Free Fatty Acid Metabolism Research Articles

CREEPING FAT-DERIVED FREE FATTY ACIDS INDUCE HYPERPLASIA OF INTESTINAL MUSCULARIS PROPRIA MUSCLE CELLS – A NOVEL LINK BETWEEN FAT AND INTESTINAL STRICTURE FORMATION IN CROHN’S DISEASE

In Crohn's disease (CD) wrapping of mesenteric fat around the bowel wall, so called 'creeping fat', is highly associated with strictures. The strongest contributor to luminal narrowing in strictures is a thickening of the human intestinal muscularis propria (MP). We hence investigated creeping fat derived factors and their effect on mechanisms of human intestinal MP smooth muscle cell (HIMC) hyperplasia. Free fatty acids (FFA) in creeping fat or non-creeping mesenteric fat organ cultures were measured via lipidomic mass spectrometry. Primary HIMC were exposed to FFA and cell proliferation was assessed. Intracellular FFA metabolism pathways and reactive oxygen species were functionally evaluated. Muscle thickness was investigated in dextran sodium sulfate (DSS) colitis with small molecule inhibition of FFA transport and a novel fat deletion mouse model. Subserosal creeping fat is associated with a markedly thickened MP. Experimental deletion of mesenteric fat (FAT-ATTAC mouse) reduced MP thickness. Human creeping fat conditioned medium strongly upregulated HIMC proliferation. Creeping fat released higher amounts of five long-chain FFA, including palmitate. Inhibition of HIMC long-chain FFA metabolism or FFA uptake into mitochondria through carnitine palmitoyltransferase (CPT)-1 reduced the palmitate induced HIMC proliferation. Blockade of conversion of palmitate into phospholipids reduced HIMC proliferation. Prophylactic inhibition of CPT-1 in experimental DSS colitis did not ameliorate inflammation, but reduced MP thickness. Creeping fat released long-chain FFA induce a selective proliferative response by HIMC. These results point to creeping fat as a novel contributor to stricture formation in CD.

Abstract P206: Renal hemodynamic and transcriptomic changes in response to salt in Dahl salt-sensitive rats

Introduction: Continuous measurements (24/7) of arterial pressure (AP) and renal blood flow (RBF), along with intermittent sampling of arterial and renal venous blood and urine in freely moving rats, have enabled the determination of sequential changes in global substrate metabolism and their association with physiological responses in the kidney. It was found that salt loading in Sprague-Dawley (SD) rats resulted in an increase in RBF and renal O 2 consumption with minimal changes in AP. At the same time, TCA cycle pathway was downregulated with reduced metabolism of free fatty acids and amino acids, while glycolysis was upregulated, in the kidney cortex (Cx). In contrast, no significant changes were observed in metabolism-related pathways in the outer medulla (OM). (PMID: 37575482). Here, we report transcriptomic responses of Dahl salt-sensitive (SS) rats to the HS diet as compared to the salt-resistant SD rats. Method: Male SS rats were fed a 0.4% NaCl (LS) diet. At 8 weeks of age, the rats were implanted with an RBF probe together with an arterial and a renal venous catheter. RBF and AP were measured continuously, and arterial and renal venous blood and urine were sampled when rats were fed the LS and on days 7, 14, and 21 after switching to the 4.0% NaCl (HS) diet. From separate groups of rats subjected to the same protocol, Cx and OM were removed for mRNAseq analysis (Novogene, Inc). Results: When switching to the HS diet, mean AP of SS rats rose from 122 ± 2 to 164 ± 5 mmHg by HS21. RBF (mL/min/g kidney weight) fell significantly over the 3 weeks of the HS diet averaging 7.2 ± 0.6 (LS) to 4.6 ± 0.7 (HS21). The gene set enrichment analysis (GSEA) of the mRNAseq data revealed that only a few metabolic pathways were enriched in HS7 in Cx such as glycosphingolipid biosynthesis, while metabolic pathways such as linolenic acid and arachidonic acid metabolism and glycosphingolipid biosynthesis were enriched in OM. Downregulation of the lysine degradation and glutathione metabolic pathways was observed in both Cx and OM in HS21. In general, fewer metabolic pathways responded to HS in SS rats compared to SD in Cx (22 in SD vs 7 in SS). Conclusion: The Cx of SS rats appear less capable of adjusting to the increased metabolic needs imposed by a HS diet compared to SD rats in contrast to the OM. The functional implications of this are being examined and the metabolomic analysis of blood, tissue and urine samples is currently in progress at the JAX lab.

Disruption of Cell Membranes and Redox Homeostasis as an Antibacterial Mechanism of Dielectric Barrier Discharge Plasma against Fusarium oxysporum.

