Alteration of retinal rod outer segment membrane fluidity in a rat model of Smith-Lemli-Opitz syndrome

Smith-Lemli-Opitz syndrome (SLOS) is caused by an inherited defect in the last step in cholesterol (Chol) biosynthesis, leading to abnormal accumulation of 7-dehydrocholesterol and decreased Chol levels. Progressive retinal degeneration occurs in an animal model of SLOS, induced by treating rats with AY9944, a selective inhibitor of the enzyme affected in SLOS. Here we evaluated alterations in the biochemical and physical properties of retinal rod outer segment (ROS) membranes in this animal model. At 1 month of AY9944 treatment, there were modest alterations in fatty acid composition, but no significant differences in cis-parinaric acid (cPA) spectroscopic parameters in ROS membranes from treated versus control rats. However, at 3 months, ROS docosahexaenoic acid (DHA) content was dramatically reduced, and cPA fluorescence anisotropy values were decreased, relative to controls. Also, 1,6-diphenyl-1,3,5-hexatriene exhibited decreased rotational motion and increased orientational order in ROS membranes from 3 month-old AY9944-treated rats, relative to controls. No significant changes in protein:lipid ratios were observed; however, rhodopsin regenerability was compromised by 3 months of treatment. These findings are consistent with reduced ROS membrane fluidity in the SLOS rat model, relative to controls, primarily due to the dramatic reduction in membrane DHA levels, rather than altered sterol composition.     

Lipidomic profiling analysis reveals the dynamics of phospholipid molecules in Arabidopsis thaliana seedling growth

High-throughput lipidomic profiling provides a sensitive approach for discovering minor lipid species. By using an advance in electrospray ionization tandem mass spectrometry, a large set of phospholipid molecular species(126 species)with high resolution were identified from Arabidopsis seedling;of them 31 species are newly identified(16 are unique in plants),including 13 species of phosphatidic acid(PA), nine phosphatidylcholine, six phosphatidylinositol and three phosphatidylserine. Further analysis of the lipidomic profile reveals dynamics of phospholipids and distinct species alterations during seedling development. PA molecules are found at the lowest levels in imbibition and follow an increasing trend during seedling growth, while phosphatidylethanolamine(PE) molecules show the opposite pattern with highest levels at imbibition and a general decreasing trend at later stages. Of PA molecular species, 34:2-, 34:3-, 36:4-, 36:5-, 38:3- and 38:4-PA increase during radicle emergence, and 34:2- and 34:3-PA reach highest levels during hypocotyl and cotyledon emergence from the seed coat. Conversely, molecular species of PE show higher levels in imbibition and decrease in later stages. These results suggest the crucial roles of specific molecular species and homeostasis of phospholipid molecules in seedling growth and provide insights into the mechanisms of how phospholipid molecules are involved in regulating plant development.     

Effects of salinity on cell growth and docosahexaenoic acid content of the heterotrophic marine microalga Crypthecodinium cohnii

The effects of salinity on cell growth and docosahexaenoic acid (DHA) content of three marine microalgal strains, Crythecodinium cohnii ATCC 30556, C. cohnii ATCC 50051 and C. cohnii RJH were investigated. The lag phases of the three strains increased with increasing salinity in Porphyridium medium. The specific growth rate of C. cohnii ATCC 30556 was the highest at 9 g L⁻¹ NaCl while the other two strains had their highest specific growth rates at 5 g L⁻¹ NaCl. The highest cell dry weight concentrations of 2.51 g L⁻¹ and 1.56 g L⁻¹ were achieved at 9 g L⁻¹ NaCl for C. cohnii ATCC 30556 and ATCC 50051, respectively, while the highest dry weight concentration of 2.49 g L⁻¹ was achieved at 5 g L⁻¹ NaCl for C. cohnii RJH. The highest cell growth yield coefficient on glucose was 0.5 g g⁻¹ for both C. cohnii ATCC 30556 and C. cohnii RJH and 0.45 g g⁻¹ for C. cohnii ATCC 50051. All three strains responded to the change of salinity by modifying their cellular fatty acid compositions. At 9 g L⁻¹ NaCl, C. cohnii ATCC 30556 had the highest total fatty acid content and DHA (C22:6) proportion. In contrast, C. cohnii ATCC 50051 and C. cohnii RJH had the highest DHA content at 5 g L⁻¹ NaCl. C. cohnii ATCC 30556 and ATCC 50051 had the highest DHA yield (131.55 and 68.24 mg L⁻¹ respectively) at 9 g L⁻¹ NaCl while C. cohnii RJH had the highest DHA yield (128.83 mg L⁻¹) at 5 g L⁻¹ NaCl. Received 27 May 1999/ Accepted in revised form 27 August 1999     

