Lipid, apolipoprotein, and lipoprotein synthesis and secretion during cellular differentiation in caco-2 cells

Abstract

Although Caco-2 cells are frequently employed for the study of enterocyte lipid metabolism, variable results have been reported regarding their ability to synthesize and secrete lipids and apolipoproteins. The major goal of this investigation is to examine the capacity of Caco-2 cells to elaborate and secrete lipids, lipoproteins, and apolipoproteins at different degrees of morphological and functional differentiation. Cells were cultured in medium with 5% fetal bovine serum (FBS), on permeable polycarbonate filters from 2 to 30 d in the presence of 14C-oleate or 35S-methionine. Cellular differentiation, as assessed by morphology (light and electron microscopy), transepithelial resistance, free fatty acid flux, and sucrase activity, progressed steadily up to 20 d of culture. Caco-2 cells esterified oleic acid mainly into phospholipids, triglycerides (TG), and smaller amounts of cholesterol esters. Lipid synthesis began as early as 2 d, and TG secretion was enhanced with increased duration of culture. However, very low efficiency of lipid export was observed at all levels of differentiation, reaching a maximum of only 6% of intracellular lipids. VLDL and LDL were the dominant lipoproteins secreted, with HDL comprising < 20% of the total. VLDL secretion increased, while LDL decreased, whereas the lipid composition of lipoproteins varied little with increasing duration of culture. Apoprotein B and A-I synthesis and secretion increased markedly from 11 to 20 d of culture. The ratio of apo B-100/B-48 decreased between 11 and 30 d, consistent with enhanced apo B editing of more mature enterocytes. Taken together, our data suggest that from 20 d of culture, Caco-2 cells are morphologically and functionally mature, capable of lipid esterification, and lipoprotein and apolipoprotein synthesis. However, despite their functional and morphological similarities to mature enterocytes, Caco-2 cells have a very limited lipid export capacity.

Gene Expression Profiling of Systems Involved in the Metabolism and the Disposition of Xenobiotics: Comparison between Human Intestinal Biopsy Samples and Colon Cell Lines

“…In the differentiated state, they mimic typical characteristics of the human small intestinal epithelium, like a well developed brush border with associated enzymes such as alkaline phosphatase and sucrase isomaltase. Nevertheless, the Caco-2 cell model is different from the small intestine in several aspects, and the phenotype of Caco-2 cells is dependent on the time in culture (Mehran et al, 1997;Engle et al, 1998). Of interest, our results reveal a relatively good correlation between Caco-2 cells and human colon with a Spearman correlation coefficient of 0.7.…”

Effect of retinoic acid on cell proliferation and differentiation as well as on lipid synthesis, lipoprotein secretion, and apolipoprotein biogenesis

“…The inserts were placed into six-well culture plates, permitting separate access to the upper and lower compartments of the monolayers. Cells were cultured for various periods, including 21 days, at which the Caco-2 cells are highly differentiated and appropriate for lipid synthesis (44,46). The medium was refreshed every second day.…”

“…The medium was refreshed every second day. 9-cis RA was then added to cells at different concentrations for 24 h. Sucrase activity was measured as a marker of cell differentiation and transepithelial resistance as a marker of monolayer integrity (Millipore, Bedford, MA) (44,46). Cell viability was assessed by Trypan blue exclusion (46).…”

Localization of the Lipid Receptors CD36 and CLA-1/SR-BI in the Human Gastrointestinal Tract: Towards the Identification of Receptors Mediating the Intestinal Absorption of Dietary Lipids

“…Probes for human CD36 and CLA-1/SR-BI were as described (Calvo et al 1997). For apoE, a 0.4-kb EcoRI-BamHI fragment derived from the fourth exon was used as a probe (Mehran et al 1997).…”