Supply Of Fatty Acids Research Articles

Claudin-3 and occludin tissue content in the glands of colonic mucosa with and without a fecal stream.

The synthesis of the proteins of the apical tight junctions (TJs) depends on a continuous supply of short-chain fatty acids (SCFAs) in colonic epithelium. No studies have evaluated the tissue contents of the TJs proteins in colon segments devoid of a fecal stream. To evaluate the contents of claudin-3 and occludin in the glands of colonic mucosa devoid of a fecal stream. Forty-five rats underwent a diversion of the fecal stream via a left side colostomy and distal mucous fistula. Three groups of 15 animals each were sacrificed at 6, 12 or 18 weeks after surgery. The presence and severity of colitis were defined by histology and inflammation grading scales, respectively. The expression of claudin-3 and occludin were evaluated by immunohistochemistry, and their contents were evaluated by computer-assisted image analysis. Mann-Whitney and Kruskal-Wallis tests were used to evaluate the results at a significance level of 5% (p < 0.05). The colonic epithelium without a fecal stream had a higher degree of inflammation. Colonic glands without a fecal stream showed a reduction in claudin-3 content independent of the time and reduction in occludin content after 12 weeks of intestinal exclusion. The content of claudin-3 and occludin were mainly reduced at the apical surfaces of the colon glands, whereas segments retaining the fecal stream were maintained. The content of claudin-3 was not reduced with time, although the levels of occludin were reduced after 6 weeks and did not vary thereafter. Deficiencies in SCFAs decreased the content of claudin-3 and occludin in colonic glands with the areas of worst inflammation, confirming the importance of an adequate supply of SCFAs in maintaining the integrity of TJ proteins.

Orthogonal Fatty Acid Biosynthetic Pathway Improves Fatty Acid Ethyl Ester Production in Saccharomyces cerevisiae.

Fatty acid ethyl esters (FAEEs) are a form of biodiesel that can be microbially produced via a transesterification reaction of fatty acids with ethanol. The titer of microbially produced FAEEs can be greatly reduced by unbalanced metabolism and an insufficient supply of fatty acids, resulting in a commercially inviable process. Here, we report on a pathway engineering strategy in Saccharomyces cerevisiae for enhancing the titer of microbially produced FAEEs by providing the cells with an orthogonal route for fatty acid synthesis. The fatty acids generated from this heterologous pathway would supply the FAEE production, safeguarding endogenous fatty acids for cellular metabolism and growth. We investigated the heterologous expression of a Type-I fatty acid synthase (FAS) from Brevibacterium ammoniagenes coupled with WS/DGAT, the wax ester synthase/acyl-coenzyme that catalyzes the transesterification reaction with ethanol. Strains harboring the orthologous fatty acid synthesis yielded a 6.3-fold increase in FAEE titer compared to strains without the heterologous FAS. Variations in fatty acid chain length and degree of saturation can affect the quality of the biodiesel; therefore, we also investigated the diversity of the fatty acid production profile of FAS enzymes from other Actinomyces organisms.

Foetal cord blood contains higher portions of n-3 and n-6 long-chain PUFA but lower portions of trans C18:1 isomers than maternal blood

Background/objectiveAn adequate supply of long-chain polyunsaturated fatty acids (LC PUFA) promotes foetal health and development, whereas generally, trans fatty acids (tFA) are considered to negatively interfere with LC PUFA metabolism. Nevertheless, to date, limited data concerning separate trans C18:1, such as t9 and t11, are available for maternal and foetal blood. Therefore, in this study the portions of individual trans C18:1, LC n-6, and n-3 PUFA in lipids of maternal and foetal plasma and erythrocyte membranes of German mother and child pairs (n=40) were analysed.ResultsPortions of linoleic acid and α-linolenic acid as LC precursors were lower (~0.4-fold); whereas the metabolites arachidonic acid (AA, n-6) and docosahexaenoic acid (DHA, n-3) were significantly higher (~2-fold) in foetal than in maternal plasma and erythrocytes. The main tFA in maternal and foetal blood were elaidic acid (C18:1t9; t9) and vaccenic acid (C18:1t11; t11). Portions of t9, t10, t11, and t12 in foetal blood lipids were lower (~0.5-fold) compared with maternal blood. In foetal lipids, t9 was higher than t11. The t9 correlated negatively with eicosapentaenoic acid (n-3) and AA in maternal and foetal lipids; whereas t11 correlated negatively only with foetal total LC n-6 (plasma and erythrocytes) and n-3 PUFA (erythrocytes). No correlation between maternal tFA and foetal PUFA was observed.Conclusions‘Biomagnification’ of LC n-6 and n-3 PUFA AA and DHA in foetal blood was confirmed, whereas single trans isomers were lower compared with maternal blood. Nevertheless, tFA intake, especially from industrial sources, should be as low as possible.

