Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0027819 (neuroblastoma)
 27,800 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Anandamide (arachidonylethanolamide; AnNH) has important neuromodulatory and immunomodulatory activities. This lipid is rapidly taken up and hydrolyzed to arachidonate and ethanolamine in many organisms. As yet, AnNH inactivation has not been studied in humans. Here, a human brain fatty-acid amide hydrolase (FAAH) has been characterized as a single protein of 67 kDa with a pI of 7.6, showing apparent Km and Vmax values for AnNH of 2.0 +/- 0.2 microM and 800 +/- 75 pmol.min-1.mg of protein-1, respectively. The optimum pH and temperature for AnNH hydrolysis were 9.0 and 37 degreesC, respectively, and the activation energy of the reaction was 43.5 +/- 4.5 kJ.mol-1. Hydro(pero)xides derived from AnNH or its linoleoyl analogues by lipoxygenase action were competitive inhibitors of human brain FAAH, with apparent Ki values in the low micromolar range. One of these compounds, linoleoylethanolamide is the first natural inhibitor (Ki = 9.0 +/- 0.9 microM) of FAAH as yet discovered. An FAAH activity sharing several biochemical properties with the human brain enzyme was demonstrated in human neuroblastoma CHP100 and lymphoma U937 cells. Both cell lines have a high affinity transporter for AnNH, which had apparent Km and Vmax values for AnNH of 0.20 +/- 0.02 microM and 30 +/- 3 pmol.min-1.mg of protein-1 (CHP100 cells) and 0.13 +/- 0.01 microM and 140 +/- 15 pmol.min-1.mg of protein-1 (U937 cells), respectively. The AnNH carrier of both cell lines was activated up to 170% of the control by nitric oxide.
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PMID:Anandamide hydrolysis by human cells in culture and brain. 982 13

The endocannabinoid anandamide (AEA) is shown to induce apoptotic bodies formation and DNA fragmentation, hallmarks of programmed cell death, in human neuroblastoma CHP100 and lymphoma U937 cells. RNA and protein synthesis inhibitors like actinomycin D and cycloheximide reduced to one-fifth the number of apoptotic bodies induced by AEA, whereas the AEA transporter inhibitor AM404 or the AEA hydrolase inhibitor ATFMK significantly increased the number of dying cells. Furthermore, specific antagonists of cannabinoid or vanilloid receptors potentiated or inhibited cell death induced by AEA, respectively. Other endocannabinoids such as 2-arachidonoylglycerol, linoleoylethanolamide, oleoylethanolamide, and palmitoylethanolamide did not promote cell death under the same experimental conditions. The formation of apoptotic bodies induced by AEA was paralleled by increases in intracellular calcium (3-fold over the controls), mitochondrial uncoupling (6-fold), and cytochrome c release (3-fold). The intracellular calcium chelator EGTA-AM reduced the number of apoptotic bodies to 40% of the controls, and electrotransferred anti-cytochrome c monoclonal antibodies fully prevented apoptosis induced by AEA. Moreover, 5-lipoxygenase inhibitors 5,8,11,14-eicosatetraynoic acid and MK886, cyclooxygenase inhibitor indomethacin, caspase-3 and caspase-9 inhibitors Z-DEVD-FMK and Z-LEHD-FMK, but not nitric oxide synthase inhibitor Nomega-nitro-l-arginine methyl ester, significantly reduced the cell death-inducing effect of AEA. The data presented indicate a protective role of cannabinoid receptors against apoptosis induced by AEA via vanilloid receptors.
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PMID:Anandamide induces apoptosis in human cells via vanilloid receptors. Evidence for a protective role of cannabinoid receptors. 1091 56

Treatment of intact human neuroblastoma CHP100 cells with anandamide (arachidonoylethanolamide, AEA) or 2-arachidonoylglycerol (2-AG) inhibits intracellular fatty acid amide hydrolase (FAAH). This effect was not associated with covalent modifications of FAAH, since specific inhibitors of farnesyltransferase, kinases, phosphatases, glycosyltransferase or nitric oxide synthase were ineffective. Electrophoretic analysis of (33)P-labelled proteins, Western blot with anti-phosphotyrosine antibodies, and glycan analysis of cellular proteins confirmed the absence of covalent modifications of FAAH. The inhibition by AEA was paralleled by an increased arachidonate release, which was not observed upon treatment of cells with linoleoylethanolamide, palmitoylethanolamide, or oleoylethanolamide. Moreover, cell treatment with AEA or 2-AG increased the activity of cyclooxygenase and 5-lipoxygenase, and the hydro(pero)xides generated from arachidonate by lipoxygenase were shown to inhibit FAAH, with inhibition constants in the low micromolar range. Consistently, inhibitors of 5-lipoxygenase, but not those of cyclooxygenase, significantly counteracted the inhibition of FAAH by AEA or 2-AG.
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PMID:Anandamide and 2-arachidonoylglycerol inhibit fatty acid amide hydrolase by activating the lipoxygenase pathway of the arachidonate cascade. 1109 52

