EXAMINING THE ROLE OF PANETH CELLS IN THE SMALL-INTESTINE BY LINEAGE ABLATION IN TRANSGENIC MICE

The Paneth cell lineage is one of four epithelial lineages derived fro m the adult mouse small intestine's multipotent stem cell, Mature Pane th cells secrete antimicrobial peptides (cryptdins), growth factors, a s well as two gene products, a secreted phospholipase A, and matrilysi n, that has been implicated as modifiers of adenoma formation in mice containing a mutation in the tumor suppressor Apc. Immature Paneth cel ls are located just above and below the cell layer, in intestinal cryp ts, that has been proposed to contain the multipotent stem cell. Panet h cells differentiate during a downward migration to the crypt base. T he location and direction of Paneth cell migration, their high density and long residency time at the crypt base, and the nature of their se creted gene products, suggest that they may influence the structure an d/or function of the stem cell niche. Paneth cell ablation can therefo re be viewed as an experimental manipulation of the cellular microenvi ronment that purportedly contains the stem cell and its immediate desc endants, Two types of ablation experiments were performed in transgeni c mice. Nucleotides -6500 to +34 of the mouse cryptdin-2 gene (CR2) we re used to express an attenuated diphtheria toxin A fragment, Light an d electron microscopic immunohistochemical analyses of several pedigre es of postnatal day 28 to 180 animals established that ablation of Pan eth cells is accompanied by an increase in the proportion of undiffere ntiated crypt base columnar cells, These cells normally co-exist with Paneth cells, The ablation does not produce a detectable effect on the proliferation or terminal differentiation programs of the other three lineages or on host-microbial interactions, Tile last conclusion is b ased on the ability of crypts to remain free of microbes detectable by Gram and Warthin-Starry stains and by retention of the normal crypt-v illus distribution of components of the diffuse gut-associated lymphoi d tissue, CR2-directed expression of simian virus 40 large T antigen a lso results in a loss of mature Paneth cells but produces a marked amp lification of crypt cells having a morphology intermediate between Pan eth and granule goblet cells, EM immunohistochemical analyses suggest that intermediate cells can differentiate to mature goblet cells but n ot to Paneth cells, as they migrate up the crypt-villus axis, Our find ings suggest that (i) stemness in the crypt is not defined by instruct ive interactions involving the Paneth cell; (ii) expressing a Paneth c ell fate may require that precursors migrate to the crypt base; (iii) antimicrobial factors produced by Paneth cells are not required to pre vent colonization of small intestinal crypts; and (iv) this lineage do es not function to maintain the asymmetric crypt-villus distribution o f components of the diffuse gut-associated lymphoid tissue.