Direct barrier discharge (DBD) plasma is a potential antibacterial strategy for controlling Fusarium oxysporum (F. oxysporum) in the food industry. The aim of this study was to investigate the inhibitory effect and mechanism of action of DBD plasma on F. oxysporum. The result of the antibacterial effect curve shows that DBD plasma has a good inactivation effect on F. oxysporum. The DBD plasma treatment severely disrupted the cell membrane structure and resulted in the leakage of intracellular components. In addition, flow cytometry was used to observe intracellular reactive oxygen species (ROS) levels and mitochondrial membrane potential, and it was found that, after plasma treatment, intracellular ROS accumulation and mitochondrial damage were accompanied by a decrease in antioxidant enzyme activity. The results of free fatty acid metabolism indicate that the saturated fatty acid content increased and unsaturated fatty acid content decreased. Overall, the DBD plasma treatment led to the oxidation of unsaturated fatty acids, which altered the cell membrane fatty acid content, thereby inducing cell membrane damage. Meanwhile, DBD plasma-induced ROS penetrated the cell membrane and accumulated intracellularly, leading to the collapse of the antioxidant system and ultimately causing cell death. This study reveals the bactericidal effect and mechanism of the DBD treatment on F. oxysporum, which provides a possible strategy for the control of F. oxysporum.

The abnormalities of free fatty acid metabolism in patients with hypertrophic cardiomyopathy, a single-center retrospective observational study

BackgroundPrevious studies have shown the importance of energy deficiency and malfunctioning mitochondria in the pathophysiology of hypertrophic cardiomyopathy (HCM). There has been a little research into the relationship between plasma free fatty acids (FFA), one of the heart’s main energy sources, and HCM. We evaluated its clinical importance in HCM to see if there was a link between plasma FFA metabolism and HCM.MethodsIn a single-center retrospective observational study, we investigated 420 HCM patients diagnosed at Beijing Anzhen Hospital between January 1, 2018, and December 31, 2022. Meanwhile, 1372 individuals without HCM (non-HCM) were recruited. 391 non-HCM patients were chosen as controls via a propensity score matching (PSM) study with a 1:1 ratio.ResultsFFA in HCM patients showed statistically significant correlations with creatinine (r = 0.115, p = 0.023), estimated GFR (r=-0.130, p = 0.010), BNP (r = 0.152, p = 0.007), LVEF (r=-0.227, p < 0.001), LVFS (r=-0.160, p = 0.002), and LAD (r = 0.112, p = 0.028). Higher FFA levels were found in HCM patients who had atrial fibrillation and NYHY functional classes III or IV (p = 0.015 and p = 0.022, respectively). In HCM patients, multiple linear regression analysis revealed that BNP and LVEF had independent relationships with increasing FFA (Standardized = 0.139, p = 0.013 and =-0.196, p < 0.001, respectively).ConclusionsAmong HCM patients, the plasma FFA concentration was lower, and those with AF and NYHY functional class III or IV had higher FFA levels, and LVEF and BNP were independently associated with increasing FFA. The findings of the study should help inspire future efforts to better understand how energy deficiency contributes to hypertrophic cardiomyopathy (HCM) development.

PCYT1A deficiency disturbs fatty acid metabolism and induces ferroptosis in the mouse retina

BackgroundInherited retinal dystrophies (IRDs) are a group of debilitating visual disorders characterized by the progressive degeneration of photoreceptors, which ultimately lead to blindness. Among the causes of this condition, mutations in the PCYT1A gene, which encodes the rate-limiting enzyme responsible for phosphatidylcholine (PC) de novo synthesis via the Kennedy pathway, have been identified. However, the precise mechanisms underlying the association between PCYT1A mutations and IRDs remain unclear. To address this knowledge gap, we focused on elucidating the functions of PCYT1A in the retina.ResultsWe found that PCYT1A is highly expressed in Müller glial (MG) cells in the inner nuclear layer (INL) of the retina. Subsequently, we generated a retina-specific knockout mouse model in which the Pcyt1a gene was targeted (Pcyt1a-RKO or RKO mice) to investigate the molecular mechanisms underlying IRDs caused by PCYT1A mutations. Our findings revealed that the deletion of Pcyt1a resulted in retinal degenerative phenotypes, including reduced scotopic electroretinogram (ERG) responses and progressive degeneration of photoreceptor cells, accompanied by loss of cells in the INL. Furthermore, through proteomic and bioinformatic analyses, we identified dysregulated retinal fatty acid metabolism and activation of the ferroptosis signalling pathway in RKO mice. Importantly, we found that PCYT1A deficiency did not lead to an overall reduction in PC synthesis within the retina. Instead, this deficiency appeared to disrupt free fatty acid metabolism and ultimately trigger ferroptosis.ConclusionsThis study reveals a novel mechanism by which mutations in PCYT1A contribute to the development of IRDs, shedding light on the interplay between fatty acid metabolism and retinal degenerative diseases, and provides new insights into the treatment of IRDs.

Does Resveratrol Improve Metabolic Dysfunction-Associated Steatotic Liver Disease (MASLD)?

