|Budget Amount *help
¥2,100,000 (Direct Cost: ¥2,100,000)
Fiscal Year 1990: ¥1,000,000 (Direct Cost: ¥1,000,000)
Fiscal Year 1989: ¥1,100,000 (Direct Cost: ¥1,100,000)
A method is presented by which murine preadipose cells may be successfully grown and converted into mature adipocytes in the complete absence of serum. ST 13 preadipocyte were grown in a 1 : 1 mixture of RPMI 1640 and Ham F12 supplemented with sodium selenite, Hepes, transferrin, insulin, thrombin, and fetuin on various substrates, including four types of collagen, fibronectin, and laminin. A highly potent adipogenic agent, 3-hydroxyciglitazone (ADD), was essential for the induction of adipose conversion of ST 13 cells grown under serum-free conditions. As was true for cells in serum-supplemented medium, ADD treatment must begin in the logarithmic growth phase to induce efficient adipose conversion. Two major differences were observed, however, between serum-free and serum-supplemented adipose conversion systems. First, ST 13 cells grown without serum require a collagenous substrate (especially collagen type IV) for adipose conversion and for maintenance of a confluent monolayer, through not for cell proliferation. Second, insulin, which plays relatively minor role in the serum-supplemented system, is required absolutely in the serum-free system. The present observation demonstrates rhat ST 13 cells are tightly held in the preadipose state under serum-free conditions and appropriate adipogenic stimuli (such as ADD plus insulin) are essential for adipose conversion. The serum-free system will be useful for the identification and investigation of regulatory factors affecting adipose conversion.