TRB3参与高脂高糖饮食妊娠大鼠糖耐量异常发病机制的研究

发布时间：2018-02-04 21:52

本文关键词： 妊娠 TRB3 AKT CHOP 高脂高糖　出处：《郑州大学》2014年硕士论文　论文类型：学位论文

【摘要】：背景与目的妊娠期糖尿病(gestational diabetes mellitus,GDM)在全球范围内比例逐年上升，带来严重的经济及公共卫生健康问题。妊娠期糖尿病女性罹患各种高危妊娠并发症风险增加，如妊娠期高血压、难产及剖宫产比例升高、巨大儿等，尤其增加了日后发展为糖尿病及心血管疾病的风险。多种因子介导的胰岛素信号传导通路缺陷、内质网应激诱导的β细胞凋亡不同程度的参与了妊娠期糖尿病的发生发展。除此之外，，氧化应激、妊娠期肥胖、高龄等危险因素也在GDM的发病机制中发挥一定作用。最新研究发现：Tribbles同源蛋白3（TRB3）通过TRB3/AKT信号通路、CHOP/TRB3信号通路广泛参与糖尿病的发病机制。但在妊娠期间TRB3与GDM发病机制的关联研究较少，且妊娠期间饮食干预能否有效减轻胰岛素抵抗、降低β细胞凋亡程度的分子机制需要我们进一步证实研究。本实验通过建立高脂高糖饮食妊娠大鼠模型，并观察大鼠模型肝脏组织TRB3和AKT基因的表达水平，观察胰腺组织TRB3及CHOP蛋白表达水平，且对比饮食干预前后的变化，初步探讨TRB3参与妊娠期糖耐量异常发病机制的研究。材料与方法选取105只雌性和30只雄性健康42日龄SD大鼠（体重约200g）。采用普通饲料适应性喂养1周后雌鼠随机分为5组:普通饮食非妊娠组（CV组15只）、普通饮食妊娠组（CG25只）、高脂高糖非妊娠组（FV15只）、高脂高糖妊娠组（FG25只）及高脂高糖妊娠后饮食干预组（DI组25只）（干预组，妊娠前给予高脂高糖饲料，妊娠后给予普通饮食干预）。雄性大鼠均用普通饲料饲养。分别喂养6周后将雄性大鼠与雌性大鼠合笼备孕，拟妊娠雌鼠与雄性大鼠以2∶1比例合笼，次晨镜检雌鼠阴道，以镜下可见精子为标准，标记为1d，并隔离妊娠大鼠。合笼1周后未孕大鼠弃之不用。妊娠第20天进行口服葡萄糖耐量试验（OGTT）检查，分别在0min,60min,120min尾静脉取血检测血糖水平。妊娠第22天时心内取血离心，检测TC、TG、FFA等指标，检测空腹血糖及胰岛素，得出HOMA-IR。应用逆转录聚合酶链反应(RT-PCR)方法测定各组大鼠肝脏组织中TRB3mRNA和AKT mRNA的表达。应用Western Blot方法检测胰腺组织中TRB3和CHOP蛋白的表达水平。妊娠第22天时CG、FG及DI3组各10只大鼠行剖宫产术取胎鼠，称取胎鼠个体及整体重量，称取胎盘重量，取血液检测胎鼠的FBG、胰岛素、血脂水平。采用SPSS19.0对数据进行统计分析。结果 (1)妊娠、高脂高糖饮食均可致大鼠HOMA-IR升高，F妊娠=2318.491，F饮食=2888.237，F交互=993.094，P均＜0.001； (2)相较于其他4组，妊娠、高脂高糖饮食均可致肝组织中TRB3mRNA的表达明显升高，（F妊娠=256.887，F饮食=1749.406，F交互=2.579，P均＜0.001）；AKT mRNA的表达显著降低（F妊娠=221.091，F饮食=1416.984，F交互=28.918，P均＜0.001）。FG组、DI组、CG组三组TRB3mRNA、AKT mRNA的表达水平差异有统计学意义（F=445.986和390.334，P均＜0.001）； (3)相较于其他4组，妊娠、高脂高糖饮食均可致大鼠胰腺组织中TRB3蛋白的表达水平明显升高，（F妊娠=475.219，F饮食=2580.320，F交互=206.142，P均＜0.001）；CHOP蛋白的表达也显著升高（F妊娠=1275.615，F饮食=4308.857，F交互=245.461，P均＜0.001）。FG组、DI组、CG组三组TRB3及CHOP蛋白的表达水平差异有统计学意义（F=764.890和1462.573，P均＜0.001）； (4)FG组体质量增加值、OGTT各时段数据检测值、血糖、胰岛素、TG、FFA、TC等均高于其他4组，差异具有统计学意义； (5)剖宫产取出的单个胎鼠重量在FG、CG、DI三组间无明显差异，但是总胎体重三组之间有差异，具有统计学意义（P0.05）。三组胎鼠的FBG、Insulin和胎盘重量无明显差异。结论 1.可通过长期高脂高糖饮食诱导妊娠期糖耐量异常大鼠模型，高脂高糖及妊娠因素相互叠加作用于妊娠期糖耐量异常甚或妊娠期糖尿病的发病机制； 2. TRB3可通过TRB3/AKT、CHOP/TRB3等信号通路广泛参与高脂高糖喂养大鼠糖耐量异常的发病机制，作用机制有一定相互交叉作用； 3.对妊娠期糖尿病进行饮食干预，可有效改善妊娠期糖尿病病情。
[Abstract]:Background and purpose
Gestational diabetes mellitus (gestational diabetes, mellitus, GDM) increased year by year in the global scope of the proportion of serious economic and public health problems. Pregnant women suffering from high-risk pregnancy increases the risk of complications, such as gestational hypertension, dystocia and cesarean section increased the proportion of macrosomia increased, especially after the development of Japan the risk of diabetes and cardiovascular disease. Insulin signal transduction defects mediated by multiple factors, endoplasmic reticulum stress induced beta cell apoptosis in different degree in the occurrence and development of gestational diabetes. In addition, oxidative stress, pregnancy, obesity, age and other risk factors also play a role in the pathogenesis of GDM. The latest research found: Tribbles homolog 3 (TRB3) through the TRB3/AKT signaling pathway, CHOP/TRB3 signaling pathway is widely involved in the pathogenesis of diabetes in pregnancy but. There is less study on the relationship between TRB3 and GDM pathogenesis, and whether the dietary intervention during pregnancy can effectively reduce insulin resistance and reduce the molecular mechanism of the degree of apoptosis of beta cells, we need further confirmation.
The high fat diet rat model of pregnancy, and to observe the expression of TRB3 and AKT gene in liver tissue of rats, to observe the expression of TRB3 and CHOP protein levels in pancreatic tissue, and compared the changes before and after the dietary intervention, a preliminary study of TRB3 participated in the study of gestational impaired glucose tolerance and pathogenesis.
Materials and methods
