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Mol Cell Endocrinol. 2015 Nov 15;416:57-69. doi: 10.1016/j.mce.2015.08.021. Epub 2015 Aug 20.

Upregulation of miR-497 induces hepatic insulin resistance in E3 rats with HFD-MetS by targeting insulin receptor.

Author information

1
Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Xi'an Jiaotong University Health Science Center, Xi'an, Shaanxi 710061, PR China; Key Laboratory of Environment and Genes Related to Diseases (Xi'an Jiaotong University), Ministry of Education of China, Xi'an, Shaanxi 710061, PR China; Department of Endocrinology, Jinshan Hospital of Fudan University, Shanghai 201508, PR China.
2
Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Xi'an Jiaotong University Health Science Center, Xi'an, Shaanxi 710061, PR China; Key Laboratory of Environment and Genes Related to Diseases (Xi'an Jiaotong University), Ministry of Education of China, Xi'an, Shaanxi 710061, PR China.
3
School of Life Sciences, Northwest University, Xi'an, Shaanxi 710061, PR China.
4
Department of Pathology and Laboratory Medicine, Metabolic Diseases Institute, University of Cincinnati, Cincinnati, OH 45237, USA.
5
Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Xi'an Jiaotong University Health Science Center, Xi'an, Shaanxi 710061, PR China; Key Laboratory of Environment and Genes Related to Diseases (Xi'an Jiaotong University), Ministry of Education of China, Xi'an, Shaanxi 710061, PR China. Electronic address: dongmin_li2015@163.com.

Abstract

OBJECTIVE:

The study aims to find regulatory microRNA(s) responsible for down-regulated insulin receptor (InsR) in the liver of HFD-MetS E3 rats with insulin resistance.

METHODS:

Firstly, hepatic insulin resistance in HFD-MetS E3 rats was evaluated by RT-qPCR, western blotting, immunohistochemistry and PAS staining. Secondly, the candidate miRNAs targeting rat InsR were predicted through online softwares and detected in the liver of HFD-MetS E3 rats with insulin resistance. Then, the expression of InsR, phosphorylated IRS-1 (pIRS-1) at Tyr632, phosphorylated AKTs (pAKTs) at Ser473 and Thr308, phosphorylated GSK-3β (p GSK-3β) at Ser9, phosphorylated GS (pGS) at Ser641 and the glycogen content were detected in CBRH-7919 cells treated with 100 nM insulin for different time periods by western blotting or PAS staining respectively, after transient transfection with miR-497 mimics or inhibitors for 24 h. Lastly, the relation between miR-497 and InsR was further determined using dual luciferase reporter assay.

RESULTS:

Elevated miR-497 was negatively related with down-regulated InsR in the liver of HFD-MetS E3 rats with insulin resistance. Comparing with the mNC group, glycogen content and the expression of InsR, pIRS-1 (Tyr632), pAKTs (Ser473 and Thr308) and pGSK-3β (Ser9) decreased significantly in CBRH-7919 cells, while pGS (Ser641) increased significantly, after transient transfection with miR-497 mimics for 24 h and treatment with 100 nM insulin for corresponding time periods, counter to those results in CBRH-7919 cells after similar procedures with miR-497 inhibitors and insulin. In addition, dual luciferase reporter assay further confirmed that miR-497 can bind to the 3'UTR of rat InsR.

CONCLUSION:

Insulin receptor is the target gene of miR-497, and elevated miR-497 might induce hepatic insulin resistance in HFD-MetS E3 Rats through inhibiting the expression of insulin receptor and confining the activation of IRS-1/PI3K/Akt/GSK-3β/GS pathway to insulin.

KEYWORDS:

Hepatic insulin resistance; High-fat-diet induced metabolic syndrome (HFD-MetS); Insulin receptor; miR-497

PMID:
26300412
DOI:
10.1016/j.mce.2015.08.021
[Indexed for MEDLINE]

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