Does small intestinal atresia affect epithelial protein expression in human newborns?

OBJECTIVES: Bowel segments distal to a congenital intestinal obstruction have been suggested to be immature. In other words, luminal components such as amniotic fluid (before birth) and/or enteral nutrition (after birth) may be required to activate intestinal epithelial protein expression, thereby influencing epithelial differentiation. We investigated cell-type-specific protein expression proximal and distal to jejunal and ileal atresias in human newborns. PATIENTS AND METHODS: We immunohistochemically studied intestinal tissue specimens of 16 newborns who had undergone surgery for jejunal or ileal atresia. Sections were taken from both the proximal and distal sides of the atresias. RESULTS: For all patients, the enterocyte-specific markers lactase, sucrase-isomaltase, sodium glucose cotransporter 1, glucose transporters 2 and 5, intestinal fatty acid-binding protein and alkaline phosphatase were expressed at a mean 3 +/- 1 days after birth, both proximal and distal to jejunal and ileal atresias. Expression of goblet cell-specific markers mucin 2 and trefoil factor 3 and that of the Paneth cell marker lysozyme was maintained at either side of the atretic segment. CONCLUSIONS: With respect to the markers used, the human small intestinal epithelium is already differentiated shortly after birth. The absence of intestinal continuity in case of a jejunal or ileal atresia does not affect epithelial protein expression. This would seem to indicate that the developing small intestinal epithelium matures independently of luminal components.

Specific responses in rat small intestinal epithelial mRNA expression and protein levels during chemotherapeutic damage and regeneration.

The rapidly dividing small intestinal epithelium is very sensitive to the cytostatic drug methotrexate. We investigated the regulation of epithelial gene expression in rat jejunum during methotrexate-induced damage and regeneration. Ten differentiation markers were localized on tissue sections and quantified at mRNA and protein levels relative to control levels. We analyzed correlations in temporal expression patterns between markers. mRNA expression of enterocyte and goblet cell markers decreased significantly during damage for a specific period. Of these, sucrase-isomaltase (-62%) and CPS (-82%) were correlated. Correlations were also found between lactase (-76%) and SGLT1 (-77%) and between I-FABP (-52%) and L-FABP (-45%). Decreases in GLUT5 (-53%), MUC2 (-43%), and TFF3 (-54%) mRNAs occurred independently of any of the other markers. In contrast, lysozyme mRNA present in Paneth cells increased (+76%). At the protein level, qualitative and quantitative changes were in agreement with mRNA expression, except for Muc2 (+115%) and TFF3 (+81%), which increased significantly during damage, following independent patterns. During regeneration, expression of each marker returned to control levels. The enhanced expression of cytoprotective molecules (Muc2, TFF3, lysozyme) during damage represents maintenance of goblet cell and Paneth cell functions, most likely to protect the epithelium. Decreased expression of enterocyte-specific markers represents decreased enterocyte function, of which fatty acid transporters were least affected.

Epithelial proliferation, cell death, and gene expression in experimental colitis: alterations in carbonic anhydrase I, mucin MUC2, and trefoil factor 3 expression.

BACKGROUND AND AIMS: To gain insight in intestinal epithelial proliferation, cell death, and gene expression during experimental colitis rats were treated with dextran sulfate sodium (DSS) for 7 days. MATERIALS AND METHODS: Proximal and distal colonic segments were excised on days 2, 5, 7, and 28. Epithelial proliferation, cell death, enterocyte gene expression (carbonic anhydrase I (CA I) and goblet cell gene expression (mucin, MUC2; trefoil factor 3, TFF3) were studied immunohistochemically and biochemically. RESULTS: Proliferative activity was decreased in the proximal and distal colon at the onset of disease (day 2). However, during active disease (days 5-7) epithelial proliferation was increased in the entire proximal colon and in the proximity of ulcerations in the distal colon. During DSS treatment the number of apoptotic cells in the epithelium of both colonic segments was increased. In the entire colon surface enterocytes became flattened and CA I negative during active disease (day 5-7). Additionally, CA I levels in the distal colon significantly decreased during this phase. In contrast, during the regenerative phase (day 28) CA I levels were restored in the distal colon and up-regulated in the proximal colon. During all disease phases increased numbers of goblet cells were observed in the surface epithelium of the entire colon. In the distal colon TFF3 expression extended to the bottom of the crypts during active disease. Finally, MUC2 and TFF3 expression was increased in the proximal colon during disease. CONCLUSION: DSS affected the epithelium by inhibiting proliferation and inducing apoptosis. DSS-induced inhibition of CA I expression indicates down-regulation of specific enterocyte functions. Accumulation of goblet cells in the surface epithelium and up-regulation of MUC2 and TFF3 expression in the proximal colon underline the importance of goblet cells in epithelial protection and repair, respectively.

