Data were analyzed using Summit software. Caspase 37 activity was quantified by using Caspase Glo 37 Assay from Promega. Cell viability was addressed by using CellTiter AQueousOne Solution Cell Proliferation Assay, a colorimetric method based on the reduction of a tetrazo lium compound selleck kinase inhibitor by NADPH or NADH produced by de hydrogenase enzymes in metabolically active cells. Levels of reduced Inhibitors,Modulators,Libraries glutathione were quantified by using GSH Glo Glutathione Assay following the ma nufacturers instructions. Nuclear and cytoplasmic protein fractions were Inhibitors,Modulators,Libraries obtained by using NE PER Nuclear and Cytoplasmic Extraction Kit. Experiments in hypoxia were performed as previously described. In the inhibition studies for the RAS downstream sig naling pathways, breast cancer cell lines MDA MB 231 and MCF 7 were seeded onto 6 well plates and 24 hours later washed with PBS and subjected to free serum standard media.

24 hours later the cells were incubated with free serum standard media containing DMSO or the following chemicals ERK kinases inhibitor U0126 . PI3K inhibi tors LY294002 and wortmannin . and AKT Inhibitors,Modulators,Libraries inhibitor GSK690693. After Inhibitors,Modulators,Libraries 16 hours incubation, RNA was collected and qRT PCR was performed. Protein extracts were also col lected for western blot analysis. Quantitative real time polymerase chain reaction Total RNA was extracted using RNEasy mini kit and mRNA levels were quantified by qRT PCR using Taqman Gene Expression Assays. SYBR Green Master Mix was used with optimized primers for GA PDH at final concentration of 300 nM. b Actin was used as reference gene for the experiments in hypoxia.

All experiments were per formed at least by triplicate. Additional cell lines Additional MSC lines with activated RAS or RAS downstream effectors were generated by infection of MSC4 with the retroviral vector pBabe hygro encoding constitutive active RAS, the membrane targeted RAF 1, and Inhibitors,Modulators,Libraries myristoylated AKT, all kindly provided by Dr. Pablo Rodriguez Viciana. Immortal human mammary epithelial cells, obtained from Dr. Rodriguez Viciana, were cultured in DMEMF 12 containing 5% horse serum and supplemented with EGF, hydrocortisone, cholera toxin and insulin, all from Sigma. HMEC expressing different oncogene combinations were generated after infection with the same retroviral vectors used for the generation of MSC lines. Breast cancer cell lines MDA MB 231 and MCF 7 were cultured in DMEM containing 10% fetal bovine serum. Human umbilical vein endothelial cells were obtained from Promocell and cultured selleck products with Endothelial Cell Growth Medium according to suppliers instructions. Public transcriptome data The expression level of Nrf2 in many types of cancer was and The Cancer Genome Atlas databases of cancer expression data.

Beyond genetic ablation, abrogation of endogenous IKK BNF ��B signaling is also achieved by the activation selleck of a G protein coupled receptor of the rhodopsin family and also a bona fide long chain fatty acid sensor coined GPR120. GPR120 activation by unsaturated FAs particularly of the omega 3 variety prevents signaling through the IKK BNF ��B pathway by physically Inhibitors,Modulators,Libraries interact ing with B2 arrestin and sequestering the major activator of the pathway, transforming growth factor B activated kinase 1 binding protein. Without sufficient TAB1 available to activate its partner protein TAK1 the downstream kinase IKK B and key transcription factor NF��B remain dormant and the transcriptional in flammatory response is not induced.

Despite the fact that GPR120 is well documented to signal through the Gqll subunit Inhibitors,Modulators,Libraries activating the protein kinase C mitogen activated Inhibitors,Modulators,Libraries protein kinase extracellular signal regulated kinase and the phosphoinositide 3 kinase AKT cascades, the relevance of either cascade in mediating the anti inflammatory properties of GPR120 remains unex plored. The presence of functional GPR120 and adequate omega 3 FAs for its activation is sufficient to lower sys temic inflammatory state and improve overall energy utilization in mice. Genetic disruption of GPR120 eliminates the ability of dietary omega 3 FAs to improve energy homeostasis in obese mice, attesting to the potency of this receptor. Importantly, GPR120 activity is also physiologically relevant in humans given the recent dis covery of functionally disrupted GPR120 mutations in obese Europeans.

