Background
FOXO3a is a widely studied transcription factor and plays an important role in a variety of biology. The
Conclusion
FOXO3a could promote lipid accumulation and inflammation in differentiated 3T3-L1 adipocytes by targeting autophagy. Our results provide a new theoretical basis for FOXO3a to regulate obesity.
Methods
The obese mouse model was successfully induced by high-fat diet. SiRNA targeting FOXO3a was transfected into differentiation of 3T3-L1 adipocytes to reduce the expression of FOXO3a. The culture medium of RAW264.7 cells was added to the differentiated 3T3-L1 adipocytes to form a co-culture system. Subsequently, ELISA or AdipoRed assay was performed to measure the expression of triglyceride (TG) and cholesterol (TC) in mouse adipose tissue or differentiation of 3T3-L1 adipocytes. Adipocyte differentiation was detected by Oil Red O-staining. Ad-mCherry-GFP-LC3II was used to detect the level of autophagy in differentiation of 3T3-L1 adipocytes. Western blotting or qRT-PCR was used to detect the expression of FOXO3a, autophagy-related proteins (beclin 1, CEBPβ, PPARγ, ACC1 and KLF4), inflammatory cytokines (TNF-α, IL-1β, IL-6 and MCP1), NF-κB signal pathway-related proteins or adipokines (Adiponectin, AdipoR1 and resistin) in differentiated 3T3-L1 or RAW264.7 cells.
Results
The expression of FOXO3a and autophagy levels were significantly increased in visceral adipose tissue of obese mice and differentiation of 3T3-L1 adipocytes. Downregulation of FOXO3a significantly inhibited the autophagy and lipid accumulation in differentiation of 3T3-L1 adipocytes. In addition, FOXO3a knockdown significantly reduced Lipopolysaccharide (LPS)-induced inflammation and adipokines release in RAW264.7 cells treated with the culture medium of 3T3-L1 adipocytes. These above activity changes could be reversed by autophagy inducer rapamycin.