Effects Of Formaldehyde On Lipid Metabolism In HepG2Cell

Objective: Animal experiments show that formaldehyde (FA) could induce hepatocyte injury,oxidative damage, and carcinogenicity which has been found in epidemiologic studies. In ourstudy, HepG2was used as experimental model. It was confirmed that FA could increasetriglyceride levels in the hepatocyte. In order to explore the possible mechanism of lipid changesand the TG accumulation in liver, we tested the expression of the protein that associated withlipid metabolism, therefore, to provide theoretical basis for preventing formaldehyde abuse andprotecting human health and environment.Methods: Human hepatocellular carcinoma cell (HepG2) were cultured in10%calf serummedium with high glucose, logarithmic growth phase cells were seeded in cell culture plates, thecells were respectively treated with0.02,0.1,0.5,2.5,12.5mmol/L concentration of FA for24hand48h, CCK-8assays were used to measure the activity of HepG2. TG concentrations weremeasured by GPO-POD enzyme method. Morphological changes of the HepG2nuclear, whichwere stained by Hoechst33258, were observed by optical microscopes; HepG2treated with0.004,0.02,0.1mmol/L concentration of FA for24h and48h. The expression of Carnitinepalmitoyl transferase1(CPT1), Sterol regulatory element-binding protern-1c (SREBP-1c),SAR1A, ADP-ribosylation factor1(Arf1), Coat protein comlex â… (COPI),Triacylglycerolhydrolase (TGH) and Microsomal triglyceride transfer protein (MTP) were tested by Westernblot. VLDL level, ApoB100and ApoE secretion in the cell culture supernatant were measured byenzyme-linked immunosorbent assay (ELISA).Results:1. The effect of FA on the activity of HepG2: after24h exposure, D(470) values in the treatmentgroups were remarkably less than the negative control group (P0.05). D (470) value decreasedwith the increasing of FA, which suggested that with the increasing of FA concentration, theinhibition of the activity of HepG2gradually intensified. The survival rate of heatocytes of0.02mmol/L group was remarkablely higher than the negative control group, which suggested that inthis concentration, FA could promote the proliferation. After48h exposure, HepG2activity in the0.1~12.5mmol/L treatment groups was significantly lower than the negative control group, which indicated that FA could depress the activity of HepG2.2.The results of the Hoechst33258stain: After24h and48h exposure, a large number of nuclearcondensation or apoptotic cells were observed in the treatment groups (0.5,2.5,12.5mmol/LFA). In the positive control group (10mmol/L CP), hyperchromatic nuclei were observed, thenumber of the cells significantly reduced, especially in the higher groups (2.5~12.5mmol/L) andthe postive control group. In the negative control group and the lower groups, the nuclei are pale,large but no condensed.3. The influences of TG after the exposure to formaldehyde in the HepG2cell:After24h exposedto the FA, the difference between0.1mmol/L FA group and the negative control group had nostatistical difference (P0.05). After48h, there were significant differences between the lowergroups (0.02,0.1mmol/L) and the negative control group(P0.05). The TG level in48h waslower than the24h, while in the positive control group, TG level in48h was higher than24h.4.The effects of CPT1and SREBP-1c expression afer the exposure of formaldehyde: After24hexposed to FA, there was no change in CPT1protein expression, but it was significantlyrestrained after48h; the SREBP-1c expression were higher than the negative control group,which suggested that FA exposure could promote HepG2intracellular fatty synthesis in theinitial stage. The inhibitory effect on the-oxidation of fatty acid gradually emerged with theprolongation of exposure time to FA.5. The effects of TGH and MTP protein expression in the hepatocytes: The TGH proteinexpression in the0.01mmol/L FA group and the oleic acid group increased significantly after24hexposure, and significantly lower than the negative control group after48h exposure. Theexpression of MTP protein in0.1mmol/L FA group were increased significantly compare withthe negative control group (P0.05).6.The effects of Arf1,COPâ… and SAR1A protein expression in the hepatocytes exposured to FA:After24h exposure to FA, the Arf1and COPâ… expression in the hepatocytes increasedsignificantly, while the Arf1expression returned to normal level and the COPâ… expressionhigher than the negative control group after48h exposure.It was suggested that the hepatocytescould decrease the accumulation of TG by increasing the expression of Arf1and COPâ… andpromote VLDL secretion. The protection function became weak as the prolongation of exposuretime to FA. SAR1A expression had no significant difference compared to the negative control group after24h and48h exposure to FA (P>0.05).7.The expression of VLDL,ApoE and ApoB100in the culture supernatant exposure to FA ofHepG2cells: The VLDL and ApoE expression had no significant difference compared to thenegative control group after24h and48h exposure to FA (P>0.05). The ApoB100concentrationof0.02and0.1mmol/L of FA and oleic acid significantly lower than the negative control groupafter48h, which may be associated with the reuptake of the ApoB100.Conclusion:1. The high levels of FA could reduce the activity of hepatocytes obviously and even death.2. The low levels of FA could disturb adipose metabolism in the hepatocytes and induce the TGlevel, this might be associated with the inhibitory of fatty acid oxidation and promote TGsynthesis.