Digit regeneration is regulated by Msx1 and BMP4 in fetal mice

The regeneration of digit tips in mammals, including humans and rodents, represents a model for organ regeneration in higher vertebrates. We had previously characterized digit tip regeneration during fetal and neonatal stages of digit formation in the mouse and found that regenerative capability correlated with the expression domain of the Msx1 gene. Using the stage 11 (E14.5) digit, we now show that digit tip regeneration occurs in organ culture and that Msx1, but not Msx2, mutant mice display a regeneration defect. Associated with this phenotype, we find that Bmp4 expression is downregulated in the Msx1 mutant digit and that mutant digit regeneration can be rescued in a dose-dependent manner by treatment with exogenous BMP4. Studies with the BMP-binding protein noggin show that wild-type digit regeneration is inhibited without inhibiting the expression of Msx1, Msx2 or Bmp4. These data identify a signaling pathway essential for digit regeneration, in which Msx1 functions to regulate BMP4 production. We also provide evidence that endogenous Bmp4 expression is regulated by the combined activity of Msx1 and Msx2 in the forming digit tip; however, we discovered a compensatory Msx2 response that involves an expansion into the wild-type Msx1 domain. Thus, although both Msx1 and Msx2 function to regulate Bmp4 expression in the digit tip, the data are not consistent with a model in which Msx1 and Msx2 serve completely redundant functions in the regeneration response. These studies provide the first functional analysis of mammalian fetal digit regeneration and identify a new function for Msx1 and BMP4 as regulators of the regenerative response.