OFFE Inhibits Adipogenesis in 3T3-L1 Adipocytes by Downregulating Key Genes

SREBP-1c and C/EBPα are key factors in lipid metabolism, and upregulation of SREBP-1c and C/EBPα induces differentiation-related lipid accumulation. To investigate the effect of OFFE on adipocyte differentiation, cells were treated with 0.2 mg/ml, 0.4 mg/ml, and 0.6 mg/ml OFFE starting from day 3 of cell differentiation until day 8 when the cells had fully differentiated into mature adipocytes. Real-time fluorescence PCR results showed that the mRNA levels of SREBP-1c and C/EBPα decreased in a concentration-dependent manner with increasing concentrations of OFFE. Compared to the control group, 0.2 mg/ml of OFFE reduced the mRNA levels of SREBP-1c and C/EBPα by 13% and 33% respectively, while 0.4 mg/ml of OFFE significantly downregulated the mRNA levels of SREBP-1c and C/EBPα to 71% and 34% respectively. Additionally, 0.6 mg/ml of OFFE further decreased the mRNA levels by 92% and 97% respectively (Figure 4a, P <0.001).

Furthermore, we examined the effect of OFFE on the mRNA expression levels of adipogenic genes GLUT-4 and FASN. The mRNA levels of GLUT-4 and FASN were downregulated in a concentration-dependent manner with increasing concentrations of OFFE. Compared to the control group, the mRNA levels of GLUT-4 and FASN were significantly downregulated by 42% and 49% respectively after treatment with 0.2 mg/ml of OFFE. The inhibitory effect of higher concentrations of OFFE was greater than that of lower concentrations, with 0.6 mg/ml of OFFE downregulating the mRNA levels of both lipid-forming genes by 75% and 87% respectively (Fig. 4a, P <0.001).

Regarding the lipid degradation gene CPT1a, both 0.2 mg/ml and 0.4 mg/ml OFFE treatments resulted in upregulation of CPT1a mRNA levels, while 0.6 mg/ml treatment downregulated the mRNA levels of CPT1a. However, none of the three concentrations showed statistically significant effects (Fig. 4b). Consistent with the qRT-PCR results, the protein level of the CPT1α gene showed an upward trend after treatment with OFFE compared to control cells (Fig. 4c), while the protein levels of FASN, C/EBPα, and SREBP-1c genes were all reduced in a dose-dependent manner (Fig. 4d, e, f, P < 0.001). These findings indicate that OFFE treatment significantly downregulates the expression of adipogenesis-related genes and inhibits adipogenesis in 3T3-L1 adipocytes.

In summary, these data suggest that OFFE treatment can significantly inhibit the expression of adipogenesis-related genes and lipid degradation-related genes, thereby inhibiting adipogenesis in 3T3-L1 adipocytes.