Lipid Loading Research Articles

Activation of receptor-independent fluid-phase pinocytosis promotes foamy monocyte formation in atherosclerotic mice

Atherosclerotic cardiovascular disease (ASCVD) is the leading cause of death worldwide. Clinical and experimental data demonstrated that circulating monocytes internalize plasma lipoproteins and become lipid-laden foamy cells in hypercholesterolemic subjects. This study was designed to identify the endocytic mechanisms responsible for foamy monocyte formation, perform functional and transcriptomic analysis of foamy and non-foamy monocytes relevant to ASCVD, and characterize specific monocyte subsets isolated from the circulation of normocholesterolemic controls and hypercholesterolemic patients. We hypothesized that activation of fluid-phase macropinocytosis contributes to foamy monocyte formation in vitro and in hypercholesterolemic mice in vivo. High resolution scanning electron microscopy (SEM) and quantification of FITC/TRITC-dextran internalization demonstrated macropinocytosis stimulation in human (THP-1) and wild type murine monocytes. Stimulation of macropinocytosis induced foamy monocyte formation in the presence of unmodified, native LDL (nLDL) and oxidized LDL (ox-LDL) in vitro. Genetic blockade of macropinocytosis (LysMCre+Nhe1f/f) inhibited foamy monocyte formation in hypercholesterolemic mice in vivo and attenuated monocyte adhesion to atherosclerotic aortas ex vivo. Mechanistic studies identified NADPH oxidase 2 (Nox2)-derived superoxide (O2⋅−) as an important downstream signaling molecule stimulating macropinocytosis in monocytes. qRT-PCR identified CD36 as a major SR that increases in response to lipid loading in monocytes and deletion of CD36 (Cd36-/-) inhibited foamy monocyte formation in hypercholesterolemic mice. Bulk RNA-sequencing characterized transcriptional differences between non-foamy and foamy monocytes versus macrophages. Finally, flow cytometry analysis of CD14 and CD16 expression demonstrated a significant increase in intermediate monocytes in hypercholesterolemic patients compared to normocholesterolemic controls. These results provide novel insights into the mechanisms of foamy monocyte formation and potentially identify new therapeutic targets for the treatment of atherosclerosis.

Functional improvement of non-infarct related coronary artery stenosis by extensive LDL-C reduction with a PCSK9 antibody: the FITTER trial, microvascular function analysis

Abstract Background Atherosclerosis and microvascular dysfunction are closely linked. Moreover, impaired microvascular function is strongly associated with increased risk on major adverse cardiac events. In patients presenting with acute coronary syndrome (ACS), microvascular function is reduced. PCSK9-inhibitors have shown to reduce plaque burden and lipid load in epicardial vessels and might exhibit similar results on microvascular vessels, thereby improving microvascular function. The FITTER trial offers the unique opportunity to test this hypothesis. Methods The FITTER trial is a multi-center, randomized, double blind, placebo controlled clinical trial for patients presenting with ACS and multi-vessel disease (MVD). After treatment of the culprit lesion, fractional flow reserve (FFR) measurement of non-infarct related artery (non-IRA) lesions was performed. Patients with intermediate, non-critical lesions (FFR: 0.67-0.85) were included in this trial. For exploratory purpose, additional physiological assessment of the non-IRA included coronary flow reserve (CFR) and microvascular resistance index (IMR) measurements. Microvascular function was evaluated using a pressure- and temperature-sensitive guidewire and a bolus thermodilution method. Participants were randomized and treated for 12 weeks with either evolocumab or placebo in addition to high-intensity statin therapy. Staged reevaluation was done at 12 weeks. After repeated physiology testing, PCI was performed if deemed necessary. Pre-defined exploratory analysis includes the absolute and relative difference in change between CFR and IMR from baseline to follow up in the non-IRA. PCSK9 inhibition might result in a relatively larger increase of CFR and larger decrease of IMR. Results Between March 2020 and August 2023, a total of 150 patients were included in the trial. In 61 patients (88.5% male, age 65.3 ± 9.1 years, 23.0% STEMI, 70.5% non-STEMI, 6.6% UAP) successful baseline and follow-up measurements of microvascular function were performed. Complete results will be available at the beginning of March 2024. The primary- and secondary endpoints will be submitted for late-breaking clinical trials selection. The results of this abstract will include a per vessel analysis of coronary artery microvascular function. Conclusion The FITTER trial will provide new insights on the potential of extensive LDL-C reduction with a PCSK9 antibody on microvascular coronary artery function in patients presenting with ACS and MVD.

