Caco-2 Cell Iron Uptake from Meat and Casein Digests Parallels in Vivo Studies: Use of a Novel in Vitro Method for Rapid Estimation of Iron Bioavailability

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Abstract
We developed a model for assessing iron bioavailability from foods which combines simulated peptic and intestinal digestion with measurement of iron uptake by Caco-2 cell monolayers. Our objective was to further validate this model by determining if meat enhances Caco-2 cell iron uptake relative to casein. Caco-2 cell monolayers were covered with Hank's balanced salt solution (HBSS) buffered with HEPES, pH 7.4. An upper chamber was created over the cells by fitting the bottom of a Costar Transwell® insert with a 12,000–14,000 molecular weight cut-off dialysis membrane. This membrane allowed low molecular weight iron complexes to diffuse into the media bathing the cells and prevented damage to the cells from the digestive enzymes. Prior to digestion, each sample (homogenate of beef, chicken, fish or casein) was mixed with 59FeCl3 to achieve an iron concentration of 10 μmol/L. Following pepsin digestion (pH 2), pH was adjusted to 7.4, pancreatic enzymes and bile extract were added to each digest, and an aliquot was then introduced into the upper chamber of the culture dish. During this intestinal digestion period, 59Fe uptake occurred from iron that dialyzed into the lower chamber. The 59Fe uptake from beef, chicken and fish digests was 300–400% of the 59Fe uptake from a casein digest. Our results parallel human absorption studies indicating that meat enhances iron absorption. The results suggest that digestion products of the meat proteins were at least partially responsible for the enhancement of iron uptake. Overall, this study supports the usefulness of our model as a means of assessing iron bioavailability.