Acid Flux Research Articles

E2F2 Reprograms Macrophage Function By Modulating Material and Energy Metabolism in the Progression of Metabolic Dysfunction-Associated Steatohepatitis.

Macrophages are essential for the development of steatosis, hepatic inflammation, and fibrosis in metabolic dysfunction-associated steatohepatitis(MASH). However, the roles of macrophage E2F2 in the progression of MASH have not been elucidated. This study reveals that the expression of macrophage E2F2 is dramatically downregulated in MASH livers from mice and humans, and that this expression is adversely correlated with the severity of the disease. Myeloid-specific E2F2 depletion aggravates intrahepatic inflammation, hepatic stellate cell activation, and hepatocyte lipid accumulation during MASH progression. Mechanistically, E2F2 can inhibit the SLC7A5 transcription directly. E2F2 deficiency upregulates the expression of SLC7A5 to mediate amino acids flux, resulting in enhanced glycolysis, impaired mitochondrial function, and increased macrophages proinflammatory response in a Leu-mTORC1-dependent manner. Moreover, bioinformatics analysis and CUT &Tag assay identify the direct binding of Nrf2 to E2F2 promoter to promote its transcription and nuclear translocation. Genetic or pharmacological activation of Nrf2 effectively activates E2F2 to attenuate the MASH progression. Finally, patients treated with CDK4/6 inhibitors demonstrate reduced E2F2 activity but increased SLC7A5 activity in PBMCs. These findings indicated macrophage E2F2 suppresses MASH progression by reprogramming amino acid metabolism via SLC7A5- Leu-mTORC1 signaling pathway. Activating E2F2 holds promise as a therapeutic strategy for MASH.

Fatty acids from nutrition sources for preterm infants and their effect on plasma fatty acid profiles

BackgroundIn preterm infants, IV administration of fat is less well tolerated compared to intake via the enteral route, often resulting in hypertriglyceridemia. It is therefore recommended that parenteral fat intake should not exceed 3.5 to 4.0 g/kg/d whereas human milk can provide up to 8 g/kg/d. It is unknown whether such hypertriglyceridemic conditions are caused by a uniform increase of all fatty acids or it is linked to an elevation of distinct fatty acids due to an unbalanced intake. Obviously, both scenarios could potentially influence the formulation of novel lipid solutions for preterm infants. Objective of this exploratory study was to compare fatty acid profiles between a) different nutritional sources and corresponding plasma samples, b) plasma of infants fed breast milk versus those receiving lipid emulsion, and c) plasma of infants with normal versus elevated triglyceride levels.MethodsForty-seven preterm infants < 36 weeks of gestation were included; fatty acid profiles were measured in serum samples and corresponding nutritional sources (breast milk and lipid emulsion) using gas chromatography/mass spectrometry.ResultsCompared to breast milk levels, plasma contained significantly lower C8:0, C10:0, C12:0, C14:0, C19:1n9, C18:3n3 (p < 0.0001). In contrast, relative abundance of C16:0, C18:0 and C20:4n6 was higher in plasma than in corresponding breast milk samples (p < 0.001) and lipid emulsion (p < 0.01). Compared to the corresponding lipid emulsion, the abundance of C18:2n6 and C18:3n3 was significantly lower in plasma (p < 0.001). Fatty acid profiles in plasma of infants fed breast milk compared to lipid emulsion were not markedly different. Hypertriglyceridemic samples showed elevated levels for C18:1n9 and C16:0 when compared with normotriglyceridemic samples.ConclusionsOur study reveals that lipid levels in plasma show both depletion and enrichment of distinct fatty acids which do not seem to be closely related to dietary intake. A more detailed understanding of fatty acid flux rates is needed, like the understanding of amino acid metabolism and is supported by the finding that hypertriglyceridemia might be a state of selective fatty acid accumulation. This would allow to develop more balanced diets for intensive care and potentially improve clinical outcomes.

