Collagen IV synthesis is restricted to the enteroendocrine pathway during multilineage differentiation of human colorectal epithelial stem cells

The human large intestine is lined by a rapidly renewing epithelial monolay er where cell loss is precisely balanced with cell production. The continuo us supply of new cells is produced by undifferentiated multipotent stem cel ls via a coordinated program of proliferation and differentiation yielding three epithelial lineages: absorptive, goblet and enteroendocrine. Cell-mat rix interactions have been suggested to be regulators of the multilineage d ifferentiation program of the colorectal crypt but the expression of matrix proteins or their receptors does not appear to have the subtlety expected for this task. We have developed an in vitro model system of intestinal epithelial stem ce lls to facilitate the direct analysis of stem cells undergoing lineage comm itment and differentiation. Using this culture system, we can now directly investigate the role of cell-matrix signalling in stem-cell decisions. In t his study collagen-IV synthesis has been followed in monolayers of multipot ent cells that have been induced to differentiate into absorptive, goblet a nd enteroendocrine cells. Our experiments demonstrate that commitment to th e enteroendocrine lineage is specifically accompanied by the expression of type-IV collagen that remains enteroendocrine-cell associated. Undifferenti ated cells, absorptive cells and goblet cells do not express collagen IV. T o confirm that the differential lineage-specific expression of collagen IV observed in the model system was representative of the in vivo situation, c ollagen-IV synthesis was analysed in isolated human colorectal crypts and t issue sections using immunocytochemistry and in situ hybridisation. These s tudies confirmed the in vitro findings, in that implementation of the enter oendocrine differentiation program involves synthesis and accumulation of a collagen-IV matrix. Thus, human colorectal enteroendocrine cells are uniqu e in the colorectal crypt in that they assemble a cell-associated collagen- IV-rich matrix not observed on other colorectal epithelial cells. This study provides the first evidence for differential matrix synthesis be tween colorectal epithelial lineages in human colorectal epithelium. The sp ecialised pericellular environment of the enteroendocrine cells might expla in some of the unique phenotypic characteristics of this cell lineage. Furt hermore, these findings suggest a potential mechanism whereby individual ep ithelial cells could modulate their cell-matrix signalling even while rapid ly migrating in heterogeneous sheets over a shared basement membrane.