Free fatty acid receptor 4-β-arrestin 2 pathway mediates the effects of different classes of unsaturated fatty acids in osteoclasts and osteoblasts

Bone is a dynamic tissue that is constantly remodelled by bone resorbing osteoclasts and bone forming osteoblasts, respectively. A breakdown in the remodelling process underlies several bone diseases such as osteoporosis. Unsaturated fatty acids (UFAs) have been shown to have beneficial effects on bone health. However, the mechanism of action of UFAs in bone remains unclear. Free fatty acid receptor 4 (FFAR4) is expressed in bone cells and preferentially binds ω−3 and ω−7 UFAs. Therefore, we sought to determine if FFAR4 influenced the action of different classes of UFAs in bone cells. FFAR4 and potential signalling pathways, β-arrestin 2 (βarr2) and G_αq, were silenced in RAW264.7 murine macrophages (pre-osteoclasts) and MC3T3-E1 murine pre-osteoblasts. Cell differentiation, activation of signalling pathways and expression of regulatory genes were evaluated. The ω−3 UFAs, docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA), and the ω−7 UFA, palmitoleic acid (PLA), were shown to require the FFAR4/βarr2 signalling pathway to inhibit osteoclast differentiation in RAW264.7 murine macrophages. The ω−6 UFA, arachidonic acid, and the ω−9 UFA, oleic acid (OA), were shown to inhibit osteoclast formation but did not use FFAR4. DHA, EPA, PLA and OA enhanced osteoblast signalling through the FFAR4/βarr2 signalling axis. This study reveals that FFAR4/βarr2 signalling may mediate the bone protective effects of different classes of UFAs in osteoclasts and osteoblasts.     

Fatty acids of membrane phospholipids in Drosophila melanogaster lines showing rapid and slow recovery from chill coma

We investigated the fatty acid compositions of phospholipids in Drosophila melanogaster lines showing rapid (CR), intermediate (CTL), or slow (CS) recovery from chill coma, which were established by artificial selection or by free recombination without selection. Compared to CTL, CS showed a low composition of dienoic acids and a small number of double bonds in the fatty acids. The ratio of unsaturated fatty acids and saturated fatty acids (UFAs/SFAs) was significantly lower in CS than in CTL. CR had higher monoenoic acid composition and lower dienoic acid composition than CTL. In addition, the amount of SFAs was lower and therefore the UFAs/SFAs ratio considerably higher in CR than in CTL. These changes in phospholipid fatty acids probably contributed to losing and maintaining the homeoviscosity of the cellular membranes in CS and CR, respectively, at low temperature and therefore produced their distinct phenotypes in recovery from chill coma.     

Effects of Diets High in Unsaturated Fatty Acids on Socially Induced Stress Responses in Guinea Pigs

Unsaturated fatty acids (UFAs), such as omega-3 and omega-6 poly- and omega-9 monounsaturated fatty acids are important nutrients and major components of neuronal cell membranes. They play a major role in modulating brain functions and physiology and may therefore diminish behavioral and physiological stress reactions in corroboration with decreased cortisol concentrations. Functionally, cortisol itself can modulate several behaviors and also the fatty acid metabolism in the long term. But only little is known about the behavioral and physiological influences of dietary UFAs in a social group, where individuals are regularly exposed to stressful situations. Therefore, the aim of this study was to determine the effects of dietary UFAs on saliva cortisol concentrations and behavioral responses in socially confronted guinea pigs. Three groups of animals were additionally supplemented with 500 mg chia seeds (high in omega-3), walnuts (high in omega-6), or peanuts (high in omega-9) per kg bodyweight each day and compared to a control group. During social confrontation saliva cortisol concentrations significantly increased in all groups, which was accompanied by a loss in bodyweight. However, cortisol levels remained lower in the chia and walnut groups compared to controls. Additionally, the walnut group displayed significantly increased locomotion, while no differences between groups were detected in socio-positive, sexual, or aggressive behaviors. Total plasma omega-3, omega-6, and omega-9 fatty acids were significantly increased in the corresponding groups, due to the dietary supplementations. However, a significant decrease in plasma omega-3 and an increase in plasma n-6 fatty acids were detected in the chia group when comparing the measurements before and after social confrontation. We conclude that both omega-3 and omega-6 polyunsaturated fatty acids can diminish behavioral and physiological stress responses to the social environment, enabling individuals to cope with social stressors, but at the expense of plasma derived omega-3 fatty acids.     