Effect of docosahexaenoic acid intake on lipid peroxidation in diabetic rat retina under oxidative stress

Docosahexaenoic acid (DHA) plays an important role in visual function but has a highly oxidation-prone chemical structure. Therefore, we investigated how dietary DHA affects the generation of lipid peroxides in rat retina under oxidative stress in diabetes with/without vitamin E (VE) deficiency. Streptozotocin-induced (50 mg i.p./kg B.W.) diabetic Sprague-Dawley (SD) rats were assigned to four groups: (i) control/VE(+), (ii) DHA/VE(+), (iii) control/VE( - ) and (iv) DHA/VE( - ), and raised for 28 days. We then measured lipid peroxide levels in the retina, serum and liver. With a normal intake of VE, dietary DHA increased only the retinal level of thiobarbituric acid-reactive substances (TBARS) slightly. In contrast, in rats with VE deficiency, dietary DHA increased serum and liver lipid peroxide levels but not in the retina. These results suggest that dietary DHA does not necessarily promote lipid peroxidation in the retina even under high oxidative stress.     

Heart arachidonic acid is uniquely sensitive to dietary arachidonic acid and docosahexaenoic acid content in domestic piglets

This study determined the sensitivity of heart and brain arachidonic acid (ARA) and docosahexaenoic acid (DHA) to the dietary ARA level in a dose–response design with constant, high DHA in neonatal piglets. On day 3 of age, pigs were assigned to 1 of 6 dietary formulas varying in ARA/DHA as follows (% fatty acid, FA/FA): (A1) 0.1/1.0; (A2) 0.53/1.0; (A3–D3) 0.69/1.0; (A4) 1.1/1.0; (D2) 0.67/0.62; and (D1) 0.66/0.33. At necropsy (day 28) higher levels of dietary ARA were associated with increased heart and liver ARA, while brain ARA remained unaffected. Dietary ARA had no effect on tissue DHA accretion. Heart was particularly sensitive, with pigs in the intermediate groups having different ARA (A2, 18.6±0.7%; A3, 19.4±1.0%) and a 0.17% increase in dietary ARA resulted in a 0.84% increase in heart ARA. Further investigations are warranted to determine the clinical significance of heart ARA status in developing neonates.     

Fatty acid composition of a new marine picoplankton species of the Chromophyta

A marine yellowish picoplankton, strain PP301, which was newly isolated from the surface seawater of the western Pacific Ocean was an eminent producer of polyunsaturated fatty acids. Its fatty acids were mostly shared by the shortest saturated form (14:0, 20–30%) and polyunsaturated forms (20:4, EPA and DHA) which accounted for about 50% of the total fatty acids. The amount of intermediate forms in 16 and 18 carbon chains were very little. This composition was consistently observed irrespective of the growth temperatures (15–35 °C).     

海洋破囊壶菌Thraustochytrium sp. FJN-10 DHA生物合成 途径相关延长酶的克隆与表达

二十二碳六烯酸(Docosahexaenoic acid,DHA)是具有各种重要生理功能的高度不饱和脂肪酸.以海洋真菌Thraustochytrium sp.FJN-10为研究对象,利用RT-PCR结合RACE,获得了两个碳链延长酶(TFD6和TFD5)的完整基因,其中TFD6 cDNA全长816 bp,编码271个氨基酸;TFD5 cDNA全长831 bp,编码276个氨基酸.将TFD6、TFD5酶切后分别连接到HindⅢ/Xba Ⅰ处理过的pYES2载体,醋酸锂法转化酿酒酵母感受态细胞,成功构建了延长酶酵母表达系统.气相色谱分析表明TFD6可延长C18:3n-6至C20:3n-6,TFD5可延长C20:5n-3至C22:5n-3.     

