INFLUENCE OF DIGITS, ECTODERM, AND RETINOIC ACID ON CHONDROGENESIS BYMOUSE INTERDIGITAL MESODERM IN CULTURE

We have cultured tissues isolated from the interdigital zones (IDZ) of the mouse footplate in the presence of the digits, ectoderm, and ah-t rans retinoic acid. The objective was to understand how these various factors influence the developmental fate of the interdigital tissues. Neutral red staining showed that these tissues normally differentiate by dying between day 12.5-14.5. However, if they were isolated from th e footplate between day 12.5-13.5 (when cell death is not overtly obvi ous in the IDZ) and maintained in organ culture, these tissues would d evelop into cartilage and soft connective tissues. In culture, chondro genesis is initiated very rapidly in the interdigital explants as reve aled by in situ hybridization with riboprobes specific for type IIA an d IIB procollagen mRNAs. The ability of interdigital tissues to form c artilage is not attributed to factors present in the serum of the cult ure medium as this phenomenon is also observed in serumless cultures. We have found that if all-trans retinoic acid, at concentrations of 10 -50 ng/ml culture medium, were added to the explants it could inhibit chondrogenesis and promote cell death. Moreover, in some of the cultur es, a single digit was left attached to the interdigital tissue, This also dramatically reduced the incidence of chondrogenesis, We have tri ed to determine whether the digits and ectoderm can produce a diffusib le factor that can prevent cartilage from developing by culturing day 12.5 interdigital tissues in ectoderm and digit conditioned media, The ectoderm conditioned medium had no effects on interdigital growth or chondrogenesis, In contrast, the size of interdigital explants culture d in the presence of digit conditioned medium was shown to be signific antly smaller than the control. These explants also produced a smaller quantity of cartilage as revealed by Alcian blue binding assay. In su m, our results showed that the fate of the interdigital tissues are no t fully determined until after day 13.5. These tissues have the potent ials to form cartilage and soft connective tissues, We tentatively pro pose that these interdigital tissues do not normally realize their his togenetic potentials because of the antichondrogenic influence of the digits and retinoic acid. (C) 1994 Wiley-Liss, Inc.