MITOGEN-INDUCED PROLIFERATION OF ISOLATED ADULT-MOUSE SCHWANN-CELLS

The proliferation of neonatal Schwann cells (SCs) in response to mitog enic agents has been well analyzed in vitro, but a limited range of mi togens have been defined. We investigated whether three identified neo natal SC mitogens [glial growth factor (GGF), platelet-derived growth factor BB (PDGF-BB), and basic fibroblast growth factor (bFGF)] are re quired to stimulate mitosis of adult SCs. Adult SCs were isolated from mouse sciatic nerves by mechanical and chemical dissociation, followi ng three experimental steps: 1) culturing the dissociated cells for 24 hr in 10% FCS-F12, medium, 2) culturing these cells in serum-free med ium for the next 48 hr, and 3) purifying adult SCs by differential adh esion. We describe a new method for preparation of SCs from peripheral nerves of adult mouse that provides 99.5% pure SCs populations at cel l yields of greater than 3 x 10(3) cells/mg of starting nerve wet weig ht within 5 culture days. Although mitosis of SCs in culture in respon se to mitogens requires the presence of serum, the complex nature of s erum renders difficult a complete analysis of mitogens required for SC s DNA synthesis, so we examined the proliferating response of adult SC s to GGF, PDGF-BB, and bFGF in serum-free medium. GGF alone had mitoge nicity for adult SCs in a dose-dependent manner, and synergistic activ ation coupling with forskolin was not observed. Neither PDGF-BB nor bF GF was mitogenic for adult SCs when used alone or with forskolin. Meas urement of intracellular cyclic AMP levels in SCs cultured with these growth factors showed that cAMP levels were not changed by GGF and bFG F, and PDGF-BB reduced the cAMP level to half of basal one. This study demonstrated that adult SCs could proliferate without serum factors a nd forskolin in response to GGF, and adult SCs were activated by GGF t hrough a signal transduction pathway separate from the cyclic AMP-depe ndent system. In addition, PDGF-BB or bFGF has no mitogenic effect on adult SCs under serum-free conditions. (C) 1995 Wiley-Liss, Inc.