Effect of preduodenal lipase inhibition in suckling rats on dietary octanoic acid (C8:0) gastric absorption and plasma octanoylated ghrelin concentration.

Part of medium chain fatty acids (MCFAs) coming from dietary triglycerides (TGs) can be directly absorbed through the gastric mucosa after the action of preduodenal lipase (lingual lipase in the rat). MCFA gastric absorption, particularly that of octanoic acid (C8:0), may have a physiological importance in the octanoylation of ghrelin, the orexigenic gastric peptide acting as an endogenous ligand of the hypothalamic growth hormone secretagogue receptor 1a (GHSR-1a). However, the amount of C8:0 absorbed in the stomach and its metabolic fate still haven't been clearly characterized. The purpose of the present study was to further characterize and quantify the importance of preduodenal lipase activity on the release and gastric absorption of dietary C8:0 and on the subsequent ghrelin octanoylation in the stomach mucosa. Fifteen days old rats received fat emulsions containing triolein or [1,1,1-(13)C]-Tri-C8:0 and a specific inhibitor of preduodenal lipase, 5-(2-(benzyloxy)ethoxy)-3-(3-phenoxyphenyl)-1,3,4-oxadiazol-2(3H)-one or BemPPOX. The fate of the (13)C-C8:0 was followed in rat tissues after 30 and 120min of digestion and octanoylated ghrelin was measured in the plasma. This work (1) demonstrates that part of C8:0 coming from Tri-C8:0 is directly absorbed at the gastric level, (2) allows the estimation of C8:0 gastric absorption level (1.3% of the (13)C-C8:0 in sn-3 position after 30min of digestion), as well as (3) the contribution of rat lingual lipase to total lipolysis and to duodenal absorption of dietary FAs (at least 30%), (4) shows no short-term effect of dietary Tri-C8:0 consumption and subsequent increase of C8:0 gastric tissue content on plasma octanoylated ghrelin concentration.

Impact of a Standard Rodent Chow Diet on Tissue n-6 Fatty Acids, Δ9-Desaturation Index, and Plasmalogen Mass in Rats Fed for One Year.

Although many studies focus on senescence mechanisms, few habitually consider age as a biological parameter. Considering the effect of interactions between food and age on metabolism, here we depict the lipid framework of 12 tissues isolated from Sprague-Dawley rats fed standard rodent chow over 1 year, an age below which animals are commonly studied. The aim is to define relevant markers of lipid metabolism influenced by age in performing a fatty acid (FA) and dimethylacetal profile from total lipids. First, our results confirm impregnation of adipose and muscular tissues with medium-chain FA derived from maternal milk during early infancy. Secondly, when animals were switched to standard croquettes, tissues were remarkably enriched in n-6 FA and especially 18:2n-6. This impregnation over time was coupled with a decrease of the desaturation index and correlated with lower activities of hepatic Δ5- and Δ6-desaturases. In parallel, we emphasize the singular status of testis, where 22:5n-6, 24:4n-6, and 24:5n-6 were exceptionally accumulated with growth. Thirdly, 18:1n-7, usually found as a discrete FA, greatly accrued over the course of time, mostly in liver and coupled with Δ9-desaturase expression. Fourthly, skeletal muscle was characterized by a surprising enrichment of 22:6n-3 in adults, which tended to decline in older rats. Finally, plasmalogen-derived dimethylacetals were specifically abundant in brain, erythrocytes, lung, and heart. Most notably, a shift in the fatty aldehyde moiety was observed, especially in brain and erythrocytes, implying that red blood cell analysis could be a good indicator of brain plasmalogens.

MAPT STUDY: A MULTIDOMAIN APPROACH FOR PREVENTING ALZHEIMER'S DISEASE: DESIGN AND BASELINE DATA.

