Differences in lipid characteristics of undifferentiated and enterocytic-differentiated HT29 human colonic cells

In this paper we compared several lipid characteristics of the homogenate and the corresponding plasma membrane in undifferentiated and differentiated HT29 human colon cancer cells, using normal human colonic cells as a reference. Electron microscopy showed that HT29 cells were morphologically undifferentiated when cultured in the presence of either glucose or inosine without glucose at early confluency. On the contrary, HT29 cells cultured at late confluency in a glucose-free medium containing inosine or grown in nude mice exhibited an enterocytic differentiation with the presence of tight junctions and an apical brush border. The cell homogenate and the plasma membrane were prepared from each cell type. The study of specific marker enzymes showed the same degree of purity in all plasma membranes, with a highly marked increase of brush border-associated hydrolases (N-aminopeptidase and alkaline phosphatase) only in the organelles isolated from differentiated HT29 and colonic cells. Respective similar increases in the amount of free cholesterol and phospholipid and in the free cholesterol:phospholipid molar ratio were found in the plasma membrane as compared with the homogenate in all HT29 cell types. This ratio, due to an increased phospholipid content in both homogenate and plasma membrane, was lowered in colonic cells. No differences in the phospholipid profile were found between the homogenates of all cell types and the plasma membrane of undifferentiated HT29 cells, with the exception of a decrease of cardiolipin in this organelle. On the contrary, the plasma membrane phospholipid composition was different from that of the corresponding homogenate in differentiated HT29 and colonic cells. The most striking changes were a highly increased sphingomyelin amount and concomitant decreases in phosphatidylethanolamine, phosphatidylserine, and cardiolipin. Moreover, differences in the percentage of phosphatidylcholine plus sphingomyelin as well as in phosphatidylcholine:sphingomyelin, phosphatidylethanolamine, and/or phosphatidylcholine molar ratios were also found. The monounsaturated:polyunsaturated fatty acid ratio in phosphatidylethanolamine was similar in differentiated HT29 and colonic cells and lower than in undifferentiated HT29 cells. A decrease in this latter ratio in phosphatidylcholine was also observed in colonic cells and HT29 cells grown in nude mice. These changes were essentially due to opposite variations in the percentage of palmitoleic acid and those of linoleic and/or arachidonic acids in both phospholipids. Thus, these data indicate that undifferentiated HT29 cells were characterized by the absence of a specific phospholipid composition in their plasma membrane, which is suggested to be related to altered phospholipid sorting. The plasma membrane phospholipid profile reversed essentially to the normal pattern when HT29 cells recovered the ability to differentiate.