi)6周齡雄性SPF級(jí)C57BL/6小鼠,用高脂飲食聯(lián)合鏈脲佐菌素(STZ)建立誘發(fā)性2型糖尿病小鼠模型(即HFD/STZ小鼠)。購(gòu)買(mǎi)6周齡雄性SPF級(jí)db/db小鼠作為自發(fā)性2型糖尿病模型。本研究選擇了誘發(fā)性和自發(fā)性2型糖尿病動(dòng)物模型作為研究對(duì)象,通過(guò)外源性補(bǔ)充或中和循環(huán)中的GDF11水平,觀察GDF11對(duì)小鼠胰島β細(xì)胞的作用及機(jī)制。每周定期測(cè)定各組小鼠的攝食量、體重及空腹血糖。干預(yù)完成后,各組小鼠行腹腔葡萄糖耐量實(shí)驗(yàn)(IPGTT),并測(cè)定基線及糖負(fù)荷30min的血漿胰島素水平。實(shí)驗(yàn)結(jié)束時(shí),ELISA檢測(cè)血清生理生化水平;ELISA檢測(cè)血清及胰腺內(nèi)胰島素、胰高血糖素水平;采用RT-PCR分析胰島PDX-1、MafA、NKX6.1、Insulin2基因轉(zhuǎn)錄水平。取胰腺組織行HE染色及免疫熒光定量分析胰島形態(tài)及細(xì)胞含量變化。Western blot檢測(cè)胰島相關(guān)蛋白表達(dá)水平。結(jié)果研究結(jié)果顯示,增加循環(huán)內(nèi)GDF11水平對(duì)正常小鼠糖脂代謝無(wú)明顯影響。但是,補(bǔ)充GDF11可顯著降低兩種糖尿病模型小鼠的空腹血糖、HbAlc及血脂水平,降低胰高血糖素濃度,改善糖耐量,同時(shí)明顯增加空腹及糖負(fù)荷30min后的胰島素水平,上調(diào)胰島PDX-1、MafA、NKX6.1、Insulin2基因表達(dá)。另外,rGDFl1和AAV-GDF11干預(yù)后可維持β細(xì)胞含量,降低β細(xì)胞凋亡率,但是對(duì)α細(xì)胞含量、β細(xì)胞大小及增殖率無(wú)明顯影響。而且,補(bǔ)充GDF11可以增加胰腺內(nèi)胰島素含量,但對(duì)胰高血糖素含量無(wú)明顯作用。另外,中和循環(huán)中GDF11對(duì)正常小鼠無(wú)明顯作用,但可進(jìn)一步加重糖尿病小鼠高血糖,損害糖耐量,抑制胰島素分泌功能及升高β細(xì)胞凋亡率。機(jī)制方面,補(bǔ)充GDF11可顯著增加P-Smad2表達(dá)水平,但對(duì)P-Smad3水平無(wú)明顯作用。同時(shí),補(bǔ)充GDF11可升高P-AKT及P-Fox01表達(dá)水平,但對(duì)P-S6K1水平無(wú)明顯影響。結(jié)論綜上所述,本研究首次證實(shí)GDF11可能通過(guò)激活TGF-β/Smad2及PI3K-AKT-FoxOl信號(hào)通路改善糖尿病小鼠糖脂代謝,抑制胰高血糖素的釋放,保護(hù)胰島β細(xì)胞功能,減少β細(xì)胞凋亡。然而,使用抗體中和血液中GDF11可誘導(dǎo)相反作用。GDF11可能在胰島β細(xì)胞功能及含量中扮演重要角色,對(duì)探討糖尿病的防治方法提供了新的思路。
[Abstract]:Background Type 2 diabetes mellitus (T2DM) is an age-related disease. With the progression of the disease, the function and content of islet 尾 cells decrease gradually. It was found that growth differentiation factor] 1 (GDF11) could improve the senescence phenotype of many organs and positively regulate the differentiation and maturation of islet 尾 cells. However, there is no literature about the effect of GDF11 on adult islet 尾-cells. Objective to investigate the effect of GDF11 on the function and content of islet 尾 cells in diabetic mice and its possible mechanism. Methods six week-old male SPF grade C57BL/6 mice were purchased and induced type 2 diabetic mice (HFD/STZ mice) were established by high fat diet combined with streptozotocin (STZ). Six week old male SPF db/db mice were purchased as a model of spontaneous type 2 diabetes mellitus. In this study, the animal models of induced and spontaneous type 2 diabetes mellitus were selected to observe the effect and mechanism of GDF11 on islet 尾 cells in mice by exogenous supplementation or neutralization of GDF11 levels in circulation. The food intake, body weight and fasting blood glucose of each group were measured regularly every week. After the intervention, the plasma insulin levels of baseline and glucose loaded 30min were measured by intraperitoneal glucose tolerance test (IPGTT),) in each group. At the end of the experiment, serum physiological and biochemical levels were detected by ELISA, insulin and glucagon levels in serum and pancreas were detected by ELISA, and PDX-1,MafA,NKX6.1,Insulin2 gene transcription level of islet was analyzed by RT-PCR. The changes of pancreatic morphology and cell content in pancreatic tissue were determined by HE staining and immunofluorescence. The expression of islet associated protein was detected by. Western blot. Results the results showed that increasing the level of GDF11 in circulation had no significant effect on the metabolism of glucose and lipid in normal mice. However, supplementation of GDF11 could significantly decrease the levels of fasting blood glucose, HbAlc and blood lipid, decrease the concentration of glucagon, improve glucose tolerance, and increase the insulin level after 30min. Upregulation of islet PDX-1,MafA,NKX6.1,Insulin2 gene expression. In addition, rGDFl1 and AAV-GDF11 could maintain 尾 cell content and decrease 尾 cell apoptosis rate, but had no significant effect on 偽 cell content, 尾 cell size and proliferation rate. In addition, supplementation of GDF11 could increase insulin content in pancreas, but had no effect on glucagon content. In addition, GDF11 had no obvious effect on normal mice in neutralization cycle, but it could further aggravate hyperglycemia, damage glucose tolerance, inhibit insulin secretion and increase the rate of 尾 cell apoptosis in diabetic mice. In terms of mechanism, supplementation of GDF11 could significantly increase the level of P-Smad2 expression, but had no effect on P-Smad3 level. At the same time, supplementation of GDF11 could increase the expression of P-AKT and P-Fox01, but had no significant effect on the level of P-S6K1. Conclusion in conclusion, GDF11 may improve glucose and lipid metabolism, inhibit glucagon release, protect islet 尾 cell function and decrease 尾 cell apoptosis by activating TGF- 尾 / Smad2 and PI3K-AKT-FoxOl signaling pathway in diabetic mice for the first time. GDF11 may play an important role in the function and content of islet 尾 cells, which provides a new idea for the prevention and treatment of diabetes mellitus.
【學(xué)位授予單位】：南方醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類(lèi)號(hào)】：R587.1

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相關(guān)期刊論文 前3條

1 李倩;郝志明;;生長(zhǎng)分化因子11:TGF-β超家族的新成員[J];生物化學(xué)與生物物理進(jìn)展;2015年07期

2 閆柄文;劉毅;裴海峰;王Y，

本文編號(hào)：2329488