Objective:To investigate the effects and mechanisms of free fatty acid(FFA) on 3T3-L1 preadipocyte differentiation. Methods:11-茁-HSD1-SiRNA expression plasmid(pGCsilencerTM H1/TetO1-11-茁-HSD1) was constructed and transfected into 3T3-L1 cells. 100 μg/ml FFA was added to the 3T3-L1 cells. Levels of adipogenesis-associated markers such as Peroxisome proliferator-activated receptorγ(PPARγ), CCAAT enhancer binding protein α(C/EBPα), lipoprotein lipase(LPL), and fatty acid synthetase(FAS) were measured. Results:When the cells treated with FFA, mRNA expressions of PPARγ,C/EBPα,LPL and FAS were up-regulated and the expression of 11-茁-HSD1 also increased. When 11-茁-HSD1 was silenced, the levels of the markers were down-regulated. Conclusion:FFA and 11-茁-HSD1 could promote adipocyte differentiation, and the effect of FFA on adipogenesis carried out through increasing the expression of 11-茁-HSD1.