Effect of docosahexaenoic acid on interleukin-2 receptor signaling pathway in lipid rafts

Recent studies have shown that polyunsaturated fatty acids (PUFA) regulated the functions of membrane receptors in T cells and suppressed T cell-mediated immune responses. But the molecular mechanisms of immune regulation are not yet elucidated. Lipid rafts are plasma membrane microdomains, in which many receptors localized. The purpose of this study was to investigate the effect of DHA on IL-2R signaling pathway in lipid rafts. We isolated lipid rafts by discontinuous sucrose density gradient ultracentrifugation, and found that DHA could change the composition of lipid rafts and alter the distribution of key molecules of IL-2R signaling pathway, which transferred from lipid rafts to detergent-soluble membrane fractions. These results revealed that DHA treatment increased the proportion of polyunsaturated fatty acids especially n−3 polyunsaturated fatty acids in lipid rafts and changed the lipid environment of membrane microdomains in T cells. Compared with controls, DHA changed the localization of IL-2R, STAT5a and STAT5b in lipid rafts and suppressed the expression of JAK1, JAK3 and tyrosine phosphotyrosine in soluble membrane fractions. Summarily, this study concluded the effects of DHA on IL-2R signaling pathway in lipid rafts and explained the regulation of PUFAs in T cell-mediated immune responses.     

Effect of docosahexaenoic acid on interleukin-2 receptor signaling pathway in lipid rafts

PUFAs have been shown to mediate immune re-sponse especially the functions of T cells[1]. Recent researches have demonstrated that PUFAs can in-crease membrane fluidity and modify the functions of membrane receptors and enzymes in T cell membra-ne[2,3]. M…     

An extraordinary degree of structural specificity is required in neural phospholipids for optimal brain function: n-6 docosapentaenoic acid substitution for docosahexaenoic acid leads to a loss in spatial task performance

This study was conducted to determine whether provision of preformed dietary docosapentaenoic acid (DPAn-6) can replace docosahexaenoic acid (DHA) for brain function as assessed by spatial task performance. A newly modified artificial rearing method was employed to generate n-3 fatty acid-deficient rats. Newborn pups were separated from their mothers at 2 days of age and given artificial rat milk containing linoleic acid (LA), or LA supplemented with 1% DHA (DHA), 1% DPAn-6 (DPA) or 1% DHA plus 0.4% DPAn-6 (DHA/DPA). The animals were then weaned onto similar pelleted diets. At adulthood, behavioural tasks were administered and then the brains were collected for fatty acid analysis. The LA and DPA groups showed a lower (63-65%) brain DHA than the dam-reared, DHA and DHA/DPA groups and this loss was largely compensated for by an increase in brain DPAn-6. The brain fatty acid composition in the DPA group was the same as that in the LA group at adulthood. In the Morris water maze, the LA and DPA groups exhibited a longer escape latency than the dam-reared and DHA groups and had a defect in spatial retention. In conclusion, DPAn-6 could not replace DHA for brain function, indicating a highly specific structural requirement for DHA.     

Seasonal changes in the microbial community of a salt marsh, measured by phospholipid fatty acid analysis

Microbial activity within the environment can have distinct geochemicaleffects, and so changes in a microbial community structure can result ingeochemical change. We examined seasonal changes in both the microbialcommunityand the geochemistry of an inter-tidal salt marsh in north-west England tocharacterise biogeochemical processes occurring at this site.Phospholipid fatty acid (PLFA) analysis of sediment samples collected atmonthly intervals was used to measure seasonal changes in microbial biomass andcommunity structure. The PLFA data were analysed using multivariate techniques(Ward's method and the Mahalanobis distance metric), and we show that the useofthe Mahalanobis distance metric improves the statistical analysis by providingdetailed information on the reasons samples cluster together and identifyingthedistinguishing features between the separate clusters. Five clusters of likesamples were defined, showing differences in the community structure over thecourse of a year.At all times, the microbial community was dominated by PLFA associated withaerobic bacteria, but this was most pronounced in summer (August). Theabundanceof branched fatty acids, a measure of the biomass of anaerobes, started toincrease later in the year than did those associated with aerobes and thefungalbiomarker 18:26 showed a brief late-summer peak.The salt marsh remained mildly oxic throughout the year despite the increase inmicrobial respiration, suggested by the large increases in the abundance ofPLFA, in the warmer months. The conditions therefore remained most favourablefor aerobic species throughout the year, explaining their continual dominanceatthis site. However, as the abundance of PLFA synthesised by anaerobesincreased,increases in dissolved Mn concentrations were observed, which we suggest weredue to anaerobic respiration of Mn(IV) to Mn(II). Overall, the geochemicalconditions were consistent with the microbial community structure and changeswithin it.     