Distinct epithelial responses in experimental colitis: implications for ion uptake and mucosal protection.

In the present study, we aimed to investigate enterocyte- and goblet cell-specific functions during the different phases of acute colitis induced with dextran sulfate sodium (DSS). Rats were treated with DSS for 7 days, followed by a 7-day recovery period. Colonic tissue was excised on days 2 (onset of disease), 7 (active disease), and 14 (regenerative phase). Enterocyte functions were studied by the expression of carbonic anhydrases (CAs), sodium/hydrogen exchangers (NHEs) and intestinal fatty acid-binding protein (iFABP) and by alkaline phosphatase (AP) activity. The expression and secretion of the mucin Muc2 and trefoil factor family peptide-3 (TFF3) were used as parameters for goblet cell function. DSS induced a downregulation of the CAs, NHEs, and iFABP in some normal-appearing surface enterocytes and in most of the flattened-surface enterocytes during disease onset and active disease. During the regenerative phase most enterocytes expressed these genes again. Quantitative analysis revealed a significant decrease in CAs, NHEs, and iFABP expression levels during onset and active disease. During the regenerative phase, the expression levels of the CAs were restored, whereas the expression levels of the NHEs and iFABP remained decreased. In contrast, enterocyte-specific AP activity was maintained in normal and flattened enterocytes during DSS-induced colitis. Goblet cells continued to express MUC2 and TFF3 during and after DSS treatment. Moreover, Muc2 and TFF3 expression and secretion levels were maintained or even increased during each of the DSS-induced disease phases. In conclusion, DSS-induced colitis was associated with decreased expression of CAs, NHEs, and iFABP. The loss of these genes possibly accounts for some of the pathology seen in colitis. The maintenance or upregulation of Muc2 and TFF3 synthesis and secretion levels implies that goblet cells at least maintain their epithelial defense and repair capacity during acute inflammation induced by DSS.

Alterations in Muc2 biosynthesis and secretion during dextran sulfate sodium-induced colitis.

To gain insight into mucin 2 (Muc2) synthesis and secretion during dextran sulfate sodium (DSS)-induced colitis, rats were treated with DSS for 7 days. Colonic segments were excised on days 0 (control), 2 (onset of disease), 7 (active disease), and 14 (regenerative phase) for histological evaluation. Explants were metabolically labeled with (35)S-labeled amino acids or [(35)S]sulfate followed by chase incubation. Homogenates were analyzed by SDS-PAGE and (35)S-labeled Muc2 was quantified. Also, total Muc2 protein and mRNA were quantified. DSS-induced crypt loss, ulcerations, and concomitant goblet cell loss were most pronounced in the distal colon. Muc2 precursor synthesis increased progressively in the proximal colon but was unaltered in the distal colon during onset and active disease. During the regenerative phase, Muc2 precursor synthesis levels normalized in the proximal colon but increased in the distal colon. Total Muc2 levels paralleled the changes seen in Muc2 precursor synthesis levels. During each disease phase, total Muc2 secretion was unaltered in the proximal and distal colon. [(35)S]sulfate incorporation into Muc2 only decreased in the proximal colon during active disease and the regenerative phase, whereas secretion of [(35)S]sulfate-labeled Muc2 increased. During the regenerative phase, Muc2 mRNA levels were downregulated in both colonic segments. In conclusion, DSS-induced loss of goblet cells was accompanied by an increase or maintenance of Muc2 precursor synthesis, total Muc2 levels, and Muc2 secretion. In the proximal colon, Muc2 became undersulfated, whereas sulfated Muc2 was preferentially secreted. Collectively, these data suggest specific adaptations of the mucus layer to maintain the protective capacities during DSS-induced colitis.