| Objective To compare the changes of CHOP and Caspase3expression in normoal glucose tolerance (NGT) rats, impaired glueose tolerance (IGT) and IGT rats with GLP-1agonists intervention. To explore protective effect and possible mechanism of GLP-1agonists on the endoplasmic reticulum stress pathway of apoptosis in myocardial cells.Methods Fifty-four male Wistar rats in4-5weeks (150g-170g) of clean grade were purchased in Physiology laboratory of Shanxi Medical University. After one week adaptive feeding (five/cage), they were divided into A group (n=18) and B group (n=36) randomly. The A group rats were fed by normal diet (13.4kJ/g, which accounted for10.2%of calories, fat and protein accounted for23.3%respectively, carbohydrate accounted for66.5%); The B group rats were given high-sugar and high-fat feed (21.8kJ/g, which accounted for56.0%of calories, fat and protein accounted for7.0%respectively, carbohydrates accounted for37.0%) and free access to water, at room temperature controlled at18-22℃, relative humidity30%-70%. Two groups of rats twice a day morning and evening feeding, daily food in take for each rat to vote for3%of body weight. When12weeks, do oral glucose tolerance test (OGTT):after fasting12-14h, cut the tail blood and rapid blood glucose meters measure fasting blood glucose (FBG), intragastric administration with50%glucose injection2g/kg body weight, after2h cut the tail blood again and measure2h postprandial blood glucose (2hPG). Building a successful model was7.8mmol/L<2hPG<11.lmmol/L and continued for more than a week.The A group, blood glucose to normal, and being setted to normoal glucose tolerance control group (NGT group, n=18). Thirty-two rats in B group were into models, it has the making model success rate of88%, and randomly divided the B group into two groups, the impaired glucose tolerance group (IGT group, n=16), Exendin-4interventionin group (Ex group, n=16). Select randomly nine rats in NGT group, eight rats in IGT group and Ex group respectively, weighing, and then do oral glucose tolerance test measure2h PG. The next day, after fasting12-14h, anesthetized rats with5%chloral hydrate by intraperitoneal injection0.6ml/100g body weight, quickly remove the heart with inc PBS wash clean and dry with filter paper. And then obtain a little apical tissues, fixed with10%formaldehyde solution, dehydrated, embedded in paraffin, regular slices, for HE staining immunohistochemical detection the Caspase3expression of myocardial. and put the remaining pancreas rapidly in liquid nitrogen, after thoroughly frozen, stored in-70℃ refrigerator. Measured the expression of CHOP mRNA by real time quantitative polymerase chain reaction (RT-PCR). The remaining rats in NGT group continue to give normal diet, the IGT group and Ex group were to continue to give high-sugar high-fat feeding and freedom drinking water. Ex group received Exendin-4(Eli Lilly and Company to provide)5ug/kg subcutaneously, twice daily, in the NGT group and the IGT group were given equal volume of saline injection. When interventing4weeks, weighing, and then measure2h PG. The next day, anesthetized rats, quickly remove the heart, then obtain a little pancreas tissues, for immunohistochemical detection the Caspase3expression of heart. and measured the expression of CHOP mRNA by RT-PCR. All results were measured by (x±s), using SPSS17.0software package, multiple groups means were compared with single factor analysis of variance, LSD test. P<0.05was considered statistically significant. Then Pearson correlations were used to determine the relationship between Caspase3and CHOP.Results1.The changes of body and2hPG between rats in each groups Before intervention, compared with the NGT group, the body weigh of rats in the IGT group and the Ex group were higher (307.44±71.85vs.171.74±12.1)(321.71±105.65vs.171.74±12.1) and the differences were also significant (both P<0.05). the2hPG of rats were higher (5.96±0.77vs.9.69±0.33)(5.96±0.77vs.9.26±0.38) the differences were also significant (both P<0.05). Compared with the IGT group, the body weigh of rats in the Ex group was higher (321.71±105.65vs.307.44±71.85) but the difference was not significant (P>0.05). After intervented with Exendin-44weeks, compared with the IGT group, the body weigh and the2hPG of rats in the Ex group were decreased (520.75±18.42vs.445.88±20.29)(10.28±0.38vs.7.19±0.34) the differences were significant (both P<0.05). Compared with the Ex group before intervented, the levels were decreased (513.12±11.86vs.321.71±105.65)(9.28±0.38vs.7.19±0.34) the differences were significant (bothP<0.05).2.The expression of Caspase3in myocardial with eath groups Before intervention, compared with the NGT group, the expression of Caspase3with rats in the IGT group and the Ex group were increased (49.27±2.14vs.30.65±1.46)(49.73±3.56vs.30.65±1.46) and the differences were significant (both P<0.05). Compared with the IGT group, the expression of Caspase3with rats in the Ex group was higher (49.73±3.56vs.49.27±2.14) but the difference was not significant (P>0.05). After intervented with Exendin-44weeks, compared with the IGT group, the expression of Caspase3in the Ex group was lower (40.79±3.81vs.53.31±3.03) the differences was significant (P<0.05). Compared with the Ex group before intervented, the levels was also lower(40.79±3.81vs.49.73±3.56) the differences were significant (P<0.05). Compared with the NGT group the level was increased (40.79±3.81vs.31.57±0.93) the difference was significant (P<0.05).3. The expression of CHOP in pancreas with eath groups Before intervention, compared with the NGT group, the expression of CHOP with rats in the IGT group and the Ex group were increased(0.44±0.13vs.0.03±0.01)(0.49±0.67vs.0.03±0.01) and the differences were significant (both P<0.05). Compared with the IGT group, the expression of CHOP with rats in the Ex group was higher (0.49±0.67vs.0.44±0.13) but the difference was not significant (P>0.05). After intervented with Exendin-44weeks, compared with the IGT and Ex group, the expression of CHOP in the Ex group was decreased (0.17±0.07vs.2.44±2.04)the differences was significant (both P<0.05). Compared with the NGT group the level was incresed (0.17±0.07vs.0.49±0.67) the difference was significant (P<0.05).4. The changes of myocardial histology in HE staining.Before intervention HE staining showed that normal cardiac muscle cells arranged in neat rows, the nucleus size of the uniform, uniform cytoplasmic staining in NGT group generally normal. Myocardial cells arrangement were not structured, nuclei vary in size, cytoplasmic staining uneven in the IGT and Ex group. After intervented with Exendin-44weeks, Ex group than IGT group in the former had inproved myocardial structure.5. The expression of CHOP and the expression of Caspase3in myocardial was significantly correlated in rats with impaired glucose tolorence (r=0.65,P<0.01).Conclusions1. In rats with impaired glucose tolerance, the expression of CHOP with rats was increased, the expression of Caspase3with rats was also increased, endoplasmic reticulum stress has happened in myocardial cells,2. Exendin-4inhibits the CHOP signaling pathways, to reduce endoplasmic reticulum stress response, and ultimately inhibit the endoplasmic reticulum-mediated apoptosis in myocardial cells. |
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