Impacts of dietary fat changes on pregnant women with gestational diabetes mellitus: a randomized controlled study.

This study aims to determine the impact of increasing polyunsaturated fatty acid intake on blood glucose, lipid metabolism, and pregnancy outcomes of pregnant women with gestational diabetes mellitus. Under constant total energy and protein intake, 84 pregnant women with gestational diabetes mellitus were randomly divided into the experimental and control groups, which were given oil-rich and conventional low-oil meals, respectively. After the dietary intervention, the intake and energy supply of fat and the three fatty acids were significantly higher in the experimental group than the control group (p<0.001). The intake and energy supply of polyunsaturated fatty acids increased significantly post-intervention in the experimental group but did not change in the control group. In both the intervention and the control group, fasting blood glucose, 2 h postprandial plasma glucose, and the insulin resistance index decreased significantly post-intervention (p<0.05); the lipid changes were consistent between groups. Pregnancy outcomes did not differ significantly between the two groups (p>0.05). An appropriate increase in polyunsaturated fatty acid intake benefits pregnant women with gestational diabetes mellitus as well as fetuses, as long as the diet therapy follows basic recommendations and total energy intake is strictly controlled.

Long-Chain Polyunsaturated Fatty Acids in Breast-Milk and Erythrocytes and Neurodevelopmental Outcomes in Danish Late-Preterm Infants

Background: The supply of long-chain polyunsaturated fatty acids (LC-PUFA) during pregnancy and early lactation has been shown to affect cognitive development in preterm infants, but the effect on early neurodevelopment of late-preterm infants has not yet been examined. Aim: To examine the fatty acid composition of late-preterm human milk and identify possible associations between infant LC-PUFA status and perinatal as well as 1-year neurobehavioral outcomes. Methods: Mother’s milk and erythrocytes (RBC) were sampled from 53 Danish late-preterm infants (33-36 weeks of gestation) 1 week and 1 month after delivery, and 3 months corrected age. Fatty acid composition was determined by gas-liquid chromatography. Neurodevelopmental outcomes were assessed by the Nicu Network Neurobehavioral Scale (NNNS) at 1 week and 1 month and the Bayley Scales (BSID-III) at 1 year corrected age. Results: We found that breast-milk content of arachidonic acid (AA) and docosahexaenoic acid (DHA) was similar to reported fatty acid compositions of term human milk. Infant RBC-AA decreased from 1 week to 1 month of age and the size of the decrease was associated with better NNNS-scores at 1 month, specifically on regulation (p=0.03). Infant RBC-AA at 1 month was also associated with a lower 1-year corrected age BSID-III score of receptive language (p=0.05) and fine motor development (p=0.03). Infant RBC-DHA did not decrease significantly after delivery and was not associated with any of the developmental outcomes. Conclusion: Breast-milk LC-PUFA content was reflected in the RBC LC-PUFA status of the infant. Early RBC-AA status was associated with both early and long-term neurobehavioral development, but not in a consistent way.

Myocardial Lipid Profiling During Time Course of High Fat Diet and its Relationship to the Expression of Fatty Acid Transporters

Background/Aims: It is well documented that increased fatty acids (FA) supply causes lipid accumulation and insulin resistance in skeletal muscles. Whether the same mechanism is present in the heart is still unclear. Therefore, the goal of our study was to determine the content of specific myocardial lipid fractions during feeding rats a high fat diet (HFD) for 5 weeks. Moreover, the relation between changes in myocardial lipid content, whole body insulin resistance and the expression of fatty acid transporters in each week of HFD was established. Methods: Gas liquid chromatography and high performance liquid chromatography were used to determine the content of lipid fractions in the left ventricle. Expression of selected proteins was estimated by Western blot technique. All measurements were made after each week of HFD. Results: As expected, lipid profile in myocardium was altered by HFD in different weeks of the study with the most intense changes in triacylglycerols, long chain fatty acid-CoA and ceramide. Furthermore, there was a significant elevation of plasmalemmal (the 4^th and the 5^th week) and mitochondrial expression (from the 3^rd to the 5^th week) of fatty acid translocase. Conclusion: High fat diet affects myocardial lipid profile in each week of its duration and causes alternations in FA metabolism in cardiomyocytes.