Ecological adaptations of amphibians to environmental changes along an altitudinal gradient (Case Study: Bufo gargarizans) from phenotypic and genetic perspectives

BackgroundOrganisms have evolved a range of phenotypic and genetic adaptations to live in different environments along an altitudinal gradient. Herein, we studied the widely distributed Chinese toad, Bufo gargarizans, as a model and used an integrated phenotype-genotype approach to assess adaptations to different altitudinal environments.ResultsComparison of populations from four altitudes (50 m, 1200 m, 2300 m, and 3400 m) showed more effective defenses among high-altitude toads. These included thickened epidermis, more epidermal capillaries and granular glands, greater gland size in skin, and higher antioxidant enzyme activities in plasma. High-altitude toads also showed increased erythrocytes and hematocrit and elevated hemoglobin concentration, potentially improving oxygen delivery. Elevated altitude led to a metabolic shift from aerobic to anaerobic metabolism, and high-altitude populations favored carbohydrates over fatty acids to fuel for energy metabolism. Differentially expressed genes were associated with adaptive phenotypic changes. For instance, expression of genes associated with fatty acid metabolism showed greater suppression at high altitude (3400 m), consistent with decreased flux of β-hydroxybutyric acid and lower free fatty acids levels. Moreover, down-regulation of genes involved in carbon metabolism processes at high altitude (3400 m) were coincident with reduced TCA cycle flux. These results suggest that high-altitude toads adopt a metabolic suppression strategy for survival under harsh environmental conditions. Moreover, the hypoxia-inducible factor signaling cascade was activated at high altitude.ConclusionsCollectively, these results advance our comprehension of adaptation to high-altitude environments by revealing physiological and genetic mechanisms at work in Chinese toads living along altitudinal gradients.

Loss of adipose ATF3 promotes adipose tissue lipolysis and the development of MASH

The crosstalk between adipose tissue and the liver is finely controlled to maintain metabolic health. Yet, how adipose tissue controls toxic free fatty acid overflow into the liver remains incompletely understood. Here, we show that adipocyte activating transcription factor 3 (ATF3) was induced in human or mouse obesity. Adipocyte Atf3−/− (Atf3Adi−/−) mice developed obesity, glucose intolerance, and metabolic dysfunction-associated steatohepatitis (MASH) in chow diet, high-fat diet, or Western diet-fed mice. Blocking fatty acid flux by inhibiting hepatocyte CD36, but not the restoration of hepatic AMPK signaling, prevented the aggravation of MASH in Atf3Adi−/− mice. Further studies show that the loss of adipocyte ATF3 increased lipolysis via inducing adipose triglyceride lipase, which in turn induced lipogenesis and inflammation in hepatocytes. Moreover, Atf3Adi−/− mice had reduced energy expenditure and increased adipose lipogenesis and inflammation. Our data demonstrate that adipocyte ATF3 is a gatekeeper in counteracting MASH development under physiological and pathological conditions.

Arabidopsis trigalactosyldiacylglycerol1 mutants reveal a critical role for phosphtidylcholine remodeling in lipid homeostasis.

Lipid remodeling plays a critical role in plant response to abiotic stress and metabolic perturbations. Key steps in this process involve modifications of phosphatidylcholine (PC) acyl chains mediated by lysophosphatidylcholine: acyl-CoA acyltransferases (LPCATs) and phosphatidylcholine: diacylglycerol cholinephosphotransferase (ROD1). To assess their importance in lipid homeostasis, we took advantage of the trigalactosyldiacylglycerol1 (tgd1) mutant that exhibits marked increases in fatty acid synthesis and fatty acid flux through PC due to a block in inter-organelle lipid trafficking. Here, we showed that the increased fatty acid synthesis in tgd1 is due to posttranslational activation of the plastidic acetyl-coenzyme A carboxylase. Genetic analysis showed that knockout of LPCAT1 and 2 resulted in a lethal phenotype in tgd1. In addition, plants homozygous for lpcat2 and heterozygous for lpcat1 in the tgd1 background showed reduced levels of PC and triacylglycerols (TAG) and alterations in their fatty acid profiles. We further showed that disruption of ROD1 in tgd1 resulted in changes in fatty acid composition of PC and TAG, decreased leaf TAG content and reduced seedling growth. Together, our results reveal a critical role of LPCATs and ROD1 in maintaining cellular lipid homeostasis under conditions, in which fatty acid production largely exceeds the cellular demand for membrane lipid synthesis.

