The effect of nonlimb tissues on forelimb regeneration in the axolotl, Ambystoma mexicanum

Abstract

AbstractPrevious studies with Notophthalmus viridescens have shown that forelimb stumps covered by flank skin do not regenerate when amputated through the graft of nonlimb skin [Tank, P.W. (1984) J. Exp. Zool., 229:143–153]. In an attempt to determine whether or not cells derived from nonlimb skin dedifferentiate and enter the regeneration blastema, four experimental series were prepared using triploid (3N) marked cells grafted into diploid (2N) hosts of the axolotl (Ambystoma mexicanum). In the first series, head skin was used to replace limb skin followed by amputation through the graft 30 days later. Fully 96% (24/25) of the limbs failed to regenerate an externally visible outgrowth and 4.4% of the blastemal mesenchyme cells were observed to be 3N based on the characteristic of three nucleoli per nucleus. In the second series, flank skin replaced limb skin and 38% (8/21) of the limbs failed to regenerate. The remaining limbs regenerated an assortment of hypomorphic, normal, abnormal, or multiple structures. Only 2.4% of the cells in the blastemal mesenchyme were observed to be 3N in the second series. In the third series, head dermis was implanted in place of limb musculature and 20% (3/15) of the limbs failed to regenerate. The other limbs in series three were either abnormal, hypomorphic, or multiple in morphology. Histological analysis revealed that 3.4% of the cells in the blastemal mesenchyme of the head dermis group were 3N. In the fourth series, flank dermis replaced limb musculature and only 7% (1/15) failed to regenerate. Ten limbs (66%) formed hypomorphic regenerates and the remainder were normal, abnormal, or multiple in morphology. Histological analysis revealed that 6.8% of the cells were 3N in the blastemal mesenchyme of limbs in series four. Sham‐operated controls for all four series regenerated normally and histological analysis of contralateral control limb regenerates revealed background 3N marker that averaged from 0.1 to 0.8%. The regenerate epidermis of limbs that were covered by 3N head skin or flank skin contained approximately 25% 3N cells, indicating that the epidermis was of nonlimb origin. The regenerate epidermis of limbs bearing 3N dermis implants did not contain marked cells. These results indicate that non‐limb cells are capable of undergoing dedifferentiation and entering the limb regeneration blastema. An explanation of regenerative failure is provided that is based on interference with short range cellular interactions by nonlimb cells.