Metabolic dysfunction-associated steatotic liver disease (MASLD) is influenced by a variety of factors, including environmental and genetic factors. The most significant outcome is the alteration of free fatty acid and triglyceride metabolism. Lipotoxicity, impaired autophagy, chronic inflammation, and oxidative stress, as well as coexisting insulin resistance, obesity, and changes in the composition of gut microbiota, are also considered crucial factors in the pathogenesis of MASLD. Resveratrol is a polyphenolic compound that belongs to the stilbene subgroup. This review summarises the available information on the therapeutic effects of resveratrol against MASLD. Resveratrol has demonstrated promising antisteatotic, antioxidant, and anti-inflammatory activities in liver cells in in vitro and animal studies. Resveratrol has been associated with inhibiting the NF-κB pathway, activating the SIRT-1 and AMPK pathways, normalizing the intestinal microbiome, and alleviating intestinal inflammation. However, clinical studies have yielded inconclusive results regarding the efficacy of resveratrol in alleviating hepatic steatosis or reducing any of the parameters found in MASLD in human patients. The lack of homogeneity between studies, low bioavailability of resveratrol, and population variability when compared to animal models could be the reasons for this.

Untargeted metabolomic profiling reveals molecular signatures associated with type 2 diabetes in Nigerians

BackgroundType 2 diabetes (T2D) has reached epidemic proportions globally, including in Africa. However, molecular studies to understand the pathophysiology of T2D remain scarce outside Europe and North America. The aims of this study are to use an untargeted metabolomics approach to identify: (a) metabolites that are differentially expressed between individuals with and without T2D and (b) a metabolic signature associated with T2D in a population of Sub-Saharan Africa (SSA).MethodsA total of 580 adult Nigerians from the Africa America Diabetes Mellitus (AADM) study were studied. The discovery study included 310 individuals (210 without T2D, 100 with T2D). Metabolites in plasma were assessed by reverse phase, ultra-performance liquid chromatography and mass spectrometry (RP)/UPLC-MS/MS methods on the Metabolon Platform. Welch’s two-sample t-test was used to identify differentially expressed metabolites (DEMs), followed by the construction of a biomarker panel using a random forest (RF) algorithm. The biomarker panel was evaluated in a replication sample of 270 individuals (110 without T2D and 160 with T2D) from the same study.ResultsUntargeted metabolomic analyses revealed 280 DEMs between individuals with and without T2D. The DEMs predominantly belonged to the lipid (51%, 142/280), amino acid (21%, 59/280), xenobiotics (13%, 35/280), carbohydrate (4%, 10/280) and nucleotide (4%, 10/280) super pathways. At the sub-pathway level, glycolysis, free fatty acid, bile metabolism, and branched chain amino acid catabolism were altered in T2D individuals. A 10-metabolite biomarker panel including glucose, gluconate, mannose, mannonate, 1,5-anhydroglucitol, fructose, fructosyl-lysine, 1-carboxylethylleucine, metformin, and methyl-glucopyranoside predicted T2D with an area under the curve (AUC) of 0.924 (95% CI: 0.845–0.966) and a predicted accuracy of 89.3%. The panel was validated with a similar AUC (0.935, 95% CI 0.906–0.958) in the replication cohort. The 10 metabolites in the biomarker panel correlated significantly with several T2D-related glycemic indices, including Hba1C, insulin resistance (HOMA-IR), and diabetes duration.ConclusionsWe demonstrate that metabolomic dysregulation associated with T2D in Nigerians affects multiple processes, including glycolysis, free fatty acid and bile metabolism, and branched chain amino acid catabolism. Our study replicated previous findings in other populations and identified a metabolic signature that could be used as a biomarker panel of T2D risk and glycemic control thus enhancing our knowledge of molecular pathophysiologic changes in T2D. The metabolomics dataset generated in this study represents an invaluable addition to publicly available multi-omics data on understudied African ancestry populations.

Therapeutic potency of A1 adenosine receptor antagonists in the treatment of cardiovascular diseases, current status and perspectives.

Cardiomyocytes form, transport, and metabolize the omnipresent metabolite adenosine. Depending upon the adenosine concentrations and the pharmacological properties of receptor subtypes, adenosine exerts (patho)physiological responses in the cardiovascular system. The objective of this review is to present different protective mechanisms of A1-adenosine receptor inhibitors in cardiovascular diseases. Literature references were collected and sorted using relevant keywords and key phrases as search terms in scientific databases such as Web of Science, PubMed and Google Scholar. A1 adenosine receptor regulates free fatty acid metabolism, lipolysis, heart rate, blood pressure, and cardiovascular toxicity. The evidence clearly supporting the therapeutic potency of pharmacological A1 adenosine receptors agonists and antagonists in modulating cardiovascular risk factor parameters and treatment of cardiovascular diseases. This review summarizes the protective role of pharmacological A1-adenosine receptor regulators in the pathogenesis of cardiovascular diseases for a better management of cardiovascular diseases.

