Expression of type XII collagen by wound epithelial, mesenchymal, and ependymal cells during blastema formation in regenerating newt (Notophthalmus viridescens) tails.

Previously we showed that type XII collagen (col XII) is highly upregulated in the regenerating newt (Notophthalmus viridescens) forelimb. Here, using immunohistochemistry and in situ hybridization, we studied the pattern of expression of col XII during early stages of adult newt tail regeneration. The results show that immunoreactivity of col XII is first seen as a thin layer beneath the wound epithelium (WE) at 3 days after amputation. Reactivity associated with the mesenchyme becomes obvious at day 4 and increases considerably between days 6 and 7 after amputation. In situ hybridization indicates that the early WE-associated reactivity and later mesenchymal reactivity are due to increased col XII gene expression by the WE and mesenchyme, respectively. At 7 days after tail amputation both wound epithelial and mesenchymal cells exhibit a strong riboprobe signal. Interestingly, a distinct riboprobe signal is also seen in the cells of the outgrowing ependymal tube at day 7 but little if any col XII immunoreactivity is present. The spatial pattern of col XII gene expression changes by day 14 after amputation in that transcription in mesenchyme is maintained at a high level, in the WE it is reduced, and in ependyma it ceases to be detectable. Local deprivation of the spinal cord significantly lowers the level of col XII message in the mesenchyme. Much of this decrease in transcription is due to minimal mesenchymal cell accumulation secondary to spinal cord ablation. The temporal and spatial patterns of expression of the col XII gene in the WE, mesenchyme, and ependyma during tail regeneration strongly suggest a role for col XII in regulating both spinal cord outgrowth and spinal cord-dependent tail regeneration.