The sphingomyelin pathway has been implicated in mediating the effect of several extracellular agents leading to important biochemical and cellular changes. The aim of this investigation is to study interleukin-1 beta (IL-1 beta) signaling in oligodendrocytes. For this purpose, the CG4 oligodendrocyte cells were differentiated and incubated with IL-1 beta. This treatment induced a time- and dose-dependent increase of the endocellular ceramide. To mimic the effect of the elevation of endogenous ceramide, the CG4 cells were treated with the ceramide analogue C2-ceramide. Cell survival, measured with the MTT assay, showed that, by increasing the concentration of ceramide, up to 40% of CG4 cells were dying within 6 h, similar data were obtained with the primary differentiated oligodendrocytes. Condensation of chromatin, nuclear fragmentation, and formation of apoptotic bodies indicated that apoptosis was the cause of death. Surprisingly, long-term exposure (72 h) to increasing concentrations of IL-1 beta, which increases intracellular ceramide, did not induce oligodendroglial cell death. These results show that an increase of intracellular ceramide is not sufficient to induce apoptosis in oligodendrocytes and that IL-1 beta signaling through the ceramide pathway in these cells can mediate functions other than programmed cell death.

The expression of a molecule recognized by anti-galactosyl ceramide antibodies (MAb) O1 on the surface membrane of human spermatozoa was investigated by biochemical and immunochemical methods. Indirect immunofluorescence shows that this molecule is preferentially localized on the middle piece of the sperm tail. Immuno-thin-layer chromatography has identified it as a glycolipid related but not identical to galactosylceramide. Consistent with a structure similar to galactosylceramide, the sperm glycolipid is capable of binding gp120. An improved ELISA has been utilized to demonstrate the specificity of binding of the antibodies and gp120 to the isolated lipid fraction. Identity of the binding site of the two ligands to the glycolipid is suggested by competition assays. On the basis of preliminary biochemical analysis this glycolipid was tentatively classified as a galactosylalkylacylglycerolipid (GalAAG), the nonsulfated form of the seminolipid, a glycolipid known to be present in the testis and germ cells of mammals. These data indicate that human sperm express a glycolipid similar in structure to the receptor for HIV described on the CD4- neural and colonic epithelial cell lines, and moreover suggest that this glycolipid could also function as HIV receptor and possibly be implicated in its transmission.

The RT-PCR analysis of RNA from progenitor and differentiated primary rat oligodendrocytes, and from the oligodendrocyte CG-4 cell line, shows the presence of the IL-1beta mRNA, the type I IL-1beta receptor and the IL-1 receptor accessory protein in these cells. In situ hybridization of a rat IL-1beta probe to primary progenitor and differentiated rat oligodendrocytes results in a positive signal. The double hybridization of the IL-1beta probe, together with an oligodendrocyte-specific differentiation marker, to sections of postnatal rat brain at different stages of differentiation is also positive. The double immuno-labelling technique utilized indicates coincidence of the signals on the brain slices. The results show that IL-1beta mRNA is constitutively expressed in rat brain oligodendrocytes from 1 day after birth onward. In agreement with this observation, CG-4 cells, primary progenitor and differentiated rat oligodendrocytes are positively stained by antibodies against IL-1beta. Postnatal brain slices from 1 and 4 day old and adult rats, labelled with a double immunofluorescence technique, are also stained by antibodies against IL-1beta. This signal coincides with that of antibodies against oligodendrocyte-specific surface markers. We conclude that IL-1beta is constitutively expressed in rat brain progenitor and differentiated oligodendrocytes.

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