Recent studies have shown that monounsaturated oleic acid induces steatosis in cultured hepatocyte steatosis in the form of nonalcoholic fatty liver disease models in vitro. However, the underlying mechanism of steatosis development is not completely understood. Therefore, we investigated the molecular mechanism of steatosis and the role of mitogen-activated protein kinase (MAPK)/toll-like receptor 4-related protein (TLR4) expression in this study. Rat hepatocyte cells were subjected to oleic acid in different concentrations (1.2-2.4 mM) for 24 hours. The cell morphological injury index and the changes in the MAPK/TLR4 signaling pathway-related proteins were evaluated. We found that the microstructure of the cells in the oleic acid treatment group was damaged, and higher phosphorylation levels of the MAPK pathway-related proteins were detected than those in the control group. In addition, the protein expression of TLR4, sterol regulatory element-binding protein-1, and fatty acid synthase were increased in the oleic acid treatment group. Our findings demonstrate that oleic acid causes toxic damage to rat hepatocyte cells, and the MAPK/TLR4 signaling pathway plays a significant role in lipid storage.

Effect of oleic acid on induction of steatosis and cytotoxicity in BRL 3A cells.

Objective To study the effect of SREBP-1c silencing on lipid metabolism and expression of inflammatory chemokines in a NAFLD model with endoplasmic reticulum stress. Method NAFLD model was established in L02 cells treated with oleic acid. SREBP-1c expression was inhibited using RNA interference with a pSilencer-1.0-U6-4476 vector. After transfection with pSilencer-1.0-U6-4476 or control vector for 0h, 24h, 48h and 72h, the extent of fatty degeneration was shown by Oil Red O staining. The mRNA and protein expression of inflammatory chemokine CCL2 and basic fibroblast growth factor-21 (FGF21) were determined by real time PCR and Western blot respectively. Results SREBP-lc silenced L02 cells showed fat droplets with smaller diameter and attenuated fatty deposition, as compared with control cells. The relative CCL2 mRN A levels in SREBP- 1 c silencing vector transfected L02 cells were 1.03 ± 0.11 for 0 h, 1.11 + 0.21 for 24 h, 0.88 + 0.16 for 48 h, and 1.05 + 0.15 for 72h, which showed no significant difference as compared with control cells (P ＞ 0.05, respectively). In addition, no difference was found between the different time points within the same group (P ＞ 0.05). However, CCL2 protein levels in SREBP- 1 c silenced cells were 1.19 + 0.15, 1.07 + 0.18, 0A8 + 0.14, and 0.05 + 0.24 after transfection for 0 h, 24h, 48h, and 72h respectively, which were significantly downregulated as compared to the control group (P ＜ 0.01). And CCL2 protein levels between different time points in SREBP-1c silenced cells were also distinct (P ＜0.01). The relative FGF21 mRNA levels in SREBP-1c silenced L-02 cells were 1.01 + 0.08, 0.91 + 0.22, 0.98 + 0.20, and 1.02 ± 0.12 for 0h, 24h, 48h, and 72h respectively, which were not statistically different as compared with the corresponding control cells. Statistic difference of FGF21 mRNA levels in SREBP-1c knockdown cells of different time points was not found (P＞ 0.05). In striking contrast, robust downregualtion of FGF21 protein in SREBP-lc silenced cells was observed, with 0.81 + 0.05, 0.66 + 0.12, 0.58 + 0.08 and 0.19 + 0.13 after transfection for 0 h, 24h, 48h and 72 h respectively, as compared to control group (P ＜ 0.01). And differences in FGF2 1 protein level between different time points in SREBP-1c silenced cells were also demonstrated (P＜0.01). Conclusion SREBP1c knockdown attenuated fatty deposition in oleic acid treated L02 cells. In addition, silencing of SREBP-1c expression reduced expressions of CCL2 and FGF21 proteins posttranscriptionally, which may play a role in endoplasmic reticulum stress induced inflammatory response in NAFLD.

Key words: 
Fatty liver;  Fibroblast growth factor 21;  Chemoldne cytokine ligand 2.

[SREBP-1c knockdown attenuated fatty degeneration in hepatic L02 cells and inhibited CCL2 and FGF21 protein expression].

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[SREBP-1c knockdown attenuated fatty degeneration in hepatic L02 cells and inhibited CCL2 and FGF21 protein expression].