MicroRNA-561 promotes acetaminophen-induced hepatotoxicity in HepG2 cells and primary human hepatocytes through downregulation of the nuclear receptor corepressor dosage-sensitive sex-reversal adrenal hypoplasia congenital critical region on the X chromosome, gene 1 (DAX-1)

Minghua Li; Yinxue Yang; Zhi-Xu He; Zhi-Wei Zhou; Tianxin Yang; Peixuan Guo; Xueji Zhang; Shu-Feng Zhou

doi:10.1124/dmd.113.052670

MicroRNA-561 promotes acetaminophen-induced hepatotoxicity in HepG2 cells and primary human hepatocytes through downregulation of the nuclear receptor corepressor dosage-sensitive sex-reversal adrenal hypoplasia congenital critical region on the X chromosome, gene 1 (DAX-1)

Drug Metab Dispos. 2014 Jan;42(1):44-61. doi: 10.1124/dmd.113.052670. Epub 2013 Oct 8.

Authors

Minghua Li¹, Yinxue Yang, Zhi-Xu He, Zhi-Wei Zhou, Tianxin Yang, Peixuan Guo, Xueji Zhang, Shu-Feng Zhou

Affiliation

¹ Department of Pharmaceutical Sciences, College of Pharmacy, University of South Florida, Tampa, Florida (M.L., Z.W.Z., S.F.Z.); Department of Hepatobiliary Surgery, General Hospital of Ningxia Medical University, Yinchuan City, Ningxia Hui Autonomous Region, China (Y.Y.); Guizhou Provincial Key Laboratory for Regenerative Medicine, Stem Cell and Tissue Engineering Research Center, Guiyang Medical University, Guiyang, Guizhou, China (Z.X.H.); Department of Internal Medicine, University of Utah and Salt Lake Veterans Affairs Medical Center, Salt Lake City, Utah (T.Y.); Nanobiotechnology Center and Markey Cancer Center, College of Pharmacy, University of Kentucky, Lexington, Kentucky (P.G.); Research Center for Bioengineering and Sensing Technology, University of Science and Technology Beijing, Beijing, China (X.Z.).

PMID: 24104199
DOI: 10.1124/dmd.113.052670

Abstract

One of the major mechanisms involved in acetaminophen (APAP)-induced hepatotoxicity is hepatocyte nuclear factor 4α (HNF4α)-mediated activation of pregnane X receptor (PXR) and constitutive androstane receptor (CAR). In the present study, we investigated the role of miR-561 and its target gene DAX-1 encoding a corepressor of HNF4α in the process of APAP-induced hepatotoxicity. We used both human hepatocellular liver carcinoma cell line (HepG2) cells and primary human hepatocytes in this study and monitored the levels of reactive oxygen species, lactate dehydrogenase, and glutathione. Our bioinformatics study suggests an association between miR-561 and DAX-1, but not HNF4α. Treatment of HepG2 cells with APAP significantly reduced the expression of DAX-1 in a concentration-dependent manner. miR-561 was induced by APAP treatment in HepG2 cells. Transfection of HepG2 cells with an miR-561 mimic exacerbated APAP-induced hepatotoxicity. HNF4α is physically associated with DAX-1 in HepG2 cells. A decreased protein level of DAX-1 by APAP treatment was also enhanced by miR-561 mimic transfection in HepG2 cells and primary human hepatocytes. The basal and APAP-induced expression of PXR and CAR was enhanced by miR-561 mimic transfection; however, transfection of HepG2 cells or primary human hepatocytes with a miR-561 inhibitor or DAX-1 small interfering RNA reversed these effects. Additionally, the chromatin immunoprecipitation assay revealed that recruitment of DAX-1 onto the PXR promoter was inversely correlated with the recruitment of peroxisome proliferator-activated receptor-α coactivator-1α and HNF4α on APAP treatment. These results indicate that miR-561 worsens APAP-induced hepatotoxicity via inhibition of DAX-1 and consequent transactivation of nuclear receptors.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Acetaminophen / adverse effects*
Adrenal Hyperplasia, Congenital / genetics
Adrenal Hyperplasia, Congenital / metabolism
Adrenal Insufficiency
Antioxidants / metabolism
Cell Line, Tumor
Chemical and Drug Induced Liver Injury / genetics*
Chemical and Drug Induced Liver Injury / metabolism
Co-Repressor Proteins / genetics*
Co-Repressor Proteins / metabolism
Constitutive Androstane Receptor
DAX-1 Orphan Nuclear Receptor / genetics*
DAX-1 Orphan Nuclear Receptor / metabolism
Down-Regulation / genetics*
Genetic Diseases, X-Linked / genetics
Genetic Diseases, X-Linked / metabolism
Glutathione / metabolism
Gonadal Dysgenesis / genetics
Gonadal Dysgenesis / metabolism
Hep G2 Cells
Hepatocyte Nuclear Factor 4 / genetics
Hepatocyte Nuclear Factor 4 / metabolism
Hepatocytes / drug effects
Hepatocytes / metabolism*
Humans
Hypoadrenocorticism, Familial
L-Lactate Dehydrogenase / metabolism
Liver / drug effects
Liver / metabolism
MicroRNAs / genetics*
Peroxisome Proliferator-Activated Receptor Gamma Coactivator 1-alpha
Pregnane X Receptor
Promoter Regions, Genetic / genetics
Reactive Oxygen Species / metabolism
Receptors, Cytoplasmic and Nuclear / genetics
Receptors, Cytoplasmic and Nuclear / metabolism
Receptors, Steroid / genetics
Receptors, Steroid / metabolism
Transcription Factors / genetics
Transcription Factors / metabolism
Up-Regulation / genetics

Substances

Antioxidants
Co-Repressor Proteins
Constitutive Androstane Receptor
DAX-1 Orphan Nuclear Receptor
HNF4A protein, human
Hepatocyte Nuclear Factor 4
MicroRNAs
NR0B1 protein, human
PPARGC1A protein, human
Peroxisome Proliferator-Activated Receptor Gamma Coactivator 1-alpha
Pregnane X Receptor
Reactive Oxygen Species
Receptors, Cytoplasmic and Nuclear
Receptors, Steroid
Transcription Factors
Acetaminophen
L-Lactate Dehydrogenase
Glutathione

Supplementary concepts

Dosage-sensitive sex reversal