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Diabetologia. 2017 Jun;60(6):1057-1065. doi: 10.1007/s00125-017-4237-z. Epub 2017 Mar 9.

MicroRNA 21 targets BCL2 mRNA to increase apoptosis in rat and human beta cells.

Sims EK1,2,3, Lakhter AJ4,5,6, Anderson-Baucum E4,7, Kono T4,6,7, Tong X4,7, Evans-Molina C4,6,7,8,9.

Author information

1
Center for Diabetes and Metabolic Diseases, Indiana University School of Medicine, 635 Barnhill Drive, MS2031, Indianapolis, IN, 46202, USA. eksims@iu.edu.
2
Department of Pediatrics, Indiana University School of Medicine, Indianapolis, IN, USA. eksims@iu.edu.
3
Herman B. Wells Center for Pediatric Research, Indiana University School of Medicine, Indianapolis, IN, USA. eksims@iu.edu.
4
Center for Diabetes and Metabolic Diseases, Indiana University School of Medicine, 635 Barnhill Drive, MS2031, Indianapolis, IN, 46202, USA.
5
Department of Pediatrics, Indiana University School of Medicine, Indianapolis, IN, USA.
6
Herman B. Wells Center for Pediatric Research, Indiana University School of Medicine, Indianapolis, IN, USA.
7
Department of Medicine, Indiana University School of Medicine, Indianapolis, IN, USA.
8
Department of Biochemistry and Molecular Biology, Indiana University School of Medicine, Indianapolis, IN, USA.
9
Richard L. Roudebush VA Medical Center, Indianapolis, IN, USA.

Abstract

AIMS/HYPOTHESIS:

The role of beta cell microRNA (miR)-21 in the pathophysiology of type 1 diabetes has been controversial. Here, we sought to define the context of beta cell miR-21 upregulation in type 1 diabetes and the phenotype of beta cell miR-21 overexpression through target identification.

METHODS:

Islets were isolated from NOD mice and mice treated with multiple low doses of streptozotocin, as a mouse model of diabetes. INS-1 832/13 beta cells and human islets were treated with IL-1β, IFN-γ and TNF-α to mimic the milieu of early type 1 diabetes. Cells and islets were transfected with miR-21 mimics or inhibitors. Luciferase assays and polyribosomal profiling (PRP) were performed to define miR-21-target interactions.

RESULTS:

Beta cell miR-21 was increased in in vivo models of type 1 diabetes and cytokine-treated cells/islets. miR-21 overexpression decreased cell count and viability, and increased cleaved caspase 3 levels, suggesting increased cell death. In silico prediction tools identified the antiapoptotic mRNA BCL2 as a conserved miR-21 target. Consistent with this, miR-21 overexpression decreased BCL2 transcript and B cell lymphoma 2 (BCL2) protein production, while miR-21 inhibition increased BCL2 protein levels and reduced cleaved caspase 3 levels after cytokine treatment. miR-21-mediated cell death was abrogated in 828/33 cells, which constitutively overexpress Bcl2. Luciferase assays suggested a direct interaction between miR-21 and the BCL2 3' untranslated region. With miR-21 overexpression, PRP revealed a shift of the Bcl2 message towards monosome-associated fractions, indicating inhibition of Bcl2 translation. Finally, overexpression in dispersed human islets confirmed a reduction in BCL2 transcripts and increased cleaved caspase 3 production.

CONCLUSIONS/INTERPRETATION:

In contrast to the pro-survival role reported in other systems, our results demonstrate that miR-21 increases beta cell death via BCL2 transcript degradation and inhibition of BCL2 translation.

KEYWORDS:

Animal – mouse; Basic science; Beta cell signal transduction; Cell lines; Islet degeneration and damage; Islets

PMID:
28280903
PMCID:
PMC5425307
DOI:
10.1007/s00125-017-4237-z
[Indexed for MEDLINE]
Free PMC Article

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