Background

Enteric antimicrobial peptides secreted from Paneth cells, including alpha-defensins (in mice named cryptdins), are key effector molecules of innate immunity in the small intestine. The importance of Paneth cells alpha-defensins emerged from studies of enteric bacterial infection in genetically modified mice, as well as from recent studies linking reduced levels of these alpha-defensins to Crohn's disease localized to the ileum. However, analysis of expression of Paneth cell alpha-defensins is incomplete. We therefore performed a comprehensive evaluation of the distribution of antimicrobial molecules along the mouse small intestinal tract to identify potential variations in regional expression.

Results

In conventionally reared mice, the repertoire of Paneth cell antimicrobials differs between duodenum and ileum. In contrast to the uniform expression of most Paneth cell antimicrobials, both cryptdin 4 and cryptdin-related sequences (CRS) 4C peptides were expressed at progressively increasing amounts (101- and 104-fold, respectively) comparing duodenum and ileum. In tissues other than the small intestine, expression of CRS peptides was noted in thymus and caecum. Most Paneth cell products were also produced in the small intestine of germ-free mice at levels similar to those in controls, however CRS4C and RegIIIgamma had reduced levels in the former (3- and 8-fold, respectively). No significant changes in expression levels of Paneth cell antimicrobial peptides was observed after oral challenge with either Salmonella enterica serovar typhimurium or Listeria monocytogenes, supporting current notions on the constitutive nature of this defensive system.

Conclusion

The repertoire of antimicrobial peptides changes along the small intestinal tract, and a subset of these molecules are up-regulated upon colonization, but not in response to enteric bacterial pathogens. The changes detected upon colonization suggest that Paneth cell antimicrobial peptides may play an important role in commensal microbial homeostasis, in addition to their proposed role in protection against infection. In addition, the differential expression of CRS4C along the small intestine suggests mechanisms of regulation that are distinct from other Paneth cell derived antimicrobial peptides.

Background

The small intestinal epithelium mediates vital functions of nutrient absorption and host defense. The spatial organization of the epithelial cells along the crypt-villus axis segregates them into regions of specialized function. However, the differences in transcriptional programming and the molecular machinery that governs the migration, adhesion, and differentiation of intestinal epithelial cell lineages in humans remain under-explored. To increase our understanding of these mechanisms, we have evaluated gene expression patterns of ileal epithelial cells isolated by laser capture microdissection from either the villus epithelial or crypt cell regions of healthy human small intestinal mucosa. Expression profiles in villus and crypt epithelium were determined by DNA microarray, quantitative real-time PCR, and immunohistochemistry based methods. The expression levels of selected epithelial biomarkers were also compared between gastrointestinal tissues.

Results

Previously established biomarkers as well as a novel and distinct set of genes believed to be linked to epithelial cell motility, adhesion, and differentiation were found to be enriched in each of the two corresponding cell populations (GEO accession: GSE10629). Additionally, high baseline expression levels of innate antimicrobials, alpha defensin 5 (HD5) and regenerating islet-derived 3 alpha (Reg3A), were detected exclusively within the small bowel crypt, most notably in the ileum in comparison to other sites along the gastrointestinal tract.

Conclusion

The elucidation of differential gene expression patterns between crypt and villus epithelial cell lineages in human ileal tissue provides novel insights into the molecular machinery that mediates their functions and spatial organization. Moreover, our findings establish an important framework of knowledge for future investigations of human gastrointestinal diseases.