Kinetics of tumour necrosis factor and prostaglandin production by murine resident peritoneal macrophages as affected by dietary n-3 polyunsaturated fatty acids.

Cell-associated and secreted tumour necrosis factor (TNF), prostaglandin (PG) E2, and 6-keto PGF1 alpha were monitored at various times following in vitro stimulation of resident peritoneal macrophages with lipopolysaccharide (LPS). Macrophages were obtained from mice maintained on diets containing 1.5 wt% n-3 polyunsaturated fatty acids (PUFA)+ 1.5 wt% n-6 fatty acids; 1.5 wt% n-6 fatty acids; or 3 wt% n-6 fatty acids, for 4 weeks. Cell-associated TNF increased transiently in the resident peritoneal macrophages from mice consuming all diets and decreased after TNF secretion had reached maximum and plateaued. Macrophages from mice consuming the n-3 PUFA contained more cell-associated TNF and secreted more TNF than macrophages from mice consuming diets containing n-6 fatty acids only, at all time-points studied. Macrophages from mice consuming the n-3 PUFA showed an earlier increase in cell-associated and secreted TNF compared with macrophages from mice consuming n-6 fatty acids only. Kinetics of maximum TNF production was not affected by the diets and dietary n-3 PUFA did not cause a prolonged increase in TNF secretion. Macrophages from mice consuming the n-3 PUFA produced less PG than macrophages from mice consuming the n-6 fatty acids only. PG secretion increased following appearance of cell-associated TNF but when PG had accumulated in the medium there was no further increase in TNF production.     

A high n-6 polyunsaturated fatty acid diet reduces muscarinic M2/M4 receptor binding in the rat brain

The aim of this study was to examine the influence of different fat diets on muscarinic acetylcholine receptor binding. Nineteen male Sprague–Dawley rats were divided into four groups and fed a diet of either high saturated fat, n-6 polyunsaturated fatty acid (PUFA), n-3 PUFA or low fat (control) for 8 weeks. Using quantitative autoradiography, [³H]pirenzepine binding to muscarinic M1/M4 receptors and [³H]AF-DX384 binding to M2/M4 receptors were measured throughout the brain in all four groups. The main findings were that compared to the low fat control group, M2/M4 receptor binding was significantly reduced in the dorsolateral, dorsomedial and ventromedial parts of the caudate putamen (61–64%, p < 0.05), anterior cingulate cortex (59%, p < 0.01), dentate gyrus and CA1–3 fields of the hippocampus (32–43%, p < 0.01) of rats on a high n-6 PUFA diet; however, no differences in M1/M4 receptor binding densities between the four groups were observed. These results suggest that a diet high in n-6 PUFA, but not of n-3 PUFAs or saturated fat, may selectively alter M2/M4 receptor-mediated signal transduction in the rat brain.     

Effects of dietary supplementation with n-6 and n-3 long-chain polyunsaturated fatty acids on serum lipoproteins and platelet function in hypertriglyceridaemic patients

Twenty-seven patients with hypertriglyceridaemia were given dietary supplementation either with evening primrose oil rich in gammalinolenic acid (GLA, 18:3 n-6) (n = 13) or a marine oil concentrate containing n-3 fatty acids (n = 14) in a double-blind cross-over design during 8 + 8 weeks with olive oil as placebo. During GLA supplementation, increases in GLA and dihomogammalinolenic acid (20:3 n-6) were found in plasma lipid esters and platelet phospholipids, whereas platelet function and serum lipoproteins were unaffected. During supplementation with n-3 fatty acids there was a significant decrease in triglycerides in all lipoprotein fractions with a slight increase in high density lipoprotein and low density lipoprotein cholesterol. A marked increase in the long-chain n-3 fatty acids was found both in plasma and platelets, mainly at the expense of the n-6 fatty acids. No pronounced effects on platelet reactivity could be demonstrated. Our results confirm a triglyceride-lowering effect of n-3 fatty acids, whereas no such effect of GLA could be demonstrated.     