OBJECTIVE: The Multidomain Alzheimer Preventive Trial (MAPT study) was designed to assess the efficacy of isolated supplementation with omega-3 fatty acid, an isolated multidomain intervention (consisting of nutritional counseling, physical exercise, cognitive stimulation) or a combination of the two interventions on the change of cognitive functions in frail subjects aged 70 years and older for a period of 3 years. Ancillary neuroimaging studies were additionally implemented to evaluate the impact of interventions on cerebral metabolism (FDG PET scans) and atrophy rate (MRIs), as well as brain amyloïd deposit (AV45 PET scans). DESIGN PATIENTS: 1680 subjects (mean age: 75.3 years; female: 64.8 %), enrolled by 13 memory clinics, were randomized into one of the following four groups: omega-3 supplementation alone, multidomain intervention alone, omega-3 plus multidomain intervention, or placebo. Participants underwent cognitive, functional and biological assessments at M6, M12, M24 and M36 visits. The primary endpoint is a change of memory function at 3 years, as assessed by the Free and Cued Selective Reminding test. All participants will be followed for 2 additional years after the 3-years intervention (MAPT PLUS extension study). INTERVENTIONS: 1/Omega-3 supplementation: two soft capsules daily as a single dose, containing a total of 400 mg docosahexaenoic acid (DHA), i.e., 800 mg docosahexaenoic acid per day, for 3 years. 2/ Multidomain intervention: collective training sessions conducted in small groups (6-8 participants) in twelve 120-minute sessions over the first 2 months (two sessions a week for the first month, and one session a week the second month) then a 60-minute session per month in the following three areas: nutrition, physical activity, and cognition until the end of the 3 years. In addition to the collective sessions, individualized preventive outpatient visits exploring possible risk factors for cognitive decline are performed at baseline, M12 and M24. BASELINE POPULATION: For cognition, the mean MMSE at baseline was 28.1 (± 1.6). About 58% and 42% of participants had a CDR score equal to 0 and 0.5, respectively. Regarding mobility status, 200 (11.9%) had a 4-m gait speed lower or equal to 0.8 m/s. According to the Fried criteria, 673 (42.1%) participants were considered pre frail, and 51 (3.2%) frail. The red blood cell DHA content was 26.1 ± 8.1 µg/g. Five hundred and three participants underwent baseline MRI. AV45 PET scans were performed in 271 individuals and preliminary results showed that 38.0% had a cortical SUVR > 1.17, which gave an indication of significant brain amyloïd deposit. DISCUSSION: The MAPT trial is presently the first largest and longest multidomain preventive trial relevant to cognitive decline in older adults with subjective memory complaints. The multidomain intervention designed for the MAPT trial is likely to be easily implemented within the general population.

Comparative effects of well-balanced diets enriched in α-linolenic or linoleic acids on LC-PUFA metabolism in rat tissues.

The intake of the essential fatty acid precursor α-linolenic acid (ALA) contributes to ensure adequate n-3 long-chain polyunsaturated fatty acid (LC-PUFA) bioavailability. Conversely, linoleic acid (LA) intake may compromise tissue n-3 PUFA status as its conversion to n-6 LC-PUFA shares a common enzymatic pathway with the n-3 family. This study aimed to measure dietary ALA and LA contribution to LC-PUFA biosynthesis and tissue composition. Rats were fed with control or experimental diets moderately enriched in ALA or LA for 8 weeks. Liver Δ6- and Δ5-desaturases were analyzed and FA composition was determined in tissues (red blood cells, liver, brain and heart). Hepatic Δ6-desaturase activity was activated with both diets, and Δ5-desaturase activity only with the ALA diet. The ALA diet led to higher n-3 LC-PUFA composition, including DHA in brain and heart. The LA diet reduced n-3 content in blood, liver and heart, without impacting n-6 LC-PUFA composition. At levels relevant with human nutrition, increasing dietary ALA and reducing LA intake were both beneficial in increasing n-3 LC-PUFA bioavailability in tissues.

A restricted cubic spline approach to assess the association between high fat fish intake and red blood cell EPA + DHA content.