Acetyl-CoA carboxylase inhibition increases retinal pigment epithelial cell fatty acid flux and restricts apolipoprotein efflux

Lipid-rich deposits called drusen accumulate under the retinal pigment epithelium (RPE) in the eyes of patients with age-related macular degeneration (AMD) and Sorsby’s fundus dystrophy (SFD). Drusen may contribute to photoreceptor and RPE degeneration in these blinding diseases. We hypothesize that stimulating β-oxidation of fatty acids could decrease the availability of lipid with which RPE cells can generate drusen. Inhibitors of acetyl-CoA carboxylase (ACC) stimulate β-oxidation and diminish lipid accumulation in fatty liver disease. In this report we test the hypothesis that an ACC inhibitor, Firsocostat, can diminish lipid deposition by RPE cells. We probed metabolism and cellular function in mouse RPE-choroid tissue and in cultured human RPE cells. We used 13C6-glucose, 13C16-palmitate, and gas chromatography-linked mass spectrometry to monitor effects of Firsocostat on glycolytic, Krebs cycle, and fatty acid metabolism. We quantified lipid abundance, apolipoprotein E (ApoE) and vascular endothelial growth factor (VEGF) release using liquid chromatography-mass spectrometry, enzyme-linked immunosorbent assays and localized ApoE deposits by immunostaining. RPE barrier function was assessed by trans-epithelial electrical resistance (TEER). Firsocostat-mediated ACC inhibition increases β-oxidation, decreases intracellular lipid levels, diminishes lipoprotein release, and increases TEER. When human serum or outer segments are used to stimulate lipoprotein release, fewer lipoproteins are released in the presence of either lipid source and Firsocostat. In a culture model of SFD, Firsocostat stimulates fatty acid oxidation, increases TEER, and decreases ApoE release. We conclude that Firsocostat remodels RPE metabolism and can limit lipid deposition. This suggests that ACC inhibition could be an effective strategy for diminishing pathologic drusen in the eyes of patients with AMD or SFD.

Obesity alters adipose tissue response to fasting and refeeding in women: A study on lipolytic and endocrine dynamics and acute insulin resistance

Fasting induces significant shifts in substrate utilization with signs of acute insulin resistance (IR), while obesity is associated with chronic IR. Nonetheless, both states substantially influence adipose tissue (AT) function. Therefore, in this interventional study (NCT04260542), we investigated if excessive adiposity in premenopausal women alters insulin sensitivity and AT metabolic and endocrine activity in response to a 60-h fast and a subsequent 48-h refeeding period. Using physiological methods, lipidomics, and AT explants, we showed that obesity partially modified AT endocrine activity and blunted the dynamics of AT insulin resistance in response to the fasting/refeeding challenge compared to that observed in lean women. AT adapted to its own excess by reducing lipolytic activity/free fatty acids (FFA) flux per mass. This adaptation persisted even after a 60-h fast, resulting in lower ketosis in women with obesity. This could be a protective mechanism that limits the lipotoxic effects of FFA; however, it may ultimately impede desirable weight loss induced by caloric restriction in women with obesity.

Biogenically synthesized green silver nanoparticles exhibit antimalarial activity

The suboptimal efficacies of existing anti-malarial drugs attributed to the emergence of drug resistance dampen the clinical outcomes. Hence, there is a need for developing novel drug and drug targets. Recently silver nanoparticles (AgNPs) constructed with the leaf extracts of Euphorbia cotinifolia were shown to possess antimalarial activity. Therefore, the synthesized AgNPs from Euphorbia cotinifolia (EcAgNPs) were tested for their parasite clearance activity. We determined the antimalarial activity in the asexual blood stage infection of 3D7 (laboratory strain) P. falciparum. EcAgNPs demonstrated the significant inhibition of parasite growth (EC50 of 0.75 µg/ml) in the routine in vitro culture of P. falciparum. The synthesized silver nanoparticles were seen to induce apoptosis in P. falciparum through increased reactive oxygen species (ROS) ROS production and activated programmed cell death pathways characterized by the caspase-3 and calpain activity. Also, altered transcriptional regulation of Bax/Bcl-2 ratio indicated the enhanced apoptosis. Moreover, inhibited expression of PfLPL-1 by EcAgNPs is suggestive of the dysregulated host fatty acid flux via parasite lipid storage. Overall, our findings suggest that EcAgNPs are a non-toxic and targeted antimalarial treatment, and could be a promising therapeutic approach for clearing malaria infection.

