STAGE-DEPENDENT AND REGION-DEPENDENT CHONDROGENESIS AND GROWTH OF CHICK WING-BUD MESENCHYME IN SERUM-CONTAINING AND DEFINED TISSUE-CULTURE MEDIA

During development, limb-bud mesenchymal cells carry out complex spati otemporal patterns of growth and differentiation. Tissue and organ cul ture facilitate analysis of environmental influences on these cell beh aviors, allowing their partial dissection into exogenous and endogenou s components. Two factors that complicate such in vitro analyses are t he heterogeneity of the cultured cells and imprecise knowledge of cult ure medium composition. Limb mesenchyme comprises a heterogenous cell population with important regional differences in cell type. Dividing the limb into subregions helps limit the cellular heterogeneity and us ing chemically defined, serum-free medium allays concerns about medium composition. In the present study, mesenchyme from different regions along the anteroposterior and proximodistal axes of stage 21-22 and st age 23-24 chick wing buds was grown in high-density microtiter culture s in chemically defined and in serum-containing medium. Four-day cultu res of the various regions were compared in terms of culture morpholog y and the accumulation of Alcian blue-positive cartilage matrix and DN A. The results demonstrate stage- and region-dependent differences in the in vitro growth, differentiation, and responsiveness of these cell s. For example, mesenchyme from the distal anterior region of the wing bud exhibited lower intrinsic chondrogenic capacity and greater respo nsiveness to serum than other regions. Patterns of in vitro chondrogen esis also suggest that, at the stages examined, distal wing-bud mesenc hyme may be less homogeneous than has been believed. A case is made fo r the suitability of serum-free medium for future in vitro studies of chick limb-bud mesenchyme. The results are considered in relation to t he process of limb development and regional expression of pattern-rela ted genes. (C) 1994 Wiley-Liss, Inc.