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Multidrug-resistant tumor cells display enhanced levels of glucosylceramide. In this study, we investigated how this relates to the overall sphingolipid composition of multidrug-resistant ovarian carcinoma cells and which mechanisms are responsible for adapted sphingolipid metabolism. We found in multidrug-resistant cells substantially lower levels of lactosylceramide and gangliosides in sharp contrast to glucosylceramide, galactosylceramide, and sphingomyelin levels. This indicates a block in the glycolipid biosynthetic pathway at the level of lactosylceramide formation, with concomitant accumulation of glucosylceramide. A series of observations exclude regulation at the enzyme level as the underlying mechanism. First, reduced lactosylceramide formation occurred only in intact resistant cells whereas cell-free activity of lactosylceramide synthase was higher compared with the parental cells. Second, the level of lactosylceramide synthase gene expression was equal in both phenotypes. Third, glucosylceramide synthase (mRNA and protein) expression and activity were equal or lower in resistant cells. Based on the kinetics of sphingolipid metabolism, the observation that brefeldin A does not restore lactosylceramide synthesis, and altered localization of lactosylceramide synthase fused to green fluorescent protein, we conclude that lactosylceramide biosynthesis is highly uncoupled from glucosylceramide biosynthesis in the Golgi apparatus of resistant cells.

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In the present study, we evaluated whether lymphocyte cytokine production during human and rat pregnancy shifts toward T helper cell type 2 (Th2) cytokine production. Therefore, blood samples were taken during the follicular and luteal phase and during pregnancy in rats and humans. Whole blood was ex vivo-stimulated with phorbol 12-myristate 13-acetate and calcium ionophore and intracellular interferon-␥ (IFN-␥) and interleukin (IL)-4 production, and the percentage of cells in the various lymphocyte populations was measured using flow cytometry. Rats and humans adapted their immune responses to pregnancy but have different strategies: During human pregnancy, the percentage of lymphocytes producing IFN-␥ was decreased, and the percentage IL-4-producing lymphocytes was not affected. The rat adapts its immune response to pregnancy by decreasing the total number of the various lymphocyte populations, and the percentage of IFN-␥-or IL-4-producing lymphocytes was not affected or increased (% IFN-␥-producing cytotoxic lymphocytes). It is speculated that during rat pregnancy, there is no need to decrease the number of IFN-␥-producing lymphocytes, as in nonpregnant rats, the total number of IFN-␥-producing lymphocytes after stimulation is relatively low, and there is no necessity for a further decrease. In nonpregnant humans, the percentage IFN-␥-producing lymphocytes is much higher and probably dangerous for pregnancy, and therefore, this percentage needs to decrease during pregnancy. In conclusion, although the data from humans concur with the Th1/ Th2 paradigm, the data from rats do not concur with this paradigm. The present studies therefore challenge the classical Th1/Th2 paradigm during pregnancy. J. Leukoc. Biol. 78: 946 -953; 2005.

The present study shows that ATP infusion induced a pro-inflammatory response leading to glomerular albuminuria exclusively in the pregnant rat. Why extracellular ATP showed this pro-inflammatory response exclusively in the pregnant condition is unclear but is probably related with relatively enhanced non-specific immunity and inflammatory reactions characteristic for the pregnant condition.