Abstract
There are three types of cell death; apoptosis, necrosis, and autophagy. The possibility that activation of the macroautophagy (autophagy) pathway may increase beta cell death is addressed in this study. Increased autophagy was present in pancreatic islets from Pdx1(+/-) mice with reduced insulin secretion and beta cell mass. Pdx1 expression was reduced in mouse insulinoma 6 (MIN6) cells by delivering small hairpin RNAs using a lentiviral vector. The MIN6 cells died after 7 days of Pdx1 deficiency, and autophagy was evident prior to the onset of cell death. Inhibition of autophagy prolonged cell survival and delayed cell death. Nutrient deprivation increased autophagy in MIN6 cells and mouse and human islets after starvation. Autophagy inhibition partly prevented amino acid starvation-induced MIN6 cell death. The in vivo effects of reduced autophagy were studied by crossing Pdx1(+/-) mice to Becn1(+/-) mice. After 1 week on a high fat diet, 4-week-old Pdx1(+/-) Becn1(+/-) mice showed normal glucose tolerance, preserved beta cell function, and increased beta cell mass compared with Pdx1(+/-) mice. This protective effect of reduced autophagy had worn off after 7 weeks on a high fat diet. Increased autophagy contributes to pancreatic beta cell death in Pdx1 deficiency and following nutrient deprivation. The role of autophagy should be considered in studies of pancreatic beta cell death and diabetes and as a target for novel therapeutic intervention.
Highlights
There are three types of cell death; apoptosis, necrosis, and autophagy [2]
We treated Pdx1 KD mouse insulinoma 6 (MIN6) cells CHO-treated Pdx1 KD MIN6 cells. These results indicate with BafA1, which inhibits the final step of lysosomal digestion that 3-MA delays Pdx1 KD-induced MIN6 cell death, but in autophagy to determine whether reduced Pdx1 expression in 3-MA-treated Pdx1 KD MIN6 cells die by caspase-3
The present study addressed the role of autophagy in beta cell survival and death
Summary
Cell Culture—Mouse insulinoma MIN63 cells were cultured in Dulbecco’s modified Eagle’s medium supplemented with 15% FBS, antibiotics Autophagy Regulates Pancreatic Beta Cell Death nologies), actin (A-2066; Sigma), and Atg Death Assay—For the last hour of incubation, 10 g/ml propidium iodide (PI) and 20 g/ml DAPI were added directly to the media. PBS and fixed with 3.7% formaldehyde for 15 min at 4 °C. Each condition reported represents Ͼ600 cells counted by randomized field selection. The percentage of cell death was calculated as the number of PI-. Stained nuclei over the total number of nuclei stained by DAPI
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