Role of dietary n-3 polyunsaturated fatty acids in type 2 diabetes: A review of epidemiological and clinical studies

The worldwide increasing prevalence of type 2 diabetes mellitus (T2DM) poses an immense public health hazard leading to a variety of complications such as cardiovascular diseases, nephropathy and neuropathy. Diet, as a key component of a healthy human lifestyle, plays an important role in the prevention and management of T2DM and its complications. The dietary n-3 polyunsaturated fatty acids (PUFAs) have been associated with various favourable functions such as anti-inflammatory effects, improving endothelial function, controlling the blood pressure, and reducing hypertriglyceridemia and insulin insensitivity. According to some epidemiological studies, a lower prevalence of T2DM was found in populations consuming large amounts of seafood products, which are rich in n-3 PUFAs. However, the evidence on the relation between fish intake, dietary n-3 PUFAs, and risk of T2DM is controversial. Therefore, this paper aimed to review the epidemiological and clinical studies on the role of dietary n-3 PUFAs in T2DM. Also, the limitations of these studies and the need for potential further research on the subject are discussed.     

Participation of corticosteroids and effects of indomethacin on the acute inflammatory response of rats fed n-6 or n-3 polyunsaturated fatty acid-rich diets

We have previously shown that both n-3 (fish oil) and n-6 (soybean oil) PUFA-rich diets reduce carrageenan-induced paw edema in rats. The present study evaluated the role of corticosteroids, and the effect of indomethacin on this response. Basal (pre-carrageenan) levels of corticosterone were elevated in both lipid diets compared to the chow diet. During inflammation, corticosterone levels increased to a similar extent in the chow and lipid diets. With 2.0 mg/kg indomethacin, edema was reduced in the chow diet and the n-3 diet, while it was not changed in the n-6 diet. In contrast, the 16.6 mg/kg dose of indomethacin induced a mild increase in edema in the chow diet but a pronounced edema increase in the lipid diets. The increase in corticosterone levels induced by carrageenan was either reduced (chow) or completely abolished (lipids) by the treatment with the higher dose of indomethacin, compared to both the control (untreated) group, and the lower dose of indomethacin. These data indicate that both acute inflammation and the response to an antiinflammatory drug were attenuated by n-3 or n-6 PUFA-rich diets. They also showed that indomethacin can have anti- or proinflammatory properties reflecting the extent of the corticosterone inhibition by indomethacin.     

Chronic n-3 polyunsaturated fatty acid diet-deficiency acts on dopamine metabolism in the rat frontal cortex: a microdialysis study

The effects of -linolenic acid diet deficiency on rat dopaminergic metabolism were investigated in the frontal cortex of male 2–3 month-old rats using the microdialysis method. Increased basal levels of dopamine metabolites were observed in the frontal cortex of awake deficient rats, without modification of dopamine levels. Moreover, using KCl perfusion which releases newly synthesized dopamine, no difference was observed in anaesthetized deficient rats versus control rats. In addition, a decrease in dopamine release was observed in anaesthetized deficient rats versus control rats after tyramine stimulation, which is known to induce release of dopamine from vesicular stores. A working model is proposed which suggests that a chronic n-3 polyunsatured fatty acids (PUFA) deficiency may lead to modifications in the internalization of dopamine in the storage pool in the frontal cortex.     

增加n-6脂肪酸对高脂诱导的大鼠胰岛素抵抗和血清游离脂肪酸谱的影响

目的探讨喂养占总热量59％的饱和脂肪酸及n-6脂肪酸代替其中20％热量后，对胰岛素抵抗和血清游离脂肪酸谱的影响。方法45只雄性Wistar大鼠分为3组。对照组喂饲普通饲料，高脂组喂饲提供59％热卡的饱和脂肪酸高脂饲料，n-6脂肪酸组喂饲高脂饲料，其中提供20％热量的饱和脂肪酸由豆油中的C18：2代替。各组共喂饲11周后测定糖耐量、空腹胰岛素、胰岛素耐量、胰岛素抵抗指数、血清瘦素、血脂、血清游离脂肪酸谱。结果①高脂组大鼠从第4周开始体重明显升高，糖负荷后血糖、糖耐量试验中葡萄糖曲线下面积、皮下注射胰岛素后血糖及胰岛素耐量试验中葡萄糖曲线下面积、血清胰岛素、胰岛素抵抗指数、血清瘦素、血清胆固醇和甘油三酯含量均较正常对照组明显升高。高脂组大鼠血清游离脂肪酸谱中，饱和脂肪酸及18烷酸明显升高，不饱和脂肪酸及18碳2烯酸、18碳3烯酸、20碳4烯酸、n-6脂肪酸和n-3脂肪酸均明显下降。②n-6脂肪酸组从第1～10周体重均较高脂组明显减轻，糖负荷后血糖和糖耐量试验中葡萄糖曲线下面积、皮下注射胰岛素后40min、90min血糖及胰岛素耐量试验中葡萄糖曲线下面积、血清胰岛素、胰岛素抵抗指数、血清胆固醇、甘油三酯和低密度脂蛋白含量均较高脂组明显降低；血清瘦素水平、高密度脂蛋白、18碳2烯酸较高脂组明显升高。结论n-6脂肪酸代替诱导大鼠胰岛素抵抗的饱和脂肪酸的20％热量后可以降低胰岛素抵抗，同时血清中18碳2烯酸提高。     

