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[Adipocytes differentiation of NIH3T3 cells induced by peroxisome proliferator activation receptor gamma 2
1Shanghai Ruijin Hospital, Shanghai Institute of Endocrinology, Shanghai the Second Medical University, Shanghai 200025, China.
Objective:
To express the mouse peroxisome proliferator activated receptor gamma 2(mPPAR gamma 2) in NIH3T3 cells mediated by the recombinant retrovirus and study its function.
Methods:
mPPAR gamma 2 gene digested from the recombinant plasmid pcDNA3/mPPAR gamma 2 and confirmed to contain the target gene segment with fluorescence-sequencing was subcloned into retrovirus vector pGCEN to generate the recombinant retrovirus pGCEN/mPPAR gamma 2. The recombinant retrovirus pGCEN/mPPAR gamma 2 and pGCEN were packaged with PA317 cells and anti-G418 clones of PA317 cells were selected. Viral supernatants were collected and used to infect NIH3T3 cells. Peroxisome proliferator activated receptor gamma 2 (PPAR gamma 2)-expressing NIH3T3 cells cultured in the differentiation media containing PPAR gamma activator ETYA were induced into adipocytes.
Results:
The recombinant retrovirus pGCEN/mPPAR gamma 2 was constructed, 5 x 10(4) CFU/ml of the viral supernatants containing pGCEN/mPPAR gamma 2 and 6 x 10(5) CFU/ml of the viral supernatants containing pGCEN were obtained. mPPAR gamma 2 was expressed in NIH3T3 cells mediated by the recombinant retrovirus. Lipid accumulation obviously existed in PPAR gamma 2-expressing NIH3T3 cells at 10 days postdifferentiation and the lipid-containing cells morphologically resembled the mature adipocytes in vivo.
Conclusion:
An adipocyte differentiation model in vitro was established. The work is the basis for further researches on the molecular mechanism of adipocyte differentiation induced by PPAR gamma 2.