Culture conditions affect the nutritional value of the copepod Acartia tonsa

Live feed are still necessary for the rearing of larval stages of several fish species, especially marine ones. Compared to Artemia, copepods are of superior quality. This is based on a suite of traits like size, movement, and nutritional value. Copepods are for example usually high in protein and fatty acids. Essential fatty acid profiles reflect to a large degree the fatty acid supply, which provides the opportunity to manipulate fatty acid profiles of, amongst others, copepods. By manipulating nutrient supply of the algae Rhodomonas salina we were able to double essential fatty acid concentrations in naupliar and copepodit life stages of the copepod Acartia tonsa. However, this lead to growth depression rather than to increased growth rates in a series of consumer species, including larval fish. The reason for the growth depression is likely to be mineral deficiencies occurring along with the nutrient manipulation of the algae.

The Omega-3 Fatty Acid Docosahexaenoic Acid Modulates Inflammatory Mediator Release in Human Alveolar Cells Exposed to Bronchoalveolar Lavage Fluid of ARDS Patients.

Background. This study investigated whether the 1 : 2 ω-3/ω-6 ratio may reduce proinflammatory response in human alveolar cells (A549) exposed to an ex vivo inflammatory stimulus (bronchoalveolar lavage fluid (BALF) of acute respiratory distress syndrome (ARDS) patients). Methods. We exposed A549 cells to the BALF collected from 12 ARDS patients. After 18 hours, fatty acids (FA) were added as docosahexaenoic acid (DHA, ω-3) and arachidonic acid (AA, ω-6) in two ratios (1 : 2 or 1 : 7). 24 hours later, in culture supernatants were evaluated cytokines (TNF-α, IL-6, IL-8, and IL-10) and prostaglandins (PGE2 and PGE3) release. The FA percentage content in A549 membrane phospholipids, content of COX-2, level of PPARγ, and NF-κB binding activity were determined. Results. The 1 : 2 DHA/AA ratio reversed the baseline predominance of ω-6 over ω-3 in the cell membranes (P < 0.001). The proinflammatory cytokine release was reduced by the 1 : 2 ratio (P < 0.01 to <0.001) but was increased by the 1 : 7 ratio (P < 0.01). The 1 : 2 ratio reduced COX-2 and PGE2 (P < 0.001) as well as NF-κB translocation into the nucleus (P < 0.01), while it increased activation of PPARγ and IL-10 release (P < 0.001). Conclusion. This study demonstrated that shifting the FA supply from ω-6 to ω-3 decreased proinflammatory mediator release in human alveolar cells exposed to BALF of ARDS patients.

Optimizing initial feeding of the Pike silversideChirostoma estor: oil droplet depletion, point of no return, growth and fatty acid utilization in larvae fed enriched rotifers

The point of no return (PNR) and disappearance of the oil droplet were measured in Chirostoma estor larvae as a function of the time of first feeding. In a separate trial, growth and survival of larvae fed rotifers enriched with Chlorella sp., cod liver oil and corn oil were assessed. Fatty acid and lipid composition of eggs, oil droplets, egg yolk, feed and larvae were also evaluated. The PNR was found between 7 and 8 days posthatching (dph). Total oil droplet depletion occurred between 7 and 11 dph, depending on the time of first feeding. Best growth and survival were obtained in larvae fed with Chlorella-enriched rotifers, followed by those fed cod liver oil-enriched rotifers. In larvae fed corn oil, Chlorella and cod liver oil-enriched rotifers, total oil droplet depletion took place on days 9, 10 and 11, respectively. There was a direct relationship between presence and duration of oil droplets and the survival of larvae under different starvation conditions. The feed source could prolong the existence of the oil droplet depending on particular dietary supply of essential fatty acids; the time of its disappearance could be a useful indicator of larval vigour and health status.