Anandamide is an endogenous compound that acts as an agonist at cannabinoid receptors. It is inactivated via intracellular degradation after its uptake into cells by a carrier-mediated process that depends upon a concentration gradient. The fate of anandamide in those cells containing an amidase called fatty-acid amide hydrolase (FAAH) is hydrolysis to arachidonic acid and ethanolamine. The active site nucleophilic serine of FAAH is inactivated by a variety of inhibitors including methylarachidonylfluorophosphonate (MAFP) and palmitylsulfonyl fluoride. In the current report, the net uptake of anandamide in cultured neuroblastoma (N18) and glioma (C6) cells, which contain FAAH, was decreased by nearly 50% after 6 min of incubation in the presence of MAFP. Uptake in laryngeal carcinoma (Hep2) cells, which lack FAAH, is not inhibited by MAFP. Free anandamide was found in all MAFP-treated cells and in control Hep2 cells, whereas phospholipid was the main product in N18 and C6 control cells when analyzed by TLC. The intracellular concentration of anandamide in N18, C6, and Hep2 cells was up to 18-fold greater than the extracellular concentration of 100 nm, which strongly suggests that it is sequestered within the cell by binding to membranes or proteins. The accumulation of anandamide and/or its breakdown products was found to vary among the different cell types, and this correlated approximately with the amount of FAAH activity, suggesting that the breakdown of anandamide is in part a driving force for uptake. This was shown most clearly in Hep2 cells transfected with FAAH. The uptake in these cells was 2-fold greater than in vector-transfected or untransfected Hep2 cells. Therefore, it appears that FAAH inhibitors reduce anandamide uptake by cells by shifting the anandamide concentration gradient in a direction that favors equilibrium. Because inhibition of FAAH increases the levels of extracellular anandamide, it may be a useful target for the design of therapeutic agents.
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PMID:The cellular uptake of anandamide is coupled to its breakdown by fatty-acid amide hydrolase. 1111 29

Endocannabinoids are lipid mediators thought to modulate central and peripheral neural functions. We report here gas chromatography-electron impact mass spectrometry analysis of human brain, showing that lipid extracts contain anandamide and 2-arachidonoylglycerol (2-AG), the most active endocannabinoids known to date. Human brain also contained the endocannabinoid-like compounds N-oleoylethanolamine, N-palmitoylethanolamine and N-stearoylethanolamine. Anandamide and 2-AG (0.16 +/- 0.05 and 0.10 +/- 0.05 nmol/mg protein, respectively) represented 7.7% and 4.8% of total endocannabinoid-like compounds, respectively. N-Palmitoyethanolamine was the most abundant (50%), followed by N-oleoyl (23.6%) and N-stearoyl (13.9%) ethanolamines. A similar composition in endocannabinoid-like compounds was found in human neuroblastoma CHP100 and lymphoma U937 cells, and also in rat brain. Remarkably, human meningioma specimens showed an approximately six-fold smaller content of all N-acylethanolamines, but not of 2-AG, and a similar decrease was observed in a human glioblastoma. These ex vivo results fully support the purported roles of endocannabinoids in the nervous system.
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PMID:Gas chromatography-mass spectrometry analysis of endogenous cannabinoids in healthy and tumoral human brain and human cells in culture. 1120 22

The endogenous cannabinoid receptor agonist anandamide (AEA) and the related compound palmitoylethanolamide (PEA) are inactivated by transport into cells followed by metabolism by fatty acid amide hydrolase (FAAH). The cellular uptake of AEA has been characterized in detail, whereas less is known about the properties of the PEA uptake, in particular in neuronal cells. In the present study, the pharmacological and functional properties of PEA and AEA uptake have been investigated in mouse Neuro-2a neuroblastoma and, for comparison, in rat RBL-2H3 basophilic leukaemia cells. Saturable uptake of PEA and AEA into both cell lines were demonstrated with apparent K(M) values of 28 microM (PEA) and 10 microM (AEA) in Neuro-2a cells, and 30 microM (PEA) and 9.3 microM (AEA) in RBL-2H3 cells. Both PEA and AEA uptake showed temperature-dependence but only the AEA uptake was sensitive to treatment with Pronase and phenylmethylsulfonyl fluoride. The AEA uptake was inhibited by AM404, 2-arachidonoylglycerol (2-AG), R1- and S1-methanandamide, arachidonic acid and olvanil with similar potencies for the two cell types. PEA, up to a concentration of 100 microM, did not affect AEA uptake in either cell line. AEA, 2-AG, arachidonic acid, R1-methanandamide, (9)-THC, and cannabidiol inhibited PEA transport in both cell lines. The non-steroidal anti-inflammatory drug indomethacin inhibited the AEA uptake but had very weak effects on the uptake of PEA. From these data, it can be concluded that PEA is transported in to cells both by passive diffusion and by a facilitated transport that is pharmacologically distinguishable from AEA uptake.
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PMID:Characterization of palmitoylethanolamide transport in mouse Neuro-2a neuroblastoma and rat RBL-2H3 basophilic leukaemia cells: comparison with anandamide. 1130 46

