Ingo Uphues

LKB1 and AMPK differentially regulate pancreatic β-cell identity

Fully differentiated pancreatic β cells are essential for normal glucose homeostasis in mammals. Dedifferentiation of these cells has been suggested to occur in type 2 diabetes, impairing insulin production. Since chronic fuel excess (“glucotoxicity”) is implicated in this process, we sought here to identify the potential roles in β‐cell identity of the tumor suppressor liver kinase B1 (LKB1/STK11) and the downstream fuel‐sensitive kinase, AMP‐activated protein kinase (AMPK). Highly β‐cell‐restricted deletion of each kinase in mice, using an Ins1‐controlled Cre, was therefore followed by physiological, morphometric, and massive parallel sequencing analysis. Loss of LKB1 strikingly (2.0‐12‐fold, E<0.01) increased the expression of subsets of hepatic (Alb, Iyd, Elovl2) and neuronal (Nptx2, Dlgap2, Cartpt, Pdyn) genes, enhancing glutamate signaling. These changes were partially recapitulated by the loss of AMPK, which also up‐regulated β‐cell “disallowed” genes (Slc16a1, Ldha, Mgst1, Pdgfra) 1.8‐ to 3.4‐fold (E<0.01). Correspondingly, targeted promoters were enriched for neuronal (Zfp206; P= 1.3×10‐33) and hypoxia‐regulated (HIF1; P= 2.5×10‐16) transcription factors. In summary, LKB1 and AMPK, through only partly overlapping mechanisms, maintain β‐cell identity by suppressing alternate pathways leading to neuronal, hepatic, and other characteristics. Selective targeting of these enzymes may provide a new approach to maintaining β‐cell function in some forms of diabetes.—Kone, M., Pullen, T. J., Sun, G., Ibberson, M., Martinez‐Sanchez, A., Sayers, S., Nguyen‐Tu, M.‐S., Kantor, C., Swisa, A., Dor, Y., Gorman, T., Ferrer, J., Thorens, B., Reimann, F., Gribble, F., McGinty, J. A., Chen, L., French, P. M., Birzele, F., Hildebrandt, T., Uphues, I., Rutter, G. A., LKB1 and AMPK differentially regulate pancreatic β‐cell identity. FASEB J. 28, 4972–4985 (2014). www.fasebj.org

Hes3 is expressed in the adult pancreatic islet and regulates gene expression, cell growth, and insulin release

Background: The transcription factor Hes3 regulates the growth of neural and brain cancer stem cells. Results: Hes3 regulates growth, gene expression, evoked insulin release in cultured insulinoma cells, and sensitivity to streptozotocin in vivo. Conclusion: Hes3 is a novel regulator of cellular functions of importance in diabetes. Significance: Introducing Hes3 and its regulators in diabetes research may provide new opportunities for the design of novel therapeutics. The transcription factor Hes3 is a component of a signaling pathway that supports the growth of neural stem cells with profound consequences in neurodegenerative disease models. Here we explored whether Hes3 also regulates pancreatic islet cells. We showed that Hes3 is expressed in human and rodent pancreatic islets. In mouse islets it co-localizes with alpha and beta cell markers. We employed the mouse insulinoma cell line MIN6 to perform in vitro characterization and functional studies in conditions known to modulate Hes3 based upon our previous work using neural stem cell cultures. In these conditions, cells showed elevated Hes3 expression and nuclear localization, grew efficiently, and showed higher evoked insulin release responses, compared with serum-containing conditions. They also exhibited higher expression of the transcription factor Pdx1 and insulin. Furthermore, they were responsive to pharmacological treatments with the GLP-1 analog Exendin-4, which increased nuclear Hes3 localization. We employed a transfection approach to address specific functions of Hes3. Hes3 RNA interference opposed cell growth and affected gene expression as revealed by DNA microarrays. Western blotting and PCR approaches specifically showed that Hes3 RNA interference opposes the expression of Pdx1 and insulin. Hes3 overexpression (using a Hes3-GFP fusion construct) confirmed a role of Hes3 in regulating Pdx1 expression. Hes3 RNA interference reduced evoked insulin release. Mice lacking Hes3 exhibited increased islet damage by streptozotocin. These data suggest roles of Hes3 in pancreatic islet function.

Alterations in β-cell calcium dynamics and efficacy outweigh islet mass adaptation in compensation of insulin resistance and prediabetes onset

Emerging insulin resistance is normally compensated by increased insulin production of pancreatic β-cells, thereby maintaining normoglycemia. However, it is unclear whether this is achieved by adaptation of β-cell function, mass, or both. Most importantly, it is still unknown which of these adaptive mechanisms fail when type 2 diabetes develops. We performed longitudinal in vivo imaging of β-cell calcium dynamics and islet mass of transplanted islets of Langerhans throughout diet-induced progression from normal glucose homeostasis, through compensation of insulin resistance, to prediabetes. The results show that compensation of insulin resistance is predominated by alterations of β-cell function, while islet mass only gradually expands. Hereby, functional adaptation is mediated by increased calcium efficacy, which involves Epac signaling. Prior to prediabetes, β-cell function displays decreased stimulated calcium dynamics, whereas islet mass continues to increase through prediabetes onset. Thus, our data reveal a predominant role of islet function with distinct contributions of triggering and amplifying pathway in the in vivo processes preceding diabetes onset. These findings support protection and recovery of β-cell function as primary goals for prevention and treatment of diabetes and provide insight into potential therapeutic targets.

Molecular phenotyping of multiple mouse strains under metabolic challenge uncovers a role for Elovl2 in glucose-induced insulin secretion.

Objective In type 2 diabetes (T2D), pancreatic β cells become progressively dysfunctional, leading to a decline in insulin secretion over time. In this study, we aimed to identify key genes involved in pancreatic beta cell dysfunction by analyzing multiple mouse strains in parallel under metabolic stress. Methods Male mice from six commonly used non-diabetic mouse strains were fed a high fat or regular chow diet for three months. Pancreatic islets were extracted and phenotypic measurements were recorded at 2 days, 10 days, 30 days, and 90 days to assess diabetes progression. RNA-Seq was performed on islet tissue at each time-point and integrated with the phenotypic data in a network-based analysis. Results A module of co-expressed genes was selected for further investigation as it showed the strongest correlation to insulin secretion and oral glucose tolerance phenotypes. One of the predicted network hub genes was Elovl2, encoding Elongase of very long chain fatty acids 2. Elovl2 silencing decreased glucose-stimulated insulin secretion in mouse and human β cell lines. Conclusion Our results suggest a role for Elovl2 in ensuring normal insulin secretory responses to glucose. Moreover, the large comprehensive dataset and integrative network-based approach provides a new resource to dissect the molecular etiology of β cell failure under metabolic stress.