Field production, purification and analysis of high-oleic acetyl-triacylglycerols from transgenic Camelina sativa

A diacylglycerol acetyltransferase, EaDAcT, from Euonymus alatus, synthesizes sn-3 acetyl triacylglycerols (acetyl-TAG) when expressed in Arabidopsis, Camelina and soybean. Compared to most vegetable oils, acetyl-TAGs have reduced viscosity and improved cold temperature properties that confer advantages in applications as biodegradable lubricants, food emulsifiers, plasticizers, and ‘drop-in’ fuels for some diesel engines. A high-oleic Camelina line was engineered to express the EaDAcT gene in order to produce acetyl-TAG oils with fatty acid compositions and physiochemical properties complementary to wild-type acetyl-TAG. The accumulation of acetyl-TAGs at 70mol% of seed TAG in field-grown high-oleic Camelina had minor or no effect on seed weight, oil content, harvest index and seed yield. The total moles of TAG increased up to 27% reflecting the ability to synthesize more acetyl-TAG from the same supply of long-chain fatty acid. Acetyl-TAG could be separated from long-chain TAG by silica column or by reverse phase chromatography. The predominant acetyl-TAG molecular species produced in high-oleic Camelina was acetyl-dioleoyl-glycerol. The crystallization temperature of high-oleic acetyl-TAG (by differential scanning calorimetry at 1.0°C/min) was reduced by 30°C compared to control TAG. The viscosity of high-oleic acetyl-TAG was 27% lower than TAG from the high-oleic control and the caloric content was reduced by 5%. Field production of T4 and T5 transgenic plants yielded over 250kg seeds for oil extraction and analysis.

Peroxisomal enzymes in the liver of rats with experimental diabetes mellitus type 2.

Diabetes mellitus is relatively frequently associated with fatty liver disease. Increased oxidative stress probably plays an important role in the development of this hepatopathy. One of possible sources of reactive oxygen species in liver is peroxisomal system. There are several reports about changes of peroxisomal enzymes in experimental diabetes, mainly enzymes of fatty acid oxidation. The aim of our study was to investigate the possible changes of activities of liver peroxisomal enzymes, other than enzymes of beta-oxidation, in experimental diabetes mellitus type 2. Biochemical changes in liver of experimental animals suggest the presence of liver steatosis. The changes of serum parameters in experimental group are similar to changes in serum of patients with non-alcoholic fatty liver disease. We have shown that diabetes mellitus influenced peroxisomal enzymes by the different way. Despite of well-known induction of peroxisomal beta-oxidation, the activities of catalase, aminoacid oxidase and NADH-cytochrome b(5) reductase were not significantly changed and the activities of glycolate oxidase and NADP-isocitrate dehydrogenase were significantly decreased. The effect of diabetes on liver peroxisomes is probably due to the increased supply of fatty acids to liver in diabetic state and also due to increased oxidative stress. The changes of metabolic activity of peroxisomal compartment may participate on the development of diabetic hepatopathy.

Abstract 15951: Palmitate, Not Oleate, is the Preferred Fatty Acid for Cardiac Triacylglycerol Synthesis

Introduction: Previous studies using cell culture models identified cyto-toxic effects of palmitate and that supplementation with oleate was protective by redirecting palmitate into triacylglycerol (TAG) stores. However, other cull culture studies reported that diacylglycerol transferase 1 (DGAT1), the last enzyme in TAG synthesis, demonstrated a preference for oleate. At present, it is not clear whether the supply of exogenous fatty acids (FA) to the heart is differentially allocated into the endogenous TAG pool. Therefore, the purpose of the present study is to examine the influence of palmitate and/or oleate on cardiac TAG incorporation. METHODS/RESULTS: Hearts were isolated from DGAT1-transgenic (DGAT) and control littermates (CON) and perfused in Langendorff mode with a mixed substrate buffer consisting of glucose, lactate, insulin, and FAs. The FA supply was varied with 0.2mM of both labeled (13C) and unlabeled (12C) FAs in 4 different experiments: 1) 13C/12C palmitate; 2) 13C/12C oleate; 3) 13C palmitate/12C oleate; 4) 13C oleate/12C palmitate. The incorporation of 13C palmitate or 13C oleate into the TAG pool was monitored by 13C NMR spectroscopy. In CON hearts (n=3), the incorporation of palmitate was ~65% higher than oleate when the perfusate contained a homogenous supply of FA. This was also observed in DGAT hearts (n=4) although the incorporation of both palmitate and oleate was ~75% higher compared to CON (P &lt;0.05). In the presence of oleate, palmitate incorporation decreased 25-30% in both CON and DGAT hearts. In contrast, oleate incorporation was diminished by ~50% and ~100% in CON and DGAT hearts, respectively, in the presence of palmitate. CONCLUSIONS: These data suggest that when palmitate and oleate are provided in equal concentrations, palmitate is more readily utilized in the synthesis of endogenous TAG stores in the heart. Furthermore, although overexpression of DGAT increases both oleate and palmitate incorporation, the DGAT1 enzyme demonstrates a preference for palmitate. These findings provide insight into the relationship between exogenous FA supply and endogenous TAG dynamics in the contracting heart.

