Modulation of Autophagy Influences the Function and Survival of Human Pancreatic Beta Cells Under Endoplasmic Reticulum Stress Conditions and in Type 2 Diabetes

Abstract

Autophagy is the major mechanism involved in degradation and recycling of intracellular components, and its alterations have been proposed to cause beta cell dysfunction. In this study, we explored the effects of autophagy modulation in human islets under conditions associated to endoplasmic reticulum (ER) stress. Human pancreatic islets were isolated by enzymatic digestion and density gradient purification from pancreatic samples of non-diabetic (ND; n = 17; age 65 ± 21 years; gender: 5 M/12 F; BMI 23.4 ± 3.3 kg/m 2 ) and T2D ( n = 9; age 76 ± 6 years; 4 M/5 F; gender: BMI 25.4 ± 3.7 kg/m 2 ) organ donors. Nine ND organ donors were treated for hypertension and 1 for both hypertension and hypercholesterolemia. T2D organ donors were treated with metformin ( 1 ), oral hypoglycemic agents ( 2 ), diet + oral hypoglycemic agents ( 3 ), insulin ( 3 ) or insulin plus metformin ( 3 ) as for antidiabetic therapy and, of these, 3 were treated also for hypertension and 6 for both hypertension and hypercholesterolemia. Two days after isolation, they were cultured for 1–5 days with 10 ng/ml rapamycin (autophagy inducer), 5 mM 3-methyladenine or 1.0 nM concanamycin-A (autophagy blockers), either in the presence or not of metabolic (0.5 mM palmitate) or chemical (0.1 ng/ml brefeldin A) ER stressors. In ND islets palmitate exposure induced a 4 to 5-fold increase of beta cell apoptosis, which was significantly prevented by rapamycin and exacerbated by 3-MA. Similar results were observed with brefeldin treatment. Glucose-stimulated insulin secretion from ND islets was reduced by palmitate (−40 to 50%) and brefeldin (−60 to 70%), and rapamycin counteracted palmitate, but not brefeldin, cytotoxic actions. Both palmitate and brefeldin induced PERK, CHOP and BiP gene expression, which was partially, but significantly prevented by rapamycin. With T2D islets, rapamycin alone reduced the amount of p62, an autophagy receptor that accumulates in cells when macroautophagy is inhibited. Compared to untreated T2D cells, rapamycin-exposed diabetic islets showed improved insulin secretion, reduced proportion of beta cells showing signs of apoptosis and better preserved insulin granules, mitochondria and ER ultrastructure; this was associated with significant reduction of PERK, CHOP and BiP gene expression. This study emphasizes the importance of autophagy modulation in human beta cell function and survival, particularly in situations of ER stress. Tuning autophagy could be a tool for beta cell protection.

Pancreatic Beta Cell Autophagy is Impaired in Type 1 Diabetes

“…Autophagy has been identified as an important player in beta cell survival and growth, contributing to cellular homeostasis and stress response [30]. While the role of autophagy in the context of type 2 diabetes has been studied extensively [31,32], its role in type 1 diabetes is relatively unexplored.…”

The Phosphatase PHLPP2 Plays a Key Role in the Regulation of Pancreatic Beta-Cell Survival

“…mTor senses nutrient availability and regulates cell homeostasis, and it has been suggested that its loss impairs β-cells homeostasis as well as insulin sensitivity in peripheral tissues [27]. Among the intracellular processes controlled by mTor, autophagy as gained attention as a possible player in the survival of pancreatic β-cells with conflicting data showing, in different experimental models, pro-or antiapoptotic effects of an increased activation of autophagic pathways [28][29][30]. In our model, the recovered prosurvival profile was paralleled by decreased expression of the autophagic marker LC3-II, supporting the hypothesis of a negative impact of a disproportionated activation of autophagic pathways on cell survival.…”

Assessment of the association of serum progranulin with autophagy in diabetic patients

“…Obesity and visceral fat are probably the main risk factors for T2D [9] and are involved in its pathophysiology as well as inflammation and insulin resistance [21]. Moreover, a study evaluating T2DM patients reported that obese subjects present higher levels of PGRN [13]. Taking in consideration the fact that BMI is a simple index of weight-for-height that is commonly used to classify overweight and obesity in adults, this study agreed with our finding revealing a significant difference of BMI between diabetic and control groups with significant positive correlation with PGRN.…”

“…Autophagy is the major mechanism involved in the degradation and recycling of intracellular components, and its alterations have been proposed to cause beta cell dysfunction [13]. However, autophagy may be enhanced under a variety of atrophic conditions, as shown by the expression of several proteins involved in lysosomal degradation, including LC3 [14].…”