Role Of ER Stress-autophagic Pathway In Lipotoxic Injury In Pancreatic β-cells And Its Underlying Mechanisms

Lipotoxicity plays an important role in obesity-related type2diabetes.Lipotoxicity causes not only insulin resistance in peripheral tissues but also injuriesin pancreatic β-cells. The pathophysiology of β-cells during lipotoxic procedureremains largely unknown. Previous studies indicated that autophagy is necessary inmaintaining the mass, structure and function of β-cells and that endoplasmicreticulum (ER) stress is one of the important mediators in β-cell injuries.Investigation on the relations between lipotoxicity and the above two, autophagy andER stress, will help us understand lipotoxicity better. Mouse insulinoma MIN6cellline was used, and was treated using pharmacological methods. The aim of this studyis to verify whether ER stress could induce autophagy in β-cells, and to explore theroles and related mechanisms of this possible pathway in lipotoxicity.1ER stress-autophagic pathway in pancreatic β-cells and its regulatorymechanismsThe cytotoxic effect of ER stress inducer thapsigargin (TG) on MIN6cells wasdose-dependent. After36h treatment with0.1μM or1μM TG, cell viability wasinhibited, and cell morphology was unnormal. The expression of LC3II, a hallmarkof autophagy activation, increased after treatment with0.1μM TG for36h. And anincrease of autophagy was also observed using transmission electron microscope.The insulin-induced p-Akt (Ser473) gradually decreased by administration of TG,while the expression of insulin receptor β (IRβ) was not affected. LY294002(50μM),PI3K inhibitor, was found to increase TG-induced LC3II expression. JNK phosphorylation was gradually activated after TG treatment for different times. Theexpression of LC3II induced by TG was inhibited by20μM JNK inhibitorSP600125. Compared with TG alone, pretreatment with5mM3-MA for1h causeda further decrease in cell viability (P<0.01), in the proportion of cell apoptosis, and inglucose-stimulated insulin secretion (GSIS)(P<0.01).2Palmitate-induced autophagy in pancreatic β-cells and its relationshipwith ER stressPalmitate (PA) inhibited cell viability of MIN6cells in a dose-dependent andtime-dependent manner. After0.5mM PA treatment for24h (P<0.01) or36h(P<0.01), both basic insulin secretion (BIS) and GSIS decreased. Treatment with PAfor36h significantly enhanced LC3II level. The expression of BiP, CHOP, and JNKphosphorylation gradually increased after PA treatment. Both TUDCA (500μM) andSP600125(20μM) exhibited an inhibitory effect on the PA-induced LC3II increase.Cell apoptosis caused by PA further increased with3-MA, but was inhibited byTUDCA or autophagy inducer rapamycin (50nM). Oil Red-O staining showed thatPA caused intracellular lipid accumulation. PA down-regulated the phosphorylationof Akt (Ser473) gradually, but did not reduce the levels of IRβ. A further increase ofLC3II expression induced by PA was observed in cells treated with50μMLY294002. Akt phosphorylation reduction was also inhibited by TUDCA orSP600125. TUDCA or rapamycin administration improved the inhibition of cellviability (both P<0.05) and the reduction of GSIS caused by PA (P<0.01, P<0.05,respectively). The expression of BiP and CHOP induced by PA was enhanced after3-MA pretreatment.In conclusion,1) ER stress-autophagic pathway in pancreatic β-cells is verified;2) ER stress, JNK activation and the suppression of insulin signaling contributed toPA-induced autophagy;3) The reduction of autophagy aggravated ER stress andrelated cell injuries in β-cells;4) Inhibition of ER stress and up-regulation ofautophagy are two possible solutions for protecting β-cells against lipotoxic injuries.