Deuterated docosahexaenoic acid protects against oxidative stress and geographic atrophy‐like retinal degeneration in a mouse model with iron overload

Oxidative stress plays a central role in age‐related macular degeneration (AMD). Iron, a potent generator of hydroxyl radicals through the Fenton reaction, has been implicated in AMD. One easily oxidized molecule is docosahexaenoic acid (DHA), the most abundant polyunsaturated fatty acid in photoreceptor membranes. Oxidation of DHA produces toxic oxidation products including carboxyethylpyrrole (CEP) adducts, which are increased in the retinas of AMD patients. In this study, we hypothesized that deuterium substitution on the bis‐allylic sites of DHA in photoreceptor membranes could prevent iron‐induced retinal degeneration by inhibiting oxidative stress and lipid peroxidation. Mice were fed with either DHA deuterated at the oxidation‐prone positions (D‐DHA) or control natural DHA and then given an intravitreal injection of iron or control saline. Orally administered D‐DHA caused a dose‐dependent increase in D‐DHA levels in the neural retina and retinal pigment epithelium (RPE) as measured by mass spectrometry. At 1 week after iron injection, D‐DHA provided nearly complete protection against iron‐induced retinal autofluorescence and retinal degeneration, as determined by in vivo imaging, electroretinography, and histology. Iron injection resulted in carboxyethylpyrrole conjugate immunoreactivity in photoreceptors and RPE in mice fed with natural DHA but not D‐DHA. Quantitative PCR results were consistent with iron‐induced oxidative stress, inflammation, and retinal cell death in mice fed with natural DHA but not D‐DHA. Taken together, our findings suggest that DHA oxidation is central to the pathogenesis of iron‐induced retinal degeneration. They also provide preclinical evidence that dosing with D‐DHA could be a viable therapeutic strategy for retinal diseases involving oxidative stress.     

A new role for apolipoprotein E: modulating transport of polyunsaturated phospholipid molecular species in synaptic plasma membranes

Phospholipids and their acyl group composition are important in providing the proper membrane environment for membrane protein structure and function. In particular, the highly unsaturated phospholipids in synaptic plasma membranes in the CNS are known to play an important role in modulating receptor function and neurotransmitter release processes. Apolipoprotein E (apoE) is a major apolipoprotein in the CNS, mediating the transport of cholesterol, phospholipids and their fatty acids, particularly in reparative mechanisms during neuronal injury. This study was performed to determine whether deficiency in the apoE gene contributes to an alteration of the phospholipids in synaptic plasma membranes. Phospholipid molecular species were identified and quantitated by HPLC/electrospray ionization-mass spectrometry. Analysis of the different phospholipid classes in membranes of apoE-deficient and C57BL/6 J mice indicated no obvious differences in the distribution of different phospholipid classes but substantial differences in composition of phospholipid molecular species. Of special interest was the prevalence of phospholipids (phosphatidylcholine, diacyl-phosphatidylethanolamine, and phosphatidylserine) with 22:6n-3 in both the sn-1 and sn-2 positions of SPM and these phospholipid species were significantly higher in apoE-deficient mice as compared to control mice. Since polyunsaturated fatty acids in neurons are mainly supplied by astrocytes, these results revealed a new role for apoE in regulating polyunsaturated phospholipid molecular species in neuronal membranes.     

Fatty acid content of the dorsal muscle—an indicator of fat quality in freshwater fish

In 56 samples of freshwater fish, most were low in fat, ≤ 5% of dry weight (D.W.), and the sum of all fatty acids (ΣFA) was about 2% of D.W. Trout, whitefish, and grayling had the highest content of the long-chained FA. of ω3 type, EPA and DHA (1·7–2·6% of D.W.). Two large, low-fat pikes with ΣFA of about12–3% of D.W. and a medium-fat whitefish had the highest ω3/ω6 ratios, 8–9, whereas the fattiest fishes, eels from two lakes and the Baltic (ΣFA =17–26% of D.W.) had lower ω3/ω6 ratios, 1·1–1·8 (ω3 and ω6 FA are two important series of FA). The results indicate that ΣA controls the content of saturated FA (SAFA) and monounsaturated FA (MUFA), whereas the polyunsaturated FA (PUFA) was independent of ΣFA after a break point of about 10%ΣFA of D.W. The P/S ratio (PUFA/SAFA) and the PUFA/ΣFA ratio decreased with increased ΣFA, whereas the ω3/ω6 ratio showed no clear correlation to ΣFA. The difference in fatty acid patterns lay between low-fat and high-fat fishes, rather than between marine and freshwater fishes. The variation, both within and between species of the separate FA is small in fish with similar ΣFA content. Also, low-fat and medium-fat fishes tend to be more dietarily favourable than high-fat fishes, when considering the latest criteria for high nutritional value to humans. Abbreviations used in the text: FA, fatty acids; ΣFA, sum of all FA; AA, arachidonic acid (20 : 4ω6); EPA, eicosapentaenoic acid (20 : 5ω3); DHA, docosahexaenoic acid (22 : 6ω3); SAFA, saturated fatty acids; MUFA, monounsaturated fatty acids; PUFA, polyunsaturated fatty acids; D.W., dry weight; F.W. fresh weight; CV, coefficient of variation; ω3 FA, series of PUFA with the first double bond located at carbon number 3; ω6 FA, series of PUFA with the first double bond located at carbon number 6. The fatty acids are described by three numbers, x:ywz, where x=number of carbon atoms, y=number of double bonds, and z=position of the first double bond counted from the methyl end of the molecule.     