Altered bile metabolome and its diagnostic potential for biliopancreatic malignancies

BackgroundDue to the difficulty of pathological sampling, the clinical differentiation between benign and malignant biliopancreatic diseases remains challenging. Endoscopic retrograde cholangiopancreatography (ERCP) is used to investigate biliary diseases, enabling the collection of bile. This study assessed potential metabolic alterations in biliopancreatic malignancies by exploring changes in the bile metabolome and the diagnostic potential of bile metabolome analysis. MethodsA total of 264 bile samples were collected from patients who were divided into a discovery cohort (n = 85) and a validation cohort (n = 179). Untargeted metabolomic analysis was used in the discovery cohort, while targeted metabolomic analysis was used in the validation cohort for further investigation of the differentially abundant metabolites. ResultsThe untargeted metabolomic analysis revealed that the metabolic changes associated with biliopancreatic malignancies occurred mainly in lipid metabolites, among which fatty acid metabolism was most significantly altered, and differentially abundant metabolites identified in the discovery cohort were mainly enriched in unsaturated fatty acid synthesis and linolenic acid synthesis pathways. Analysis of free fatty acid (FFA) metabolism in the validation cohort revealed that the FFA levels and related indicators verified the abnormal fatty acid metabolism associated with biliopancreatic malignancies. The combined model for biliopancreatic malignancies based on the fatty acid indexes and clinical test results improved the diagnostic performance of current clinical level. Then, we used machine learning to define three different FFA metabolic clusters of biliopancreatic malignancies, and survival analysis showed significant differences in prognostic outcomes among the three clusters. ConclusionsThis study found metabolic alterations in biliopancreatic malignancies based on bile samples, which may provide new insights for the clinical diagnosis and prognostic assessment of biliopancreatic malignancies.

APOE genotype dictates lipidomic signatures in primary human hepatocytes

Apolipoprotein E (APOE) genetic variants are most notably known for their divergent impact on the risk of developing Alzheimer’s disease. While APOE genotype has been consistently shown to modulate lipid metabolism in a variety of cellular contexts, the effect of APOE alleles on the lipidome in hepatocytes is unknown. In this study, we investigated the contribution of APOE alleles to lipidomic profiles of donor-derived primary human hepatocytes from 77 subjects. Lipidomic data obtained by liquid chromatography-mass spectrometry were analyzed across ε2/ε3, ε3/ε3, and ε3/ε4 genotypes to reveal how APOE modulates lipid relative levels over age and between groups. Hepatic APOE concentration, measured by ELISA, was assessed for correlation with lipid abundance in subjects grouped as per APOE genotype and sex. APOE genotype-specific differential lipidomic signatures associated with age for multiple lipid classes but did not differ between sexes. Compared to ε2/ε3, ε3/ε4 hepatocytes had higher abundance of acylcarnitines (AC) and acylphosphatidylglycerol (AcylPG) as a class, as well as higher medium and long-chain ACs, AcylPG, phosphatidylglycerol (PG), bis(monoacylglycerol)phosphate (BMP), monoacylglycerol (MG) and diacylglycerol (DG) species. The ε3/ε4 hepatocytes also exhibited a higher abundance of medium and long-chain ACs compared to the ε3/ε3 hepatocytes. Only in the ε3/ε4 hepatocytes, APOE concentration was lower and showed a negative correlation with BMP levels, specifically in females. APOE genotype dictates a differential lipidome in primary human hepatocytes. The lipids involved suggest mitochondrial dysfunction with accompanying alterations in neutral lipid storage, reflective of a general disturbance of free fatty acid metabolism in human hepatocytes with the ε4 allele.

B Vitamins, Glucoronolactone and the Immune System: Bioavailability, Doses and Efficiency.

The present review deals with two main ingredients of energy/power drinks: B vitamins and glucuronolactone and their possible effect on the immune system. There is a strong relationship between the recommended daily dose of selected B vitamins and a functional immune system. Regarding specific B vitamins: (1) Riboflavin is necessary for the optimization of reactive oxygen species (ROS) in the fight against bacterial infections caused by Staphylococcus aureus and Listeria monocytogenes. (2) Niacin administered within normal doses to obese rats can change the phenotype of skeletal fibers, and thereby affect muscle metabolism. This metabolic phenotype induced by niacin treatment is also confirmed by stimulation of the expression of genes involved in the metabolism of free fatty acids (FFAs) and oxidative phosphorylation at this level. (3) Vitamin B5 effects depend primarily on the dose, thus large doses can cause diarrhea or functional disorders of the digestive tract whereas normal levels are effective in wound healing, liver detoxification, and joint health support. (4) High vitamin B6 concentrations (>2000 mg per day) have been shown to exert a significant negative impact on the dorsal root ganglia. Whereas, at doses of approximately 70 ng/mL, sensory symptoms were reported in 80% of cases. (5) Chronic increases in vitamin B12 have been associated with the increased incidence of solid cancers. Additionally, glucuronolactone, whose effects are not well known, represents a controversial compound. (6) Supplementing with D-glucarates, such as glucuronolactone, may help the body's natural defense system function better to inhibit different tumor promoters and carcinogens and their consequences. Cumulatively, the present review aims to evaluate the relationship between the selected B vitamins group, glucuronolactone, and the immune system and their associations to bioavailability, doses, and efficiency.

Investigation of seasonal changes in lipid synthesis and metabolism-related genes in the oviduct of Chinese brown frog (Rana dybowskii).