Octadecanoid-derived signals in plants

Signalling pathways that regulate the expression of plant genes are poorly understood. Recent evidence indicates that octadecanoid-derived signals may be parts of signalling pathways in plants in which C(18) fatty acids, liberated from membranes in response to environmental or developmental cues, are converted to intracellular signalling molecules such as jasmoric acid and traumatin. These events are analogous to prostaglandin and leukotriene synthesis in animals.     

细菌不饱和脂肪酸合成研究进展

脂肪酸不仅是细菌细胞膜组分，还是许多生物活性物质的合成原料。不饱和脂肪酸(unsaturated fatty acid, UFA)具有更低的相变温度，是细菌调节细胞膜流动性的重要分子，因此UFA合成途径是重要的抗菌药物筛选靶点。细菌可利用厌氧途径合成UFA，其中模式生物大肠杆菌利用经典的FabA-FabB途径合成UFA，但不同细菌中UFA合成的厌氧途径具有多样性，相关催化酶类也不尽相同；细菌还可以利用需氧途径合成UFA，利用脂肪酸脱饱和酶直接将饱和脂肪酸(saturated fatty acid, SFA)转化为不饱和脂肪酸，而不同脱饱和酶会生成不同结构的UFA，在逆境耐受、致病力等多方面发挥重要作用；细菌还可以利用单加氧酶，将脂肪酸合成途径中癸酰酰基载体蛋白(acyl carrier protein, ACP)转化为顺-3-癸烯酰ACP，并最终合成UFA。细菌脂肪酸合成相关的其他酶类在UFA合成或不同种类UFA调节中也发挥着重要作用。本文系统地总结了细菌UFA合成途径与相关酶类的多样性研究进展，旨在为进一步了解细菌UFA合成机制，并以此为靶点开发抗菌药物等方面提供理论支撑。     

Monounsaturated but not polyunsaturated fatty acids are required for growth of the deep-sea bacterium Photobacterium profundum SS9 at high pressure and low temperature

There is considerable evidence correlating the production of increased proportions of membrane unsaturated fatty acids (UFAs) with bacterial growth at low temperatures or high pressures. In order to assess the importance of UFAs to microbial growth under these conditions, the effects of conditions altering UFA levels in the psychrotolerant piezophilic deep-sea bacterium Photobacterium profundum SS9 were investigated. The fatty acids produced by P. profundum SS9 grown at various temperatures and pressures were characterized, and differences in fatty acid composition as a function of phase growth, and between inner and outer membranes, were noted. P. profundum SS9 was found to exhibit enhanced proportions of both monounsaturated (MUFAs) and polyunsaturated (PUFAs) fatty acids when grown at a decreased temperature or elevated pressure. Treatment of cells with cerulenin inhibited MUFA but not PUFA synthesis and led to a decreased growth rate and yield at low temperature and high pressure. In addition, oleic acid-auxotrophic mutants were isolated. One of these mutants, strain EA3, was deficient in the production of MUFAs and was both low-temperature sensitive and high-pressure sensitive in the absence of exogenous 18:1 fatty acid. Another mutant, strain EA2, produced little MUFA but elevated levels of the PUFA species eicosapentaenoic acid (EPA; 20:5n-3). This mutant grew slowly but was not low-temperature sensitive or high-pressure sensitive. Finally, reverse genetics was employed to construct a mutant unable to produce EPA. This mutant, strain EA10, was also not low-temperature sensitive or high-pressure sensitive. The significance of these results to the understanding of the role of UFAs in growth under low-temperature or high-pressure conditions is discussed.     

