On the role of ectoderm in limb development

Isolated limb buds of chick embryos in stages 17 to 21 were stripped of ectoderm by means of focused ultrasound or treatment with EDTA (ethylenediaminetetraacetic acid). Stripped buds were examined for the presence of residual ectoderm and grown as grafts to the coelom, flank, chorioallantoic membrane, or wing bud stump; others were sectioned for histological examination. Four hundred and three grafts prepared by EDTA treatment were examined. These did not form terminal limb parts if completely freed of ectoderm before grafting. Those which were deliberately contaminated with ectodermal cells or considered likely to have residual ectoderm, however, formed skeletal parts characteristic of all proximodistal levels of the limb in a significant percentage of cases even though the ridge was not present as a morphological entity at the time of grafting. Of 68 limb buds prepared by ultrasonation, two or, possibly, three formed terminal limb parts when grafted. These developed from among 41 limb buds which were diagnosed as essentially free of ectodermal cells. Of 27 “contaminated” buds, none formed distal limb structures. On the basis of these results it may be concluded that: (1) terminal limb parts do not develop from limb buds which have been completely freed of ectoderm by means of EDTA treatment; (2) distal limb parts can form from EDTA-stripped limb buds containing residual ectodermal cells, even though the apical ridge is not present as a morphological entity; (3) some limb buds may form terminal limb parts in the complete absence of ectodermal cells after stripping by means of focused ultrasound; (4) distal parts do not form from buds which contain residual ectoderm after ultrasonation.