A peculiar physiological characteristic of the Chinese brown frog (Rana dybowskii) is that its oviduct dilates during pre-brumation rather than during the breeding season. This research aimed to examine the expression of genes connected with lipid synthesis and metabolism in the oviduct of R. dybowskii during both the breeding season and pre-brumation. We observed significant changes in the weight and size of the oviduct between the breeding season and pre-brumation. Furthermore, compared to the breeding season, pre-brumation exhibited significantly lower triglyceride content and a marked increase in free fatty acid content. Immunohistochemical results revealed the spatial distribution of triglyceride synthase (Dgat1), triglyceride hydrolase (Lpl and Hsl), fatty acid synthase (Fasn), and fatty acid oxidases (Cpt1a, Acadl, and Hadh) in oviductal glandular cells and epithelial cells during both the breeding season and pre-brumation. While the mRNA levels of triglycerides and free fatty acid synthesis genes (dgat1 and fasn) did not show a significant difference between the breeding season and pre-brumation, the mRNA levels of genes involved in triglycerides and free fatty acid metabolism (lpl, cpt1a, acadl, acox and hadh) were considerably higher during pre-brumation. Furthermore, the R. dybowskii oviduct's transcriptomic and metabolomic data confirmed differential expression of genes and metabolites enriched in lipid metabolism signaling pathways during both the breeding season and pre-brumation. Overall, these results suggest that alterations in lipid synthesis and metabolism during pre-brumation may potentially influence the expanding size of the oviduct, contributing to the successful overwintering of R. dybowskii.

APOE genotype‐specific lipidomic signatures in primary human hepatocytes

AbstractBackgroundApolipoprotein E (APOE) is primarily responsible for cholesterol homeostasis in the periphery and in the brain through independently regulated pathways, as it does not cross the blood‐brain barrier (BBB). Carriers of the APOE ε4 allele have a 3‐ to 15‐fold higher risk of developing Alzheimer’s disease (AD). Recently, we demonstrated that hepatic APOE4 was associated with alterations in AD‐relevant brain functions, suggesting clinical significance of plasma APOE, 90% of which is derived from the liver. How APOE genotype affects the hepatic lipidome is yet unknown. In this study, we assessed APOE genotype‐specific lipidomic profiles of primary human hepatocytes and their association with hepatic APOE.MethodWe conducted a lipidomic analysis of primary human hepatocytes isolated from 77 subjects by use of Ultra Performance Liquid Chromatography‐Tandem Mass Spectrometry. Lipid class levels over age across ε2/ε3, ε3/ε3 and ε3/ε4 genotypes were assessed using Pearson correlation analysis and between‐group comparisons for lipid class and species levels, controlled for demographic variables, were performed using one‐way ANOVA with Tukey HSD correction for multiple comparisons. We assessed hepatic APOE levels by ELISA in order to compare APOE expression between APOE genotypes and investigate correlations with lipid levels.ResultWe found APOE allele‐specific lipidomic patterns over age and between groups. APOE genotype differentially modulated the relationship between age and lipid levels across multiple lipid classes. The ε4 carriers had higher acylcarnitines (AC) and acylphosphatidylglycerol (AcylPG) as a class, as well as higher medium and long‐chain ACs, AcylPG, bis(monoacylglycerol)phosphate (BMP), monoacylglycerol and diacylglycerol species compared to ε2 carriers. Hepatocyte APOE levels decreased with age in the whole cohort, but correlated with monoacylglycerols (MG) and phosphatidylcholine (PC) only in ε4 carriers. We observed no differences in APOE hepatocyte levels specific to genotype or sex of the donor.ConclusionAPOE genotype dictates a differential lipidome in primary human hepatocytes but has no effect on APOE levels. The lipids involved suggest mitochondrial dysfunction with accompanying alterations in neutral lipid storage, reflective of a general disturbance of free fatty acid metabolism in human hepatocytes with the ε4 allele.

IL-6 Mediates a Switch in Free Fatty Acid Metabolism from the Liver to the Bone Marrow to Support HSC Expansion during Infection