Inhibitors,Modulators,Libraries The anti inflammatory actions of GPR120 in response to long chain FAs, in particular omega 3 FAs, have been elegantly demonstrated in adipo cytes and macrophages, but little is known regarding these actions in other tissues throughout the body. Recently, GPR120 expression and the GPR120 B2 arrestin TAB1 interaction were identified in the hypothalamus, but its role in the CNS remains poorly understood. Given the anti inflammatory actions of GPR120 in the periphery and the conservation the IKK BNF ��B pathway in the CNS, we hypothesize that GPR120 may modulate hypo thalamic function by altering the inflammatory status of this tissue. We test this hypothesis at the Inhibitors,Modulators,Libraries cellular level using a hypothalamic neuronal model, rHypoE 7, isolated from the rat.

Methods Cell culture The hypothalamic all targets cell line from the embryonic rat, rHypoE 7, was isolated and immortalized as previously de scribed. Cell lines were maintained and grown to confluency in Dulbeccos Modified Eagle Medium supplemented with 5% fetal bovine serum, 1% penicillin, and 1% streptomycin and seeded into 60 mm cul ture plates 24 hrs prior to experimental treatments. Production of cDNA and quantitative real time reverse transcriptase PCR Total cellular RNA was isolated as previously described using the guanidium thiocyanate phenol chloroform extrac tion method.

In Sweden, with a population of 9 million, there are approximately 300 new those GBM cases per year. Histolo gically, GBM is characterized by the presence of necrotic areas in the brain tissue surrounded by anaplastic cells and hyperplastic blood vessels, and a disparate genetic signature, which illustrates the heterogeneity of this tumor. Inhibitors,Modulators,Libraries Current treatments such as surgical resec tion, radiation, and chemotherapy are relatively ineffec tive due to the aggressive nature of GBM. Despite increasing molecular knowledge of GBM tumors, few new therapeutic strategies have been offered these patients during the past decades and these patients still have a poor prognosis with a median survival of 12 15 months. Recent studies have reported the presence of an active Human Cytomegalovirus infection in 90 100% of GBM tumors.

HCMV belongs to the herpesviridea Inhibitors,Modulators,Libraries family and main tains latency in pre monocytic cells after a primary infection. Viral reactivation can occur when latently infected monocytes undergo differentiation to macro phages or dendritic cells, which involves stimulation with inflammatory cytokines. Thus, it is possible that the inflammatory environment in the glioblastoma tumor triggers reactivation of latent HCMV. Although there is compelling evidence that the HCMV protein US28 may be oncogenic, this virus has not been considered to be oncogenic. Rather, HCMV proteins confer a variety of biological functions that potentially affect tumor biology and contribute to cancer progression.

HCMV proteins for example interact with p53, Rb and PTEN and influence cell cycle regulation, induce chromosomal Inhibitors,Modulators,Libraries aberrations, cellular differentiation, migration, angiogenesis and confer immune evasion mechanisms. Recent studies have demonstrated that HCMV US28 induces cyclo oxygenase 2 expression, production of vascular endothelial growth factor and tumor formation in vivo, via activa tion of nuclear factor kappa B, STAT3 phos phorylation and accumulation of beta cathenin in the cell nucleus. Furthermore, we found that phosphorylated STAT 3 was correlated with poor survival of GBM patients. Thus, this virus may either be an epi phenomenon of glioblastomas or may through its sophisticated Inhibitors,Modulators,Libraries strategies that affect tumor biology and provide immune evasion strategies contribute to cancer progression.

We recently demonstrated that targeting HCMV infection in medulloblastoma tumors with an anti viral drug Inhibitors,Modulators,Libraries and a COX 2 inhibitor prevented tumor growth by 72% in an animal www.selleckchem.com/products/ABT-888.html model. These observa tions suggest that HCMV may contribute to the tumor genesis of medulloblatsoma tumors, and that the virus may be a novel target for cancer therapy. We hypothe sized that if the virus affects tumor progression, a low grade HCMV infection in glioblastomas may be asso ciated with longer patient survival. In this case control study, we investigated if the grade of HCMV infection in GBM tumors was associated with survival over 18 months.