Nutrition and immunity: the immunoregulatory effect of n-6 essential fatty acids is mediated through prostaglandin E

Suppression of cell-mediated immune responses by essential fatty acids (EFA) was demonstrated in mice maintained on a standard laboratory diet for rodents. Daily oral administration of EFA at doses ranging from 125 to 750 mg/kg body weight significantly suppressed local host-versus-graft and graft-versus-host reactions as measured by popliteal lymph node assay. Studies employing immune sera directed against E-type prostaglandin demonstrated that n-6 EFA-induced suppression was mediated through prostaglandin E1. In titration experiments the effect of n-6 EFA on the reactions was dose dependent with enhancement of the responses at low concentrations and suppression at high concentrations.     

Neither the density nor function of striatal dopamine transporters were influenced by chronic n-3 polyunsaturated fatty acid deficiency in rodents

We hypothesized that the chronic dietary deficiency of n-3 polyunsaturated fatty acids (n-3 PUFAs) might affect the density and/or function of dopamine transporters (DAT), which have a major role in regulating the synaptic level of dopamine. This hypothesis was tested by investigating DAT in the striatum using three complementary methods in control and deficient rats. The density of DAT was determined by quantitative autoradiography using [(125)I]PE2I, a specific ligand of this transporter. Functional investigations were performed (i) in vitro by measuring [(3)H]dopamine uptake on synaptosomes, and (ii) in vivo using intracerebral microdialysis. The results demonstrated that neither the density nor the function of DAT were influenced by n-3 PUFA deficiency in the striatum. This suggests lower sensitivity to n-3 PUFA deficiency in the striatum than that previously observed in the frontal cortex.     

Fatty acid profile of plasma and liver lipids in mice depleted in long-chain polyunsaturated (n-3) fatty acids

Considering the numerous features of the metabolic syndrome found in rats depleted in long-chain polyunsaturated (n-3) fatty acids and in the perspective of further work conducted in (n-3)-depleted mice, the fatty acid profile of plasma and liver lipids was assessed in both male and female control and second-generation (n-3)-depleted mice. In addition to gender differences, the major alteration found in the (n-3)-depleted animals consisted in the expected severe depletion of plasma triacylglycerols and phospholipids, as well as liver phospholipids, in C20:5(n-3), C22:5(n-3) and C22:6(n-3). In plasma triacylglycerols, the weight percentages of C18:2(n-6) and C18:3(n-6) were lower in (n-3)-depleted mice than in control animals. In both plasma and liver phospholipids, however, the weight percentages of long-chain polyunsaturated (n-6) fatty acids (C20:4(n-6) and C22:4(n-6)) were higher in (n-3)-depleted mice than in control animals. The C16:1(n-7)/C16:0 and C18:1(n-9)/C18:0 ratio in both plasma and liver phospholipids were also increased in female (n-3)-depleted mice but not so in male animals. Highly significant correlations were found between the weight percentage of each fatty acid in liver versus plasma phospholipids. Taken as a whole, these findings indicate that second-generation mice depleted in (n-3) fatty acids represent a suitable model, in terms of the remodelling of the fatty acid profile in plasma and liver lipids, to investigate the metabolic and functional consequences of such a depletion.     

