linoleic-acid and Neuroblastoma

High-risk neuroblastoma (HR-NB) has a significantly lower survival rate compared to low- and intermediate-risk NB (LIR-NB) due to the lack of risk classification diagnostic models and effective therapeutic targets. The present study aims to characterize the differences between neuroblastomas with different risks through transcriptomic and metabolomic, and establish an early diagnostic model for risk classification of neuroblastoma.Plasma samples from 58 HR-NB and 38 LIR-NB patients were used for metabolomics analysis. Meanwhile, NB tissue samples from 32 HR-NB and 23 LIR-NB patients were used for transcriptomics analysis. In particular, integrative metabolomics and transcriptomic analysis was performed between HR-NB and LIR-NB. A total of 44 metabolites (P < 0.05 and fold change > 1.5) were altered, including 12 that increased and 32 that decreased in HR-NB. A total of 1,408 mRNAs (P < 0.05 and |log

Topics: Arginine; Humans; Linoleic Acid; Metabolomics; Neuroblastoma; Transcriptome; Vitamin A

Parkinson's disease (PD) is the second most prevalent neurodegenerative disease after Alzheimer's disease. The principal pathological feature of PD is the progressive loss of dopaminergic neurons in the ventral midbrain. This pathology involves several cellular alterations: oxidative stress, mitochondrial dysfunction, loss of proteostasis, and autophagy impairment. Moreover, in recent years, lipid metabolism alterations have become relevant in PD pathogeny. The modification of lipid metabolism has become a possible way to treat the disease. Because of this, we analyzed the effect and possible mechanism of action of linoleic acid (LA) on an SH-SY5Y PD cell line model and a PD mouse model, both induced by 6-hydroxydopamine (6-OHDA) treatment. The results show that LA acts as a potent neuroprotective and anti-inflammatory agent in these PD models. We also observed that LA stimulates the biogenesis of lipid droplets and improves the autophagy/lipophagy flux, which resulted in an antioxidant effect in the in vitro PD model. In summary, we confirmed the neuroprotective effect of LA in vitro and in vivo against PD. We also obtained some clues about the novel neuroprotective mechanism of LA against PD through the regulation of lipid droplet dynamics.

Topics: Animals; Autophagy; Cell Line, Tumor; Humans; Linoleic Acid; Lipid Droplets; Mice; Neuroblastoma; Neurodegenerative Diseases; Oxidopamine; Parkinson Disease

The 24(S)-hydroxycholesterol (24S-OHC), which plays an important role in maintaining brain cholesterol homeostasis, has been shown to possess neurotoxicity. We have previously reported that 24S-OHC esterification by ACAT1 and the resulting lipid droplet (LD) formation are responsible for 24S-OHC-induced cell death. In the present study, we investigate the functional roles of 24S-OHC esters and LD formation in 24S-OHC-induced cell death, and we identify four long-chain unsaturated fatty acids (oleic acid, linoleic acid, arachidonic acid, and DHA) with which 24S-OHC is esterified in human neuroblastoma SH-SY5Y cells treated with 24S-OHC. Here, we find that cotreatment of cells with 24S-OHC and each of these four unsaturated fatty acids increases prevalence of the corresponding 24S-OHC ester and exacerbates induction of cell death as compared with cell death induced by treatment with 24S-OHC alone. Using electron microscopy, we find in the present study that 24S-OHC induces formation of LD-like structures coupled with enlarged endoplasmic reticulum (ER) lumina, and that these effects are suppressed by treatment with ACAT inhibitor. Collectively, these results illustrate that ACAT1-catalyzed esterification of 24S-OHC with long-chain unsaturated fatty acid followed by formation of atypical LD-like structures at the ER membrane is a critical requirement for 24S-OHC-induced cell death.

