The Effects Of GnRH And GnIH On Pancreatic Islet Cells

As a kind of metabolic disease,type 2 diabetes mellitus(T2DM)has become an increasingly serious public health problem.More and more studies have found that the body's energy metabolism and reproductive function are closely related,also affect each other.Hypothalamus synthesizes gonadotropin releasing hormone(GnRH)and gonadotropin inhibitory hormone(GnIH),and they are the key hormones regulating reproduction.The receptors of GnRH and GnIH were found in tissues and organs which closely related to metabolism and reproduction.Previous in situ pancreatic perfusion experiments in rats showed that GnRH promoted glucagon secretion under high glucose conditions,and GnIH also had the effect of regulating feeding.These findings suggest that GnRH and GnIH may be associated with the pancreas-the key organs regulating energy metabolism.Therefore,we studied the effects of GnRH and GnIH on islet cells.Part 1: The effects of GnRH on pancreatic islet cells.We found the receptor of GnRH(GnRHR)expressed in mouse pancreatic islet alpha cell line alpha TC1-6 and beta cell line beta TC-6 by real-time PCR and western blot.Then we studied whether GnRH promoted the dedifferentiation and transdifferentiation of beta cells into alpha cells under high glucose conditions,which led to high level of glucagon.The results showed that GnRH did not change the localization and expression of NKX6.1,a characteristic transcription factor of beta cells,in both beta TC-6 cells and pancreatic primary beta cells under high levels of saturated free fatty acid palmitate and glucose conditions.GnRH did not promote proliferation in both alpha TC1-6 cells and beta TC-6 cells.Part 2: The effects of GnIH on pancreatic islet cells.We found the expression of GnIH receptor(GPR147)in mouse alpha TC1-6 cells and beta TC-6 cells by real-time PCR.Immunohistochemical studies also confirmed the expression of GPR147 in both mouse alpha TC1-6 cells and pancreatic islets.The fluorescent in situ hybridization(FISH)combined with immunofluorescence results also showed that the GPR147 mRNA and glucagon are colocalized in alpha TC1-6 cells.Cell Counting Kit-8(CCK-8)was used to detect the effects of GnIH(also named RFRP-3 in mammals)on the survival and proliferation of alpha TC1-6 cells.We found that GnIH promoted alpha TC1-6 cells survival under high glucose and serum starvation.Phosphorylated protein assay showed that GnIH promoted the expression of p-AKT and p-ERK 1/2.Treatment with inhibitors for either AKT(MK-2206)or ERK 1/2(PD98059),blocked the survival effects of RFRP-3.After administration of GPR147 antagonist RF9,the activation of PI3K/AKT and ERK 1/2 pathways were blocked.The secretion of glucagon in alpha TC1-6 cells was tested by ELISA.There was no significant difference between the low or high glucose group and the GnIH group.We also found that GnIH could promote the proliferation of beta TC-6 cells under high levels of palmitate and glucose conditions,but had no effect on PI3K/AKT and ERK1/2 signaling pathways.Studies on the dedifferentiation of beta TC-6 cells showed that GnIH had no effect on the localization and distribution of the characteristic transcription factors NKX6.1 and PDX1.GnIH promoted the expression of PCNA both in alpha TC1-6 and beta TC-6 cells.In conclusion,it is difficult for mouse islet cell line beta TC-6 cells to dedifferentiate under high levels of palmitate and glucose conditions.Rat islet primary beta cells can partially dedifferentiate under high levels of palmitate and glucose conditions.GnRH and GnIH have no effect on the dedifferentiation of the above two cells.GnIH promotes the survival and proliferation of mouse alpha TC1-6 cells and beta TC-6 cells.GnIH activates PI3K/AKT and ERK1/2 signaling pathways in alpha TC1-6 cells,but not in beta TC-6 cells.This study revealed the expression and effects of GnRH and GnIH in pancreatic islet cells,and lays a foundation for further study of their effects on primary islet cells and islet cell lines.