Revisiting delta-6 desaturase regulation by C18 unsaturated fatty acids,depending on the nutritional status

Dietary polyunsaturated fatty acids (PUFA) play a key role in regulating delta-6 desaturase (D6D), the key enzyme for long-chain PUFA biosynthesis. Nevertheless, the extent of their effects on this enzyme remains controversial and difficult to assess. It has been generally admitted that C18 unsaturated fatty acids (UFAs) regulate negatively delta-6 desaturase (D6D). This inhibition has been evidenced in regard to a high glucose/fat free (HG/FF) diet used in reference. However, several nutritional investigations did not evidence any inhibition of desaturases when feeding fatty acids.Because the choice of the basal diet appeared to be of primary importance in such experiments, our goal was to reconsider the specific role of dietary UFAs on D6D regulation, depending on nutritional conditions. For that, sixteen adult Wistar rats were fed purified linoleic acid, α-linolenic acid or oleic acid, included in one of two diets at 4% by weight: an HG/FF or a high starch base (HS) where the pure UFAs replaced a mixed vegetable oil. Our results showed first that D6D specific activity was significantly greater when measured in presence of an HG/FF than with an HS/4% vegetable oil diet. Secondly, we found that linoleic and alpha-linolenic acids added to HG/FF reduced the specific activity of D6D. In contrast, when pure UFAs were added to an HS base, D6D specific activities remained unchanged or increased. Concordant results were obtained on D6D mRNA expression.Altogether, this study evidenced the importance of the nutritional status in D6D regulation by C18 UFAs: when used as control, HG/FF diet stimulates D6D compared with a standard control diet containing starch and 4% fats, leading to an overestimation of the D6D regulation by UFAs. Then, UFAs should be considered as repressors for unsaturated fatty acid biosynthesis only in very specific nutritional conditions.     

Monounsaturated but Not Polyunsaturated Fatty Acids Are Required for Growth of the Deep-Sea Bacterium Photobacterium profundum SS9 at High Pressure and Low Temperature

There is considerable evidence correlating the production of increased proportions of membrane unsaturated fatty acids (UFAs) with bacterial growth at low temperatures or high pressures. In order to assess the importance of UFAs to microbial growth under these conditions, the effects of conditions altering UFA levels in the psychrotolerant piezophilic deep-sea bacterium Photobacterium profundum SS9 were investigated. The fatty acids produced by P. profundum SS9 grown at various temperatures and pressures were characterized, and differences in fatty acid composition as a function of phase growth, and between inner and outer membranes, were noted. P. profundum SS9 was found to exhibit enhanced proportions of both monounsaturated (MUFAs) and polyunsaturated (PUFAs) fatty acids when grown at a decreased temperature or elevated pressure. Treatment of cells with cerulenin inhibited MUFA but not PUFA synthesis and led to a decreased growth rate and yield at low temperature and high pressure. In addition, oleic acid-auxotrophic mutants were isolated. One of these mutants, strain EA3, was deficient in the production of MUFAs and was both low-temperature sensitive and high-pressure sensitive in the absence of exogenous 18:1 fatty acid. Another mutant, strain EA2, produced little MUFA but elevated levels of the PUFA species eicosapentaenoic acid (EPA; 20:5n-3). This mutant grew slowly but was not low-temperature sensitive or high-pressure sensitive. Finally, reverse genetics was employed to construct a mutant unable to produce EPA. This mutant, strain EA10, was also not low-temperature sensitive or high-pressure sensitive. The significance of these results to the understanding of the role of UFAs in growth under low-temperature or high-pressure conditions is discussed.     