Exploring the effect of lipid loading on acidogenic fermentation of food waste: Insights from microbiome, quorum sensing and carbon metabolic pathway

The high lipid loading in food waste (FW) is widely regarded as one of the main bottlenecks in the traditional anaerobic digestion, but the effect of lipid on FW acidogenic fermentation has been historically ignored. This study attempted to reveal the influence of lipid loading on volatile fatty acids (VFAs) production and the corresponding synthesis mechanisms during FW acidogenic fermentation. The VFAs production reached 27.10 ± 1.00 g COD/L when the lipid loading comprised 25 % of the FW dry weight, increasing by 36.88 % compared to the CK group. Lactobacillus, with a relative abundance of 59.43 %, was the predominant genus under lipid loading. In quorum sensing (QS) system, the relative abundance of luxS that is involved in the biosynthetic pathway of the Autoinducer-2 (AI-2) signal molecule increased from 0.066 % to 0.10 % in the presence of lipid. The relative abundance of Genetic Information Processing increased from 16.36 % to 24.25 % under lipid loading, with up-regulation of Translation and Replication and repair. Lipid loading not only promoted the conversion of acetyl-CoA to acetic acid, as evidenced by a 1.36-fold increase in pta gene abundance, but also concurrently suppressed the consumption of acetic acid, reflected in the 96.83 % and 77.16 % decreases in ALDH and aldB abundance, thus accumulating acetic acid. Carbon emission was also constrained under lipid loading, in which the conversion from formate to CO2 and passing Tricarboxylic acid (TCA) cycle were restricted over 60 %. Together, lipid loading in FW represents a potential strategy for enhancing VFAs production in FW acidogenic fermentation.

Magnetic resonance reveals early lipid deposition in murine prediabetes as predictive marker for cardiovascular injury

People with diabetes have an increased cardiovascular risk and a poorer outcome after myocardial infarction (MI). However, the exact underlying mechanisms are still unclear, as is the question of which non-invasive measures could be used to predict the altered risk for the patient at early stages of the disease and adapt personalized treatment. Here, we used a holistic magnetic resonance approach to monitor longitudinally not only the main target heart, but also liver, peripheral/skeletal muscle, bone marrow, and hematopoiesis during disease development and subsequent MI. In prediabetic mice, we found a strong accumulation of lipids in all organs which preceded even a significant whole-body weight gain. Intramyocellular lipids (IMCLs) were most sensitive to reveal in vivo very early alterations in tissue properties during the prediabetic state. Subsequent induction of MI led to a persistent impairment of contractile function in septal/posterior segments of prediabetic hearts which correlated with their lipid load prior MI. At the same time, prediabetic cardiomyocytes exhibited sarcomere function at its limit resulting in overload and lower compensatory contractility of the healthy myocardium after MI. In summary, we identified IMCLs as very early marker in murine prediabetes and together with the cardiac lipid load as predictive for the functional outcome after MI.

Methanogenic degradation of long-chain fatty acids associated with syntrophic microbial dynamics during thermophilic AnMBR treatment of lipid-rich dairy wastes

Anaerobic membrane reactors (AnMBRs) have been applied to effectively treat dairy wastes because they can address the drawbacks of sludge flotation and biomass washout. However, when dairy wastes contain considerable proportion of lipids, any anaerobic systems will likely encounter challenges with long-chain fatty acid (LCFA) inhibition. In this study, a thermophilic AnMBR was operated to treat dairy processing wastewater (DPW) plus ice cream waste (ICW). To evaluate the impact of LCFAs, the lipid-rich ICW was gradually introduced at different levels (0, 5, 10, and 20 %) into the AnMBR, along with a stepwise increase in lipid loading rate (LLR) from 0.22 to 11.24 g lipid/L/d. The results of biomethanation revealed that the AnMBR could be successfully operated when the LLR was maintained at 5.73 g lipid/L/d with a lipid/VS ratio of 50.22 % in the feedstock. However, at ultra-high LLR of 11.24 g lipid/L/d (20 % ICW added), a significant inhibitory effect on methane yields was observed, resulting in a reduction of 56.90 %. Dynamic analysis of the LCFAs profiled that a lower degradation rate was an important factor contributing to LCFA accumulation, and the inhibition concentration of various LCFAs was identified. Based on microbial evolution and co-occurrence network results, potential syntrophic partnerships between LCFA-degrading bacteria and methanogens were proposed. The responsible enzyme genes in the LCFA-degrading, acetoclastic, and hydrogenotrophic methanogenic pathways played crucial roles in LCFA inhibition and degradation. These findings offer practical insights for the efficient methanogenic treatment of lipid-rich wastes.