The hematopoietic stem cell (HSC) pool is tightly controlled in both normal and stressed conditions. Under normal conditions, HSCs reside quiescently in their hypoxic niche with minimal mitochondrial activity, instead favouring glycolysis to meet their energy needs. During acute infection HSCs switch to oxidative phosphorylation over glycolysis due to increased energy requirements. This metabolic switch enables HSCs to expand and differentiate into downstream progenitors to increase the immune cell pool. We recently showed that fatty acid (FA) uptake and metabolism play a central role in providing HSCs with sufficient energy for their expansion during acute infection. However, how infection drives changes in available FAs for HSC expansion is not known. This study investigated the role of liver in regulating lipid availability for HSC expansion. We have previously shown that circulating free fatty acids (FFAs) within the serum of mice increased in response to infection using both lipopolysaccharide (LPS) (6 hours) and S. Typhimurium (72 hours) models. This correlated with an expansion of bone marrow HSCs 16 hours after LPS and 72 hours after S. Typhimurium confirmed using flow cytometry. To investigate real-time fatty-acid uptake by hematopoietic cells in response to infection, an in vivo transplant model was used. CD45.1 lineage negative, CD117-positive cells tagged with firefly luciferase (LK+FF) were transplanted into CD45.2 mice. After transplantation was confirmed, mice were injected with LPS for 16 hours. Mice were then subsequently injected with a luciferin molecule conjugated to a long-chain FFA (FFA-luc), which is visible via bioluminescent imaging if the FFA-luc is taken up into the transplanted LK+FF engrafted cells. Live animal imaging using this method confirmed that long-chain FFA is taken up by hematopoietic cells in response to LPS in vivo. As the liver is the master regulator of FA in the serum we isolated livers from control and LPS (6 hour) treated mice. RNAseq revealed down-regulation of genes involved in FA uptake and metabolism as well genes involved in ketogenesis and up-regulation in genes involved in glucose uptake. Further kinetic studies showed that FA uptake and metabolism genes were downregulated within 2 hours of LPS treatment. To understand if this was a direct LPS effect on hepatocytes which express the receptor for LPS (TLR4) or mediated through unknown factors, we isolated primary hepatocytes and treated them with LPS or media conditioned with serum from control or LPS treated mice. Only the media conditioned with serum from LPS treated mice downregulated genes associated with FA uptake and metabolism. To identify the factors driving this metabolic switch in hepatocytes, a cytokine array of serum from control or LPS treated mice (90 minutes) was performed. This proteome cytokine array revealed a significant upregulation of several cytokines and chemokines, including IL-6, TNFα and CXCL1 and CXCL2. Primary mouse hepatocytes were then treated with factors identified in the array for 2 and 4 hours. RT-qPCR confirmed that IL-6 downregulated FA uptake and metabolism genes in primary hepatocytes. To understand if this was a LPS specific response or induce by infection mice were treated with S. Typhimurium for various time points and livers extracted. RT-qPCR confirmed that S. Typhimurium treated mice down-regulated genes associated with FA uptake and metabolism. To study the role of IL-6 in regulating liver FA metabolism in response to LPS, mice were treated with IL-6 neutralising antibody (MP5-20F3). Pre-treatment of mice with MP5-20F3 inhibited LPS induced downregulation of liver FA uptake and metabolic genes, which lead to reversing LPS-induced increase in serum FA. In conclusion we have shown a role for IL-6 in switching fatty acid metabolism from the liver to the bone marrow to allow HSC expansion during infection.

Metabolomic Differences in Connective Tissue Disease-Associated Versus Idiopathic Pulmonary Arterial Hypertension in the PVDOMICS Cohort.

Patients with connective tissue disease-associated pulmonary arterial hypertension (CTD-PAH) experience worse survival and derive less benefit from pulmonary vasodilator therapies than patients with idiopathic PAH (IPAH). We sought to identify differential metabolism in patients with CTD-PAH versus patients with IPAH that might underlie these observed clinical differences. Adult participants with CTD-PAH (n=141) and IPAH (n=165) from the Pulmonary Vascular Disease Phenomics (PVDOMICS) study were included. Detailed clinical phenotyping was performed at cohort enrollment, including broad-based global metabolomic profiling of plasma samples. Participants were followed prospectively for ascertainment of outcomes. Supervised and unsupervised machine learning algorithms and regression models were used to compare CTD-PAH versus IPAH metabolomic profiles and to measure metabolite-phenotype associations and interactions. Gradients across the pulmonary circulation were assessed using paired mixed venous and wedged samples in a subset of 115 participants. Metabolomic profiles distinguished CTD-PAH from IPAH, with patients with CTD-PAH demonstrating aberrant lipid metabolism with lower circulating levels of sex steroid hormones and higher free fatty acids (FAs) and FA intermediates. Acylcholines were taken up by the right ventricular-pulmonary vascular (RV-PV) circulation, particularly in CTD-PAH, while free FAs and acylcarnitines were released. In both PAH subtypes, dysregulated lipid metabolites, among others, were associated with hemodynamic and RV measurements and with transplant-free survival. CTD-PAH is characterized by aberrant lipid metabolism that may signal shifted metabolic substrate utilization. Abnormalities in RV-PV FA metabolism may imply a reduced capacity for mitochondrial beta oxidation within the diseased pulmonary circulation.