Membrane electrolysis distillation for volatile fatty acids extraction from pH-neutral fermented wastewater

Volatile fatty acids (VFAs) serve as building blocks for a wide range of chemicals, but it is difficult to extract VFAs from pH-neutral wastewater using evaporation methods because of the ionized form. This study presents a new membrane electrolysis distillation (MED) process that extracts VFAs from such fermentation solutions. MED uniquely integrates pH regulation and joule heating to facilitate the efficient evaporation of VFAs. This integration occurs alongside a hydrophobic membrane that ensures effective gas-liquid phase separation. Operating solely on electricity, MED achieved an acid flux rate of 12.03 g/m2/h at 6V. In contrast, the control results without the joule heating or pH swing only obtained a 0.23 g/m2/h and 0.32 g/m2/h flux, respectively. In addition, a physicochemical model was developed to assess the impacts of temperature on membrane surface pH. This system enhances resource recovery from waste streams and helps achieve a circular carbon economy.

Relative contributions of mobility and partitioning to volatile fatty acid flux during electrodialysis

Economically valuable volatile fatty acids (VFAs) are sustainably produced via fermentation processes. To use VFAs downstream, they must be recovered using technologies like electrodialysis (ED). Solute transport properties (i.e., partition coefficient (K), diffusion coefficient (D), and permeability (P=KD)) govern flux in ED. Therefore, to advance understanding of VFA flux through anion exchange membranes (AEMs) in ED, we aimed to elucidate the relative contributions of VFA partitioning and mobility to their flux. Accordingly, for VFAs of different sizes (C1–C5) and inorganic anions (Cl–, Br–), we measured their fluxes during ED, permeabilities, and partition coefficients, and calculated the diffusion coefficients. We then evaluated the correlations between flux and transport properties and between transport properties and anion physicochemical properties. Results showed VFA flux had a strong correlation with permeability (R2=0.94, p<0.01), consistent with flux described by the Nernst-Planck equation. Further, while there was a negative correlation between VFA flux and partition coefficient (R2=0.46, p=0.21), there was a positive correlation between VFA flux and diffusion coefficient (R2=0.95, p<0.01) which showed VFA mobility governed VFA flux. We observed a negative correlation between VFA diffusion coefficient and carbon–chain length which was attributed to steric hindrance, and a positive correlation between partition coefficient and carbon chain-length which we attributed to hydrophobicity and polarizability. This work provides fundamental insight on interactions between VFAs and AEMs which affect anion flux during ED.

29. A model to simulate metabolic fluxes of amino acids in the small intestine of pigs

29. A model to simulate metabolic fluxes of amino acids in the small intestine of pigs

Enhancement of chemoresistance by claudin-1-mediated formation of amino acid barriers in human lung adenocarcinoma A549 cells

Claudin-1 (CLDN1) is highly expressed in human lung adenocarcinoma-derived A549 cells and is involved in the augmentation of chemoresistance. However, the mechanism of chemoresistance is not fully understood. In the tumor microenvironment, cancer cells are exposed to stress conditions such as hypoxia and malnutrition. Here, we investigated the effect of CLDN1 expression on amino acid (AA) flux and chemoresistance using A549 cells. The expression of L-type AA transporters, LAT1 and LAT3, was decreased by CLDN1 silencing in A549 spheroids. A reduction in extracellular AA concentration increased the expression of these AA transporters in two-dimensional (2D) cultured cells. The paracellular AA flux except for Ser, Thr, Tyr, Ala, and Gly was enhanced by CLDN1 silencing. These results suggest that CLDN1 forms a paracellular barrier to some AAs, leading to the elevation of LAT1/3 expression in spheroids. The production of reactive oxygen species in the mitochondria and cytosol was decreased by CLDN1 silencing in spheroids, resulting in downregulation of the expression of nuclear factor erythroid 2-related factor 2 (Nrf2) and its target antioxidant genes. CLDN1 silencing enhanced the cytotoxicity of anticancer drugs including doxorubicin and cisplatin, which was blocked by sulforaphane, an inducer of Nrf2 signaling. Similarly, the anticancer-induced toxicity was enhanced by Nrf2 silencing. In 2D cultured cells, the anticancer-induced toxicity was attenuated by AA deficiency and sulforaphane. We suggest that CLDN1 forms an AA barrier in spheroids, leading to the augmentation of Nrf2-dependent chemoresistance in A549 cells.

Molecular signatures of longissimus dorsi differ between dairy cattle based on prepartum muscle reserves and branched-chain volatile fatty acid supplementation.

