Stimulation of chick neuroblast proliferation in culture by brain extracts

Chick neuroblasts from 6-day-old embryos were grown on a collagen substrate in minimum Eagle's medium supplemented with 5% horse serum. We found that [3H]thymidine incorporation in these cultures decreased drastically from day 2 to day 3 after seeding. Addition of brain extract from 8-day-old chick embryos (CBe8) to the nutrient medium after 24 hours of culture elicited a very important stimulation of [3H]thymidine incorporation. Extracts of brains from adult animals were much less effective. It was shown that the increase of tritiated-thymidine incorporation reflected proliferation of neuroblasts in the culture, at least between day 2 and 4. The activity of the brain extracts was destroyed by trypsin treatment, which suggests that the factor responsible for the stimulation of the proliferation is a polypeptide. The CBe8 and the brain extract of adult chicken (CBa) were fractionated by gel filtration. An active fraction was eluted at a volume indicating an apparent molecular weight of roughly 70,000. At that stage of the study results suggest that a polypeptide (tentatively called neuroblast proliferation factor, NPF) present in the embryonic chick brain extract, is a proliferation factor for chick neuroblasts in primary culture.