STABLE, POSITION-RELATED RESPONSES TO RETINOIC ACID BY CHICK LIMB-BUDMESENCHYMAL CELLS IN SERUM-FREE CULTURES

Retinoic acid (RA) has dramatic effects on limb-skeletal patterning in vivo and may well play a pivotal role in normal limb morphogenesis. R A's effects on the expression-of pattern-related genes in the developi ng limb are probably mediated by cytoplasmic RA-binding proteins and n uclear RA-receptors. Little is known, however, about how RA modifies s pecific cellular behaviors required for skeletal morphogenesis. Earlie r studies supported a role for regional differences in RA concentratio n in generating the region-specific cell behaviors that lead to patter n formation. The present study explores the possibility that position- related, cell-autonomous differences in the way limb mesenchymal cells respond to RA might have a role in generating pattern-related cell be havior. Mesenchymal cells from different proximodistal regions of stag e 21-22 and 23-24 chick wing-buds were grown ip chemically defined med ium and exposed to 5 or 50 ng/ml of RA for 4 days in high-density micr otiter cultures. The effects of RA on chondrogenesis in these cultures clearly differed depending on the limb region from which the cells we re isolated. Regional differences in RA's effects on growth over 4 day s in these cultures were less striking. The region-dependent responses of these cells to RA proved relatively stable in culture despite ongo ing cytodifferentiation. This serum-free culture model will be useful in exploring the mechanisms underlying the region-dependent responsive ness of these cells io RA.