Topics: Acetyl-CoA C-Acetyltransferase; Arachidonic Acid; Brain; Cell Death; Cell Line, Tumor; Endoplasmic Reticulum; Esterification; Gene Expression Regulation; Humans; Hydroxycholesterols; Linoleic Acid; Lipid Droplets; Neuroblastoma; Neurons; Oleic Acid

Neurodegenerative diseases are linked to neuronal cell death and impairment of neurite outgrowth. An edible mushroom, Pleurotus giganteus was found to stimulate neurite outgrowth in vitro but the chemical constituents and the underlying mechanism is yet to be elucidated. The chemical constituents of P. giganteus (linoleic acid, oleic acid, cinnamic acid, caffeic acid, p-coumaric acid, succinic acid, benzoic acid, and uridine) were tested for neurite outgrowth activity. Uridine (100 μM) was found to increase the percentage of neurite-bearing cells of differentiating neuroblastoma (N2a) cells by 43.1 ± 0.5%, which was 1.8-fold higher than NGF (50 ng/mL)-treated cells. Uridine which was present in P. giganteus (1.80 ± 0.03 g/100g mushroom extract) increased the phosphorylation of extracellular-signal regulated kinases (ERKs) and protein kinase B (Akt). Further, phosphorylation of the mammalian target of rapamycin (mTOR) was also increased. MEK/ERK and PI3K-Akt-mTOR further induced phosphorylation of cAMP-response element binding protein (CREB) and expression of growth associated protein 43 (GAP43); all of which promoted neurite outgrowth of N2a cells. This study demonstrated that P. giganteus may enhance neurite outgrowth and one of the key bioactive molecules responsible for neurite outgrowth is uridine.

Topics: Animals; Benzoic Acid; Biomarkers; Caffeic Acids; Cell Line, Tumor; Cinnamates; Coumaric Acids; Cyclic AMP Response Element-Binding Protein; Enzyme-Linked Immunosorbent Assay; Extracellular Signal-Regulated MAP Kinases; GAP-43 Protein; Linoleic Acid; MAP Kinase Signaling System; Mice; Neurites; Neuroblastoma; Oleic Acid; Phosphorylation; Pleurotus; Propionates; Proto-Oncogene Proteins c-akt; Signal Transduction; Succinic Acid; Uridine

Conjugated linoleic acid (CLA) plays important roles in physiological conditions. The aim of present study was to explore the effects of CLA on the cleavage of amyloid precursor protein (APP) and the potential mechanism involved. The effects of CLA on intracellular APP, BACE1 (β-site APP Cleaving Enzyme1, BACE1), a disintegrin and metalloprotease (ADAM10) and extracellular sAPPα (soluble) were analyzed by RT-PCR, Western blot and ELISA in SH-SY5Y cells. Our study indicated that CLA significantly decreased the expression of BACE1 and increased the extracellular secretion of sAPPα, but not affected the levels of APP and ADAM10. The study also revealed that the nuclear receptor peroxisome proliferators activated receptor γ (PPARγ) played an important role in the CLA-induced intracellular BACE1 decrease, as well as the extracellular sAPPα increase through knockdown of PPARγ transcription using siRNA. We hypothesize that CLA acts as an agonist or ligand, which binds with PPARγ and leads to the increase in APP cleavage via α-secretase-mediated pathway and the decrease in the deposition of Aβ.

Topics: ADAM Proteins; ADAM10 Protein; Amyloid beta-Protein Precursor; Amyloid Precursor Protein Secretases; Analysis of Variance; Aspartic Acid Endopeptidases; Cell Line, Tumor; Cell Survival; Dose-Response Relationship, Drug; Enzyme-Linked Immunosorbent Assay; Gene Expression Regulation, Neoplastic; Humans; Linoleic Acid; Membrane Proteins; Neuroblastoma; PPAR gamma; RNA, Messenger

We investigated the ability of different hydroperoxides generated by lipoxygenase isozymes to induce programmed cell death (PCD) in human cells. Erythroleukemia K562 and neuroblastoma CHP100 cells were used, because they showed high basal activity of lipoxygenase. The hydroperoxides generated by 5-, 12-, or 15-lipoxygenases from linoleate, linolenate, or arachidonate, and the corresponding hydroxides, were able to induce PCD in both cell types, in a concentration- and time-dependent manner. After 24 h, K562 and CHP100 cells showed 2.5- to 3.5-fold more apoptotic bodies than the untreated controls. PCD elicited by lipoxygenase products was independent of intracellular glutathione concentration, and did not require mRNA transcription or protein synthesis. On the other hand, lipoxygenase products evoked an immediate and sustained rise in cytoplasmic calcium (within seconds), followed by mitochondrial uncoupling (within hours). Unlike the hydro(pero)xides, the terminal products of the arachidonate cascade (i.e., leukotrienes, prostaglandins and thromboxane) were not cytotoxic.

