The Expression Of GnRH In Pancreas And Intestine In Hyperlipidemic Gestational Rats

Some clinical data confirm that the GnRH analogues are related to glucose metabolism. GnRH proceed a radical change during gestation, and studies showed treatment with long-acting triptorelin acetate made the prevalence of gestational diabetes mellitus (GDM) increased significantly in women who was undergoing vitro fertilization pregnancies. GnRH and its receptor showed widespread expression in digestive system, including exocrine pancreas, jejunum, ileum, colon and so on. Whether GnRH secreted by pancreas and intestine is changed during gestation? Whether the change is related to glucoregulation during gestation, and whether it plays a critical role in insulin resistance and GDM during gestation by acting on GnRH receptor in pancreas and intestine through effecting the secreting of islet hormones, intestinal absorption and motor function, or regulating secretion of other gastro-intestinal hormones, such as glucagon, glucagons like peptid-1, all these issues need to be investigated. The aim of this study is to explor the function of GnRH secreted by pancreas and intestine in the pathophysiology mechanism of GDM. OBJECTIVE:The purposes of this study were to identify the changes of the expression of GnRH, GnRH receptor, proglucagon and GLP-1 receptor in pancreas and intestine in gestational rat with diet-induced hyperlipidia, and to seek the relationships among the expression of GnRH, GnRH receptor, proglucagon and GLP-1 receptor in pancreas and intestine.METHOD:1. Immunohistochemical method was used to show the protein expression and the distribution of GnRH and its receptor in pancreas in gestational rat with diet-induced hyperlipidia at the time of 14 days after gestation.2. RIA was used to identify insulin during OGTT/meal test in gestational rat with diet-induced hyperlipidia at the time of 13 or 14 days after gestation.3. Real-time quantitative PCR was used to detect the changes of the mRNA expressions of GnRH, GnRH receptor, proglucagon and GLP-1 receptor in pancreas, jejunum, ileum and colon in gestational rat with diet-induced hyperlipidia at the time of 14 days after gestation.4. ELISA was used to detect the changes of the protein expression of GnRH in pancreas in gestational rat with diet-induced hyperlipidia at the time of 14 days after gestation.5. Western Blot was used to detect the changes of the protein expression of glucagon in pancreas in gestational rat with diet-induced hyperlipidia at the time of 14 days after gestation.RESULTS:1. The expression of GnRH in pancreas in hyperlipidemic gestational ratsA. OGTT showed insulin AUC in Hch+Gestation rats and Hch rats was higher than that in Control rats(204.60±79.06 vs 129.71±11.33mIU/L , P<0.05 ),(230.25±13.19 vs 129.71±11.33 mIU/L , P<0.05 ). ISI in Hch+Gestation rats was lower than that both in Control rats(2.21±0.32 vs 4.95±0.56, P<0.01)and Gestation rats(2.21±0.32 vs 4.64±0.90, P<0.01), while ISI in Hch rats was lower than Control rats (2.84±0.52 vs 4.95±0.56, P<0.01). Fasting insulin in Hch+Gestation rats was no difference with that in Hch rats. Fasting insulin in Gestation rats was higher than that in Control rats(21.68±2.55 vs 14.35±0.86,P<0.05).B. Immunohistochemical staining showed there are GnRH and GnRH receptor expressed in rat pancreas. The percentage of GnRH positive cell in Hch+Gestation rats was higher than in Control rats(P<0.05).C. In pancreas, GnRH relative mRNA level in Hch+Gestation rats was 1.76 fold higher than that in Control rats(P<0.01), 1.53 fold higher than that in Gestation rats (P<0.01)and 1.4 fold higher than that in Hch rats(P<0.05). GnRH protein in Hch+Gestation rats was higher than that in Control rats (7.06±0.28 vs 5.40±0.41 mg/L,P<0.05),higher than that in Gestation rats (7.06±0.28 vs 5.95±0.38 mg/L,P<0.05) and higher than that in Hch rats (7.06±0.28 vs 5.97±0.30 mg/L,P<0.05).D. PG relative mRNA level in Hch+Gestation was 1.77 fold higher than that in Control rats(P<0.01),1.54 fold higher than that in Hch rats(P=0.01),PG relative mRNA levels in Gestation rats was 1.46 fold higher than that in Control rats ( P<0.05 ) . The glucagon in Hch+Gestation rats was 3.44 fold higher than in that Control rats(P<0.01)and 2.9 fold higher than that in Hch rats (P<0.01), the glucagon in Gestation rats was 2.57 fold higher than in that Control rats(P<0.01). E. A significant positive correlation between the expression of GnRH mRNA and PG mRNA was observed in rat's pancreas, r =0.521,P=0.01.2. The expression of GnRH in intestine in hyperlipidemic gestational ratsA. Meal test showed Hch+Gestation rats insulin AUC was higher than that in Control rat(s213.88±8.51 vs 155.87±12.39 mIU/L,P<0.01 )and that in Gestation rats (213.88±8.51 vs 169.27±6.55 mIU/L,P<0.01). Insulin AUC in Hch rats was higher than that in Control rats(188.52±16.70 vs 155.87±12.39 mIU/L,P<0.05 ); HOMA-IR in Hch+Gestation rats was higher than that in Control rats(7.66±0.93 vs 4.14±0.47,P<0.01), HOMA-IR in Hch rats was higher than that in Control rats(6.59±0.33 vs 4.14±0.47,P<0.01).B. In jejunum, GnRH relative mRNA level in Gestation rats was 2.68 fold higher than in Control rats (P<0.01), 1.45 fold higher than in Gestation rats(P<0.05)and 1.49 fold higher than in Hch rats(P<0.05). GnRH relative mRNA level in Hch rats was 1.86 fold higher than in Control rats(P<0.05). PG relative mRNA level in Hch+Gestation rats was 1.78 fold higher than in Control rats(P<0.05),1.53 fold higher than in Gestation rats(P<0.05). PG relative mRNA level in Hch rats was 1.52 fold higher than in Control rats (P<0.05), GnRHR relative mRNA level in Gestation rats was 1.85 fold higher than in Control rats(P<0.05). A significant positive correlation between the expression of GnRH mRNA and PG mRNA was observed in rat's jejunum, r =0.699,P<0.01.C. In ileum, GnRH relative mRNA level in Hch+Gestation rats was 2.18 fold higher than in Control rats(P<0.05). PG relative mRNA level was no difference among rats. A significant positive correlation between the expression of GnRH mRNA and PG mRNA was observed in rat's ileum, r =0.755,P<0.01.D. In Colon, GnRH relative mRNA level was no difference among rats. PG relative mRNA level in Hch+Gestation rats was 2.07 fold higher than in Control rat(sP<0.05),PG relative mRNA level in Gestation rats was 2.06 fold higher than in Control rats(P<0.05). PG relative mRNA level in Hch rats was 2.11 fold higher than in Control rats(P<0.05). A significant positive correlation between the expression of GnRH mRNA and PG mRNA was observed in rat's colon, r =0.513,P<0.01.CONCLUSION:Pancreatic GnRH level in Hch+Gestation rats was higher than that in Control and Hch rats, while insulin resistance was more severe than Hch rats and Gestation rats. The mechanisms of these changes were unclear. But a significant positive correlation between the expression of GnRH mRNA and PG mRNA was observed in rat's pancreas, which might suggest pancreatic GnRH might influence glucagon secretion in hyperlipidemia rat during gestation. Significant positive correlations between the expression of GnRH mRNA and PG mRNA were observed in rat's jejunum, ileum and colon. These might also suggest intestinal GnRH may interact with intestinal PG, which influence glucose metabolism.