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To explore interleukin-6 (IL-6) production and characterize lipid accumulation in L02 hepatocytes induced by sodium oleate. L02 hepatocytes were incubated with 0, 37.5, 75, 150, 300, 600, or 1,200 μmol/L sodium oleate for 24 h, and the supernatant was collected to detect the concentration of IL-6. L02 hepatocytes were incubated with 300, 150, 75, or 0 μmol/L sodium oleate for 0-24 h. The supernatant was collected for detection of IL-6 and free fatty acids. L02 hepatocytes treated with 300 μmol/L sodium oleate for 0-24 h were stained with Oil Red O. With extended sodium oleate incubation time, IL-6 levels increased, and free fatty acids decreased. After 24 h incubation, IL-6 levels increased as sodium oleate increased from 37.5 to 300 μmol/L (
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Humanos , Relación Dosis-Respuesta a Droga , Hepatocitos/metabolismo , Interleucina-6/metabolismo , Metabolismo de los Lípidos , Ácido Oléico/administración & dosificación , Factores de TiempoRESUMEN
5-Hydroxymethylfurfural (5-HMF), a water-soluble compound extracted from wine-processed Fructus corni, is a novel hepatic protectant for treating acute liver injury. The present study was designed to investigate the protective effect of 5-HMF in human L02 hepatocytes injured by D-galactosamine (GalN) and tumor necrosis factor-α (TNF-α) in vitro and to explore the underlying mechanisms of action. Our results showed that 5-HMF caused significant increase in the viability of L02 cells injured by GalN/TNF-α, in accordance with a dose-dependent decrease in apoptotic cell death confirmed by morphological and flow cytometric analyses. Based on immunofluorescence and Western blot assays, we found that GalN/TNF-α induced ER stress in the cells, as indicated by the disturbance of intracellular Ca(2+) concentration, the activation of protein kinase RNA (PKR)-like ER kinase (PERK), phosphorylation of eukaryotic initiation factor 2 alpha (eIF2α), and expression of ATF4 and CHOP proteins, which was reversed by 5-HMF pre-treatment in a dose-dependent manner. The anti-apoptotic effect of 5-HMF was further evidenced by balancing the expression of Bcl-2 family members. In addition, the knockdown of PERK suppressed the expression of phospho-PERK, phospho-eIF2α, ATF4, and CHOP, resulting in a significant decrease in cell apoptosis after the treatment with GalN/TNF-α. 5-HMF could enhance the effects of PERK knockdown, protecting the cells against the GalN/TNF-α insult. In conclusion, these findings demonstrate that 5-HMF can effectively protect GalN/TNF-α-injured L02 hepatocytes against ER stress-induced apoptosis through the regulation of the PERK-eIF2α signaling pathway, suggesting that it is a possible candidate for liver disease therapy.
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Humanos , Apoptosis , Cornus , Química , Estrés del Retículo Endoplásmico , Factor 2 Eucariótico de Iniciación , Genética , Metabolismo , Furaldehído , Farmacología , Galactosamina , Metabolismo , Hepatocitos , Biología Celular , Metabolismo , Hígado , Biología Celular , Metabolismo , Extractos Vegetales , Farmacología , Sustancias Protectoras , Farmacología , Transducción de Señal , Factor de Necrosis Tumoral alfa , Genética , Metabolismo , eIF-2 Quinasa , Genética , MetabolismoRESUMEN
The effects of PEG10 on hydrogen peroxide (H2O2)-induced apoptosis in human normal liver cell line L02 were investigated. The PEG10 gene was transfected into L02 cells by lipofectamine,the positive clone was screened by G418 and defined as L02/PEG10,while the cell transfected with empty expression vector (pEGFP-N1) was defined as L02/vector. L02/vector and parental L02 cells served as control. RT-PCR and Western blotting were employed to detect the expression of target genes. H2O2 (50-400 mmol/L) was administered to induce the apoptosis of L02 cells. Cells viability was measured by MTT and the morphological changes of apoptotic cells were determined by fluorescence microscopy using hoechst33342 nuclei staining. DNA fragmentation was observed by agarose gel electrophoresis. PEG10 mRNA and protein levels in L02/PEG10 ceils were significantly increased as compared with those in the control cells. After treatment with 400 mmol/L H2O2 for 24 h,the cellular growth inhibition rate of L02/PEG10 cells was significantly lower (58.2%) than that of L02 (92.5%) and L02/vector (88%). Distinct morphological changes characteristic of cell apoptosis such as karyopyknosis and conglomeration were not observed in L02/PEG10. Ladder-like DNA fragmentation in a dose-dependent manner was observed in both L02 and L02/vector cell lines,but not in L02/PEG10. PEG10 over-expression significantly inhibited cytotoxicity induced by H2O2 on human normal liver cell line L02 by antagonizing H2O2-induced apoptosis.
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Objective To study the protective effect on CCl4-injured L-02 hepatocytes of polysaccharides from Hyriopsis cumingii(HCP) and try to explain the mechanism.Methods The L-02 hepatocytes were incubated and then injured by CCl4.The levels of AST,ALT,MDA and SOD in cultural supernatant were detected by general methods.Cell viability was assayed by MTT method.Results The polysaccharides(25,250 and 1000?g?L-1)could reduce the levels of AST,ALT,MDA in cultural supernatant which increased by CCl4.It also could boost the hepatocytes viability and elevate the level of SOD which reduced by CCl4.Conclusion The results suggest that polysaccharides from Hyriopsis cumingii have direct protective activity on hepatocytes injured in vitro.It might be associated with the anti-oxidative activity of polysaccharides from Hyriopsis cumingii.