Vitamin D-mediated intestinal calcium transport: Effects of essential fatty acid deficiency and spin label studies of enterocyte membrane lipid fluidity

Abstract

Vitamin D-3 and its metabolites regulate the transport of calcium across the intestinal epithelial cell via a mechanism which is as yet unknown. The purpose of this study was to evaluate the effect of an essential fatty acid deficiency on vitamin D-stimulated intestinal calcium transport as measured by both in vivo and in vitro techniques. We also describe in this report a procedure for the isolation of chick intestinal epithelial cell brush border and basal lateral membranes and an assessment of the effect of dietary vitamin D on the lipid fluidity of these membranes. An essential fatty acid deficiency in both vitamin D-replete and deficient chicks resulted in a decrease in intestinal mucosal levels of linoleic acid, with a compensatory increase in the levels of the short chain fatty acid, myristic acid, and the unsaturated fatty acids, palmitoleic and eicosatrienoic acids. An essential fatty acid deficiency did not affect the ability of vitamin D-deficient chicks to respond to vitamin D with a 2-fold increase in serum calcium and a 4–5-fold increase in intestinal calcium transport, measured in vivo. However, an essential fatty acid deficiency resulted in an inability of vitamin D to increase calcium flux in vitamin D-deficient chick ileum as measured under in vitro conditions. Dietary vitamin D resulted in no detectable change in the protein composition in either the brush border or basal lateral membranes as evidenced by SDS-polyacrylamide electrophoresis. In addition, vitamin D did not alter the levels of brush border membrane cholesterol or lipid phosphorus (0.27 ± 0.03 and 0.19 ± 0.01 μmol/mg protein, respectively). Brush border and basal lateral membranes were labeled with the 5-nitroxide stearate spin probe I(12,3). The polarity of the environment of the probe in the brush border membranes is much greater than that of the basal lateral membranes. In addition, the lipid environment of the brush border membrane is much less fluid ( S = 0.650) than that of the basal lateral membrane ( S = 0.583). The data concerning membrane lipid fluidity is qualitatively similar to fluorescence polarization studies of rat intestinal epithelial cell membranes and confirms the concept that a given cell may contain plasma membrane regions having discrete lipid structures/fluidities. Dietary vitamin D had no detectable effect on the lipid fluidity or polarity in either the brush border or basal lateral membranes. The results do not support a role for an alteration in essential fatty acid composition or gross changes in the lipid fluidity of the brush border or basal lateral membranes as mechanisms by which vitamin D regulates intestinal calcium transport.