Abstract:
Objective To observe the effects of di(2-ethylhexyl)phthalate (DEHP) and its metabolite mono(2-ethylhexyl)phthalate (MEHP) on lipid metabolism of human hepatoma cell line HepG2 cells, and explore the mechanism of relevant liver injury.
Methods HepG2 cells were treated with different concentrations of DEHP (31.25, 62.5, 125, 250, 500, and 1 000 μmol/L), MEHP (3.125, 6.25, 12.5, 25, 50, 100, and 250 μmol/L), complete medium (negative control), and 0.1% DMSO (solvent control) for 24 h, 48 h, and 72 h, respectively, to detect the cell viability by MTT assay. HepG2 cells were treated with different concentrations of DEHP (10.0 and 250.0 μmol/L), MEHP (4.0 and 100.0 μmol/L), complete medium (negative control), 0.1% DMSO (solvent control), and 0.5 mmol/L oleic acid (positive control) for 48 h, respectively, to observe the lipid accumulation of HepG2 cells by oil red O staining. HepG2 cells were treated with different concentrations of DEHP (2.0, 10.0, 50.0, and 250.0 μmol/L), MEHP (0.8, 4.0, 20.0, and 100.0μmol/L), complete medium (negative control), 0.5 mmol/L oleic acid, and 0.5μg/mL Brefeldin A for 24 h and 48 h, respectively, to detect the protein expressions of sterol regulatory element binding protein 1c (SREBP-1c), carnitine palmitoyltransferase 1A (CPT1A), malonyl-CoA decarboxylase (MLYCD), and adipose tissue triglyceride hydrolase (ATGL) by Western blot.
Results The DEHP (250.0-1 000.0 μmol/L) and MEHP (50.0-250.0 μmol/L) treatments decreased the cell viability of the experimented HepG2 cells compared with the negative control group (P < 0.05). After exposure for 48 h, the 10.0 and 250.0 μmol/L DEHP groups and the 4.0 and 100.0 μmol/L MEHP groups showed visible lipid droplets accompanied by lipid droplet fusion. Compared with the negative control group, the expression of CPT1A was down-regulated in the HepG2 cells treated with 250.0μmol/L DEHP or 100.0μmol/L MEHP for 24h and 50.0-250.0μmol/L DEHP or 4.0-100.0μmol/L MEHP for 48h compared with the negative control group (P < 0.05); the expression of MLYCD was down-regulated in the HepG2 cells treated with 10.0-250.0 μmol/L DEHP or 4.0-100.0μmol/L MEHP for 48 h (P < 0.05); the expression of ATGL was down-regulated in the HepG2 cells treated with 250.0μmol/L DEHP or 20.0-100.0μmol/L MEHP for 24 h and 10-250.0μmol/L DEHP or 0.8-100.0μmol/L MEHP for 48 h (P < 0.05).
Conclusion DEHP and MEHP can cause lipid accumulation in HepG2 cells, which may be related to the inhibition of fatty acid beta-oxidation and triglyceride hydrolysis.
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