FADS gene variants modulate the effect of dietary fatty acid intake on allergic diseases in children

Background The association between dietary fatty acid intake and the development of atopic diseases has been inconsistent. This could be due to inter‐individual genetic differences in fatty acid metabolism. Objective The aim of the current study was to assess the influence of FADS1 FADS2 gene cluster polymorphisms on the association between dietary fatty acid intake and atopic diseases and allergic sensitization in 10‐year‐old children. Methods The analysis was based on data from two German prospective birth cohort studies. Data on margarine and fatty acid intake were collected using a food frequency questionnaire. Information on atopic diseases was collected using a questionnaire completed by the parents. Specific IgE against common food and inhalant allergens were measured. Six variants of the FADS1 FADS2 gene cluster (rs174545, rs174546, rs174556, rs174561, rs174575 and rs3834458) were tested. Logistic regression modelling, adjusted for gender, age, maternal education level and study centre, was used to analyse the association between fatty acid intake and atopic diseases stratified by genotype. Results No significant association was found between the six FADS single nucleotide polymorphisms (SNPs) and allergic diseases or atopic sensitization. The total n‐3/total n‐6 ratio was positive associated with an increased risk of hayfever in homozygous major allele carriers ranging from an adjusted odds ratios of 1.25 (95%‐CI: 1.00–1.57) to 1.31 (95%‐CI: 1.01–1.69) across the six tested SNPs although this association was not significant anymore after correcting for multiple testing. Daily margarine intake was significantly associated with asthma [1.17 (1.03–1.34) to 1.22 (1.06–1.40)] in individuals carrying the homozygous major allele. This association was also significant after correcting for multiple testing. Conclusions & Clinical Relevance The association between dietary intake of fatty acids and allergic diseases might be modulated by FADS gene variants in children. Cite this as: M. Standl, S. Sausenthaler, E. Lattka, S. Koletzko, C.‐P. Bauer, H.‐E. Wichmann, A. von Berg, D. Berdel, U. Krämer, B. Schaaf, S. Röder, O. Herbarth, N. Klopp, B. Koletzko and J. Heinrich for the GINIplus and LISAplus Study Group, Clinical & Experimental Allergy, 2011 (41) 1757–1766.     

Essential fatty acid metabolism in long term primary cultures of rat cardiomyocytes: a beneficial effect of n-6:n-3 fatty acids supplementation

In long term (21 days) primary cultures of neonatal rat cardiomyocytes, utilized as a model of in vitro senescence, we investigated the dual effect of the time length in culture and of the supplementation with n-6:n-3 fatty acid mixtures on linoleic (LA) and alpha-linolenic acid (ALA) metabolism. Cardiomyocytes were divided into groups receiving: (1) control medium; (2) control medium plus n-3 fatty acids; (3) and (4) control medium plus n-6 and n-3 fatty acids in the ratio 1:2 or 2:1, respectively. In control cells. senescence caused a reduction in the conversion of LA and ALA, and the decrease in their metabolites was bypassed by the different supplementations. The fatty acid composition of cardiomyocyte lipids was therefore affected by both senescence and supplementation, as evidenced by the n-6:n-3 fatty acid ratio and the unsaturation index (U.I.) in cellular lipids. The final result of ageing in culture and of fatty acid supplementations was in all the groups of cells but one (n-6:n-3, 2:1) an unbalance in the n-6:n-3 fatty acid ratio. All the supplementations were able to counteract the decrease in the U.I. observed with senescence, but only the n-6:n-3 (2:1) was able to do so by increasing the cellular content of the fatty acids which are precursors of anti-aggregation eicosanoids without altering the n-6:n-3 fatty acid ratio.     

Leptin replacement therapy but not dietary polyunsaturated fatty acid alleviates HIV protease inhibitor-induced dyslipidemia and lipodystrophy in mice

A major complication associated with the use of protease inhibitors (PIs) in treatment of HIV-infected patients is lipid abnormalities including dyslipidemia, lipodystrophy, and liver steatosis. Previous studies revealed that these abnormalities are associated with PI-induced accumulation of activated sterol regulatory element binding proteins (SREBPs) in the nucleus of liver and adipose tissues, resulting in constitutive activation of lipid metabolism genes. This study used the mouse model to determine the potential of polyunsaturated fatty acid (PUFA) diet or leptin replacement therapy to alleviate these PI-induced metabolic abnormalities. Results showed that feeding C57BL/6 mice with a PUFA-rich diet failed to normalize plasma cholesterol and triglyceride levels in ritonavir-treated mice. The PUFA-rich diet also had no effect on ritonavir-induced interscapular fat accumulation and liver steatosis. In contrast, daily administration of leptin significantly reversed the elevated plasma cholesterol level induced by ritonavir. Leptin replacement therapy also significantly reduced the ritonavir-induced interscapular fat mass and improved liver steatosis. Taken together, these data suggest that PI-induced lipid abnormalities, especially dyslipidemia, lipodystrophy, and liver steatosis, may be reduced with leptin replacement therapy.     