BACKGROUND AND AIMS: Fish, especially fatty fish, are the main contributor to eicosapentaenoic (EPA) and docosahexaenoic (DHA) intake. EPA and DHA concentrations in red blood cells (RBC) has been proposed as a cardiovascular risk factor, with <4% and >8% associated with the lowest and greatest protection, respectively. The relationship between high fat fish (HFF) intake and RBC EPA + DHA content has been little investigated on a wide range of fish intake, and may be non-linear. We aimed to study the shape of this relationship among high seafood consumers. METHODS AND RESULTS: Seafood consumption records and blood were collected from 384 French heavy seafood consumers and EPA and DHA were measured in RBC. A multivariate linear regression was performed using restricted cubic splines to consider potential non-linear associations. Thirty-six percent of subjects had an RBC EPA + DHA content lower than 4% and only 5% exceeded 8%. HFF consumption was significantly associated with RBC EPA + DHA content (P [overall association] = 0.021) adjusted for sex, tobacco status, study area, socioeconomic status, age, alcohol, other seafood, meat, and meat product intakes. This relationship was non-linear: for intakes higher than 200 g/wk, EPA + DHA content tended to stagnate. Tobacco status and fish contaminants were negatively associated with RBC EPA + DHA content. CONCLUSION: Because of the saturation for high intakes, and accounting for the concern with exposure to trace element contaminants, intake not exceeding 200 g should be considered.

Mechanisms involved in lipid accumulation and apoptosis induced by 1-nitropyrene in Hepa1c1c7 cells.

1-Nitropyrene (1-NP) is a nitro-polycyclic aromatic hydrocarbon (nitro-PAH) present in diesel exhaust and bound to particular matter in urban air. We show that 1-NP and the referent PAH benzo(a)pyrene (BP) induce apoptosis and a lipid accumulation dependent on cytochrome P450 1A1-metabolites in mouse hepatoma cells, whereas 1-amino-pyrene had no effect. The caspase inhibitor, N-benzyloxycarbonyl-Val-Ala-Asp(O-Me) fluoromethyl ketone (Z-VAD-fmk), inhibits 1-NP-induced apoptosis, but failed to alter 1-NP-triggered lipid accumulation determined by Nile red staining. We further show that cholesterol and fatty acid contents are modified after nitro-PAH exposure and that 1-NP-induced cholesterol level is partially involved in related apoptosis. In parallel, the activity of the stearoyl-CoA desaturase 1 (SCD1), determined by fatty acid analysis, and its expression are reduced by 1-NP. The role of SCD1 in 1-NP-induced apoptosis is demonstrated in cells down-expressing SCD1, in which an increased apoptosis is observed, whereas the SCD1 overexpression elicits the opposite effects. In contrast, changes in SCD1 gene expression have no effect on the induced lipid accumulation. Moreover, 1-NP increases the activity of the AMP-dependent protein kinase (AMPK) leading to a caspase-independent apoptosis. Overall, our study demonstrates that the 1-NP-induced apoptosis is caspase- and AMPK-dependent, and is associated to a decrease of SCD1 expression which results in an alteration of lipid homeostasis.

The consumption of food products from linseed-fed animals maintains erythrocyte omega-3 fatty acids in obese humans.

Based on mechanistic and epidemiological data, we raise the question of the relationship between qualitative dietary polyunsaturated fatty acids (PUFA) changes and increase in obesity. In this double-blind trial, we studied the effects on 160 overweight volunteers (body mass index, BMI >30) of a 90 days experimental diet rich principally in animal fat with a low PUFA/saturated fatty acid (SFA) ratio but a low n-6/n-3 ratio, using animal products obtained from linseed-fed animals. The control diet provided less animal fat, a higher PUFA/SFA ratio and a higher n-6/n-3 ratio. Both diets excluded seafood. In the experimental group, we observed a significant increase in red blood cell (RBC) alpha-linolenic acid content and a slight increase in EPA and DHA derivatives, while in the control group we observed a significant reduction in EPA and DHA content. Between groups now, the difference in the three n-3 fatty acids changes in RBC was significant. This demonstrates that plasma EPA and DHA levels can be maintained without fish if products from linseed-fed animals are used. During the diets, we noted a significant reduction in weight, BMI and hip circumference within both groups of volunteers. However, no significant difference was observed between the control group and the experimental group. Interestingly, 150 days after the end of the trial (i.e., day 240), we noted a significant weight gain in the control group, whereas no significant weight gain was observed in the experimental group. This was also observed for the BMI and hip circumference. Moreover, significant differences in BMI (P < 0.05) and weight (P = 0.05) appeared between the two groups, showing in both cases a smaller increase in the experimental group. During the 90 days trial, we did not observe any differences between groups in terms of total cholesterol, HDL cholesterol, LDL cholesterol or triglycerides, suggesting that the saturate content and the P/S ratio are not as important as the n-6 and n-3 fatty acid composition.