The cloned 5-HT3 receptor from NCB-20 neuroblastoma cells was expressed in Xenopus oocytes and the effect of the endogenous cannabinoid ligand, anandamide, was investigated on the function of this receptor. The oocytes expressing the cloned 5-HT3 receptors were voltage-clamped at -70 mV. Anandamide, at the concentration range of 0.1-100 microM, reversibly inhibited 1 microM 5-HT induced currents. The inhibition of 5-HT induced currents by anandamide was concentration-dependent with an EC50 of 3.7 microM and slope value of 0.94. This inhibitory effect was not dependent on the membrane potential and anandamide did not have an effect on the reversal potential of 5-HT-induced currents. In the presence of 10 microM anandamide, the maximum 5-HT-induced response was also inhibited and the respective EC50 values were 3.4 microM and 3.1 microM in the absence and presence of anandamide, indicating that anandamide acts as a noncompetitive antagonist on 5-HT3 receptors. CB1 receptor antagonist SR-141716A (1 microM) and pertussis toxin (5 microg/ml) did not cause a significant change on the inhibition of 5-HT responses by anandamide. The effect of anandamide was not changed by preincubating the oocytes with 0.2 mM 8-Br-cAMP, a membrane-permeable analog of cAMP, or Sp-cAMPS (0.1 mM), a membrane-permeable protein kinase A activator. These results suggest that the effect of anandamide is independent of the activation of cAMP pathway and not mediated by the activation of PTX sensitive G-proteins. In conclusion, we demonstrated that the endogenous cannabinoid anandamide inhibits the function of 5-HT3 receptors expressed in Xenopus oocytes in a cannabinoid-receptor independent and noncompetitive manner.
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PMID:Endogenous cannabinoid, anandamide, acts as a noncompetitive inhibitor on 5-HT3 receptor-mediated responses in Xenopus oocytes. 1232 42

1. The ability of the endogenous fatty acid amide, cis-oleamide (ODA), to bind to and activate cannabinoid CB(1) and CB(2) receptors was investigated. 2. ODA competitively inhibited binding of the nonselective cannabinoid agonist [(3)H]CP55,940 and the selective CB(1) antagonist [(3)H]SR141716A to rat whole-brain membranes with K(i) values of 1.14 microm (0.52-2.53 microm, Hill slope=0.80, n=6) and 2.63 microm (0.62-11.20 microm, Hill slope=0.92, n=4), respectively. AEA inhibited [(3)H]CP55,940 binding in rat whole-brain membranes with a K(i) of 428 nm (346-510 nm, Hill slope=-1.33, n=3). 3. ODA competitively inhibited [(3)H]CP55,940 binding in human CB(1) (hCB(1)) cell membranes with a K(i) value of 8.13 microm (4.97-13.32 microm, n=2). In human CB(2) transfected (hCB(2)) HEK-293T cell membranes, 100 microm ODA produced only a partial (42.5+/-7%) inhibition of [(3)H]CP55,940 binding. 4. ODA stimulated [(35)S]GTPgammaS binding in a concentration-dependent manner (EC(50)=1.64 microm (0.29-9.32 microm), R(2)=0.99, n=4-9), with maximal stimulation of 188+/-9% of basal at 100 microm. AEA stimulated [(35)S]GTPgammaS binding with an EC(50) of 10.43 microm (4.45-24.42 microm, R(2)=1.00, n=3, 195+/-4% of basal at 300 microm). Trans-oleamide (trans-ODA) failed to significantly stimulate [(35)S]GTPgammaS binding at concentrations up to 100 microm. 5. ODA (10 microm)-stimulated [(35)S]GTPgammaS binding was reversed by the selective CB(1) antagonist SR141716A (IC(50)=2.11 nm (0.32-13.77 nm), R(2)=1.00, n=6). 6. The anatomical distribution of ODA-stimulated [(35)S]GTPgammaS binding in rat brain sections was indistinguishable from that of HU210. Increases of similar magnitude were observed due to both agonists in the striatum, cortex, hippocampus and cerebellum. 7. ODA (10 microm) significantly inhibited forskolin-stimulated cyclic AMP (cAMP) accumulation in mouse neuroblastoma N1E 115 cells (P=0.02, n=11). ODA-mediated inhibition was completely reversed by 1 microm SR141716A (P<0.001, n=11) and was also reversed by pretreatment with 300 ng ml(-1) pertussis toxin (P<0.001, n=6). 8. These data demonstrate that ODA is a full cannabinoid CB(1) receptor agonist. Therefore, in addition to allosteric modulation of other receptors and possible entourage effects due to fatty acid amide hydrolase inhibition, the effects of ODA may be mediated directly via the CB(1) receptor.
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PMID:Oleamide is a selective endogenous agonist of rat and human CB1 cannabinoid receptors. 1469 Oct 53

