Beryllium nitrate inhibits fibroblast migration to disrupt epimorphic regeneration

Abstract

Epimorphic regeneration proceeds with or without formation of a blastema as observed for the limb and skin respectively. Although not easily perturbed, inhibiting epimorphic regeneration provides a means to interrogate the cellular and molecular mechanisms that regulate it. In this study, we show that exposing amputated limbs to beryllium nitrate disrupts blastema formation and causes severe patterning defects in limb regeneration. In contrast, exposing full-thickness skin wounds to beryllium only causes a delay in skin regeneration. Transplanting full-thickness skin from ubiquitous GFP-expressing axolotls to wildtype hosts we demonstrate that beryllium inhibits fibroblast migration during limb and skin regeneration in vivo. Moreover, we show that beryllium inhibits cell migration in vitro using axolotl and human fibroblasts. Interestingly, beryllium did not act as an immunostimulatory agent as it does in Anurans and mammals nor did it affect keratinocyte migration, proliferation or re-epithelialization suggesting that the effect of beryllium is cell-type specific. While we did not detect an increase in cell death during regeneration in response to beryllium, it did disrupt cell proliferation in mesenchymal cells. Taken together, our data shows that normal blastema organogenesis cannot occur without timely infiltration of local fibroblasts and supports the importance of positional information to instruct pattern formation during regeneration. In contrast, non-blastemal based skin regeneration can occur despite early inhibition of fibroblast migration and cell proliferation.