The effect of dietary supplementation with n-3 polyunsaturated fatty acids on the synthesis of interleukin-1 and tumor necrosis factor by mononuclear cells

We examined whether the synthesis of interleukin-1 or tumor necrosis factor, two cytokines with potent inflammatory activities, is influenced by dietary supplementation with n-3 fatty acids. Nine healthy volunteers added 18 g of fish-oil concentrate per day to their normal Western diet for six weeks. We used a radioimmunoassay to measure interleukin-1 (IL-1 beta and IL-1 alpha) and tumor necrosis factor produced in vitro by stimulated peripheral-blood mononuclear cells. With endotoxin as a stimulus, the synthesis of IL-1 beta was suppressed from 7.4 +/- 0.9 ng per milliliter at base line to 4.2 +/- 0.5 ng per milliliter after six weeks of supplementation (43 percent decrease; P = 0.048). Ten weeks after the end of n-3 supplementation, we observed a further decrease to 2.9 +/- 0.5 ng per milliliter (61 percent decrease; P = 0.005). The production of IL-1 alpha and tumor necrosis factor responded in a similar manner. Twenty weeks after the end of supplementation, the production of IL-1 beta, IL-1 alpha, and tumor necrosis factor had returned to the presupplement level. The decreased production of interleukin-1 and tumor necrosis factor was accompanied by a decreased ratio of arachidonic acid to eicosapentaenoic acid in the membrane phospholipids of mononuclear cells. We conclude that the synthesis of IL-1 beta, IL-1 alpha, and tumor necrosis factor can be suppressed by dietary supplementation with long-chain n-3 fatty acids. The reported antiinflammatory effect of these n-3 fatty acids may be mediated in part by their inhibitory effect on the production of interleukin-1 and tumor necrosis factor.     

Effect of 22-32 carbon n-3 polyunsaturated fatty acids on superoxide production in human neutrophils: synergism of docosahexaenoic acid with f-met-leu-phe and phorbol ester.

Docosahexaenoic (22:6 n-3) and eicosapentaenoic acid (20:5 n-3) stimulated the oxygen-dependent respiratory burst in intact neutrophils in a dose-dependent manner as measured by either superoxide dismutase (SOD)-inhibitable cytochrome c reduction and lucigenin-dependent chemiluminescence. A number of longer chain hexaenoic acids isolated from ram testis (22 to 32 carbon fatty acids) showed a diminishing response with increasing carbon chain length. 22:6 acted synergistically to enhance the responses to two other neutrophil agonists, f-met-leu-phe (FMLP) and the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA). Whereas 22:6-induced chemiluminescence was markedly inhibited by pre-treatment with cytochalasin B, the response to FMLP was augmented, while TPA-induced activation was largely unaffected by cytochalasin B. 22:6-induced activation of neutrophils is independent of protein kinase C, as 22:6, unlike TPA, did not cause the membrane translocation of this enzyme. Furthermore, the putative protein kinase C inhibitor H-7 [1-(5-isoquinolinylsulphonyl)-2-methylpiperazine] had little effect on 22:6-induced chemiluminescence. In contrast, 22:6-induced activation was extremely sensitive to the calmodulin antagonist W-7 [N-(6-aminohexyl)-5-chloro-1-naphthalenesulphonamide], indicating that calmodulin-dependent enzymes may be involved in the responses to 22:6. These results suggest that different mechanisms are involved in the 22:6-, FMLP- and TPA-induced activation of neutrophil NADPH-oxidase.     

Effects of essential fatty acid deficiency on prostaglandin E2 production and cell-mediated immunity in a mouse model of leprosy.