Production of branched-chain aroma compounds by Propionibacterium freudenreichii: links with the biosynthesis of membrane fatty acids.

AIMS: Short branched-chain fatty acids (BCFAs) are cheese flavour compounds, which result from the conversion of branched-chain amino acids (BCAAs). In Swiss cheese, the production of short BCFAs is mainly performed by Propionibacterium freudenreichii and is strain dependent. Our aim was to investigate the possible links between the biosynthesis of short BCFAs and membrane BCFAs in P. freudenreichii. METHODS AND RESULTS: Short and membrane BCFAs were analysed by gas chromatography-mass spectrometry. Two strains differing in their capacities to release short BCFAs were selected. Tri-deuterated-labelled leucine was used in both strains as a precursor of short extracellular iso-BCFAs and of membrane iso-BCFAs. The proportions of anteiso : iso BCFAs synthesized varied as function of the BCAAs provided in the growth medium, from 72 : 28 to 100 : 0, with leucine and valine, and with isoleucine as sole BC precursors, respectively. The branching pattern of short BCFAs exactly matched that of membrane BCFAs, whatever the exogenous BCAAs provided. CONCLUSIONS: The biosynthesis of short BCFAs is closely related to that of membrane BCFAs in P. freudenreichii. SIGNIFICANCE AND IMPACT OF THE STUDY: The biosynthesis of short BCFAs in P. freudenreichii depends more on the strain than on the presence of exogenous BC precursors.

Substitution of dietary oleic acid for myristic acid increases the tissue storage of α-linolenic acid and the concentration of docosahexaenoic acid in the brain, red blood cells and plasma in the rat.

Various strategies have been developed to increase the cellular level of (n-3) polyunsaturated fatty acids in animals and humans. In the present study, we investigated the effect of dietary myristic acid, which represents 9% to 12% of fatty acids in milk fat, on the storage of α-linolenic acid and its conversion to highly unsaturated (n-3) fatty acid derivatives. Five isocaloric diets were designed, containing equal amounts of α-linolenic acid (1.3% of dietary fatty acids, i.e. 0.3% of dietary energy) and linoleic acid (7.0% of fatty acids, i.e. 1.5% of energy). Myristic acid was supplied from traces to high levels (0%, 5%, 10%, 20% and 30% of fatty acids, i.e. 0% to 6.6% of energy). To keep the intake of total fat and other saturated fatty acids constant, substitution was made with decreasing levels of oleic acid (76.1% to 35.5% of fatty acids, i.e. 16.7% to 7.8% of energy) that is considered to be neutral in lipid metabolism. After 8 weeks, results on physiological parameters showed that total cholesterol and low-density lipoprotein-cholesterol did not differ in the diets containing 0%, 5% and 10% myristic acid, but were significantly higher in the diet containing 30% myristic acid. In all the tissues, a significant increasing effect of the substitution of oleic acid for myristic acid was shown on the level of both α-linolenic and linoleic acids. Compared with the rats fed the diet containing no myristic acid, docosahexaenoic acid significantly increased in the brain and red blood cells of the rats fed the diet with 30% myristic acid and in the plasma of the rats fed the diet with 20% myristic acid. Arachidonic acid also increased in the brain of the rats fed the diet with 30% myristic acid. By measuring Δ6-desaturase activity, we found a significant increase in the liver of the rats fed the diet containing 10% of myristic acid but no effect at higher levels of myristic acid. These results suggest that an increase in dietary myristic acid may contribute in increasing significantly the tissue storage of α-linolenic acid and the overall bioavailability of (n-3) polyunsaturated fatty acids in the brain, red blood cells and plasma, and that mechanisms other than the single Δ6-desaturase activity are involved in this effect.

In rat hepatocytes, myristic acid occurs through lipogenesis, palmitic acid shortening and lauric acid elongation.

