Effect of TAT-obestatin on proliferation, differentiation, apoptosis and lipolysis in 3T3-L1 preadipocytes.

It has been reported that obestatin regulates adipocyte metabolism via receptors on the cell surface. We wondered whether obestatin can interact with intracellular components that activated signalling pathways in adipocytes. Because obestatin (human) only presents one lysine (at position 10), which cannot penetrate the cell membrane, therefore, we used a cell-permeable peptide TAT (49-57) as a vector to carry obestatin across the cell membrane. The goal of this study was to further understand the function of obestatin after penetrating the cell membrane. Our results showed that TAT-obestatin could cross the 3T3-L1 cell membrane in the absence of cytotoxicity. TAT-obestatin showed no effect on the proliferation of 3T3-L1 preadipocytes. In contrast, obestatin significantly stimulated proliferation at a dose of 10(-11)  M and 10(-13)  M. In addition, TAT-obestatin demonstrated a more potent inhibitory effect on cell apoptosis induced by serum starvation than that of obestatin. During the progress of adipocyte differentiation, TAT-obestatin and obestatin had no effect on adipogenesis. In the lipolysis assay, TAT-obestatin significantly increased glycerol and free fatty acid release from 3T3-L1 adipocytes after 3 h treatment but showed no significant effect on lipolysis after 24 h and 48 h of treatment. In contrast, obestatin (10(-7)  M) had no effect on glycerol release after 3, 24 and 48 h of treatment. The difference between the effect of TAT-obestatin and obestatin on adipocytes metabolism indicated that TAT-obestatin may trigger intracellular signalling as well as signalling at the cell membrane.