Some Pathogenetic Aspects of Endocardial Endothelium Damage of Rats as a Result of Stress Effect Complicated by Hypercholesterolemia (original research)

The endothelial dysfunction is a predictor of occurrence for many diseases of cardiovascular system and a key link in their pathogenesis, formation and progression of clinical manifestations. Despite the large number of experimental and clinical researches, separate links of pathogenesis of heart endothelial cell damage under stress and in case of its combination with hypercholesterolemia require the further examination. The aim of the work was to investigate the role of nitric oxide, prostaglandins and antioxidants in the pathogenesis of endocardial endothelium damage (by the content of free fatty acids in it and the number of exfoliated cells) as a result of emotional and pain stress and stress complicated by hypercholesterolemia. An electro-impulse model was used to reproduce the stress. Alimentary hypercholesterolemia was modeled by keeping animals on an atherogenic diet for 2 months. To establish the role of individual pathogenetic links in the mechanisms of endotheliocyte damage, animals were administered the following pharmacological drugs: L-arginine (a substrate for the synthesis of nitric oxide), a prostaglandin synthesis blocker indomethacin, and prostenon (a synthetic analog of prostaglandin E2, which has an antioxidant effect). The state of endocardial endothelium was examined by using the light microscopy, analyzing the smears-imprints from macropreparations of ventricles by counting the number of endothelial cells in them. The content of free fatty acids was determined by the radiochemical method. It was shown that L-arginine significantly limits the damaging effect of the studied pathogenic factors on endocardial endothelium, reducing the number of exfoliated cells. The use of prostenon gives a slight, statistically unreliable positive effect. The use of indomethacin increases the damage of endothelial cells, which indicates the cytoprotective effect of prostaglandins under stress. All three studied preparations have no significant effect on the metabolism of free fatty acids both in case of “pure” emotional and pain stress and under stress complicated by hypercholesterolemia. The hypercholesterolemia of alimentary origin significantly limits the cytoprotective effect of L-arginine and prostenon on endocardial endothelium under stress action. In relation to indomethacin, in this situation, an increase of cell desquamation is observed, which indicates a decrease of prostaglandins protective effect.

Novel approach using [18F]FTHA-PET and de novo synthesized VLDL for assessment of FFA metabolism in a rat model of diet induced NAFLD

The worldwide prevalence of Non-alcoholic Fatty Liver Disease (NAFLD) raises concerns about associated risk factors, such as obesity and type 2 Diabetes Mellitus, for leading causes of disability and death. Besides Magnetic Resonance Imaging (MRI) and Spectroscopy (MRS), functional imaging with Positron Emission Tomography (PET) could contribute to a deeper understanding of the pathophysiology of NAFLD. Here we describe a novel approach using the PET tracer [18F]FTHA, which is an analog of long-chain free fatty acids (FFA) and is taken up by tissues to enter mitochondria or to be incorporated into complex lipids for further export as very-low-density lipoprotein (VLDL). Male Sprague Dawley rats, after 6 weeks on a high-fat diet (HFD), were used as a model of diet induced NAFLD, while a standard diet (SD) served as a control group. Liver fat was estimated by MR spectroscopy at a 9.4T system for phenotyping. To measure hepatic FFA uptake, rats underwent 60min dynamic [18F]FTHA-PET scans after unrestricted access to food (HFD: n=6; SD: n=6) or overnight (≤16h) fasting (HFD: n=6; SD: n=5). FFA removal was assessed from incorporated 18F-residual in de novo synthesized VLDL out of plasma. MRS of the liver confirmed the presence of NAFLD (>5.6% fat). Under non-fasting conditions, hepatic [18F]FTHA uptake was significantly increased in NAFLD: SUVmean (p=0.03) within [0; 60] min interval, SUVmean (p=0.01) and SUVmax (p=0.03) within [30; 60] min interval. SUVs for hepatic uptake under fasting conditions were not significantly different between the groups. Analysis of FFA removal demonstrated elevated values of 18F-residue in the VLDL plasma fraction of the healthy group compared to the NAFLD (p=0.0569). Our novel approach for assessing FFA metabolism using [18F]FTHA demonstrated differences in the hepatic FFA uptake and FFA incorporation into VLDL between healthy and NAFLD rats. [18F]FTHA-PET could be used to study metabolic disturbances involved in the progression of NAFLD.

Medium-chain fatty acid oxidation is independent of l-carnitine in liver and kidney but not in heart and skeletal muscle.

Medium-chain fatty acid (MCFA) consumption confers a wide range of health benefits that are highly distinct from long-chain fatty acids (LCFAs). A major difference between the metabolism of LCFAs compared with MCFAs is that mitochondrial LCFA oxidation depends on the carnitine shuttle, whereas MCFA mitochondrial oxidation is not. Although MCFAs are said to range from 6 to 14 carbons long based on physicochemical properties in vitro, the biological cut-off length of acyl chains that can bypass the carnitine shuttle in different mammalian tissues is unknown. To define the range of acyl chain length that can be oxidized in the mitochondria independent of carnitine, we determined the oxidative metabolism of free fatty acids (FFAs) from 6 to 18 carbons long in the liver, kidney, heart, and skeletal muscle. The liver oxidized FFAs 6 to 14 carbons long, whereas the kidney oxidized FFAs from 6 to 10 carbons in length. Heart and skeletal muscle were unable to oxidize FFAs of any chain length. These data show that while the liver and kidney can oxidize MCFAs in the free form, the heart and skeletal muscle require carnitine for the oxidative metabolism of MCFAs. Together these data demonstrate that MCFA oxidation independent of carnitine is tissue-specific.NEW & NOTEWORTHY This work demonstrates that the traditional concept of mitochondrial medium-chain fatty acid oxidation as unregulated and independent of carnitine applies only to liver metabolism, and to kidney to a lesser extent, but not the heart or skeletal muscle. Thus, the benefits of dietary medium-chain fatty acids are set by liver metabolic activity and peripheral tissues are unlikely to receive direct benefits from medium-chain fatty acid metabolism, but rather metabolic byproducts of liver's medium-chain oxidative metabolism.

