Establishment of a novel hepatic steatosis cell model by Cas9/sgRNA-mediated DGKθ gene knockout

Jingjing Zhang, Junli Zhao, Xiaojing Zheng, Kai Cai, Qinwen Mao, Haibin Xia*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

3 Scopus citations

Abstract

To investigate the role of diacylglycerol kinase θ (DGKθ) in lipid metabolism and insulin resistance, the present study generated an in vitro hepatic steatosis cell model by knockout of the DGKθ gene in liver cancer cell line HepG2 using CRISPR/Cas9 technology. The cell line was characterized by Oil Red O staining and shown to exhibit increased intracellular lipid accumulation, compared with that in wild-type liver cancer cell line HepG2. The gene expression levels of signaling proteins in pathways involved in lipid metabolism, insulin resistance and gluconeogenesis were also examined. The DGKθ-knockout HepG2 cells showed increased mRNA and protein expression levels of lipid synthesis-related genes, fatty acid synthase, peroxisome proliferator-activated receptor-γ and sterol regulatory element-binding protein-1c, and decreased expression levels of the lipolysis-related gene, carnitine palmitoyltransferase1A. These changes may account for the increased intracellular lipid content of this cell line. The DGKθ-knockout HepG2 cells also exhibited an increased phosphorylation level of protein kinase Cϵ and decreased phosphorylation levels of insulin receptor substrate 1, mechanistic target of rapamycin and protein kinase B (also known as Akt). These changes have been reported to mediate insulin resistance. Taken together, an in vitro hepatic steatosis cell model was established in the present study, providing a valuable tool for understanding the pathogenesis of nonalcoholic fatty liver disease and associated insulin resistance, and for developing treatment strategies for this disease.

Original languageEnglish (US)
Pages (from-to)2169-2176
Number of pages8
JournalMolecular medicine reports
Volume17
Issue number2
DOIs
StatePublished - Feb 2018

Keywords

  • CRISPR/Cas9
  • Diacylglycerol kinase θ
  • Insulin resistance
  • Lipid accumulation
  • Nonalcoholic fatty liver disease
  • Type 2 diabetes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Medicine
  • Molecular Biology
  • Genetics
  • Oncology
  • Cancer Research

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