In fibro blasts, the trend of percentages was instead opposite. The two lists of selected probes were analysed by the In genuity Pathway Analysis software. Results obtained towards on melanoma cells are reported in Additional file 1 B and 1 C. IPA results obtained for BJ fibroblasts are reported in Additional file 2 B and 2 C. D6 treatment Inhibitors,Modulators,Libraries affects cell death and proliferation bio functions The top bio functional categories identified by IPA among the genes modulated in treated melanoma cells are listed in Table 2, where the p values range and number of mole cules involved are reported for each category. The lowest p values were found for the Cell Death cat egory with 194 molecules involved. Cell death is indeed the primary effect detected on melanoma cells after D6 treatment.

Moreover, Inhibitors,Modulators,Libraries a variable number FC value of 368. 61. HSPA6 codifies for the Hsp70B, a highly Inhibitors,Modulators,Libraries stress inducible protein. Additional pathways such as 4 Cell cycle G2/M DNA damage checkpoint regulation, 5 p53 Inhibitors,Modulators,Libraries signalling, 10 Her editary breast cancer signalling, 11 ATM signalling, 26 Role of BRCA1 in DNA damage response, and 27 Role of of molecules differentially modulated by D6 involved func tional categories strictly correlated with cell proliferation processes such as cellular function and maintenance, cell cycle and cell growth and proliferation. D6 induces stress response pathways and down regulates cell proliferation pathways Table 3 lists the most significant pathways that IPA found to be enriched with the input genes in melan oma cells. For each pathway, the respective nominal p value, along with all the input molecules are reported.

A general trend of up regulation for pathways involving Inhibitors,Modulators,Libraries cell stress response was evident . con versely, pathways that control cell proliferation appeared inhibitor purchase down regulated. The first three most significant pathways, 1 Aldosterone signalling, 2 Protein ubiquitination, and 3 NRF2 mediated oxidative stress response as well as the 21 Endoplasmic reticulum stress pathway appear to be up regulated, depicting a strong activation of stress induced molecular responses that involves over expression of heat shock proteins and activation of protein degradation processes. Among up regulated HSPs, HSPA6 is the most over expressed transcript in melanoma treated cells with a CHK proteins in cell cycle checkpoint control are all related to DNA repair mechanisms and cell death triggering, evidencing a DNA damage as cell response to D6 treatment. The up regulation of pathway 5 p53 signalling, which acts in response to cell injury or DNA damage by controlling cell proliferation and driving cells to apoptosis, is noteworthy and points out a central role of this regulatory protein in the D6 anticancer effect on melanoma cells.

furthermore, this was observed in all NSCLC cell lines examined. Bfl 1overex pression has been reported to underlie resistance to var ious apoptotic stimuli, and we previously observed that Bfl 1 is frequently over expressed in lung cancer cell lines. Therefore, we speculated that Bfl 1 might regulate the sensitivity selleck Ganetespib of NSCLC to gemcitabine. As was expected, direct Bfl 1 suppression using siRNA sen sitized cells to gemcitabine induced apoptosis in A549 cells. In a study conducted by Brien et al. in B lympho blastoid and diffuse large B cell lymphoma cell lines, Bfl 1 silencing was found to potently induce apoptosis, and sensitize cells to rituximab mediated cell death and apoptosis by doxorubicin, vincristine, cisplatin, or flu darabine.

In the present study using lung cancer cell lines, Inhibitors,Modulators,Libraries although siRNA mediated Bfl 1 suppression per se did not affect cell viability, it sensitized cells to gemcitabine. Therefore, the present and our previous findings suggest that Bfl 1 is a feasible molecular target for enhancing Inhibitors,Modulators,Libraries the efficacy of gemcitabine in lung cancer. Furthermore, in view of the fact that BC itself alone has a cytotoxic effect, it would appear that BC is likely to be better than siRNA at chemosensitizing cells. Although previous studies have reported that NF B regulates the expression of Bcl xL and/or Bcl 2 in various cell types, we observe little changes in the level of Bcl xL and Bcl 2 in cells treated with gemcitabine at low concentra tions inducing NF B activation either by RT Inhibitors,Modulators,Libraries PCR and real time RT PCR.