Effects of myonectin on porcine intramuscular adipocyte differentiation and exogenous free fatty acid utilization

As an important factor secreted by skeletal muscle, myonectin can regulate lipid metabolism and energy metabolism, but its role in the utilization of peripheral free fatty acids (FFAs) by porcine intramuscular fat cells remains to be further investigated. In this study, porcine intramuscular adipocytes were treated with recombinant myonectin and palmitic acid (PA), either alone or in combination, and then were examined for their uptake of exogenous FFAs, intracellular lipid synthesis and catabolism, and mitochondrial oxidation of fatty acids. The results showed that myonectin decreased the area of lipid droplets in intramuscular adipocytes (p < 0.05) and significantly increased (p < 0.05) the expression levels of hormone-sensitive lipase (HSL) and lipoprotein lipase (LPL). Moreover, myonectin can up-regulate the expression of p38 mitogen-activated protein kinase (p38 MAPK). Myonectin significantly promoted the uptake of peripheral FFAs (p < 0.01), improved (p < 0.05) the expression of fatty transport protein 1 (FATP1) and fatty acid binding protein 4 (FABP4) in intramuscular adipocytes. Myonectin also significantly increased (p < 0.05) the expression levels of fatty acid oxidation markers: transcription factor (TFAM), uncoupling protein-2 (UCP2) and oxidative respiratory chain marker protein complex I (NADH-CoQ) in mitochondria of intramuscular adipocytes. In summary, myonectin promoted the absorption, transport, and oxidative metabolism of exogenous FFAs in mitochondria, thereby inhibiting lipid deposition in porcine intramuscular adipocytes.

Metabolism of free fatty acids in patients with gastrointestinal diseases depending on the body mass index

Background. The purpose of the study is to analyze the content of free fatty acids (FFA) in the blood serum of overweight and underweight patients with gastrointestinal diseases. Mate­rials and methods. Thirty-one patients with gastrointestinal diseases were examined, 19 (61.3 %) men and 12 (38.7 %) women with a median age of 39 (27; 48) years. Depending on the results of the body composition study with the multifunctional monitor TANITA MC-780MA (Japan), they were divided into the following groups: overweight patients (body mass index (BMI) more than 25.0 kg/m2) and underweight persons (BMI below 18.5 kg/m2). Determination of the FFA spectrum in the blood serum was carried out using a gas chromatograph with a flame ionization detector Chromatek-Crystal 5000. The control group consisted of 16 practically healthy people. Statistical processing of the results was carried out using the Statistica 6.1 application program package. Results. Patients with a change in body weight reported a statistically significant decrease in the median total content of short-chain saturated FFA (C4:0) mainly due to a decrease in the butyric acid content by 67 times (p = 0.001) with increased BMI and by 114 times (p = 0.002) with decreased BMI compared to controls. At the same time, the analysis of the serum spectrum of saturated FFA with an average carbon chain length showed a probable increase in the content of capric acid by 14 times (p &lt; 0.01), undecyl acid by 19 times (p &lt; 0.01), and lauric acid by 25 times (p &lt; 0.001) in patients with changes in body weight. The content of caproic acid increased in the group with excessive body weight by 3.9 times (p = 0.046) compared to controls and by 2.6 times (p &gt; 0.05) compared to underweight patients. The total content of monounsaturated FFA in the blood serum of patients with increased and decreased BMI increased significantly by 32 times (p &lt; 0.001) and 19 times (p = 0.001), respectively, compared to controls. The total content of polyunsaturated FFA in the blood serum of patients of both groups was found in a trace amount. Overweight patients had a 38-fold (p &lt; 0.01) increase in the trans-FFA elaidic acid (C18:1(trans-9)) compared to controls, while underweight patients had a decrease in linoelaidic acid (C18:2(trans-9,12)) by 11.7 times (p &lt; 0.05). Correlation analysis revealed a positive relationship between BMI and increased serum content of saturated caproic acid (r = 0.39; p = 0.019); monounsaturated acids — palmitoleic (r = 0.33; р = 0.045) and heptadecenoic (r = 0.35; р = 0.034); trans-configuration fatty acids — elaidic (r = 0.43; p = 0.009) and linoelaidic (r = 0.43; p = 0.007). Conclusions. Patients with gastrointestinal diseases had an imbalance in the fatty acid spectrum of the blood serum. Against the background of the tendency to decrease the total content of short-chain saturated FFA, a significant increase in the content of monounsaturated FFA and trans-FFA was noted, especially with increased BMI. In all patients, the trace amount of polyunsaturated FFA fractions was found. The obtained results can be used to improve the differential treatment of metabolic disorders in patients with gastrointestinal tract pathology.