Topics: alpha-Linolenic Acid; Apoptosis; Arachidonic Acid; Calcium; Cell Survival; Dose-Response Relationship, Drug; Enzyme Activation; Glutathione; Humans; Isoenzymes; K562 Cells; Leukemia, Erythroblastic, Acute; Leukotrienes; Linoleic Acid; Lipid Peroxides; Lipoxygenase; Mitochondria; Neuroblastoma; Prostaglandins; Thromboxane B2; Time Factors; Tumor Cells, Cultured; Uncoupling Agents

We have studied the biosynthesis of long chain polyunsaturated fatty acids (LC-PUFA) from their precursors in cultured cells undergoing physiological modifications, or under the influence of lipid-lowering drugs or ethanol. The formation of arachidonic acid (AA, 20:4 n-6) from the percursor linoleic acid (LA, 18:2 n-6) in the neuroblastoma cells SK-N-BE is enhanced at early stages of differentiation, and declines when differentiation is complete, in concomitance with maximal accumulation of AA in cell lipids. In the monocytic cells THP-1, the biosynthesis of LC-PUFA is also enhanced by treatment with the HMGCoA reductase inhibitor simvastatin (S), an effect which is reverted by mevalonate and other intermediates of cholesterol synthesis. Maximal activation of LC-PUFA synthesis by S occurs at concentrations lower than those required for maximal inhibition of cholesterol synthesis. In the hepatoma cells HepG2, ethanol decreases the biosynthesis of LC-PUFA while potentiating the incorporation of acetate into cholesterol. LC-PUFA synthesis appears thus to be modulated in the course of cell differentiation and complex interactions between LC-PUFA and cholesterol synthesis occur, as judged from data obtained through pharmacological manipulations.

Topics: Arachidonic Acid; Carcinoma, Hepatocellular; Cell Differentiation; Cell Line; Cholesterol; Enzyme Inhibitors; Ethanol; Fatty Acids, Unsaturated; Humans; Hydroxymethylglutaryl-CoA Reductase Inhibitors; Linoleic Acid; Linoleic Acids; Liver Neoplasms; Lovastatin; Mevalonic Acid; Monocytes; Neuroblastoma; Simvastatin; Tumor Cells, Cultured

Nordihydroguaiaretic acid (NDGA; a lipoxygenase inhibitor), LY-270766 (an inhibitor of 5-lipoxygenase), and the diacylglycerol lipase inhibitor RG 80267 completely eliminated potassium-evoked release of [3H]-noradrenaline ([3H]NA) from the human neuroblastoma clone SH-SY5Y with IC50 values of 10, 15, and 30 microM, respectively. In contrast, these inhibitors only partially inhibited carbachol-evoked release and had little effect on the calcium ionophore A23187-evoked release of NA in this cell line. Arachidonic acid partially inhibited potassium- and A23187-evoked release but did not reverse the inhibition of potassium-evoked release observed in the presence of RG 80267. These studies suggest that arachidonic acid (or its lipoxygenase products) are not important intermediates in the regulation of exocytosis in SH-SY5Y. This conclusion is strengthened by our studies in which SH-SY5Y cells were grown in medium supplemented with bovine serum albumin-linoleic acid (50 microM). Under these conditions there was a selective increase in content of membrane polyunsaturated fatty acids of the omega 6 series, including arachidonic acid; however, these changes did not effect potassium-, veratridine-, carbachol-, or calcium ionophore-evoked release of [3H]NA.

