Endocytosis of cholera toxin by human enterocytes is developmentally regulated

Abstract

Many secretory diarrheas including cholera are more prevalent and fulminant in young infants than in older children and adults. Cholera toxin (CT) elicits a cAMP-dependent chloride secretory response in intestinal epithelia, which accounts for the fundamental pathogenesis of this toxigenic diarrhea. We have previously reported that the action of this bacterial enterotoxin is excessive in immature enterocytes and under developmental regulation. In this study, we tested the hypothesis that enhanced endocytosis by immature human enterocytes may, in part, account for the excessive secretory response to CT noted in the immature intestine and that enterocyte endocytosis of CT is developmentally regulated. To test this hypothesis, we used specific inhibitors to define endocytic pathways in mature and immature cell lines. We showed that internalization of CT in adult enterocytes is less and occurs via the caveolae/raft-mediated pathway in contrast to an enhanced immature human enterocyte CT uptake that occurs via a clathrin pathway. We also present evidence that this clathrin pathway is developmentally regulated as demonstrated by its response to corticosteroids, a known maturation factor that causes a decreased CT endocytosis by this pathway.

Hydrocortisone induces changes in gene expression and differentiation in immature human enterocytes

“…H4 is a previously characterized primary human fetal small intestinal cell line that we used in numerous studies defining the role of human enterocytes in the development of intestinal host defenses (8,9,13,19,20). T84, a well-characterized, human colon carcinoma cell line that comprises highly differentiated crypt-like cells, has been used as a mature human intestinal model (8,20,28). H4 cells were routinely maintained in Dulbecco's modified Eagles medium (DMEM) supplemented with 10% FBS and human recombinant insulin (0.5 U/ml).…”

“…33). Among many endogenous biological mediators, hydrocortisone (HC), epidermal growth factor, and transforming growth factor-␤ (TGF-␤) promote growth and differentiation (34), modulate the inflammatory response (9), and reduce an excessive secretory response to cholera toxin in human and rat immature enterocytes (7,19,20).…”

Function of the intestinal epithelium and its dysregulation in inflammatory bowel disease

“…9 These complex pathways are possibly maturity-dependent, with increased endocytosis demonstrated within fetal enterocytes. 100 During pinocytosis the solubility of the antigen, regardless of size, has been found to be most important, with kinetic studies showing that the IEC has significantly slower uptake than professional APCs. 101 IECs are also able to take up antigen by receptormediated endocytosis, delivering molecules to endolysomal compartments, or for transport of intact macromolecules across the cell.…”

ADP-ribosylation factors regulate the development of CT signaling in immature human enterocytes

“…Recently we have extended these observations to experimental models of human intestinal development (23,25). In these studies, we have also provided evidence that CT induces an enhanced secretory reaction mediated in part by a developmental upregulation of the cAMP response in immature vs. mature human enterocytes (23), and this enhanced response to CT is largely due to an excessive uptake of CT by a developmentally regulated clathrin-endocytic pathway (25).…”