Effects of 2-hydroxyoleic acid on the structural properties of biological and model plasma membranes

Genetic hypertension is associated with alterations in lipid metabolism, membrane lipid composition and membrane-protein function. 2-Hydroxyoleic acid (2OHOA) is a new antihypertensive molecule that regulates the structure of model membranes and their interaction with certain peripheral signalling proteins in vitro. While the effect of 2OHOA on elevated blood pressure is thought to arise through its influence on signalling proteins, its effects on membrane lipid composition remain to be assessed. 2OHOA administration altered the lipid membrane composition of hypertensive and normotensive rat plasma membranes, and increased the fluidity of reconstituted liver membranes from hypertensive rats. In spontaneously hypertensive rats (SHR), treatment with 2OHOA increased the cholesterol and sphingomyelin content while decreasing that of phosphatidylserine-phosphatidylinositol lipids. In addition, monounsaturated fatty acid levels increased as well as the propensity of reconstituted membranes to form HII-phases. These data suggest that 2OHOA regulates lipid metabolism that is altered in hypertensive animals, and that it affects the structural properties of liver plasma membranes in SHR. These changes in the structural properties of the plasma membrane may modulate the activity of signalling proteins that associate with the cell membrane such as the Galphaq/11 protein and hence, signal transduction.     

Fatty-acid biosynthesis in a branched-chain alpha-keto acid dehydrogenase mutant of Streptomyces avermitilis

Fatty-acid biosynthesis by a branched-chain alpha-keto acid dehydrogenase (bkd) mutant of Streptomyces avermitilis was analyzed. This mutant is unable to produce the appropriate precursors of branched-chain fatty acid (BCFA) biosynthesis, but unlike the comparable Bacillus subtilis mutant, was shown not to have an obligate growth requirement for these precursors. The bkd mutant produced only straight-chain fatty acids (SCFAs) with membrane fluidity provided entirely by unsaturated fatty acids (UFAs), the levels of which increased dramatically compared to the wild-type strain. The levels of UFAs increased in both the wild-type and bkd mutant strains as the growth temperature was lowered from 37 degrees C to 24 degrees C, suggesting that a regulatory mechanism exists to alter the proportion of UFAs in response either to a loss of BCFA biosynthesis, or a decreased growth temperature. No evidence of a regulatory mechanism for BCFAs was observed, as the types of these fatty acids, which contribute significantly to membrane fluidity, did not alter when the wild-type S. avermitilis was grown at different temperatures. The principal UFA produced by S. avermitilis was shown to be delta 9-hexadecenoate, the same fatty acid produced by Escherichia coli. This observation, and the inability of S. avermitilis to convert exogenous labeled palmitate to the corresponding UFA, was shown to be consistent with an anaerobic pathway for UFA biosynthesis. Incorporation studies with the S. avermitilis bkd mutant demonstrated that the fatty acid synthase has a remarkably broad substrate specificity and is able to process a wide range of exogenous branched chain carboxylic acids into unusual BCFAs.     

Effects of 2-hydroxyoleic acid on the structural properties of biological and model plasma membranes

Genetic hypertension is associated with alterations in lipid metabolism, membrane lipid composition and membrane-protein function. 2-Hydroxyoleic acid (2OHOA) is a new antihypertensive molecule that regulates the structure of model membranes and their interaction with certain peripheral signalling proteins in vitro. While the effect of 2OHOA on elevated blood pressure is thought to arise through its influence on signalling proteins, its effects on membrane lipid composition remain to be assessed. 2OHOA administration altered the lipid membrane composition of hypertensive and normotensive rat plasma membranes, and increased the fluidity of reconstituted liver membranes from hypertensive rats. In spontaneously hypertensive rats (SHR), treatment with 2OHOA increased the cholesterol and sphingomyelin content while decreasing that of phosphatidylserine-phosphatidylinositol lipids. In addition, monounsaturated fatty acid levels increased as well as the propensity of reconstituted membranes to form H_II-phases. These data suggest that 2OHOA regulates lipid metabolism that is altered in hypertensive animals, and that it affects the structural properties of liver plasma membranes in SHR. These changes in the structural properties of the plasma membrane may modulate the activity of signalling proteins that associate with the cell membrane such as the Gαq/11 protein and hence, signal transduction.     

