Growth inhibitor produced by embryonic spinal cord cells in culture: Studies on its properties and actions

Abstract

Cord inhibitor is a low molecular weight product of cultured embryonic chick spinal cord cells which in our experiments interfered with the growth and development of muscle and skin cells in culture. Partially purified preparations, obtained by ultrafiltration, gel-filtration chromatography, and agarose electrophoresis, were stable to heating to temperatures of 100°C and resistant to the action of DNase 1 and RNase 1. Trypsin and Pronase treatment resulted in the loss of inhibitor activity. Cord inhibitor activity on muscle cell cultures was not overcome by nutritional supplementation with glutamine, amino acids, fetal calf serum, or vitamins to twice the amounts usually present. Supplementation with carnitine and pyruvate also did not protect target cultures. Three agents related to the cyclic nucleotide system, theophylline, epinephrine, and dibutyryl cyclic adenosine monophosphate, also were without effect in concentrations from 10 −5 to 10 −7 m. Aspirin, 8 × 10 −5 to 8 × 10 −7 M, did not affect inhibitor production by cord cells or its action on target muscle cells. Serial passage in culture of skin fibroblasts led to their development of resistance to the cord inhibitor. Resistance developed during the first week of cell culture. Three heterologous continuous cell lines also were relatively resistant to the action of the cord inhibitor. Spinal cord cells in primary culture were resistant to the cord inhibitor. Resistant fibroblasts, or lysates prepared from them, did not diminish the titer of cord inhibitor activity remaining after a 4.5-h incubation at 37°C. We suggest that the cord inhibitor may be a peptide containing two to four amino acid residues.