Protective effects of docosahexaenoic acid in staurosporine-induced apoptosis: involvement of phosphatidylinositol-3 kinase pathway

Docosahexaenoic acid (22:6n-3, DHA) is highly enriched in neuronal membranes and is considered to be essential for proper brain function. We have previously demonstrated in Neuro 2A cells that DHA as a membrane component protects cells from apoptotic death induced by serum deprivation (Kim et al. 2000). In the present study we demonstrate that staurosporine (ST) induces apoptosis in Neuro 2A cells and DHA enrichment prior to the ST treatment significantly inhibits the apoptotic cell death, as evidenced by the reduction of caspase-3 activity, cleavage of pro-caspase-3 to active caspase-3, DNA strand-breaking and laddering. Enrichment of cells with other fatty acids such as oleic and arachidonic acids did not exert such an effect, indicating that the antiapoptotic effect was specific to DHA enrichment. Among the several protein kinase inhibitors, only phosphatidylinositol 3-kinase (PI3-K) inhibitors, wortmanin, and LY-294002 abolished the protective effect of DHA in ST-induced apoptosis. Concurrently, ST-treatment significantly decreased the phosphorylation status of Akt at Ser-473 and Thr-308 as well as Akt activity, and this reduction was partially prevented by DHA enrichment. The extent of the antiapoptotic effect of DHA correlated with a time-dependent increase in the phosphatidylserine (PS) content upon DHA enrichment. When cells were enriched with DHA in serine-free medium, the PS increase diminished and the DHA effect on caspase-3 activation as well as Akt phosphorylation in ST-induced apoptosis was no longer apparent, suggesting that DHA's role in accumulating membrane PS is an important component for the observed protection. In summary, DHA enrichment uniquely protects ST-induced apoptosis in a PS- and PI3-K-dependent manner. From these data, we suggest that the antiapoptotic effect of DHA is mediated at least in part through the PI3-K/Akt pathway, facilitated by DHA-induced PS accumulation.     

Cloning and sequencing of clustered genes involved in fatty acid biosynthesis from the docosahexaenoic acid-producing bacterium, Vibrio marinus strain MP-1

A cluster of genes involved in fatty acid biosynthesis (fab) was isolated from docosahexaenoic acid (DHA)-producing Vibrio marinus strain MP-1. This fab gene cluster included five genes highly homologous to the Escherichia coli counterparts, and their order in the cluster was the same with that of the E. coli fab gene cluster except that the latter included the additional fabH gene. These fab genes should be involved in early steps of DHA biosynthesis in V. marinus strain MP-1.     

Neonatal lamb vigour is improved by feeding docosahexaenoic acid in the form of algal biomass during late gestation

To determine whether feeding a sustainable, algal source of docosahexaenoic acid (DHA) to sheep during late pregnancy would improve neonatal lamb vigour, 48 English mule ewes, of known conception date, were divided into four treatment groups. For the last 9 weeks of gestation, ewes received one of two dietary supplements: either a DHA-rich algal biomass providing 12 g DHA/ewe per day, or a control supplement based on vegetable oil. The four dietary treatment groups (n = 12) were: control supplement for the duration of the trial (C), DHA supplement from 9 to 6 weeks before parturition (3 week), DHA supplement from 9 to 3 weeks before parturition (6 week) and DHA supplement for the duration of the trial (9 week). Dietary supplements were fed alongside grass silage and commercial concentrate. There was a tendency for gestation length to be extended with increased duration of DHA supplementation (P = 0.08). After parturition, the concentrations of eicosapentaenoic acid (EPA) and DHA in ewe and lamb plasma and colostrum were elevated in line with increased periods of DHA supplementation. Lambs from the 6-week and 9-week groups stood significantly sooner after birth than lambs from the C group (P < 0.05). These data show that neonatal vigour may be improved by the supplementation of maternal diets with DHA-rich algal biomass and that this beneficial effect depends upon the timing and/or duration of DHA allocation.