MicroRNA-29a is up-regulated in beta-cells by glucose and decreases glucose-stimulated insulin secretion

doi:10.1016/j.bbrc.2012.08.082

Biochemical and Biophysical Research Communications

Volume 426, Issue 2, 21 September 2012, Pages 266-272

https://doi.org/10.1016/j.bbrc.2012.08.082 Get rights and content

Abstract

Chronically elevated levels of glucose impair pancreatic beta-cell function while inducing beta-cell proliferation. MicroRNA-29a (miR-29a) levels are increased in several tissues in diabetic animals and mediate decreased insulin-stimulated glucose-transport of adipocytes. The aim was to investigate the impact of glucose on miR-29a levels in INS-1E beta-cells and in human islets of Langerhans and furthermore to evaluate the impact of miR-29a on beta-cell function and proliferation. Increased glucose levels up-regulated miR-29a in beta-cells and human and rat islets of Langerhans. Glucose-stimulated insulin-secretion (GSIS) of INS-1E beta-cells was decreased by forced expression of miR-29a, while depletion of endogenous miR-29a improved GSIS. Over-expression of miR-29a increased INS-1E proliferation. Thus, miR-29a up-regulation is involved in glucose-induced proliferation of beta-cells. Furthermore, as depletion of miR-29a improves beta-cell function, miR-29a is a mediator of glucose-induced beta-cell dysfunction. Glucose-induced up-regulation of miR-29a in beta-cells could be implicated in progression from impaired glucose tolerance to type 2 diabetes.

Highlights

► MicroRNA-29a (miR-29a) levels are increased by glucose in human and rat islets and INS-1E cells. ► miR-29a increases proliferation of INS-1E beta-cells. ► Forced expression of miR-29a decreases glucose-stimulated insulin secretion (GSIS). ► Depletion of beta-cell miR-29a improves GSIS. ► miR-29a may be a mediator of glucose toxicity in beta-cells.

Introduction

Type 2 diabetes mellitus is a complex metabolic disorder involving two core defects: insulin resistance and beta-cell dysfunction, both of which are present in pre-diabetic states and act in concert to progressively exacerbate glucose intolerance. Elevated plasma glucose levels (post-prandially or chronically) lead to decreased glucose-stimulated insulin secretion (GSIS) and beta-cell dysfunction [1]. The mechanisms underlying glucose-induced beta-cell dysfunction, or glucose toxicity, are incompletely understood, but involve beta-cell exhaustion from continued insulin release as well as mitochondrial dysfunction [2], [3], [4], however, an increased glucose level is also a powerful beta-cell mitogen [5].

MicroRNAs (miRNAs) are regulators of gene expression at the post-transcriptional level and function by partially binding to the 3′untranslated region (UTR) of their target gene transcripts either mediating transcript degradation or translational inhibition [6], [7], [8]. Several miRNAs have been implicated in beta-cell function: miR-9, miR-30d, miR-124, miR-133a and miR-375 are involved in glucose-dependent regulation of insulin transcription and/or insulin release in beta-cells by targeting beta-cell transcription factors and/or transcripts involved in insulin exocytosis [9], [10], [11], [12], [13], [14], [15]. Furthermore, fatty acid induced increase in miR-34a levels may cause beta-cell dysfunction [16].

miR-29a is up-regulated by glucose in skeletal muscle, liver and white adipose tissue, where it leads to insulin resistance [17], [18], [19]. We hypothesized that miR-29a could be similarly regulated by glucose in pancreatic beta-cells and mediate glucose-induced dysfunction. Thus, the aim of this study was to determine the glucose-dependent regulation of miR-29a in beta-cells, as well as effects on GSIS.

Our results indicate that glucose-mediated up-regulation of miR-29a in beta-cells mediates beta-cell dysfunction and increased beta-cell proliferation, while inhibition of miR-29a improves GSIS. Thus, the up-regulation of miR-29a by glucose could be a link between glucose-induced proliferation and beta-cell dysfunction.

Section snippets

Cell culture and nucleofection

INS-1E cells (gift from Claes Wollheim, Geneva, Switzerland) were cultured in RPMI as described previously [20]. Cells (4 × 10⁶) were nucleofected using a Nucleofector (Amaxa, Lonza, Copenhagen, Denmark) with miR-29a LNA knock-down or scrambled LNA oligonucleotide (Exiqon, Vedbaek, Denmark), or with miRIDIAN miR-29a mimic or the miRIDIAN negative control #2 (Dharmacon, ThermoFisher Scientific, Slangerup, Denmark). Nucleofected cells were seeded in poly-lysine treated culture plates, and 24 h

Regulation of miR-29a levels by glucose in human and rat islets of Langerhans and INS-1E cells

The expression levels of miR-29a in human islets of Langerhans and INS-1E cells treated with low (5 mM) or high glucose (11 mM and 20 mM respectively) for 48 h were assessed by real-time RT-Q-PCR or northern blotting. The expression levels of mature miR-29a in INS-1E cells (Fig. 1A–C), rat and human pancreatic islets (Fig. 1D–E) increased 1.7 ± 0.1 (P < 0.001), 1.5 ± 0.3 (P < 0.05) and 1.23 ± 0.06-fold (P < 0.05), respectively, in response to increased glucose levels, and in INS-1E cells miR-29a up-regulation

Discussion

Prolonged exposure of beta-cells to high levels of glucose decreases GSIS [2], [30]. The current results show that over-expression of miR-29a, up-regulated by glucose in human islets of Langerhans and in INS-1E beta-cells, decreased GSIS markedly in INS-1E cells that normally exhibit a robust insulin secretion response. Inhibition of miR-29a increases GSIS, suggesting that endogenous miR-29a exert a tonic inhibition on GSIS. Even though inhibition of miR-29a increased GSIS in cells cultured in

Acknowledgments

We are very grateful for the skilled technical assistance of Kirsten Olesen, Roskilde University and Vibeke Nielsen, Novo Nordisk A/S. LTD and JHN have been and TRC and ML are employed by and own stocks in Novo Nordisk A/S, a pharmaceutical company selling diabetes products. These studies were supported by the Danish Research Council for Technology and Production and the Danish microRNA Consortium.

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MicroRNA-29a is up-regulated in beta-cells by glucose and decreases glucose-stimulated insulin secretion

Abstract

Highlights

Introduction

Section snippets

Cell culture and nucleofection

Regulation of miR-29a levels by glucose in human and rat islets of Langerhans and INS-1E cells

Discussion

Acknowledgments

J. Biol. Chem.

J. Biol. Chem.

Mol. Cell Endocrinol.

Biochem. Biophys. Res. Commun.

J. Biol. Chem.

Beta-cell failure as a complication of diabetes

Rev. Endocr. Metab. Disord.

Mitochondrial dysfunction and type 2 diabetes

Science

Functional and morphological alterations of mitochondria in pancreatic beta cells from type 2 diabetic patients

Diabetolog

Role of mitochondria in beta-cell function and dysfunction

Adv. Exp. Med. Biol.

Glucose infusion in mice: a new model to induce beta-cell replication

Diabetes

Micromanagers of gene expression: the potentially widespread influence of metazoan microRNAs

Nat. Rev. Genet.

Illuminating the silence: understanding the structure and function of small RNAs

Nat. Rev. Mol. Cell Biol.

MicroRNAs and cell differentiation in mammalian development birth defects

Res. C. Embryo. Today

Regulation of the expression of components of the exocytotic machinery of insulin-secreting cells by microRNAs

Biol. Chem.

A pancreatic islet-specific microRNA regulates insulin secretion

Nature

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Proc. Natl. Acad. Sci. USA

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PLoS One