Topics: Arachidonic Acid; Calcimycin; Carbachol; Cyclohexanones; Eicosanoids; Exocytosis; Humans; Linoleic Acid; Linoleic Acids; Lipase; Lipoxygenase Inhibitors; Masoprocol; Neuroblastoma; Norepinephrine; Organic Chemicals; Potassium; Tumor Cells, Cultured

Following earlier observations that increasing the polyunsaturated fatty-acid (PUFA) content of N1E-115 neuroblastoma cells elevated basal and adenosine (Ado)-stimulated intracellular cyclic AMP (cAMP) formation, we carried out studies to determine the mechanism(s) by which PUFA exerted their modulatory effects. Basal increases in cAMP in the PUFA-enriched (PUFA+) cells were evident with short (60 sec) exposure to a phosphodiesterase inhibitor (Ro 20-1724), and increased to a maximum at 20 min; they were not observed in the absence of Ro 20-1724. Forskolin-stimulated cAMP formation in the presence of the Ro compound was 2- to 3-fold higher in the PUFA+ cells. Basal elevations in cAMP were reduced by approximately 70% by exposing the PUFA+ cells to Ado deaminase (ADA) or to an Ado antagonist, and were further increased by inhibiting ADA, which suggested that they could be producing endogenous Ado that activated stimulatory Ado receptors. However, this did not appear to involve PUFA-mediated stimulation of 5'-nucleotidase activity or inhibition of [3H]Ado uptake. Overall, the results of this study indicated that multiple mechanisms are involved in PUFA modulation of cAMP formation.

Topics: 4-(3-Butoxy-4-methoxybenzyl)-2-imidazolidinone; 5'-Nucleotidase; Adenosine; Basal Metabolism; Colforsin; Cyclic AMP; Dipyridamole; Linoleic Acid; Linoleic Acids; Neuroblastoma; Neurons; Pentostatin; Receptors, Purinergic; Tumor Cells, Cultured

Sodium nitroprusside (SNP) stimulates cGMP formation to a greater extent in 20,000 g supernatant fractions of the human neuroblastoma clones NB1-G and SH-SY5Y than in the human astrocytoma clone D384. This suggests that these cell lines contain the soluble form of guanylate cyclase. Arachidonic, 8,11,14- and 11,14,17-eicosatrienoic acids inhibit SNP (10(-4) M)-stimulated cGMP formation more potently than the C18 unsaturated fatty acids linolenic and linoleic acids in D384 and NB1-G. In contrast the C20 saturated fatty acid, arachidic acid had little effect even at 10(-4) M concentration. In addition arachidonic and 8,11,14-eicosatrienoic acids inhibited basal guanylate cyclase activity, in NB1-G, over the same concentration range as they inhibited SNP-stimulated cGMP formation. No evidence could be obtained for the stimulation of guanylate cyclase by arachidonic acid in either NB1-G or D384. These results provide further support for suggestions that arachidonic acid or its metabolites may be important regulators of cGMP formation in the nervous system.

Topics: 8,11,14-Eicosatrienoic Acid; Arachidonic Acid; Arachidonic Acids; Astrocytoma; Cyclic GMP; Fatty Acids, Unsaturated; Ferricyanides; Guanylate Cyclase; Humans; Linoleic Acid; Linoleic Acids; Linolenic Acids; Neuroblastoma; Nitroprusside; Tumor Cells, Cultured

The effects of membrane polyunsaturated fatty acids (PUFA) on opiate peptide-mediated inhibition of basal and prostaglandin E1-stimulated cyclic AMP formation were examined in intact N1E-115 neuroblastoma cells. Addition of opiate peptides such as methionine 5-enkephalin (metEnk) to control cultures and to cultures that had been supplemented for 48 hr with 50 microM linoleic acid resulted in dose-dependent decreases in cAMP formation; these decreases were blocked by naloxone. Maximum inhibition of basal cyclase activity was 50-55% in both control and PUFA-enriched cells; however, half-maximal inhibition required ten times more metEnk in supplemented cultures than in controls. This is consistent with our observation that the affinity of binding of [tyrosyl-3',5'-3H(N)](2-D-alanine-5-D-leucine)enkephalin ([3H]DADLE) to intact PUFA-enriched cells was lower than that to control cells. Receptor density was not modified as a result of supplementation. Addition of prostaglandin E1 (PGE1) to the cells produced rapid dose-dependent increases in cAMP formation. Maximum responses were higher in PUFA-enriched than in control cells (1924 and 972 pmol cAMP formed/mg protein respectively). Also, the apparent value for EC50 for PGE1 was consistently lower in supplemented cultures. MetEnk reduced PGE1-stimulated cAMP formation by 45-55% in both control and supplemented cells, and values for IC50 were similar (approximately 30 nM) in both. In the presence of the opiate peptide, values for EC50 for PGE1 were similar in control and PUFA-enriched cultures (0.07 and 0.09 microM respectively). The data from these studies suggest that membrane PUFA increase the efficiency of coupling of receptors that stimulate cAMP formation and decrease the efficiency of those that mediate inhibition.