It is possible that Bfl 1, a direct tar get of NF B, might function as Inhibitors,Modulators,Libraries an Inhibitors,Modulators,Libraries important and sen sitive anti apoptotic Bcl 2 family protein reflecting the alteration of NF B activity in NSCLC cells, particularly in terms of response to gemcitabine. Although Bfl 1 is an anti apoptotic bcl 2 family mem ber and primarily prevents apoptosis, its protective effect depends on cell type and apoptotic stimulus. Furthermore, Bfl 1 can be converted into a pro apopto tic molecule when processed by proteasome or TNF receptor signaling in pro B cells. In this system, the Bfl 1 C terminal domain was important for the pro apoptotic function of Bfl 1, because it was required for Bfl 1 ubiquitination and its localization at the mitochon drial membrane.

Our group, previously, demon strated that anti apoptotic Bfl 1 is converted to a pro apoptotic selleck chem protein when fused with GFP, and that the 29 amino acids of its C terminal region are sufficient to induce apoptotic cell death via the mitochondrial path way and caspase activation in 293T cells. An amphipathic tail anchoring domain of the Bfl 1 C term inal region has been implicated in the targeting of Bfl 1 to the mitochondrial membrane to induce apoptosis. These findings led us to hypothesize that the Bfl 1 C terminal region fused with GFP could be harnessed as a gene therapy in combination with che motherapeutic agents to achieve management of cancer.

Cyclin A1 may have a role in carcinogenesis, as it has been found to be over expressed neverless in acute Inhibitors,Modulators,Libraries myeloid leukemia and var ious other tumour types, however, its role in can cer is still particularly obscure. In somatic non testicular tissues, cyclin A1 is not expressed or is expressed at very low basal levels. After genotoxic insult, cyclin A1 mRNA is upregulated Inhibitors,Modulators,Libraries in vitro and in vivo. At a molecular level, human CDK2/cyclin A1 complexes interact with members of the Ku family and phosphory late Ku70, a pivotal player in the non homolo gous end joining double strand break repair pathway. Furthermore, under genotoxic condi tions the kinase activity of CDK2/cyclin A1 complex increases, while the relative kinase activity of CDK2/ cyclin A2 decreases and the CDK2/cyclin A1 complex out competes with CDK2/cyclin A2 for Ku70 binding.

Moreover, it has recently been found that CDK2 phosphorylation Inhibitors,Modulators,Libraries status and structure changes upon the cyclin A family member with which it is bound suggesting a non redundant function between CDK2/ cyclin A1 and CDK2/cyclin A2 complexes. Finally cyclin A1 knockout mice and Xenopus embryos exhibited a clear defect in DNA repair and are more prone to undergo apoptosis. Taken together these data support that during geno toxic stress differential transcriptional levels and activity of cyclin A family members may redirect CDK2 toward DNA repair resulting in a modulation of NHEJ. Since one of the most relevant effects of CDK inhibitors is the downregulation of cell cycle related cyclins, we investi gated if the inhibition of DNA repair mechanisms by Roscovitine may also occur through the modulation of the expression levels of cyclin A family members.

Phy siological CDK inhibition, Inhibitors,Modulators,Libraries in fact, results in cyclin downregulation through the inhibition of E2F family transcription factors, which drive and regulate cell cycle related cyclin transcription. Given that the promoter of the Inhibitors,Modulators,Libraries cyclin A1 gene, CCNA1, is different from the other cell cycle related cyclins, not being under the regulation of E2Fs, here we analyzed the effects of Roscovitine on cyclin A1 expression and modulation of DNA repair mechanisms. We demonstrated that www.selleckchem.com/products/Enzastaurin.html under DNA dama ging conditions cyclin A1 is strongly upregulated and localizes to the nucleus. Although Roscovitine alone was not sufficient to reduce the basal levels of cyclin A1, in contrast to cell cycle related cyclins, Roscovitine treat ment could abolish the DNA damage induced cyclin A1 upregulation, reducing NHEJ and significantly hindering DNA repair over time. Results DNA damage induces a switch in the respective levels of A family cyclins We first compared mRNA levels of both members of the cyclin A family after treatment with increasing doses of Doxorubicin, a well known inducer of DNA DSBs.

Isaiah J. Fidler. Equal protein quantities were electrophoresed and West ern blotted as described. To confirm equal sellckchem protein loading blots were either reprobed with actin or equal amounts of lysates were loaded selleck products in duplicate lanes in the same gel and separated after transfer to be probed for actin separately. Immunohistochemistry Formalin fixed,paraffin embedded tissue blocks of the human kidney tumor samples were sectioned at 4 5 micron thickness,mounted on charged glass slides and baked for one hour at 60 C. Slides were deparaffinized with 3 changes of xylene,and the endogenous peroxidases were quenched with hydrogen peroxide,followed by a series of ethanol rinses. Slides were rehydrated and prepared for antigen retrieval with citrate buffer and blocked with 10% goat serum diluted in PBS.

