MCAM knockdown inhibits 3T3-L1 fibroblasts differentiation to adipocytes

VILA, MC; GABRIELLI, M

Mar del Plata

Congreso; LXI Reunión Científica Anual de SAIC; 2016

Sociedad Argentina de Investigación Clínica

Obesity isa chronic disease associated to a dysfunction of the adipose tissue. The studyof adipose tissue development and function may help to find new approaches forthe treatment of this pathology. 3T3-L1 fibroblasts are one of the most usedcell models to study adipogenesis. In the search for novel genes involved inthis process we have previously identified MCAM (melanoma cell adhesion molecule),which mRNA increases during 3T3-L1 fibroblasts differentiation to adipocytesand its levels correlate with the extent of the differentiation achieved. MCAMis a transmembrane protein of the Ig-family which has been involved indifferent processes such as cell adhesion, signal transduction and celldifferentiation. To assess the role of MCAM and the importance of theupregulation of its mRNA during 3T3-L1 cells differentiation, we analyzed theeffects of downregulating MCAM mRNA. 3T3-L1 fibroblasts were transduced withlentivirus containing an shRNA sequence directed to MCAM mRNA. We found that MCAM-knockdowncells showed impaired differentiation as evaluated by Oil-Red-O staining ofcytosolic lipids. This was associated with lower levels of peroxisome proliferatoractivated receptor gamma (PPARγ) mRNA, the master gene in adipogenesis, andperilipin mRNA, which is a protein present in the lipid droplet. We have previouslyobserved that the increase in PPARγ mRNA level precedes the increment of MCAMmRNA. Moreover, MCAM mRNA is further increased by treatment with pioglitazone,a PPARγ agonist, suggesting that MCAM may be regulated by PPARγ. Consistentlywe recently observed that treatment of mature adipocytes with 25 ng/ml TNFα for48 h, downregulated PPARγ and also decreased MCAM mRNA. In keeping with this,we searched chromatin immunoprecipitation databases and found two putativebinding sites for PPARγ near the MCAM gene. Our results indicate that upregulationof MCAM is important in adipogenesis and may be regulated by PPARγ.