Association of mesenchyme with attenuated basement membranes during morphogenetic stages of newt limb regeneration

Abstract

The interface between epithelium and mesenchyme may be involved in inductive interactions which occur during development. This interface within the growth bud, or blastema, of a regenerating limb has been examined to determine whether changes in basement-membrane structures are visible in regions of putative epithelial-mesenchymal inductive interaction. Regenerating forelimbs of adult newts were fixed by perfusion with osmotically balanced aldehydes. Late-bulb to early-digit stage regenerates were collected and processed either for light and transmission electron microscopy or for scanning electron microscopy. Light microscopy confirmed that regions characterized by increased numbers of subepithelial mesenchymal cells were covered by a diffusely stained basement membrane. Transmission electron microscopy of these regions revealed two structural components of the basement membrane. The thin basal lamina was continuous in all regions of all stages examined, but it was attenuated apically in areas of mesenchymal cell accumulation. The thicker underlying reticular lamina was markedly attenuated in these regions near the blastemal apex. Scanning electron microscopy of de-epithelialized blastemas revealed that, apically, the reticular lamina formed only a delicate lacelike network. On the base of the blastema, it formed a dense fibrillar meshwork which was further organized into a geometric pattern in the adjacent stump skin. Cumulatively, these observations suggest that physical contact between epithelial and mesenchymal cells is not essential at these stages, but that regions of putative epithelial-mesenchymal interaction are characterized by a distinctly diminished reticular lamina. Structural changes in basement-membrane components may be related to termination of local inductive events.