Involvement of Sphingosine 1-Phosphate in Nerve Growth Factor-Mediated Neuronal Survival and Differentiation

Abstract

Sphingolipid metabolites, such as ceramide and sphingosine-1-phosphate (SPP), are emerging as a new class of second messengers involved in cellular proliferation, differentiation, and apoptosis. Nerve growth factor (NGF), a neurotrophic factor for pheochromocytoma PC12 cells, induced a biphasic increase in the activity of sphingosine kinase, the enzyme that catalyzes the formation of SPP. This activation was blocked by K252a, an inhibitor of tyrosine kinase A (trkA). A rapid 1.7-fold increase was followed by a marked prolonged increase reaching a maximum of fourfold to fivefold stimulation with a concomitant increase in SPP levels and a corresponding decrease in endogenous sphingosine levels. Levels of ceramide, the precursor of sphingosine, were only slightly decreased by NGF in serumcontaining medium. However, NGF decreased the elevation of ceramide induced by serum withdrawal. Treatment of PC12 cells with SPP did not induce neurite outgrowth or neurofilament expression, yet it enhanced neurofilament expression elicited by suboptimal doses of NGF. Moreover, SPP also protected PC12 cells from apoptosis induced by serum withdrawal. To further substantiate a role for SPP in the cytoprotective actions of NGF, we found that N,N-dimethylsphingosine, a competitive inhibitor of sphingosine kinase, also induced apoptosis and interfered with the survival effect of NGF. These effects were counteracted by exogenous SPP. Moreover, other structurally related compounds, such as dihydrosphingosine 1-phosphate and lysophosphatidic acid, had no significant protective effects. Our results suggest that activation of sphingosine kinase and subsequent formation of SPP may play an important role in the differentiation and survival effects induced by NGF.

Enhancement of radiosensitivity by combined ceramide and dimethylsphingosine treatment in lung cancer cells

“…On the basis of these findings, one can speculate that combining radiotherapy with the regulation of endogenous ceramide levels may have synergistic effects on selected tumors. Thus, modifications in the sphingomyelin/ceramide signaling pathway are expected to be a potential tool to increase the therapeutic efficiency of radiation treatment of tumors (Chmura et al, 1997;Edsall et al, 1997Edsall et al, , 1998Susin et al, 1997;Cuvillier et al, 1998;Kleuser et al, 1998;Kolesnick et al, 1998;Zhivotovsky et al, 1999). Many inhibitors of ceramide synthesis such as DL-PDMP (DL-threo-1-phenyl 2-decanoyl amino-3-morpholino-1-propanol․ HCl), B13, and imipramine have already been tried in humans and animals.…”

“…In addition, the treatment with exogenous short chain homologues of ceramide mimics endogenously generated ceramide and induces apoptosis. However, the mechanisms that convey activation signals to the enzymes responsible for ceramide production are poorly defined, and the pivotal executioner of ceramide to the apoptotic response still remains arguable (Chmura et al, 1997;Susin et al, 1997;Edsall et al, 1997Edsall et al, , 1998Cuvillier et al, 1998;Kleuser et al, 1998;Kolesnick et al, 1998).…”

Sphingosine 1-phosphate as a therapeutic agent

“…27 Intracellular S1P mobilizes Ca 2+ from internal stores independently of inositol trisphosphate, 24,28 as well as triggering diverse signaling pathways that lead to cell proliferation 29,30 and suppression of apoptosis. 11,12,23,30,31 Forced expression of SPHK1 in NIH 3T3 fibroblasts increased the proportion of cells in S phase of the cell cycle by promoting the G 1 -S transition; it also reduced the doubling time of these cells, an effect that was especially marked under low-serum conditions, indicating that intracellular S1P may be an important regulator of cell growth. 32 Furthermore, overexpression of human SPHK1 in NIH 3T3 fibroblasts resulted in their acquisition of a transformed phenotype, as determined by assays of focus formation, colony growth in soft agar, and the ability to form tumors in NOD/SCID mice, 33 suggesting that the corresponding wild-type gene may act as an oncogene.…”

“…11,12,14,23,31,52 Although the intracellular targets of S1P remain to be identified, several lines of evidence support the second messenger role of S1P: (1) dihydro-S1P binds to and activates all of the identified S1PRs, but it does not mimic all of the effects of S1P, especially those related to cell survival; 30 (2) yeast cells do not possess GPCRs, yet the abundance of phosphorylated long-chain sphingoid bases regulates environmental stress responses, cell proliferation, and cell survival in a manner reminiscent of the function of S1P in mammalian cells; (3) in plants, which do not express S1PRs, S1P regulates Ca 2+ homeostasis and ion channels; 53 and (4) microinjection of S1P into, or photolysis of caged S1P within, mammalian cells induces both Ca 2+ mobilization 54 and cell proliferation. 30 S1P has been implicated as a second messenger in cellular proliferation and survival, 10 as well as in the protection against ceramide-mediated apoptosis.…”

M1 muscarinic receptors block caspase activation by phosphoinositide 3-kinase- and MAPK/ERK-independent pathways

“…67 Moreover, the MAPK/ERK and the PI 3-kinase pathways were not essential for the survival effect of NGF on apoptosis induced by ceramide in PC12 cells. 64 One possible signaling pathway candidate for mediating the muscarinic effect on caspases might be sphingosine-1-phosphate, which was shown to play an important role in the survival effect of NGF on PC12 cells, 68 and can be induced by at least the m2 and m3 muscarinic receptor subtypes. 69 The finding that inhibition of the PI 3-kinase pathway partially attenuates the muscarinic survival effect on the viability of the cells but not on caspase inhibition, raises the question of the mechanism whereby these cells die under these conditions.…”