Topics: Adenylyl Cyclase Inhibitors; Alprostadil; Cell Division; Cell Membrane; Cyclic AMP; Endorphins; Enkephalin, Leucine; Enkephalin, Leucine-2-Alanine; Enkephalin, Methionine; Fatty Acids, Unsaturated; Fluorescence Polarization; Kinetics; Linoleic Acid; Linoleic Acids; Naloxone; Neuroblastoma; Receptors, Opioid; Tumor Cells, Cultured

Receptor-mediated cyclic GMP formation in N1E-115 murine neuroblastoma cells appears to involve oxidative metabolism of arachidonic acid. Evidence in support of this includes the blockade of this response by lipoxygenase inhibitors, e.g., eicosatetraynoic acid (ETYA) or other metabolic perturbants, e.g., methylene blue. It was recently discovered that the lipoxygenase products 15-hydroxyeicosatetraenoic (15-HETE) acid and 12-HETE, like ETYA, were inhibitors of M1 muscarinic receptor-mediated cyclic GMP formation. In the present report, the effects of monoHETEs are explored in more detail, particularly with regard to the function of the muscarinic receptor. Like 12-HETE and 15-HETE (IC50 = 13 and 11 microM, respectively), 5-HETE inhibited the cyclic GMP response to the muscarinic receptor (IC50 = 10 microM). All three of these monoHETEs were shown also to be inhibitors of the cyclic GMP responses to receptors stimulated by carbachol, histamine, thrombin, neurotensin, and bradykinin. 15-HETE was shown to inhibit the muscarinic receptor-mediated response in a complex manner (apparent noncompetitive and uncompetitive components; IC50 = 18 and 2 microM, respectively). 15-HETE did not inhibit either the M1 muscarinic receptor-stimulated release of [3H]inositol phosphates from cellular phospholipids or the M2 muscarinic receptor-mediated inhibition of hormone (prostaglandin E1)-induced AMP formation. It seemed possible that the monoHETEs could enter into biochemical pathways for arachidonate in N1E-115 cells. [3H]Arachidonate and the three [3H]-monoHETEs all rapidly labeled the membrane lipids of intact N1E-115 cells, with each [3H]eicosanoid producing a unique labeling profile. [3H]15-HETE labeling was noteworthy in that 85% of the label found in the phospholipids was in phosphatidylinositol (PI;t1/2 to steady state = 3 min). Exogenous 15-HETE inhibited the labeling of PI by [3H]arachidonate (IC50 = 28 microM) and elevated unesterified [3H]arachidonate levels. Thus, the mechanism of blockade of receptor-mediated cyclic GMP responses by monoHETEs is likely to be more complex than the simple inhibition of cytosolic mechanisms, e.g., generation of a putative second messenger by lipoxygenase, and may involve also alterations of membrane function accompanying the redistributions of esterified arachidonate.