Inhibitors,Modulators,Libraries After incubation Inhibitors,Modulators,Libraries Inhibitors,Modulators,Libraries with phospho Hsp27 antibody in PBS 0. 05% BSA overnight,slides were rinsed in PBS and incubated with anti goat second ary antibody,and incubated with DAB following vender instruc tions. Slides were counterstained in Mayers hematoxylin,dehydrated,cleared,and coverslipped. Slides were photo graphed with a Zeiss Axioskop light microscope and Axio cam digital camera Two dimensional gel electrophoresis and spot analysis Proteins were extracted from frozen tissue as previously described. A total protein concentration of 600g in IPG Rehydration Buffer containing 15 mM DTE Inhibitors,Modulators,Libraries and 0. 5% ampholytes pH 3 10 was loaded on 17 cm IPG strips pH 3 10 non linear from Bio Rad using passive rehydration at 20 C.

Isoelectric focusing was performed using the Pro tean IEF cell for 65,000 Vh.

After equilibration IPG strips were loaded on uniform 11% polyacrylamide Inhibitors,Modulators,Libraries bis acrylamide gels and electro phoresed at 20 mA constant current Inhibitors,Modulators,Libraries at 10 C. Gels were stained with Colloidal Coomassie blue Inhibitors,Modulators,Libraries and scanned with an Epson 1680 Scanner as described previously. Spot quantification and statistical analysis of differences in spot values were done as previously described. The protein spots were matched between gels using the All to One warping strategy using the Delta 2D gel analysis soft ware from Decodon GmbH. One RCC sample gel was selected as the reference gel and all replicates of all conditions were matched to this gel using the exact warping Inhibitors,Modulators,Libraries method between each gel pair with defined vectors from sample to Master gel.

In order to ensure that the same spot area was quantified in all Inhibitors,Modulators,Libraries gels,a master gel was created by fusing all gel images with the maximum intensity option selected in Delta2D. Subse quently,the spots Inhibitors,Modulators,Libraries in the master gel were detected,using optimized spot detection parameters with exact spot out lines. In some compound libraries cases spot outlines were manually edited to separate spots or to eliminate background interference. The detected spots from the www.selleckchem.com/products/crenolanib-cp-868596.html Master gel were then trans ferred to all other gels,instead of individually quantifying each gel,which yielded different spot outlines.

In some cancer moreover patients, the level of CECs is significantly higher than that of healthy individuals, and this Afatinib Inhibitors,Modulators,Libraries increased level has been identified as a surrogate marker of angiogenesis and anti www.selleckchem.com/products/ldk378.html angiogenic drug activity. Inhibitors,Modulators,Libraries The present study has shown that baseline CEC levels are markedly higher among pancreatic carcinoma patients than in healthy individuals. Our results also support the hypothesis that CEC levels are associated with clinical outcome in pancreatic carcinoma patients undergoing gemcitabine chemotherapy, and may be a prognostic factor for this disease. A previous study found that the baseline level of CECs, identified as CD45 CD31 CD34 by flow cytometry, was inversely associated with OS in patients who had gemcitabine refractory metastatic pancreatic carcinoma and were treated with bevacizumab plus erlotinib.

CEC detection by flow cytometry requires careful discrimination between blood cell popu lations with overlapping phenotypes showing hallmarks of T cells and platelets. These Inhibitors,Modulators,Libraries cells Inhibitors,Modulators,Libraries populations show distinct regulation during cancer therapy, and their concomitant analysis may offer extended prognostic and predictive information. Our study Inhibitors,Modulators,Libraries also found the baseline level Inhibitors,Modulators,Libraries of CECs, as well as the levels of HGF, IL 6, and IL 10, which are associated with gemcitabine resistance or stemness, Inhibitors,Modulators,Libraries to be significantly higher Inhibitors,Modulators,Libraries among PD patients.