Anandamide (arachidonoylethanolamide or AEA) is an endocannabinoid that acts at vanilloid (VR1) as well as at cannabinoid (CB1/CB2) and NMDA receptors. Here, we show that AEA, in a dose-dependent manner, causes cell death in cultured rat cortical neurons and cerebellar granule cells. Inhibition of CB1, CB2, VR1 or NMDA receptors by selective antagonists did not reduce AEA neurotoxicity. Anandamide-induced neuronal cell loss was associated with increased intracellular Ca(2+), nuclear condensation and fragmentation, decreases in mitochondrial membrane potential, translocation of cytochrome c, and upregulation of caspase-3-like activity. However, caspase-3, caspase-8 or caspase-9 inhibitors, or blockade of protein synthesis by cycloheximide did not alter anandamide-related cell death. Moreover, AEA caused cell death in caspase-3-deficient MCF-7 cell line and showed similar cytotoxic effects in caspase-9 dominant-negative, caspase-8 dominant-negative or mock-transfected SH-SY5Y neuroblastoma cells. Anandamide upregulated calpain activity in cortical neurons, as revealed by alpha-spectrin cleavage, which was attenuated by the calpain inhibitor calpastatin. Calpain inhibition significantly limited anandamide-induced neuronal loss and associated cytochrome c release. These data indicate that AEA neurotoxicity appears not to be mediated by CB1, CB2, VR1 or NMDA receptors and suggest that calpain activation, rather than intrinsic or extrinsic caspase pathways, may play a critical role in anandamide-induced cell death.
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PMID:Anandamide-induced cell death in primary neuronal cultures: role of calpain and caspase pathways. 1537 83

We have shown recently that in human T lymphocytes, leptin stimulates activity and expression of the endocannabinoid-degrading enzyme fatty acid amide hydrolase (FAAH), through STAT3 (signal transducer and activator of transcription 3) and its CRE (cAMP response element)-like transcriptional target in the FAAH promoter [Maccarrone, M., Di Rienzo, M., Finazzi-Agro, A., & Rossi, A. (2003) J. Biol. Chem. 278, 13318-13324]. We have also shown that progesterone, alone or additively with leptin, up-regulates the FAAH gene in human T-cells, through the Ikaros transcription factor [Maccarrone, M., Bari, M., Di Rienzo, M., Finazzi-Agro, A., & Rossi, A. (2003) J. Biol. Chem. 278, 32726-32732]. Here, we extend these observations to immortalized human lymphoma U937 cells, where stimulation of FAAH by leptin (up to approximately 300% of the controls) involves binding to a leptin receptor (Kd = 2.0 +/- 0.1 nm, Bmax = 382 +/- 5 fmol.mg protein(-1), apparent molecular mass of approximately 110 kDa), and stimulation by progesterone involves an intracellular receptor of approximately 120 kDa. Unlike FAAH, the other proteins of the endocannabinoid system are not modulated by the two hormones. Interestingly, human neuroblastoma CHP100 cells also have a leptin receptor (approximately 110 kDa, Kd = 2.2 +/- 0.2 nm, Bmax = 339 +/- 8 fmol.mg protein(-1)), a progesterone receptor (approximately 120 kDa), STAT3 and Ikaros, yet their FAAH is not activated by leptin or progesterone. These data, corroborated by transient expression and electrophoretic mobility-shift assays, demonstrate an unprecedented cell-specific regulation of the FAAH gene, which has important implications for the control of tone and activity of AEA along the neuroimmune axis.
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PMID:Differential regulation of fatty acid amide hydrolase promoter in human immune cells and neuronal cells by leptin and progesterone. 1560 54


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