Increased Expression of Laminin-1 and Collagen (IV) Subunits in the Aganglionic Bowel ofls/ls,but Notc-ret−/− Mice

Extracellular matrix molecules, including laminin, affect the development of enteric neurons and accumulate in the aganglionic colon ofls/lsmice. Quantitative Northern analysis revealed that mRNAs encoding the β1 and γ1 subunits of laminin and collagens α1(IV) and α2(IV) are increased in the colons ofls/lsmice. Transcripts of laminin α1 were evaluated quantitatively with reverse transcription and the competitive polymerase chain reaction (RT–cPCR). The abundance of laminin α1 transcripts was developmentally regulated, but greater in thels/lsthan the wild-type colon at each age examined.In situhybridization revealed that transcripts in the colon encoding laminin α1 and β1 and collagen α2(IV) were initially expressed in the endoderm, but by E15, expression shifted to cells of the colonic mesenchyme (ls/ls> wild type) where crest-derived cells migrate. The expression of laminin α1 was examined in the totally aganglionic intestine of E15 and newbornc-ret−/− mice, to determine whether an increase occurs when neurogenesis fails independently of thels/lsdefect. RT–cPCR revealed no difference from control in mRNA encoding laminin α1 in thec-ret−/− colon in either E15 or newborn animals. The accumulation of immunohistochemically demonstrable laminin that is prominent in the newbornls/lscolon could not be detected in that ofc-ret−/− animals. These observations suggest that transcripts encoding laminin-1 and collagen (IV) are increased in the colon and surrounding pelvic mesenchyme ofls/lsmice because of an intrinsic lesion, rather than a secondary consequence of aganglionosis. The data are compatible with the hypothesis that the increased expression of laminin-1 contributes to the failure of crest-derived cells to complete their colonization of thels/lscolon.