Dairy cattle with high (HM) versus low muscle (LM) reserves exhibit distinct temporal changes in longissimus dorsi muscle depth (LDD) in late gestation. Branched-chain volatile fatty acids (BCVFA) supplementation increased blood glucose levels. We hypothesized that differences in HM and LM reflect distinct muscle metabolism and BCVFA supplementation altered metabolic pathways. At 42 d before expected calving (BEC) Holstein dairy cows were enrolled in a 2 x 2 factorial study of diet and muscle reserves, by assigning to control (CON) or BCVFA supplemented diets and LDD of HM (>4.6 cm) or LM (≤4.6 cm) groups: HM-CON (n=13), HM-BCVFA (n=10), LM-CON (n=9), and LM-BCVFA (n=9). Longisumus dorsi was biopsied at 21 d BEC, total RNA isolated and protein coding gene expression measured with RNA-seq. Between HM and LM 713 genes were differentially expressed and 481 between BCVFA and CON (P<0.05). Transcriptional signatures indicated differential distribution of Type II fibers between groups, with MYH1 greater in LM and MYH2 greater in HM cattle (P<0.05). Signatures of LM cattle relative to HM indicated greater activation of autophagy, ubiquitin-proteasome, and Ca2+-calpain pathways. HM cattle displayed greater expression of genes that encode extracellular matrix proteins and factors that regulate their proteolysis and turnover. BCVFA modified transcriptomes by increasing expression of genes that regulate fatty acid degradation and flux of carbons into the TCA cycle as acetyl CoA. Molecular signatures support distinct metabolic strategies between LM and HM cattle, and that BCVFA supplementation increased substrates for energy generation.

Amino acid influx via LAT1 regulates iron demand and sensitivity to PPMX-T003 of aggressive natural killer cell leukemia

Aggressive natural killer cell leukemia (ANKL) is a rare hematological malignancy with a fulminant clinical course. Our previous study revealed that ANKL cells proliferate predominantly in the liver sinusoids and strongly depend on transferrin supplementation. In addition, we demonstrated that liver-resident ANKL cells are sensitive to PPMX-T003, an anti-human transferrin receptor 1 inhibitory antibody, whereas spleen-resident ANKL cells are resistant to transferrin receptor 1 inhibition. However, the microenvironmental factors that regulate the iron dependency of ANKL cells remain unclear. In this study, we first revealed that the anti-neoplastic effect of PPMX-T003 was characterized by DNA double-strand breaks in a DNA replication-dependent manner, similar to conventional cytotoxic agents. We also found that the influx of extracellular amino acids via LAT1 stimulated sensitivity to PPMX-T003. Taken together, we discovered that the amount of extracellular amino acid influx through LAT1 was the key environmental factor determining the iron dependency of ANKL cells via adjustment of their mTOR/Myc activity, which provides a good explanation for the different sensitivity to PPMX-T003 between liver- and spleen-resident ANKL cells, as the liver sinusoid contains abundant amino acids absorbed from the gut.

The effectiveness of selection in a species affects the direction of amino acid frequency evolution.

Nearly neutral theory predicts that species with higher effective population size (N e ) are better able to purge slightly deleterious mutations. We compare evolution in high-N e vs. low-N e vertebrates to reveal which amino acid frequencies are subject to subtle selective preferences. We take three complementary approaches, two measuring flux and one measuring outcomes. First, we fit non-stationary substitution models of amino acid flux using maximum likelihood, comparing the high-N e clade of rodents and lagomorphs to its low-N e sister clade of primates and colugos. Second, we compare evolutionary outcomes across a wider range of vertebrates, via correlations between amino acid frequencies and N e . Third, we dissect the details of flux in human, chimpanzee, mouse, and rat, as scored by parsimony - this also enables comparison to a historical paper. All three methods agree on which amino acids are preferred under more effective selection. Preferred amino acids tend to be smaller, less costly to synthesize, and to promote intrinsic structural disorder. Parsimony-induced bias in the historical study produces an apparent reduction in structural disorder, perhaps driven by slightly deleterious substitutions. Within highly exchangeable pairs of amino acids, arginine is strongly preferred over lysine, and valine over isoleucine, consistent with more effective selection preferring a marginally larger free energy of folding. These two preferences match differences between thermophiles and mesophilic relatives. These results reveal the biophysical consequences of mutation-selection-drift balance, and demonstrate the utility of nearly neutral theory for understanding protein evolution.

