Mca04g

SCs were isolated from the sciatic nerves of 3-d-old Sprague-Dawley rats and cultured as previously described (Feltri et al., 1992 (link), 1994 (link)). In brief, sciatic nerves were dissected and dissociated in 1% collagenase and 2.5% trypsin. Pelleted cells were plated on 100-mm² tissue culture plates coated with poly-l-lysine, in DMEM, supplemented with 10% heat-inactivated fetal calf serum, 2 mM l-glutamine, 2 µM forskolin, and 2 ng/ml β-neuregulin-1. Fibroblast growth was inhibited using 10-µM Ara-C, and by complement killing using anti-Thy1.1 antibodies (MCA04G; Serotec) and 400 µl of rabbit complement (234400; EMD Millipore). The cells were re-fed every 3–4 d and subcultured every 7 d. For pharmacological treatment, SCs were plated at 35 × 10⁴ on poly-l-lysine–coated 6-well plates and treated with 20 µM Furin inhibitor I (Dec-RVKR-CMK; EMD Millipore) for 48 h. Cells were next stained or scraped and proteins were extracted on ice with lysis buffer as described in the Western blot analysis section.