Results from animal and in vitro studies suggest that essential fatty acid (EFA) deficiency enhances cell-mediated immunity by reducing production of prostaglandins with immunosuppressive actions. However, direct experimental evidence that EFA deficiency enhances T-lymphocyte function in vivo has not been obtained. In this study, athymic (nu/nu) mice were infected in the footpads with Mycobacterium leprae and fed a linoleic acid-free diet. These mice, and infected nu/nu mice on control diets, were given an adoptive transfer of M. leprae-primed, T-cell-enriched lymphocytes. After 2 weeks, M. leprae bacilli were harvested from the recipient mice and bacterial viability was determined by the BACTEC system. M. leprae recovered from recipient mice fed control diets displayed little reduction in metabolic activity. In contrast, M. leprae from recipient mice fed the EFA-deficient (EFAD) diet exhibited markedly reduced viability. In vitro, donor cells from M. leprae-primed mice secreted elevated levels of gamma interferon upon exposure to the bacilli. These cells also exhibited an enhanced proliferative response, which was reduced by exogenous prostaglandin E2 (PGE2). In addition, M. leprae-infected granuloma macrophages (Mphi) from EFAD recipient nu/nu mice secreted significantly less PGE2 than granuloma Mphi from mice on control diets. These data suggest that enhanced levels of Mphi-generated PGE2, induced by M. leprae or its constituents, could act as an endogenous negative modulator of the immune response occurring in the microenvironment of the lepromatous granuloma.     

Platelet activating factor increases ciliary activity in the hamster oviduct through epithelial production of prostaglandin E2

We investigated the signal transduction mechanisms associated with an increase in ciliary beat frequency (CBF) produced by platelet activating factor (PAF) in oviductal ciliated cell cultures. In the range of concentrations similar to that produced by preimplantation embryos, PAF increased the CBF in a dose-dependent manner. The addition of PAF and prostaglandin E2 (PGE2) to the cultures produced a synergic increase of ciliary beating, suggesting that PAF and PGE2 signal transduction pathways may be associated. To demonstrate this hypothesis, cyclooxygenase-2 (COX-2) was selectively blocked by a specific inhibitor, NS-398, and the PAF-induced CBF increase was abolished. Moreover, a phospholipase A2 (PLA2) inhibitor, AACOCF3, blocks the PAF-induced CBF increase. PGE2 production by oviductal epithelial cells is stimulated by PAF, and WEB-2086, a PAF-receptor blocker, specifically blocks the PAF-induced PGE2 production. Using the fluorescent indicator fura-2, we measured the effect of PAF on intracellular Ca2+ concentration ([Ca2+]i) in individual ciliated cells. PAF induced a transient increase of [Ca2+]i that was blocked by WEB-2086 or by removal of extracellular Ca2+. We propose a mechanism for PAF-mediated signal transduction in the ciliated cells of the oviductal epithelium. Minimal doses of PAF trigger Ca2+ mobilization in tandem with increased PLA2 activity and a COX-2-mediated increase in PGE2. Local PGE2 production by the oviductal mucosa suggests the presence of an autocrine loop controlling ciliary activity.     

Etomoxir, a fatty acid oxidation inhibitor, increases food intake and reduces hepatic energy status in rats

Etomoxir, an inhibitor of fatty acid oxidation, increases food intake and reported hunger in humans. Work with animal models suggests that other inhibitors of fatty acid oxidation stimulate feeding behavior by acting on the liver. In the following study, we assessed whether etomoxir would increase food intake in rats and to what degree the effects of etomoxir on feeding were associated with changes in hepatic energy status. The effects of etomoxir on hepatic energy status were assessed by measuring liver ATP, ADP, phosphorylation potential, and glycogen content. Blood glucose, free fatty acids, and ketone bodies were also measured to determine the availability of circulating fuels following etomoxir treatment. Etomoxir and methyl palmoxirate (MP; another inhibitor of fatty acid oxidation) increased food intake. Etomoxir, like MP, also reduced hepatic ATP/ADP ratio and phosphorylation potential. In combination with 2,5-anhydro-D-mannitol (an analogue of fructose that produces an increase in feeding by action on the liver), etomoxir synergistically increased food intake and reduced hepatic ATP/ADP ratio. In summary, etomoxir increased food intake and decreased hepatic energy status in the rat. This suggests that etomoxir stimulates feeding by action on the liver.     