The origin of myristic acid in mammalian cells and the regulation of its endogenous cellular low concentration are not known. Another intriguing question is the potential metabolic properties of endogenous myristic acid as compared with exogenous myristic acid. In the present paper, we hypothesised and demonstrated that, in liver cells, in addition to the usual fatty acid synthase (FAS) pathway that produces predominantly palmitic acid and minor amounts of myristic acid, part of endogenous cellular myristic acid also comes from a shortening of palmitic acid, likely by peroxisomal ß-oxidation and from lauric acid by elongation. From a nutritional point of view, C16:0 is universally found in natural fats and its shortening to myristic acid could contribute to a non-negligible source of this fatty acid (FA) in the organism. Then, we measured the distribution of endogenously synthesised myristic acid in lipid species and compared it with that of exogenous myristic acid. Our results do not support the hypothesis of different metabolic fates of endogenous and exogenous myristic acid and suggest that whatever the origin of myristic acid, its cellular concentration and lipid distribution are highly regulated.

Mycobacterium tuberculosis Des protein: an immunodominant target for the humoral response of tuberculous patients.

The phoA gene fusion methodology permitted the identification of a new Mycobacterium tuberculosis exported protein, Des. This protein has significant sequence similarities to plant acyl-acyl carrier protein desaturases, which are enzymes involved in general fatty acid biosynthesis as well as in mycolic acid biosynthesis in mycobacteria. As shown by Western blot and enzyme-linked immunosorbent assay experiments, the Des protein is a major B-cell antigen that was recognized by all the tuberculous M. tuberculosis- and M. bovis-infected human patients tested.

Inhibiting delta9-desaturase activity impairs triacylglycerol secretion in cultured chicken hepatocytes.

The relationship between endogenous oleic acid produced by hepatic delta9-desaturase and the secretion of VLDL-triglycerides was investigated in a primary culture of chicken hepatocytes. When the fatty acid compositions of the secreted and intracellular triglycerides (TG) (or triacylglycerols) were compared, an imbalance between monoenes and saturated fatty acids was observed, with the secreted TG being significantly more unsaturated than the intracellular TG. The addition of a mixture of cyclopropenic fatty acids (specific inhibitors of fatty acid desaturation) to the culture medium of cells 24 h before measurement of their delta9-desaturase activity and TG secretion rate caused a significant impairment of both desaturase activity and TG secretion, without affecting total TG synthesis. However, the addition of oleic acid to the culture medium of cells treated with cyclopropenic fatty acids restored the TG secretion rate. Palmitic acid did not restore the TG secretion rate and linoleic acid partly restored the TG secretion rate. Finally, even in the presence of oleic acid in the culture medium of secreting cells, those which had been treated with cyclopropenic fatty acids had a significantly lower TG secretion rate than nontreated cells. Taken together, these results show that TG secretion is highly dependent on the delta9-desaturase activity and suggest that oversecretion of VLDL-TG in chickens and subsequent fattening could originate in a high hepatic delta9-desaturation of saturated fatty acids.

Relationship between hepatic fatty acid desaturation and lipid secretion in the estrogenized chicken.

Desaturation of fatty acids is thought to facilitate their incorporation into glyceride and their subsequent secretion as lipoproteins. However, in the laying hen, the dramatic increase in hepatic lipogenesis is often paralleled by a liver steatosis that may affect egg production and even result in death. The balance between lipid secretion and storage, in relation to the fatty acid desaturation process, was therefore investigated in young male estrogenized chicken. Estrogen stimulation resulted in a dramatic increase in VLDL concentration (40.4 mg/ml versus 0.158 mg/ml in control) and hepatic lipid content (8.61 g/liver versus 1.47 g/liver in control). In estrogenized chickens, VLDL, total liver, and microsomes contained relatively more monounsaturated and less saturated and polyunsaturated fatty acids, whereas hepatic delta 9 desaturase activity was twofold higher. Moreover, in these birds, the proportion of monoenoic fatty acids was greater in VLDL (55%) than in the liver (50%), which was indicative of their preferential secretion. Therefore, under the influence of estrogen, fatty acid synthesis and desaturation are associated with and increased VLDL secretion, which limits the degree of hepatic accumulation of triglyceride and the risk of subsequent steatosis.

