Professor Shiga University of Medical Science Otsu, Shiga, Japan
Abstract: Phospholipids are amphiphilic molecules, consisting of a hydrophilic head group and a hydrophobic region containing two acyl chains, and are divided into classes based on the backbone and head group structures. Phospholipids, together with cholesterol, are essential structural components of mammalian cell membranes, and play important roles in numerous cellular processes, including membrane protein regulation, membrane trafficking, cell proliferation, and intracellular signaling. To elucidate the physiological and pathological roles of phospholipids, we have recently developed enzymatic fluorometric assays for quantification of all major phospholipid classes, including phosphatidylcholine (PC), phosphatidylethanolamine (PE), phosphatidylserine (PS), phosphatidic acid (PA), phosphatidylinositol (PI), phosphatidylglycerol (PG), cardiolipin (CL), and sphingomyelin (SM). These assays, using combinations of specific enzymes and a fluorogenic substrate, allow simple, sensitive and high-throughput quantification. In this study, by applying these novel assays to human intestinal Caco-2 cells, we examined the relationship of cellular phospholipid compositions with cell growth and differentiation. The functions and morphology of Caco-2 cells change with cell differentiation to form apical membranes (Fig. 1). We assessed the phospholipid class compositions of the cells and their plasma membranes, namely apical and basal membranes. Along with cell differentiation, the cellular PC, PI and SM ratios were increased, but the PE, PA and PG + CL ratios were decreased. In addition, the apical membranes contained higher ratios of PE, PA and PG + CL, but lower ratios of PC, PS, PI, SM and cholesterol than the basal membranes. Using mass spectrometry, we demonstrated the differences in the phospholipid acyl chain composition between apical membranes and basal membranes. We also showed that the mRNA expression levels of enzymes related to phospholipid synthesis and degradation changed during cell differentiation. These findings suggest that the phospholipid metabolism in Caco-2 cells is strictly controlled by cell growth and differentiation.
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