Effects of n-3 polyunsaturated fatty acids on COX-2 and PGE2 protein levels in articular cartilage chondrocytes

Introduction Previous studies within our laboratory have shown that supplementation with n‐3 polyunsaturated fatty acids (PUFAs), but not other fatty acids, has a beneficial effect on reducing the expression and activity of degradative and inflammatory factors known to cause damage and destruction of cartilage in arthritic diseases ( Curtis et al. 2000 , 2002 ). Cyclooxygenase (COX), also known as prostaglandin H synthase, catalyses the rate‐limiting step in the formation of inflammatory prostaglandins (PGs) ( Hla & Neilson 1992 ). PG synthesis requires conversion of arachidonic acid to PGH₂ by either the constitutive COX‐1 or by COX‐2, which is induced by inflammatory and mitogenic stimuli ( Smith et al. 2000 ). Prostaglandin E₂ (PGE₂) is produced from PGH₂ and has been shown to have a number of functions including both anti‐ and pro‐inflammatory actions ( Christman et al. 1991 ). The aim of this project was to investigate the effects of n‐3 PUFAs on cyclooxygenase‐2 and prostaglandin E₂ protein levels in articular cartilage chondrocytes. Materials and methods Articular cartilage was obtained both from 7‐day‐old bovine metacarpo‐metatarsophalangeal joints and from human patients undergoing total knee replacement surgery for osteoarthritis (Llandough Hospital, S.Wales, UK). Both explant and monolayer cultures were set up in DMEM with or without 10–300 µg/ml n‐3[eicosapentaenoic acid (EPA)] or n‐6[arachidonic acid (AA)] PUFAs (minimum 8 h, at 37°C, in 5%CO₂) and in the absence or presence of IL‐1 (10 ng/ml) for a further 4 days. Results Total RNA was extracted and RT‐PCR performed using oligonucleotide primers specific to COX‐2 (Invitrogen, UK). COX‐2 mRNA was found to be absent or only present at very low levels in both bovine and human control cultures. After treatment with IL‐1, this expression greatly increased. However, supplementation of IL‐1‐treated cultures with n‐3 PUFA (EPA) resulted in a loss of COX‐2 mRNA expression. In contrast, supplementation with n‐6 PUFA (AA) had no effect. Western blot analysis, using a polyclonal antibody specific to COX‐2 (Santa Cruz Biotechnology Inc., USA), showed that COX‐2 protein was absent in all control samples and the IL‐1 induction of bovine COX‐2 protein could also be reduced when supplemented with n‐3 PUFAs but not n‐6 PUFAs. Using a commercially available PGE₂ ELISA Immunoassay kit, it was possible to analyse the PGE₂ levels present in explant media from bovine or human samples. In one example, in a 74‐year‐old female patient, PGE₂ protein was very low in the cartilage cultured without IL‐1 treatment or PUFA supplementation (control). When the cartilage was treated with IL‐1, there was a huge induction of PGE₂ levels by as much as 60 fold. This induction is in turn reduced greatly with the supplementation of n‐3 PUFA (EPA) but not with n‐6 PUFA (AA). Discussion It has long been accepted that COX‐2 plays an important role in inflammation. COX‐2 has been found in joints affected by arthritic diseases ( Hla & Neilson 1992 ) and in cultures induced by IL‐1. The current study has shown that both the IL‐1‐induced COX‐2 message and protein levels can be reduced with supplementation by n‐3 PUFAs but not n‐6 PUFAs. This work has also shown that with a decrease in COX‐2 protein levels, there is also a corresponding decrease in prostaglandin E₂ levels caused by n‐3 PUFA supplementation.     

Inhibition of elevation of the monkey plasma fatty acid level during stress by administration of prostaglandin

Investigation of the level and composition of free fatty acids in the blood plasma of monkeys revealed definite changes after stress (immobilization) which were prevented by a single injection of prostaglandin E₂ immediately before exposure to the stressor. The role of prostaglandins as a factor in the feedback mechanism limiting the lipolytic effect of neuromediators and hormones, secreted intensively during stress, is analyzed.Central Institute of Research into Regulation of the Heart and Circulation, Academy of Sciences of East Germany, Berlin-Buch. (Presented by Academician of the Academy of Medical Sciences of the USSR N. A. Yudaev.) Translated from Byulleten' Eksperimental'noi Biologii i Meditsiny, Vol. 79, No. 4, pp. 55–58, April. 1975.