Effect of insulin on triacylglycerol synthesis and secretion by chicken hepatocytes in primary culture.

A primary culture system of chicken hepatocytes was developed to study the effects of genetic, hormonal and nutritional factors on hepatic triglyceride (TG) secretion in the chicken, and the effect of insulin on TG synthesis and secretion examined. TG synthesis and secretion was measured using [3H]-glycerol incorporation into cellular and secreted TG. An additional step consisting of brief incubation of the monolayer with trypsin solution to improve harvesting the medium immediately adjacent to the cell monolayer, is also proposed. In our culture system, TG secretion occurred at least up to 75 hr of culture, showing a maximum between 40 and 60 hr of culture. No significant effect of insulin could be observed after 24 hr of culture. A clear stimulatory effect was observed with 10(-9) M insulin concentration after 48 hr. The mean ratio of the secretion rates in the presence or absence of insulin was 2.20 +/- 0.30 (SEM, n = 4). In contrast, the 10(-6) M concentration of insulin had no effect on TG secretion. The use of an additional trypsinization step enhanced the findings obtained by simple removal of the culture medium: a clear stimulatory effect of insulin on TG synthesis was observed after both 24 and 48 hr of culture. Analysis of TG fatty acid composition showed an imbalance of monoene versus saturated fatty acids between cellular and secreted TG, secreted TG being more monounsaturated than cellular TG. These results were obtained in the absence of exogenous fatty acid. An oleic acid supplement in the culture medium did not significantly influence TG secretion. In summary, a primary culture system for chicken hepatocyte was developed. A high level of TG secretion was seen in these cells with or without exogenous fatty acid and this secretion was stimulated by insulin. It was concluded that chicken hepatocytes in primary culture provide a useful model for studying regulation of TG secretion.

Stearoyl-CoA desaturase activity in primary culture of chicken hepatocytes. Influence of insulin, glucocorticoid, fatty acids and cordycepin.

1. Stearoyl-CoA desaturase (delta 9-desaturase) activity was measured in chicken primary hepatocytes, as a function of time in culture. 2. When using fasted donor animals, the desaturase activity was low at the beginning of culture and then increased steadily to a maximum value between 30 and 70 hr of culture. When hepatocyte cultures were prepared from fed animals, enzyme activity was high at the beginning of culture and maintained thereafter at similar values to those obtained in cultured hepatocytes from fasted animals after 30 hr of culture. 3. Insulin significantly enhanced enzyme activity when added to the culture medium at a 10(-9) M concentration, and a small stimulating effect was also observed with 10(-6) M dexamethasone. 4. Linoleic acid (0.5 mM) added to the culture medium as albuminic complex partly inhibited delta 9-desaturase activity. 5. Cordycepin (3' deoxyadenosine) decreased enzyme activity when present at a 3 micrograms/ml concentration in the culture medium. 6. Taken together, the induction of enzyme activity in culture, its impairment by cordycepin and response to insulin and linoleic acid strongly suggest that synthesis and translation of the delta 9-desaturase mRNA occur in chicken hepatocytes in primary culture, and that this cellular model may be a useful tool for further studies on delta 9-desaturase regulatory mechanisms.

Lipogenesis in turkeys and chickens: a study of body composition and liver lipogenic enzyme activities.

1. Carcase composition, fat deposition and the activities of three liver lipogenic enzymes were compared in turkeys and chickens fed ad libitum on two different isocaloric diets, respectively adapted to chickens (C) and to turkeys (T). Diets differed in their protein content, being higher by 60 g/kg in diet T. 2. Chickens were much fatter than turkeys and exhibited higher activities of acetyl-coenzyme A carboxylase (EC 6.4.1.2) and malic enzyme (EC 1.1.1.40). 3. The carcase composition of turkeys was not influenced by the type of diet administered, while chickens fed on diet C were fatter than chickens fed on diet T. Compared to diet T, diet C enhanced malic enzyme activity, whatever the species and age. 4. A good correlation between abdominal fat and total fatness was observed in both species but especially in turkeys. 5. In conclusion, hepatic lipogenesis is much lower in turkeys than in chickens.