Effect of glutaryl-CoA dehydrogenase gene silencing and high-concentration lysine on the viability of BRL hepatocytes.
- Author:
Jin-Zhi GAO
1
;
Cai ZHANG
;
Qin YI
;
Yan-Qin YING
;
Xiao-Ping LUO
Author Information
- Publication Type:Journal Article
- MeSH: Amino Acid Metabolism, Inborn Errors; pathology; therapy; Animals; Apoptosis; Brain Diseases, Metabolic; pathology; therapy; Caspase 3; metabolism; Cell Survival; Cells, Cultured; Fluorescent Antibody Technique; Gene Silencing; Glutaryl-CoA Dehydrogenase; deficiency; genetics; Hepatocytes; pathology; Lysine; metabolism; Rats
- From: Chinese Journal of Contemporary Pediatrics 2017;19(9):1014-1019
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVETo investigate the effect of glutaryl-CoA dehydrogenase (GCDH) gene silencing and accumulation of lysine metabolites on the viability of hepatocytes.
METHODSBRL cells were divided into normal control group, negative control group, and GCDH silencing group. The shRNA lentiviral vector for silencing GCDH gene was constructed, and the BRL hepatocytes in the GCDH silencing group and the negative control group were infected with this lentivirus and negative control virus respectively, and then cultured in a medium containing 5 mmol/L lysine. Immunofluorescence assay was used to measure the infection efficiency of lentivirus. Western blot was used to measure the expression of GCDH protein. MTT assay was used to evaluate cell viability. Hoechest33342 staining was used to measure cell apoptosis. Western blot was used to measure the expression of Caspase-3, an index of cell apoptosis.
RESULTSThe lentivirus constructed effectively silenced the GCDH gene in hepatocytes (P<0.01). MTT assay and Hoechest 33342 staining showed no significant differences in cell viability and apoptosis between groups (P>0.05). There was also no significant difference in the expression of Caspase-3 protein between groups (P>0.05).
CONCLUSIONSGCDH gene silencing and accumulation of lysine metabolites may not cause marked hepatocyte injury.
