[55] Inhibitors of cerebroside metabolism

Radin, Norman S.; Vunnam, Ranga R.

doi:10.1016/s0076-6879(81)72057-3

Cited by 29 publications

(14 citation statements)

References 16 publications

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“…We now observe that C 6 -ceramide induces endogenous ceramide generation, likely occurring due to recycling of the C 6 -ceramide sphingosine backbone, which requires the catalytic activity of ceramide synthase, as was recently demonstrated by Ogretmen et al (21). On the other hand, PDMP has been established as an inhibitor of ceramide glucosyltransferase and therefore inhibits ceramide metabolism (24). H 2 O 2 , C 6 -ceramide, PDMP, or GCS was utilized to target these three enzymatic groups, respectively, and each successfully modulated ceramide accumulation and apoptosis in our cell system.…”

Section: Discussionmentioning

confidence: 99%

“…1C). PDMP potently prevents the synthesis of glucosylceramide from uridine diphosphate-glucose and ceramide by inhibiting UDP-glucose-ceramide glucosyltransferase, thereby elevating endogenous ceramide levels (24). Figure 1, A and B, demonstrates TLC separation of 32 P-labeled ceramide extracted from cells treated with C 6 -ceramide.…”

Section: Ceramide Elevation Induces Apoptosis In A549 Cellsmentioning

confidence: 99%

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Ceramide accumulation precedes caspase-3 activation during apoptosis of A549 human lung adenocarcinoma cells

Ravid

Tsaba

Gee

et al. 2003

American Journal of Physiology-Lung Cellular and Molecular Phys

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Ceramide, the basic structural unit of sphingolipids, controls the balance between cell growth and death by inducing apoptosis. We have previously shown that accumulation of ceramide, triggered by hydrogen peroxide (H2O2) or by short-chain ceramide analogs, induces apoptosis of lung epithelial cells. Here we elucidate the link between caspase-3 activation, at the execution phase, and ceramide accumulation, at the commitment phase of apoptosis in A549 human lung adenocarcinoma cells. The induction of ceramide accumulation by various triggers of ceramide generation, such as H2O2, C6-ceramide, or UDP-glucose-ceramide glucosyltransferase inhibitordl-threo-1-phenyl-2-decanoylamino-3-morpholino-1-propanol, triggered the activation of caspase-3. This ceramide elevation also induced the cleavage of the death substrate poly(ADP-ribose) polymerase and was followed by apoptotic cell death. Ceramide-mediated apoptosis was blocked by a general caspase inhibitor, Boc-d-fluoromethylketone, and by overexpression of the antiapoptotic protein Bcl-2. Notably, overexpression of Bcl-2 reduced the basal cellular levels of ceramide and prevented the induction of ceramide generation by C6-ceramide, which implies ceramide generation as a possible target for the antiapoptotic effects of Bcl-2.

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Section: Discussionmentioning

confidence: 99%

Section: Ceramide Elevation Induces Apoptosis In A549 Cellsmentioning

confidence: 99%

Ceramide accumulation precedes caspase-3 activation during apoptosis of A549 human lung adenocarcinoma cells

Ravid

Tsaba

Gee

et al. 2003

American Journal of Physiology-Lung Cellular and Molecular Phys

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“…It has been previously demonstrated that the synthesis of GlcCer from Cer is required to sustain both normal axonal growth (4,5,13) and also axonal growth stimulated by growth factors (17) in hippocampal neurons; these studies were performed using FB 1 , an inhibitor of (DH)-Cer synthesis (32), and PDMP, an inhibitor of GlcCer synthesis (33,34). Incubation with either inhibitor at 48 h in culture resulted in a decrease in the length of the axonal plexus and a reduction in the number of axonal branch points compared with untreated cells at 72 h in culture.…”

Section: -Msdh-cer Reduces the Mass Of Totalmentioning

confidence: 99%

1-Methylthiodihydroceramide, a Novel Analog of Dihydroceramide, Stimulates Sphinganine Degradation Resulting in Decreased de Novo Sphingolipid Biosynthesis