A total of 105 female and 30 male healthy 42 day old SD rats (weighing about 200g). The normal diet after 1 weeks of feeding female rats were randomly divided into 5 groups: normal diet and non pregnancy group (CV Group 15), normal diet pregnancy group (CG25), high fat and high glucose non pregnancy group (FV15), high fat and high glucose pregnancy group (FG25) and high fat and high glucose diet after pregnancy intervention group (25 rats in group DI (intervention group), given the high fat and high glucose diet before pregnancy, pregnancy after given normal diet intervention). Male rats were fed with normal diet. After 6 weeks of feeding respectively. Male rats and female rats mated to prepare pregnant, intends to 2 to 1 the proportion of mated female rats and male rats during pregnancy, the next morning microscopic examination of female rats with vaginal microscopically sperm as the standard, labeled 1D, and isolated pregnant rat cage. After 1 weeks of non pregnant rats abandoned.
The oral glucose tolerance test (OGTT) was performed on the twentieth day of pregnancy, and blood glucose levels were measured in 0min, 60min, 120min caudal veins respectively.
The twenty-second day of pregnancy when the heart blood centrifugation, detection of TC, TG, FFA and other indicators, detection of fasting blood glucose and insulin, HOMA-IR. obtained by reverse transcriptase polymerase chain reaction (RT-PCR) method for determination of the expression of TRB3mRNA and AKT mRNA in liver of rats in each group. The expression level of TRB3 and CHOP protein in pancreatic tissue was detected by Western Blot method.
On the twenty-second day of pregnancy, 10 rats in CG, FG and DI3 groups were taken cesarean section to get the fetuses, and the weight and weight of placenta were weighed. The placental weight was weighed, and the levels of FBG, insulin and blood lipid were detected by blood.
SPSS19.0 is used to analyze the data.
Result
(1) pregnancy, high fat and high sugar diet can increase HOMA-IR in rats, F pregnancy =2318.491, F diet =2888.237, F interaction =993.094, P < 0.001;
(2) compared to the other 4 groups of pregnancy, the expression of high fat diet can be induced by TRB3mRNA in liver tissue was significantly elevated (F =256.887 F =1749.406 of pregnancy, diet, F interaction =2.579, P < 0.001); the expression of AKT mRNA decreased significantly (F =221.091 F =1416.984 of pregnancy, diet, F interaction =28.918 P < 0.001) in.FG group, DI group, CG group and three TRB3mRNA group, there was significant difference in the expression level of AKT mRNA (F=445.986 and 390.334, P < 0.001);
(3) compared to the other 4 groups of pregnancy, the expression level of high fat diet can be induced by TRB3 protein in rat pancreatic tissue was significantly elevated (F =475.219 F =2580.320 of pregnancy, diet, F interaction =206.142, P < 0.001); the expression of CHOP protein was also significantly increased (F =1275.615 F =4308.857 pregnancy diet. F, interactive =245.461, P < 0.001) in.FG group, DI group, there was significant difference in the expression level of CG group three groups of TRB3 and CHOP proteins (F=764.890 and 1462.573, P < 0.001);
(4) the value of body mass increase in FG group, data detection value at each time of OGTT, blood glucose, insulin, TG, FFA, TC were all higher than those of the other 4 groups, and the difference was statistically significant.
(5) there was no significant difference in the weight of single fetuses taken from cesarean section among FG, CG and DI groups, but the total fetal weight was different between the three groups (P0.05). There was no significant difference between the three groups of FBG, Insulin and placental weight in the three groups of fetuses.
conclusion
1., we can induce a gestational impaired glucose tolerance rat model through long-term high-fat and high carbohydrate diet. The effects of high fat, high sugar and pregnancy factors on the pathogenesis of gestational impaired glucose tolerance or gestational diabetes mellitus were also observed.
2. TRB3 can be widely involved in the pathogenesis of impaired glucose tolerance in rats fed by high fat and high glucose through TRB3/AKT, CHOP/TRB3 and other signal pathways, and the mechanisms of action are interacted.
3. the diet intervention of gestational diabetes can effectively improve the condition of gestational diabetes.

【学位授予单位】：郑州大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R714.256

【共引文献】