Topics: Animals; Arachidonic Acid; Arachidonic Acids; Cell Line; Cyclic GMP; Hydroxyeicosatetraenoic Acids; Inositol Phosphates; Kinetics; Linoleic Acid; Linoleic Acids; Mice; Neuroblastoma; Receptors, Muscarinic

In cultured neuroblastoma cells (N1E-115), the metabolism of the essential fatty acid, linoleic acid (18:2 (n-6)), to arachidonic acid (20:4(n-6)) can be altered by other fatty acids in a manner supporting a concerted action of the modulating fatty acid on the desaturation and chain elongation enzymes. In further examination of mechanisms involved, cultured glioma (C-6) or neuroblastoma-glioma hybrids (NG-108-15) cells showed similar patterns of activation by some fatty acids (e.g., 20:3(n-6) and 20:4(n-6)), and inhibition (e.g., 18:3(n-3) or 22:6(n-3)) or no effect (e.g., 18:1(n-9), 20:3(n-3)) by others. In contrast, only inhibition by 20:4(n-6) was seen in cultured HeLa cells, suggesting that the intracellular interactions may not be universal in all cell lines. For fatty acids that activate 20:4(n-6) formation, the lag observed when substrate and activator were administered simultaneously was eliminated by preincubation with activator. Maximal activation occurred within 4 h for neuroblastoma and 2 h for glioma; in each cell line activation declined steadily for 10 h after removal of the activator. Inhibition of protein synthesis did not alter activation. As 98% of the fatty acid incorporated was esterified to triacylglycerol or phospholipid and only the triacylglycerol mass expanded, several manipulations to potentially alter the flow of acyl chains between these lipid pools were evaluated using dual-label and pulse-chase experiments. Results suggested that competition between 18:2(n-6) utilization for esterification to phospholipid and the desaturation-chain elongation sequence as well as a more direct and specific interaction of certain fatty acids with the enzymes may influence 20:4(n-6) formation. A model to explain these observations is discussed.

Topics: Arachidonic Acid; Arachidonic Acids; Cell Line; Fatty Acids; Fatty Acids, Essential; Glioma; HeLa Cells; Humans; Kinetics; Linoleic Acid; Linoleic Acids; Neuroblastoma; Phospholipids; Triglycerides

The role of membrane polyunsaturated fatty acids (PUFAs) in the regulation of basal adenylate cyclase activity was examined in intact N1E-115 neuroblastoma cells. Addition of linoleic acid (50 microM) to the culture medium for 48 h resulted in a significant increase in phospholipid PUFA content and in a two- to fivefold increase in basal accumulation of cyclic AMP (cAMP). Both phenomena were reversed on removal of linoleate from the medium. PUFA enrichment stimulated cell proliferation by approximately 20% without altering the relative proportion of cellular protein. The supplemented cells synthesized significantly larger amounts of prostaglandin (PG) E and D than did the controls; however, blockade of PG synthesis by indomethacin or ibuprofen did not alter cAMP formation. Supplemented cells contained higher levels of malondialdehyde (MDA) than did controls, and MDA formation was reduced by coculture with alpha-tocopherol; however, its inclusion in the medium did not affect cAMP accumulation. Linoleate-supplemented cells responded to cyclase-activating agonists to the same extent as did control cells. Responses to inhibitory agonists (e.g., isoproterenol and carbamylcholine) were altered, but not to a sufficient extent to account for the PUFA-dependent increases in basal adenylate cyclase activity.

Topics: Adenylyl Cyclases; Animals; Cell Division; Cells, Cultured; Cyclic AMP; Fatty Acids, Unsaturated; Linoleic Acid; Linoleic Acids; Lipid Peroxides; Membranes; Mice; Neuroblastoma; Protein Biosynthesis; Receptors, Cell Surface

Addition of linoleic acid (50 microM) to culture medium significantly increases levels of polyunsaturated fatty acids (PUFA) in membrane phospholipids of NIE-115 neuroblastoma. Basal levels of cyclic AMP are elevated significantly in supplemented cells. Exogenous prostaglandins (PG) PGE1 and PGD2 stimulate cAMP formation in NIE-115 neuroblastoma. Supplemented cells produce higher levels of PGE and PGD than do control cultures. Inclusion of cyclooxygenase inhibitors in culture medium does not block elevation of cyclic nucleotide in supplemented cells. Endogenous PG production and receptor activation cannot account for increased cAMP in EFA-supplemented neuroblastoma.

Topics: Animals; Clone Cells; Cyclic AMP; Fatty Acids, Unsaturated; Linoleic Acid; Linoleic Acids; Membrane Lipids; Mice; Neuroblastoma; Prostaglandins