Echten‐Deckert¹,

Giannis²,

Schwarz³

et al. 1998

Journal of Biological Chemistry

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1-Methylthiodihydroceramide (10 M) decreased de novo ceramide biosynthesis by about 90% in primary cultured cerebellar neurons. Accordingly, de novo formation of sphingomyelin and of glycosphingolipids, all of which contain ceramide in their backbone, was reduced in a time-and concentration-dependent manner by up to 80%. Complex sphingolipid synthesis was restored upon addition of dihydroceramide or ceramide, in micromolar concentrations, to the culture medium, suggesting that none of the glycosyltransferases involved in glycosphingolipid biosynthesis is inhibited by this analog. Assays of the enzymes catalyzing sphinganine biosynthesis, as well as its N-acylation to form dihydroceramide, revealed that they were also not affected. In contrast, there was a 2.5-fold increase in the activity of sphinganine kinase. Reduction of de novo sphingolipid biosynthesis by 1-methylthiodihydroceramide is therefore due to its ability to deplete cells of newly formed free sphinganine. As a consequence of depletion of sphinganine levels, 1-methylthiodihydroceramide disrupted axonal growth in cultured hippocampal neurons in a manner similar to that reported for direct inhibitors of sphingolipid synthesis; thus, there was essentially no axon growth after incubation with 1-methylthiodihydroceramide between days 2 and 3, and co-incubation with short acyl chain analogs of ceramide (5 M) antagonized the inhibition of growth. Interestingly, the D-erythro and the L-threo isomere were equally effective, but the corresponding free base as well as other structurally related compounds did not affect either sphingolipid biosynthesis or neuronal growth.

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“…FB 1 , an inhibitor of dihydroceramide synthase (Merrill et al 1996) was obtained from the Division of Food Science and Technology (CSIR, Pretoria, South Africa). d -Threo-1-phenyl-2-decanoylamino-3-morpholino-1-propanol (PDMP), an inhibitor of glucosylceramide synthase (Radin and Vunnam 1981) was from Matreya (Pleasant Gap, PA). FB 1 and PDMP were dissolved in Hepes buffer (20 mM, pH 7.4) and added to cultures to give a final concentration of 10 M and 50 M, respectively (Schwarz et al 1995).…”

Section: Cell Culture and Reagentsmentioning

confidence: 99%

Determination of the Localization of Gangliosides Using Anti-ganglioside Antibodies: Comparison of Fixation Methods

Schwarz

Futerman

1997

J Histochem Cytochem.

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SUMMARYMany studies have examined the localization of gangliosides using anti-ganglioside antibodies, although widely differing conclusions have been reached. We now demonstrate that the apparent localization of gangliosides can be greatly influenced by the fixation method. Using monoclonal antibody (MAb) A2B5 (which reacts with a variety of gangliosides), hippocampal neurons were labeled at the cell surface when incubated with the antibody before fixation, but when incubated after fixation the cells displayed a variety of labeling patterns, depending on the fixation method. Biochemical analysis demonstrated that some of the fixatives (particularly acetone and methanol) significantly reduced or completely depleted cellular gangliosides, implying that the immunoreactivity observed with A2B5, and with other antibodies, was not due to gangliosides. When neurons were incubated with an anti-GD1b antibody prefixation, uniform labeling of the plasma membrane was observed, but after ganglioside depletion using biochemical inhibitors of ganglioside synthesis no cell surface labeling was detected. However, even in cells depleted of gangliosides, labeling of both the cell surface and intracellular compartments was observed when the anti-GD1b antibody was applied after fixation. Moreover, after fixation, antibodies to GM4 and GD2 reacted with hippocampal neurons, although these gangliosides are absent from these neurons. In contrast, the JONES antibody (which reacts with 9-O -acetylated GD3) labeled neurons with a similar pattern, essentially irrespective of the fixation method. These observations demonstrate that great care must be taken in assigning gangliosides to specific cell populations or to intracellular locations solely on the basis of use of anti-ganglioside antibodies, and suggest that optimal fixation conditions must be established for each anti-ganglioside antibody. (J Histochem Cytochem 45:611-618, 1997)

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[55] Inhibitors of cerebroside metabolism

Cited by 29 publications

References 16 publications

Ceramide accumulation precedes caspase-3 activation during apoptosis of A549 human lung adenocarcinoma cells

Ceramide accumulation precedes caspase-3 activation during apoptosis of A549 human lung adenocarcinoma cells

1-Methylthiodihydroceramide, a Novel Analog of Dihydroceramide, Stimulates Sphinganine Degradation Resulting in Decreased de Novo Sphingolipid Biosynthesis

Determination of the Localization of Gangliosides Using Anti-ganglioside Antibodies: Comparison of Fixation Methods

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