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Basal nutrition promotes human intestinal epithelial (Caco-2) proliferation, brush border enzyme activity, and
1Department of Surgery, Yale University School of Medicine, New Haven, CT USA.
Objectives:
Provision of nutrients to the apical membrane of intestinal epithelial cells by the enteral route is critical for normal gut mucosal function and for the sheet migration required for mucosal healing. The present work attempts to determine whether supplemental nutrient delivery to the basal epithelial surface is important for intestinal epithelial biology. Since attempts to regulate intestinal epithelial cell biology by manipulation of parenteral nutrition solutions have met with some success, we hypothesized that basally delivered nutrients might also be important for intestinal epithelial biology.
Design:
To test this hypothesis, we compared the brush border enzyme activity, proliferation, and motility of human intestinal epithelial (Caco-2) cells cultured on a type I collagen substrate either on cell culture dishes with culture medium above the apical side of the cell monolayer or in culture inserts on 0.45-mu semipermeable membranes with culture medium beneath the monolayers as well as above them. Proliferation was assessed by serial hematocytometric counts over 13-day period. Doubling times were calculated by logarithmic transformation of cell counts 48 hrs apart. The specific activity of the brush border enzymes, dipeptidyl dipeptidase and alkaline phosphatase, was assayed by the digestion of synthetic chromogenic substrates in protein-matched aliquots of cell lysates. Sheet migration was quantitated by the expansion of Caco-2 monolayers across collagen. Motility was dissociated from the proliferative component of monolayer expansion by blocking proliferation with mitomycin C.
Setting:
Laboratory for gastrointestinal mucosal biology.
Subjects:
A well-differentiated subclone of cells derived from the established human Caco-2 colonic epithelial cell line.
Measurements And Main Results:
Basal nutrient delivery promoted Caco-2 proliferation, brush border enzyme activity, monolayer expansion, and cell motility. Proliferation was actually increased by 694 +/- 9.89% (n = 90, p < .0001) in cells nourished apically and basally compared with a 314 +/- 3.31% increase (n = 90, p < .0001) in those cells receiving only apical nutrition. The addition of basal nutrient delivery to the cell culture system augmented both alkaline phosphatase and dipeptidyl dipeptidase specific activity by 116 +/- 5.4% and 256 +/- 14.0%, respectively (p < .0001, n = 6 for each group). The effects of basal nutrient delivery were maintained after mitomycin blockade of proliferation for both alkaline phosphatase (392 +/- 89.8% of control, n = 3, p < .0005) and dipeptidyl dipeptidase (374 +/- 79.1% of control, n = 3, p < .005), suggesting that the increased digestive enzyme-specific activity reflected differentiation rather than indirect effects of slowing of proliferation. Epithelial sheet migration increased by 389 +/- 8.8% and proliferation-blocked cell motility also increased by 76.5 +/- 1.56% (p < .0005, n = 12 for each) compared with apical nutrient delivery only.
Conclusions:
These results suggest that although apical nutrition may be critical for intestinal epithelial cell biology, nutrient delivery to the basal surface of intestinal epithelial cell membranes may also promote intestinal epithelial differentiation, proliferation, and mucosal healing.