Chronic Epinephrine-induced Endoplasmic Reticulum Stress And Oxidative Stress Affect The Secretion And Fate Of Pancreatic β-cell

Aim and significance: Under long-term high stress state,the increase of adrenaline secretion causes endocrine disorder.The adrenergic receptor on the plasma membrane of pancreatic β-cell is mainly α2 adrenergic receptor(α2-AR),and acute adrenergic secretion is reduced mainly by binding to it.However,subsequent studies have found that under the long-term action of epinephrine,the secretory ability of pancreaticβ-cell is improved and the secretion of insulin is compensated to resist the inhibitory effect of epinephrine.However,these studies were detected immediately after chronic epinephrine,and the secretory ability of pancreatic β-cell after the removal of epinephrine was not detected,so it was impossible to know whether the change was temporary or irreversible.The correct synthesis of insulin is the basis of insulin secretion.Insulin biosynthesis needs to initiate a series of complex responses in the endoplasmic reticulum,which is very sensitive to intracellular homeostasis and extracellular stimulation.Changes of external environment easily trigger endoplasmic reticulum stress,which induces oxidative stress.Therefore,this study will clarify the relationship between chronic epinephrine and endoplasmic reticulum stress and their effects on the secretory ability and fate of pancreatic β-cell,and determine whether this effect is reversible.The discovery of endoplasmic reticulum stress,oxidative stress and targets of insulin secretion disorder in pancreatic β-cell provides a basis for the prevention and treatment of insulin secretion disorder caused by stress,and has certain theoretical application value for healthy breeding of livestock and improving the metabolic health of pets.Methods: We used glucose-stimulated insulin secretion assay(GSIS)to evaluate insulin secretion and synthesis of the isolated islets and MIN6 of two pancreatic β-cell models,which were treated for 5 days,namely the 100 n M epinephrine treatment for3 days,and then normal culture for 2 days.At the same time,the growth rate and activity of MIN6 were detected.By RNA-seq macroscopic screening of chronic epinephrine-activated signaling pathways,predicting whether endoplasmic reticulum stress is the most important issue in MIN6.Because endoplasmic reticulum stress can induce oxidative stress to damage the mitochondria,we used antioxidant enzyme related kits and ATP content kits to detecte and evaluate respectively the intracellular redox state of MIN6 and its ATP synthesis ability under high glucose stimulation.Subsequently,the predicted results,changes in redox state,mitochondrial damage were verified by transmission electron microscopy(TEM),fluorescence quantitative PCR(q RT-PCR)at the molecular level.Moreover,we examined a2-AR expression by q RT-PCR to see if desensitization occurs and is involved in affecting MIN6 secretion capacity and fate.Results: During the five days of the experiment,the insulin secretion of MIN6 increased by 33% and the total insulin content increased by 43%,while the insulin releasation of islet increased by 20%.Epinephrine for 1 day,MIN6 growth fell by56.5%,the number of cells was significantly different,and the difference rate the difference rate of cells number increased.The following two days,the adaptive recovery of the cells growth rate was the same as that of the control group.After one day of epinephrine removal,the cells growth rate doubled,exceeding the control group,the difference rate of cells number decreased sharply,and the cells number difference disappeared.But this growth rate can not continue.When adrenaline removal for two days,the cells growth rate fell,as in the control group.RNA-seq found that endoplasmic reticulum stress and oxidative stress occur when MIN6 was treated for five days.The results of TEM show,the rough endoplasmic reticulum expanded significantly and its local degranulate.In the endoplasmic reticulum pool,a large number of flocculants gather were retained,insulin granules were increased.Statisticsing the mitochondrial density in the TEM photoes showed that the mitochondrial number recovered in the experimental group compared to the third day,not significantly different from the control group,and was consistent with the mt DNA q RT-PCR results.Antioxidant enzyme related kit showed that the total antioxidant capacity of MIN6 increased by 66% and the activity of glutathione peroxidase decreased by 21%.The ATP content test showed that the intracellular ATP content under high glucose stimulation was doubled.The q RT-PCR detection found that at 3days of experimental treatment,transcription of Bip,Ire1 a,Xbp1u,Xbp1 s,Atf4,Chop,Ctnnb1,Ccnd1,Pdx-1,Pink and Prkn was up-regulated,transcription of Bcl2 and a2-AR was down-regulated,while at 5 days of experimental treatment,transcription of Bip,Atf4,Chop,Gadd34 and PGC1α was up-regulated,transcription of Ire1α,Nrf2,Ucp2 and a2-ARwas down-regulated.Conclusion: To sum up,chronic epinephrine reprograms the secretion function of pancreatic β-cell and affects the fate of pancreatic β-cell by inducing endoplasmic reticulum stress,which triggers oxidative stress.In addition,α2-AR occurs desensitization to participate in the influence of MIN6 secretion and fate.