MORPHOGENESIS AND DIFFERENTIATION OF GRAFTED BLASTEMAS FORMED IN-VITRO FROM AMPUTATED HINDLIMBS OF LARVAL XENOPUS-LAEVIS

The present study was designed to test the morphogenetic potency of li mb blastemas formed in vitro from amputated limbs of larval Xenopus la evis. Hindlimbs of larvae at stage 55 (according to Nieuwkoop and Fabe r [1956] Normal Table of Xenopus laevis (Daudin)) were amputated throu gh the tarsalia, excised at the base of the thigh and cultured in Leib ovitz's L-15 supplemented with 2% FCS. After 8-10 days, 50% of the cul tured limbs formed a conic blastema on the amputation surface. However , on the excision surface no blastema was present. Three different par ts (blastema, blastema with the shank region and proximal part of the limb) of the cultured limbs were then grafted to the axial musculature or to the hindlimb of stage 57 host larvae. Results showed that the b lastema formed in vitro were true autodifferentiating regeneration bla stemas, since they were able to form well-differentiated autopodia not only when grafted with the shank region to a neutral territory (axial musculature) or to the limb territory, but also when transplanted alo ne to the two environments. The morphological complexity (no. of toes) of the autopodia differentiated from the grafted blastemas was superi mposable to that observed in vivo. Moreover, as in vivo, the entire re generation process was nerve-independent. In fact, the regeneration bl astemas, formed in vitro in the complete absence of nerves, could grow and differentiate also when grafted to denervated host Limbs. The gra fted proximal parts of the cultured Limbs never formed a regenerate. ( C) 1996 Wiley-Liss, Inc.