Current studies indicate that ceramide is involved in the regulation of important cell functions, namely cell growth, differentiation, and apoptosis. In the present study, the possible role of ceramide in the differentiation of neuroblastoma Neuro2a cells was investigated. The following results were obtained. (a) Ceramide content of Neuro2a cells, induced to differentiate by retinoic acid (RA) treatment rapidly increased after addition of RA, was maintained at high levels in RA-differentiated cells and returned to the starting levels with removal of RA and reversal of differentiation; under the same conditions, the sphingosine content remained unchanged. (b) After a short pulse with [H]sphingomyelin or [H]sphingosine or L-[H]serine, the metabolic formation of ceramide was markedly higher and more rapid in RA-differentiated than undifferentiated cells. (c) Inhibitors of ceramide biosynthesis (Fumonisin B1, β-chloroalanine and L-cycloserine) diminished the extent of the differentiating effect of RA and concomitantly Cer content decreased. (d) The activity of neutral sphingomyelinase increased after addition of RA, maintained high levels in RA-differentiated cells, and returned to the initial levels with removal of RA. (e) Experimental conditions that cause an elevation of ceramide content (treatment with sphingosine or ceramide or C-ceramide or bacterial sphingomyelinase) inhibited cell proliferation and stimulated neurite outgrowth; dihydro-analogues of sphingosine, ceramide, and C-ceramide had no effect on differentiation. (f) treatment with Fumonisin B1 completely inhibited sphingosine-induced differentiation. These data suggest a specific bioregulatory function of ceramide in the control of Neuro2a cell growth and differentiation and pose the general hypothesis of a mediator role of ceramide in the differentiation of cells of neural origin.

Drug resistance elicited by cancer cells still constitutes a huge problem that frequently impairs the efficacy of both conventional and novel molecular therapies. Chemotherapy usually acts to induce apoptosis in cancer cells; therefore, the investigation of apoptosis control and of the mechanisms used by cancer cells to evade apoptosis could be translated in an improvement of therapies. Among many tools acquired by cancer cells to this end, the de-regulated synthesis and metabolism of sphingolipids have been well documented. Sphingolipids are known to play many structural and signalling roles in cells, as they are involved in the control of growth, survival, adhesion, and motility. In particular, in order to increase survival, cancer cells: (a) counteract the accumulation of ceramide that is endowed with pro-apoptotic potential and is induced by many drugs; (b) increase the synthesis of sphingosine-1-phosphate and glucosylceramide that are pro-survivals signals; (c) modify the synthesis and the metabolism of complex glycosphingolipids, particularly increasing the levels of modified species of gangliosides such as 9-O acetylated GD3 (αNeu5Ac(2-8)αNeu5Ac(2-3)βGal(1-4)βGlc(1-1)Cer) or N-glycolyl GM3 (αNeu5Ac (2-3)βGal(1-4)βGlc(1-1)Cer) and de-N-acetyl GM3 (NeuNH(2)βGal(1-4)βGlc(1-1)Cer) endowed with anti-apoptotic roles and of globoside Gb3 related to a higher expression of the multidrug resistance gene MDR1. In light of this evidence, the employment of chemical or genetic approaches specifically targeting sphingolipid dysregulations appears a promising tool for the improvement of current chemotherapy efficacy.