Long-chain fatty acids mediate hepatic metabolic flux in preruminating dairy calves fed flaxseed oil, high oleic soybean oil, or milk fat

Nutrition and physiological state affect hepatic metabolism. Our objective was to determine if feeding flaxseed oil (∼50% C18:3n-3 cis), high oleic soybean oil (∼70% C18:1 cis-9), or milk fat (∼50% C16:0) alters hepatic expression of PC, PCK1, and PCK2 and the flow of carbons from propionate and pyruvate into the TCA cycle in preruminating calves. Male Holstein calves (n = 40) were assigned to a diet of skim milk with either: 3% milk fat (MF; n = 8), 3% flaxseed oil (Flax; n = 8), 3% high oleic soybean oil (HOSO; n = 8), 1.5% MF + 1.5% high oleic soybean oil (MF-HOSO; n = 8), or 1.5% MF + 1.5% flaxseed oil (MF-Flax; n = 8) from d 14 to d 21 postnatal. At d 21 postnatal, a liver biopsy was taken for gene expression and metabolic flux analysis. Liver explants were incubated in [U-13C] propionate and [U-13C] pyruvate to trace carbon flux through TCA cycle intermediates or with [U-14C] lactate, [1-14C] palmitic acid, or [2-14C] propionate to quantify substrate oxidation to CO2 and acid soluble products. Compared with other treatments, plasma C18:3n-3 cis was 10 times higher and C18:1 cis-9 was 3 times lower in both flax (Flax and MF-Flax) treatments. PC, PCK1, and PCK2 expression and flux of [U-13C] pyruvate as well as [U-13C] propionate were not different between treatments. PC expression was negatively correlated with the enrichment of citrate M+5 and malate M+3, and PCK2 was negatively correlated with citrate M+5, suggesting that when expression of these enzymes is increased, carbon from pyruvate enters the TCA cycle via PC mediated carboxylation, and then OAA is converted to phosphoenolpyruvate via PCK2. Acid soluble product formation and PC expression were reduced in HOSO (MF-HOSO and HOSO) treatments compared with flax (MF-Flax and Flax), indicating that fatty acids regulate PC expression and carbon flux, but that fatty acid flux control points are not connected to PC, PCK1, or PCK2. In conclusion, fatty acids regulate hepatic expression of PC, PCK1, and PCK2, and carbon flux, but the point of control is distinct.

Quantification and interpretation of postprandial whole-body protein metabolism using stable isotope methodology: a narrative review.

Stable isotopes are routinely applied to determine the impact of factors such as aging, disease, exercise, and feeding on whole-body protein metabolism. The most common approaches to quantify whole-body protein synthesis, breakdown, and oxidation rates and net protein balance are based on the quantification of plasma amino acid kinetics. In the postabsorptive state, plasma amino acid kinetics can easily be assessed using a constant infusion of one or more stable isotope labeled amino acid tracers. In the postprandial state, there is an exogenous, dietary protein-derived amino acid flux that needs to be accounted for. To accurately quantify both endogenous as well as exogenous (protein-derived) amino acid release in the circulation, the continuous tracer infusion method should be accompanied by the ingestion of intrinsically labeled protein. However, the production of labeled protein is too expensive and labor intensive for use in more routine research studies. Alternative approaches have either assumed that 100% of exogenous amino acids are released in the circulation or applied an estimated percentage based on protein digestibility. However, such estimations can introduce large artifacts in the assessment of whole-body protein metabolism. The preferred estimation approach is based on the extrapolation of intrinsically labeled protein-derived plasma bioavailability data obtained in a similar experimental design setting. Here, we provide reference data on exogenous plasma amino acid release that can be applied to allow a more accurate routine assessment of postprandial protein metabolism. More work in this area is needed to provide a more extensive reference data set.

