Glucose regulation of pancreatic islet gene expression : microarray analysis and study on the role of Hypoxia-Inducible factors 1 and 2
(2011)
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- Authors
- Supervisors
- Jonas, Jean-Christophe
- Abstract
- (en) Pancreatic β-cells secrete insulin in response to a rise in plasma concentration of glucose and other nutrients to maintain glucose homeostasis of the organism. The ability of β-cells to respond to an acute glucose challenge depends on the adequate level of expression of several genes, including preproinsulin, Glut2, glucokinase etc. This level of expression is modulated by meals: it is reduced during fasting and restored rapidly upon feeding. Interestingly, both chronic hypoglycaemia and chronic hyperglycemia induce the loss of β-cell differentiation, alter the stimulus-secretion coupling, and increase the rate of β-cell apoptosis, along with increased expression of genes under-expressed in normal β-cells. Similarly, in vitro, rat β-cell function and survival are optimally preserved by culture in the presence of 10 mM glucose (G10) and markedly impaired by culture in either lower (G2-G5) or higher (G30) glucose concentrations. It seems therefore that glucose stimulation exerts beneficial effects on the β-cell phenotype between G2 and G10. In contrast, supraphysiological glucose concentrations (between G10 and G30) are deleterious for β-cell function and survival, a concept termed as “glucotoxicity”. The latter plays an important role in the alteration of the functional β-cell mass in type 2 diabetes (T2D), and contributes to the progressive worsening of glucose intolerance in these patients. However, the precise molecular mechanisms behind such phenotypical plasticity are poorly understood. The purpose of my thesis was to gain new and more complete insights about the molecular mechanisms underlying the plasticity of the β-cell phenotype, with a special focus on the role of high glucose-induced hypoxia and Hypoxia Inducible Factors (HIF) 1 and 2 in the glucotoxic alterations of β-cell gene expression. In the first part of these studies, I tested the effects of increasing glucose concentrations (G2, G5, G10 and G30) during 18h culture on the transcriptome of rat islets. Cluster analysis of the glucose-affected genes revealed 10 mRNA profiles with unidirectional up- or downregulation that differ in glucose threshold and the maximal effective concentration, and 8 complex V-shaped or inverted V-shaped profiles with a minimum or maximum in G5 or G10. Further functional categorization of these clusters allowed us to identify 4 biological pathways enriched in genes co-expressed in response to changes in glucose levels, namely: glycolysis, cholesterol biosynthesis, the Integrated Stress Response (ISR), and the Unfolded Protein Response (UPR). Besides, I bring new information about the β-cell antioxidant machinery and showed its complex regulation by glucose. These pathways may contribute to the complex modulation of the β-cell phenotype by glucose, and some of them may participate to the induction of β-cell dysfunction and apoptosis after prolonged culture in the presence of extreme glucose concentrations. The observation that glucose increased the mRNA levels of most glycolytic enzymes and several HIF-target genes predominantly between G10 and G30 was the starting point of the second part of this work. Thus, I tested whether these changes in gene expression resulted from high glucose-induced β-cell hypoxia and subsequent activation of HIF1 and/or HIF2. Our results demonstrate that glucose, in a concentration-dependent manner, increases the formation of pimonidazole-protein adducts, an indicator of hypoxia, and enhances the nuclear accumulation of HIF1 and HIF2, in parallel to the stimulation of the expression of HIF target genes in INS-1E cells and rat islets. Interestingly, the mechanism of this activation involves the acceleration of mitochondrial metabolism and Ca2+ influx. In addition, I found that HIF1 and 2 isoforms are not redundant and that both contribute to the glucose induction of glycolytic genes while Adrenomedullin (Adm) expression depends exclusively on HIF2. We finally present evidence of HIF activation in the islets of diabetic Leprdb/db mice. Our results suggest that high glucose-induced β-cell hypoxia may be an important mechanism that contributes to the deterioration of the β-cell phenotype under prolonged hyperglycemia. These important findings will help understanding the molecular mechanisms underscoring the plasticity of the β-cell phenotype and may help in the development of more efficient therapeutic strategies for T2D.
- Affiliations
Citations
Bensellam, M. (2011). Glucose regulation of pancreatic islet gene expression : microarray analysis and study on the role of Hypoxia-Inducible factors 1 and 2. https://hdl.handle.net/2078.5/152472