ATP-citrate lyase regulates cellular senescence via an AMPK- and p53-dependent pathway.

ATP citrate lyase (ACLY) is a key enzyme that is involved in de novo lipogenesis by catalyzing conversion of cytosolic citrate into acetyl CoA and oxaloacetate. Up-regulation of ACLY in various types of tumors enhances fatty acid synthesis and supplies excess acetyl CoA for histone acetylation. However, there is evidence that its enzymatic activity alone is insufficient to explain ACLY silencing-mediated growth arrest in tumor cells. In this study, we found that ACLY knockdown in primary human cells triggers cellular senescence and activation of tumor suppressor p53. Provision of acetyl CoA to ACLY knockdown cells did not alleviate ACLY silencing-induced p53 activation, suggesting an independent role for ACLY activity. Instead, ACLY physically interacted with the catalytic subunit of AMP-activated protein kinase (AMPK) and inhibited AMPK activity. The activation of AMPK under ACLY knockdown conditions may lead to p53 activation, ultimately leading to cellular senescence. In cancer cells, ACLY silencing-induced p53 activation facilitated DNA damage-induced cell death. Taken together, our results suggest a novel function of ACLY in cellular senescence and tumorigenesis.

MicroRNA-451 exacerbates lipotoxicity in cardiac myocytes and high-fat diet-induced cardiac hypertrophy in mice through suppression of the LKB1/AMPK pathway.

In some patients with type 2 diabetes mellitus (DM) without hypertension, cardiac hypertrophy and attenuated cardiac function are observed, and this insult is termed diabetic cardiomyopathy. To date, microRNA (miRNAs or miR) functions in diabetic cardiomyopathy remain to be elucidated. To clarify the functions of miRNAs involved in diabetic cardiomyopathy caused by type 2 DM. C57BL/6 mice were fed a high-fat diet (HFD) for 20 weeks, which induced obesity and type 2 DM. miRNA microarray analyses and real-time polymerase chain reaction revealed that miR-451 levels were significantly increased in the type 2 DM mouse hearts. Because excess supply of saturated fatty acids is a cause of diabetic cardiomyopathy, we stimulated neonatal rat cardiac myocytes with palmitic acid and confirmed that miR-451 expression was increased in a dose- and time-dependent manner. Loss of miR-451 function ameliorated palmitate-induced lipotoxicity in neonatal rat cardiac myocytes. Calcium-binding protein 39 (Cab39) is a scaffold protein of liver kinase B1 (LKB1), an upstream kinase of AMP-activated protein kinase (AMPK). Cab39 was a direct target of miR-451 in neonatal rat cardiac myocytes and Cab39 overexpression rescued the lipotoxicity. To clarify miR-451 functions in vivo, we generated cardiomyocyte-specific miR-451 knockout mice. HFD-induced cardiac hypertrophy and contractile reserves were ameliorated in cardiomyocyte-specific miR-451 knockout mice compared with control mice. Protein levels of Cab39 and phosphorylated AMPK were increased and phosphorylated mammalian target of rapamycin (mTOR) was reduced in cardiomyocyte-specific miR-451 knockout mouse hearts compared with control mouse hearts. Our results demonstrate that miR-451 is involved in diabetic cardiomyopathy through suppression of the LKB1/AMPK pathway.

