Effect of collagen I and aortic carboxypeptidase-like protein on 3T3-L1 adipocyte differentiation

Aortic carboxypeptidase-like protein (ACLP) is a secreted protein expressed in preadipocytes and down-regulated during adipogenesis. Results from previous studies on the influence of ACLP overexpression on adipogenesis vary from no effect to complete inhibition. We hypothesized that ACLP may modulate adipogenesis in the presence of collagen I, a protein to which it binds. We compared control (pLXSN) 3T3-L1 preadipocytes with 3T3-L1 preadipocytes stably overexpressing ACLP (pLXSN-ACLP) that were grown in standard vs collagen I-coated dishes. Aortic carboxypeptidase-like protein overexpression, via retroviral transduction, resulted in a 3.2-fold increase in ACLP cellular levels and a 2.1-fold increase in ACLP levels released into medium. Aortic carboxypeptidase-like protein overexpression did not inhibit differentiation in standard dishes. In collagen I-coated dishes compared with standard dishes, control preadipocytes, when induced to differentiate, exhibited the same increase in triacylglycerol accumulation, but showed a significantly higher induction of fatty acid synthase (1.6-fold more), peroxisome proliferator-activated receptor γ (1.4-fold more), and CCAAT/enhancer-binding protein α (1.4-fold more). Aortic carboxypeptidase-like protein overexpression significantly reduced this enhanced induction of fatty acid synthase, peroxisome proliferator-activated receptor γ, and CCAAT/enhancer-binding protein α by 65%, 59%, and 66%, respectively, but had no effect on the accumulation of triacylglycerol during differentiation. Finally, studies on proadipogenic insulin signaling in ACLP-overexpressing preadipocytes demonstrated that insulin-stimulated Akt phosphorylation was significantly decreased by 27% in cells cultured in collagen I-coated dishes vs standard dishes. Our data suggest that ACLP inhibits certain aspects of 3T3-L1 adipogenesis in a collagen I-rich environment.