Construction and characterization of peroxisome proliferator-activated receptor-gamma co-activator 1 alpha (PGC-1α over-expressing cell line derived from human hepatocyte carcinoma HepG2 cells)

Abstract

Aims. The aim was develop stable human cell line stable over-expressing transcription co-activator peroxisome pro- liferator-activated receptor gamma co-activator 1a (PGC-1a) with restored hepatospecific functions and increased expression of major xenobiotic metabolizing enzymes. Methods. Six clones of HepG2-PGC-1a and one control clone HepG2-pcDNA3 were isolated and analyzed for secretion of hepato specific markers, fibrinogen, albumin and alphal-antitrypsin. Expression levels of protein and mRNA of hepatocyte nuclear factor (HNF4a), pregnane X receptor (PXR) and aryl hydrocarbon receptor (AhR) were determined. We measured basal and ligand inducible expression of CYP1A1 and CYP3A4. Results. Stably transfected cell line HepG2-PGC-1a derived from HepG2 cells over-expressing PGC-1a displayed increased secretion of fibrinogen, but not albumin or alphal-antitrypsin compared to parent HepG2 cells. We found increased levels of HNF4a, PXR and AhR proteins but not their mRNAs in HepG2-PGC1 cells. Basal expression of CYP3A4 protein in HepG2-PGC-1a cells was increased but rifampicin-inducible expression of CYP3A4 protein was lowered in comparison with parent HepG2 cells. Induction of CYP3A4 mRNA varied between 1.3 - 1.9 fold in individual clones. Expression of TCDD-inducible CYP1A1 protein was lower in HepG2-PGC-1a cells than in parent HepG2 cells. Induction of CYP1A1 mRNA by TCDD in HepG2-PGC-1a cells was comparable with that in parent HepG2 cells and ranged between 103 - 198 fold. Conclusion. Stable expression of PGC-1a in HepG2 cells restores several hepatospecific functions, such as secretion of fibrinogen, expression of HNF4a1 and xenoreceptors PXR and AhR. However, the expression and induction of key drug-metabolizing enzymes (CYP1A1 and CYP3A4) were not improved.