Regulation of fatty acid binding proteins by hypoxia inducible factors 1α and 2α in the placenta: Relevance to pre-eclampsia

Pre-eclampsia is characterized by placental hypoxia and dyslipidemia. Arachidonic and docosahexanoic acids are essential maternal nutrients for fetal development. They are transported via placental trophoblast cells by membrane and cytosolic fatty acid binding proteins. Others report the expressions of these proteins which are increased in hypoxic trophoblasts. Using bioinformatics, BeWo cells, reporter assays, quantitative real-time PCR and immunoblotting we tested the hypothesis that hypoxia inducible factors 1α (HIF-1α) and/or 2α (HIF-2α) regulate the expressions of FABP1, FABP3, FABP4 and FATP2 proteins. Three hypoxia responsive elements (HRE) were identified in FABP1 which cumulatively responded strongly to HIF-1α and weakly to HIF-2α. FABP3 expression partially responded to HIF-1α. Two putative HRE were validated in FABP4 both of which responded weakly to HIF-1α and HIF-2α. FATP2 protein expression reacted positively to hypoxia. Thus, fetal essential fatty acid supply via the placenta is protected under hypoxia. It will be interesting to determine if our findings are replicated in human pre-eclamptic placenta.

The intracellular pathogen Orientia tsutsugamushi responsible for scrub typhus induces lipid droplet formation in mouse fibroblasts

Mammalian cells store excess fatty acids in the form of triglycerides within lipid droplets. The intracellular bacterium Orientia tsutsugamush is the causative agent of severe human rickettiosis. We found that O. tsutsugamushi infection induces the formation of lipid droplets in mouse L-929 fibroblasts. In infected cells, a parallel increase in the number of lipid droplets and pathogens was observed. Interestingly, the pathogen-infection induced the accumulation of triglycerides even without external supply of fatty acids. These results suggest that O. tsutsugamushi alters lipid metabolism of host cells to induce lipid droplets.

Implication of modified molecular structure of lipid through heat-related process to fatty acids supply in Brassica carinata seed

This study was conducted to explore the effect of different autoclave heating times (30, 60 and 90min) on fatty acids supply and molecular stability in Brassica carinata seed. Multivariate spectral analyses and correlation analyses were also carried out in our study. The results showed that autoclaving treatments significantly decreased the total fatty acids content in a linear fashion in B. carinata seed as heating time increased. Reduced concentrations were also observed in C18:3n3, C20:1, C22:1n9, monounsaturated fatty acids (MUFA), polyunsaturated fatty acids (PUFA), omega 3 (ω-3) and 9 (ω-9) fatty acids. Correspondingly, the heated seeds showed dramatic reductions in all the peak intensities within lipid-related spectral regions. Results from agglomerative hierarchical cluster analysis (AHCA) and principal component analysis (PCA) indicated that the raw oilseed had completely different structural make-up from the autoclaved seeds in both CH3 and CH2 asymmetric and symmetric stretching region (ca. 2999–2800cm−1) and lipid ester CO carbonyl region (ca. 1787–1706cm−1). However, the oilseeds heated for 30, 60 and 90min were not grouped into separate classes or ellipses in all the lipid-related regions, indicating that there still exhibited similarities in lipid biopolymer conformations among autoclaved B. carinata seeds. Moreover, strong correlations between spectral information and fatty acid compositions observed in our study could imply that lipid-related spectral parameters might have a potential to predict some fatty acids content in oilseed samples, i.e. B. carinata. However, more data from large sample size and diverse range would be necessary and helpful to draw up a final conclusion.

Abstract 325: Dissociation of Oxidative Stress from High Fatty Acid-Induced Cardiac Myocyte Death