Abstract 2050: Slc43a3 And Intracellular Fatty Acid Trafficking

Background: Fatty acids (FAs) have important roles in many cellular functions. Dysregulation of FA flux is both a characteristic and effector of many prevalent clinical conditions, including diabetes mellitus, obesity as well as insulin resistance and/or metabolic syndrome. Adipocytes take up long chain fatty acids through diffusion and protein mediated transport, whereas fatty acid efflux is considered to occur by diffusion. Our previous study suggests the possibility of a protein facilitated/regulated process of FA efflux by demonstrating that the organic anion transport inhibitor DIDS strongly inhibits FA efflux in lipolytically-stimulated adipocytes, leading to an accumulation of intracellular FAs, while not affecting glycerol efflux. We hypothesize that there is potential regulator(s) for FA efflux. Methods and Results: In a screen of subcutaneous adipose samples from obese patients for expression levels of transporters without known function, Slc43a3 (a member of the solute carrier transporter family) shown increased expression after bariatric surgery. In addition, Slc43a3 regulates fatty acid flux in adipocytes. To study the mechanism of Slc43a3 function, clonal lines of preadipocyte OP9 cells were generated, in which green fluorescent protein has been knocked-in at the C-terminus of endogenous Slc43a3 using CRISPR/Cas9. In parallel, Slc43a3 knockout (KO) line has also been generated. Examination of the cells showed that expression of Slc43a3 induced in the cytoplasm of the adipocytes during differentiation. Slc43a3KO adipocytes accumulate bigger lipid droplets and less small lipid droplets. To study the functional significance of Slc43a3 in vivo, adipose tissue specific KO of Slc43a3 were generated with floxed Slc43a3 and tamoxifen induced adiponectin-Cre recombinase. The Slc43a3 KO mice have less body weight and less % body fat on a normal chow diet; however, no differences are noted when the mice are switched to high fat diet, indicating the lack of lipid signaling molecules in the Slc43a3 KO mice with normal chow diet. Functional analysis shows a blunted stimulated lipolysis in the Slc43a3 KO mice. Conclusions: Both in vitro and in vivo data confirmed an unique role of Slc43a3 in regulating fatty acid flux in adipose tissue.

Mitochondrial enzyme FAHD1 reduces ROS in osteosarcoma

This study investigated the impact of overexpressing the mitochondrial enzyme Fumarylacetoacetate hydrolase domain-containing protein 1 (FAHD1) in human osteosarcoma epithelial cells (U2OS) in vitro. While the downregulation or knockdown of FAHD1 has been extensively researched in various cell types, this study aimed to pioneer the exploration of how increased catalytic activity of human FAHD1 isoform 1 (hFAHD1.1) affects human cell metabolism. Our hypothesis posited that elevation in FAHD1 activity would lead to depletion of mitochondrial oxaloacetate levels. This depletion could potentially result in a decrease in the flux of the tricarboxylic acid (TCA) cycle, thereby accompanied by reduced ROS production. In addition to hFAHD1.1 overexpression, stable U2OS cell lines were established overexpressing a catalytically enhanced variant (T192S) and a loss-of-function variant (K123A) of hFAHD1. It is noteworthy that homologs of the T192S variant are present in animals exhibiting increased resistance to oxidative stress and cancer. Our findings demonstrate that heightened activity of the mitochondrial enzyme FAHD1 decreases cellular ROS levels in U2OS cells. However, these results also prompt a series of intriguing questions regarding the potential role of FAHD1 in mitochondrial metabolism and cellular development.

Metabolic rewiring promotes anti-inflammatory effects of glucocorticoids.

Glucocorticoids represent the mainstay of therapy for a broad spectrum of immune-mediated inflammatory diseases. However, the molecular mechanisms underlying their anti-inflammatory mode of action have remained incompletely understood1. Here we show that the anti-inflammatory properties of glucocorticoids involve reprogramming of the mitochondrial metabolism of macrophages, resulting in increased and sustained production of the anti-inflammatory metabolite itaconate and consequent inhibition of the inflammatory response. The glucocorticoid receptor interacts with parts of the pyruvate dehydrogenase complex whereby glucocorticoids provoke an increase in activity and enable an accelerated and paradoxical flux of the tricarboxylic acid (TCA) cycle in otherwise pro-inflammatory macrophages. This glucocorticoid-mediated rewiring of mitochondrial metabolism potentiates TCA-cycle-dependent production of itaconate throughout the inflammatory response, thereby interfering with the production of pro-inflammatory cytokines. By contrast, artificial blocking of the TCA cycle or genetic deficiency in aconitate decarboxylase 1, the rate-limiting enzyme of itaconate synthesis, interferes with the anti-inflammatory effects of glucocorticoids and, accordingly, abrogates their beneficial effects during a diverse range of preclinical models of immune-mediated inflammatory diseases. Our findings provide important insights into the anti-inflammatory properties of glucocorticoids and have substantial implications for the design of new classes of anti-inflammatory drugs.