Univariate Cox model analysis further demonstrated that PS, clin ical stage, CRP levels, Inhibitors,Modulators,Libraries and CEC levels are all associated with the survival of pancreatic carcinoma patients, while multivariate Cox analysis showed that Inhibitors,Modulators,Libraries CEC and IL 10 levels are strongly associated with survival.

The number of CECs detectable Inhibitors,Modulators,Libraries in individuals Inhibitors,Modulators,Libraries Inhibitors,Modulators,Libraries has previously been found to be associated with the plasma levels Inhibitors,Modulators,Libraries of VCAM 1 and VEGF in cancer patients. Our findings further show that, Inhibitors,Modulators,Libraries in addition to VEGF, CEC levels are strongly associated with the expression levels of IL 8, IL 10, and HGF in pancreatic carcinoma patients. These molecules, among others, play important roles in tumor biology and have been implicated in several cellular phenotypes.

Chemokines, including IL 8 and IL 10, are small peptides involved in controlling cell migration, particularly in leukocytes, during inflammation and the immune response.

Chemo kines are also important dasatinib IC50 in tumor biology as they influ ence tumor growth, invasion, metastasis, and angiogenesis.

http://www.selleckchem.com/products/arq-197.html For instance, VEGF, HGF and IL 8 signifi cantly read more stimulate the proliferation, migration, and inva sion of cancer cells. CEC are shed from vessels and this process may be amplified by an aberrant vascular turn over/remodeling associated with high local levels of VEGF required for CEC survival. The chemokine SDF 1 has likewise been found to enhance the produc tion of IL 8 by pancreatic cells in a paracrine manner.

No other serum enzymes and proteins were significantly elevated by LCL85. LCL85 suppresses colon carcinoma metastatic potential in an experimental lung metastasis mouse model in vivo To determine the efficacy of LCL85 in suppression of me tastasis in vivo, we used an experimental metastasis mouse model. Colon26 cells, then a highly metastatic colon carcinoma cell line, were injected i. v. to mice. Tumor bearing mice were treated with LCL85 over time. Lung metastasis was then analyzed. LCL85 significantly suppressed colon26 lung metastasis in a dose dependent manner. Although LCL85 Inhibitors,Modulators,Libraries possesses direct anti tumor cytotox icity that might contribute to the observed tumor suppression, it is possible that LCL85 might also sensitize the Inhibitors,Modulators,Libraries tumor cells to apoptosis induction by FasL of host immune cells, particularly CD8 CTLs.

We then dissected tumor bearing lungs and made single cell suspension with collagenase. Staining cells with CD8 and FasL specific mAbs revealed that CD8 T cells in tumor free mice are essentially FasL. In contrast, ap proximately 31% of tumor infiltrating Inhibitors,Modulators,Libraries CD8 T cells are FasL. CD8 cells in tumor free mice are all FasL. Therefore, LCL85 might sensitize colon carcinoma cells to host FasL CTL mediated tumor suppression. LCL85 suppresses spontaneous breast cancer metastasis in vivo To further determine the function of LCL85 in suppres sion of cancer metastasis, we used a complimentary breast cancer lung metastasis mouse model. Murine breast cancer Inhibitors,Modulators,Libraries 4 T1 cells were injected to the mammary fat pad. Tumor bearing mice were treated with LCL85 over time and both primary tumor growth and lung metastasis were examined.

LCL85 significantly suppressed the primary mammary tumor growth in vivo as measured by tumor size and tumor weight. Interestingly, the spontaneous lung metastasis was also significantly sup pressed by LCL85. The observation that LCL85 suppresses spontaneous breast cancer lung me tastasis is significant. However, it is possible that the decreased lung metastasis Inhibitors,Modulators,Libraries was due to the decreased primary tumor growth. To deter mine whether LCL85 directly suppresses spontaneous metastasis, 4 T1 cells were injected to mouse mammary fat pad. Primary tumors were surgically removed 15 days after tumor cell injection. Mice were treated with LCL85 over time after surgery. This procedure thus mimics human breast cancer patient treatment.

Analysis of lungs indicated that LCL85 inhibitor AZD9291 significantly suppresses breast can cer spontaneous lung metastasis. Taken together, our data demonstrated that LCL85 at a subtoxic dose is effective in suppression of colon and breast cancer metastasis. Discussion Ceramide mediates apoptosis through multiple mecha nisms. It has been reported that ceramide mediates Fas receptor clustering, capping and activation to promote Fas mediated apoptosis.