Metabolic syndrome, featured by obesity and diabetes, is an independent risk factor for cardiovascular disease. The high availability of free fatty acids characteristic of obesity has been proposed to increase fatty acid oxidation, impair mitochondrial function, stimulate reactive oxygen species (ROS) production and eventually lead to oxidative stress, which then would cause myocyte damage and heart dysfunction. However, the role of oxidative stress as the initial cause of obesity-induced heart failure is controversial. We evaluated the time-dependent effect of fatty acids in vitro in adult rat cardiac myocytes (treated with palmitate and oleate, the major saturated and unsaturated fatty acids in the plasma, respectively), and in vivo in mice fed a high-fat diet (fat calories = 60%). Palmitate and oleate dose-dependently stimulated mitochondrial respiration and superoxide flash activity, a respiration-coupled bursting ROS production event in single mitochondria. The flash frequency increased 2-fold (0.1 mM, the physiological level in plasma) and 3-fold (0.3 mM, the plasma level found in obese subjects) after only 2 hr incubation and up to 24 hr, increase that could be blocked by the CPT1 inhibitor etomoxir (100uM). Surprisingly, there was no obvious mitochondrial oxidative stress, although the high level of fatty acids caused mPTP opening and cell death at 48 hr. Further, aconitase activity was normal and NAD/NADH ratio was decreased, indicating a more reduced status rather than oxidative stress. Intriguingly, mitochondrial membrane potential was slightly lowered after only 2 hr of treatment. Finally, the high-fat diet-fed mice developed heart dysfunction after 12 weeks of treatment, but oxidative stress only appeared after 18 weeks. In the light of these results, we hypothesize that high fatty acid supply induced multiple adaptive and mal-adaptive responses in cardiac myocytes. The increased mitochondrial respiration and bursting ROS production is offset by a mild uncoupling and increased reducing equivalents. Therefore, oxidative stress is unlikely an initial cause of myocyte death and heart dysfunction in obesity, but rather a consequence of mitochondrial dysfunction, although the exact cause of this dysfunction remains to be elucidated.

Lipid storage by adipose tissue macrophages regulates systemic glucose tolerance.

Proinflammatory pathways in adipose tissue macrophages (ATMs) can impair glucose tolerance in obesity, but ATMs may also be beneficial as repositories for excess lipid that adipocytes are unable to store. To test this hypothesis, we selectively targeted visceral ATMs in obese mice with siRNA against lipoprotein lipase (LPL), leaving macrophages within other organs unaffected. Selective silencing of ATM LPL decreased foam cell formation in visceral adipose tissue of obese mice, consistent with a reduced supply of fatty acids from VLDL hydrolysis. Unexpectedly, silencing LPL also decreased the expression of genes involved in fatty acid uptake (CD36) and esterification in ATMs. This deficit in fatty acid uptake capacity was associated with increased circulating serum free fatty acids. Importantly, ATM LPL silencing also caused a marked increase in circulating fatty acid-binding protein-4, an adipocyte-derived lipid chaperone previously reported to induce liver insulin resistance and glucose intolerance. Consistent with this concept, obese mice with LPL-depleted ATMs exhibited higher hepatic glucose production from pyruvate and glucose intolerance. Silencing CD36 in ATMs also promoted glucose intolerance. Taken together, the data indicate that LPL secreted by ATMs enhances their ability to sequester excess lipid in obese mice, promoting systemic glucose tolerance.

Lipid droplet-associated proteins in high-fat fed mice with the effects of voluntary running and diet change

ObjectiveThe relation between lipid accumulation and influence of exercise on insulin sensitivity is not straightforward. A proper balance between lipid droplet synthesis, lipolysis, and oxidative metabolism would ensure low local intramyocellular fatty acid levels, thereby possibly protecting against lipotoxicity-associated insulin resistance. This study investigated whether the accumulation of triglycerides and lipid droplets in response to high availability of fatty acids after high-fat feeding would parallel the abundance of intramyocellular perilipin proteins, especially PLIN5. The effects on these variables after diet change or voluntary running exercise intervention in skeletal muscle were also investigated. MethodsDuring a 19-week experiment, C57BL/6J mice were studied in six different groups: low-fat diet sedentary, low-fat diet active, high-fat diet sedentary, high-fat diet active and two groups which were high-fat sedentary for nine weeks, after which divided into low-fat sedentary or low-fat active groups. Myocellular triglyceride concentration and perilipin protein expression levels were assessed. ResultsWe show that, concurrently with impaired insulin sensitivity, the expression level of PLIN5 and muscular triglyceride concentration increased dramatically after high-fat diet. These adaptations were reversible after the diet change intervention with no additional effect of exercise. ConclusionsAfter high-fat diet, lipid droplets become larger providing more surface area for PLIN5. We suggest that PLIN5 is an important regulator of lipid droplet turnover in altered conditions of fatty acid supply and consumption. Imbalances in lipid droplet metabolism and turnover might lead to lipotoxicity-related insulin resistance.