Hypoxia-induced alterations of lipid metabolism in the normal human hepatic L02 cell line / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To investigate the effects of hypoxia on lipid metabolism in the normal human hepatic cell line L02 and to investigate the underlying molecular mechanisms.</p><p><b>METHODS</b>L02 cells were exposed to hypoxic conditions (experimental groups: at 1% O2, 5% CO2, 94% N2 for 3, 6, 12, 24, or 48 hours) or normoxic conditions (control group: at 21% O2). Lipid droplet accumulation and triglyceride content were measured in each group by oil red O staining and biochemical assay, respectively. The mRNA expression levels of hypoxia-inducible factor (HIF)-2a and sterol regulatory element binding protein (SREBP)-1c were detected by reverse transcription-polymerase chain reaction. Protein expression levels of HIF-2a, adipose differentiation-related protein (ADRP), and Fas were tested by Western blot analysis.</p><p><b>RESULTS</b>Lipid droplet accumulation and the triglyceride content were significantly higher in the hypoxia group than the normoxia group. In addition, the hypoxia groups had significantly down-regulated mRNA expression of SREBP-1c (12h: 0.236+/-0.043, 24 h: 0.287+/-0.044, 48 h: 0.342+/-0.049 vs. normoxia: 0.503+/-0.037; F = 28.37, P less than 0.01) and FAS protein (12 h: 0.562+/-0.054, 24 h: 0.674+/-0.062, 48 h: 0.682+/-0.057 vs normoxia: 0.857+/-0.069; F = 16.08, P less than 0.01). In normoxic cells, little or no expression of HIF-2a protein was detected by Western blot. In hypoxic cells, HIF-2a protein expression peaked at 6h (0.973+/-0.067). ADRP protein expression was significantly higher in hypoxia groups than in the normoxia group (12 h: 0.319+/-0.043, 24 h: 0.732+/-0.056 and 48 h: 0.873+/-0.066 vs. 0.211+/-0.019; all, P less than 0.05.</p><p><b>CONCLUSION</b>Exposure to hypoxic conditions might induce lipidosis in normal human hepatic cells by stimulating HIF-2a and ADRP expression.</p>

Effect of hepatitis B virus X protein on expression of lipid metabolism-related genes in HepG2 cells / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To investigate the effect of Hepatitis B Virus X Protein (HBx) on the expression of lipid metabolism-related genes and its role in pathogenesis of hepatocyte fatty degeneration.</p><p><b>METHODS</b>Hepatitis B Virus X gene eukaryon expression vector pIRES2-eGFP-HBx was transfected into HepG2 cells to establish HepG2/HBx cell model for HBx expression. HepG2 cells transfected with pIRES2-eGFP (HepG2/pIRES2 cell) and non-transfected were used as controls. At 24, 48 and 72 hours after transfection, the expression of green fluorescent protein (GFP) was observed by fluorescence microscope and the triglyceride(TG) content was detected. RT-PCR and Western blot were applied to detect the levels of sterol regulatory element binding protein-1 (SREBP-1), liver x receptor alpha (LXRalpha) mRNA and the levels of HBx, LXRalpha and fatty acid synthase (FAS) protein. At 24, 48 and 72 hours after transfection, the expression of GFP was found in HepG2/HBx and HepG2/pIRES2 cells, and increased gradually. The expression of HBx was detected only in HepG2/HBx cells, and was increased with time after transfection (F = 32.21, P less than 0.01). These suggested successful obtaining of HepG2-HBx cell model for HBx expression.</p><p><b>RESULTS</b>At 24h, 48h and 72h after transfection, the expression levels of LXRalpha mRNA (0.386+/-0.055, 0.505+/-0.071, 0.649+/-0.058 ) and SREBP-1 mRNA (0.395+/-0.055, 0.548+/-0.047, 0.795+/-0.058), as well as the levels of LXRalpha protein(0.178+/-0.036, 0.263+/-0.047, 0.347+/-0.058) and FAS protein(0.436+/-0.055, 0.608+/-0.053, 0.827+/-0.046) in HepG2-HBx group were dramatically higher than those in the controls at the same time points (all P less than 0.05/0.01), and were gradually increased with time (all P less than 0.05/0.01). A positive correlationship was observed between HBX protein level and the LXRalpha, SREbP-1 mRNA and LXRalpha, FAS protein levels. The difference of TG content between HepG2/HBx group and control groups was not statistically significant (P more than 0.05).</p><p><b>CONCLUSIONS</b>HBx-LXRalpha-SREBP-1/FAS pathway suggested regulating transcription and expression of lipid metabolism-related genes, which might be one of the important molecular mechanism causing hepatocyte fatty degeneration.</p>