Increased unsaturation of lipids in cytoplasmic lipid droplets in DAOY cancer cells in response to cisplatin treatment

Increases in ¹H nuclear magnetic resonance spectroscopy (NMR) visible lipids are a well-documented sign of treatment response in cancers. Lipids in cytoplasmic lipid droplets (LDs) are the main contributors to the NMR lipid signals. Two human primitive neuroectodermal tumour cell lines with different sensitivities to cisplatin treatment were studied. Increases in NMR visible saturated and unsaturated lipids in cisplatin treated DAOY cells were associated with the accumulation of LDs prior to DNA fragmentation due to apoptosis. An increase in unsaturated fatty acids (UFAs) was detected in isolated LDs from DAOY cells, in contrast to a slight decrease in UFAs in lipid extracts from whole cells. Oleic acid and linoleic acid were identified as the accumulating UFAs in LDs by heteronuclear single quantum coherence spectroscopy (HSQC). ¹H NMR lipids in non-responding PFSK-1 cells were unchanged by exposure to 10 μM cisplatin. These findings support the potential of NMR detectable UFAs to serve as a non-invasive marker of tumour cell response to treatment.     

Inhibition of receptor-mediated calcium influx in T cells by unsaturated non-esterified fatty acids.

The effect of omega-3, omega-6 and omega-9 unsaturated fatty acids (UFAs) on receptor-mediated Ca2+ entry was investigated in a T-cell line (JURKAT) by using anti-CD3 antibodies (OKT3) to induce intracellular Ca2+ [( Ca2+]i) increase and Ca2+ influx. All the UFAs, as well as Ni2+ ions and 12-O-tetradecanoylphorbol 13-acetate, decreased the OKT3-induced sustained [Ca2+]i increase to basal levels. Although non-esterified fatty acids activate protein kinase C (PKC) [McPhail, Clayton & Snyderman (1984) Science 224, 622-624; Murakami, Chan & Routtenberg (1986) J. Biol. Chem. 261, 15424-15429], studies using H-7 and analysis of the PKC-dependent phosphorylation of 19 and 80 kDa marker substrates ruled out the involvement of PKC in UFA-induced inhibition of Ca2+ entry. Flow-cytometry analysis showed that UFAs do not interfere with antibody-receptor binding. BSA (0.2%, w/v) reversed the effect of UFAs after these fatty acids have decreased the OKT3-induced [Ca2+]i increase to basal levels. The relevance of these findings and possible mechanisms for inhibition by UFAs of receptor-mediated Ca2+ influx were discussed.     

Interactions of cholesterol and 7‑ketocholesterol with unsaturated fatty acids of different unsaturation degree – The monolayer study

Unsaturated fatty acids (UFAs) are known to lower the level of sterols in blood, which accounts for their cardioprotective effect. To understand the molecular basis of this effect, Langmuir monolayer studies have been performed. A series of UFAs differing in the length of the fatty acid chain and the number of double bonds (oleic acid, OA; linoleic acid, LA; stearidonic acid, SDA; eicosanoic acid, EA) were mixed with cholesterol and its more toxic oxidized derivative, 7‑ketocholesterol (7-KC), abundantly present in atheroma plaques. Strong attractive UFA-sterol interactions were attributed to the formation of “surface complexes”, in which sterol molecules are bound, thereby reducing the amount of free sterol molecules. It has been found that strength of interactions increases with the degree of unsaturation of the acyl chain in UFA molecule. The most attractive interactions correspond to mixtures with SDA containing 70 mol% of 7-KC and 50 mol% of cholesterol. In both cases, the formation of high stability complexes of, respectively, 2:1 and 1:1 sterol/SDA stoichiometry has been proposed. Other complexes of lower stability and 1:2 stoichiometry were postulated for chol (or 7-KC)/LA systems. The complexes of the lowest stability correspond to chol (or 7-KC) mixtures with OA and EA of 1:1 stoichiometry. In all the cases, the interactions of 7-KC with UFAs are more energetically favorable versus cholesterol. The elongation of the hydrophobic chain of UFAs decreased the interactions with the studied sterols. The obtained results can be related to different conformations of the fatty acids chains.