Effects of dietary fish to rapeseed oil ratio on steatosis symptoms in Atlantic salmon (Salmo salar L) of different sizes

Choline is recognized as an essential nutrient for Atlantic salmon at all developmental stages. However, its dietary requirement is not well defined. Choline plays a critical role in lipid transport, and the clearest deficiency sign is intestinal steatosis. The present work, aiming to find whether lipid source and fish size may affect steatosis symptoms, was one of a series of studies conducted to identify which production-related conditions may influence choline requirement. Six choline-deficient diets were formulated varying in ratios of rapeseed oil to fish oil and fed to Atlantic salmon of 1.5 and 4.5 kg. After eight weeks, somatic characteristics were observed, and the severity of intestinal steatosis was assessed by histological, biochemical, and molecular analyses. Fatty acid composition in pyloric intestine, mesenteric tissue, and liver samples was also quantified. The increasing rapeseed oil level increased lipid digestibility markedly, enhancing lipid supply to the fish. Moreover, small fish consumed more feed, and consequently had a higher lipid intake. In conclusion, the results showed that choline requirement depends on dietary lipid load, which depends on the fatty acid profile as well as the fish size.

A Lipid-Structured Model of Atherosclerosis with Macrophage Proliferation

Atherosclerotic plaques are fatty deposits that form in the walls of major arteries and are one of the major causes of heart attacks and strokes. Macrophages are the main immune cells in plaques and macrophage dynamics influence whether plaques grow or regress. Macrophage proliferation is a key process in atherosclerosis, particularly in the development of mid-stage plaques, but very few mathematical models include proliferation. In this paper we reframe the lipid-structured model of Ford et al. (J Theor Biol 479:48–63, 2019. https://doi.org/10.1016/j.jtbi.2019.07.003) to account for macrophage proliferation. Proliferation is modelled as a non-local decrease in the lipid structural variable. Steady state analysis indicates that proliferation assists in reducing eventual necrotic core lipid content and spreads the lipid load of the macrophage population amongst the cells. The contribution of plaque macrophages from proliferation relative to recruitment from the bloodstream is also examined. The model suggests that a more proliferative plaque differs from an equivalent (defined as having the same lipid content and cell numbers) recruitment-dominant plaque in the way lipid is distributed amongst the macrophages. The macrophage lipid distribution of an equivalent proliferation-dominant plaque is less skewed and exhibits a local maximum near the endogenous lipid content.

Moutan cortex radicis extract alleviates lipid accumulation by modulating endoplasmic reticulum stress in high-fat diet-fed mice

To analyse the effects of Moutan cortex radicis extract (ME) on lipid metabolism disorder via regulation of lipid accumulation and endoplasmic reticulum stress (ERS). C57BL/6 mice were fed an HFD for 11 weeks and then an HFD plus 200 mg/kg ME for 2 weeks. We found that ME reduced hepatic CHOL levels and lipid droplet deposition, and down-regulated PPARγ, SCD1, GRP78 and C/EBP compared to the HFD group (P < 0.05). In the oleic acid-palmitic acid-induced AML12 cell lipid load model, ME addition led to significant reduction in the intracellular TG content, lipid deposition, and mRNA levels of PERK, IRE1, ATF4, CHOP, C/EBP-α, LXRα and ACC (P < 0.05). These findings indicate that ME might be a promising treatment strategy to reverse the lipid metabolism disorder induced by an HFD and that the potential mechanism is associated with a reduction in hepatic lipid accumulation and amelioration of ERS via inhibiting PERK/IRE1-CEBP signaling pathways.

Liraglutide alleviates high-fat diet-induced kidney injury in mice by regulating the CaMKKβ/AMPK pathway

Objective Liraglutide, a glucagon-like peptide-1 receptor agonist, has been shown to regulate blood sugar and control body weight, but its ability to treat obesity-related nephropathy has been poorly studied. Therefore, this study was designed to observe the characteristics and potential mechanism of liraglutide against obesity-related kidney disease. Methods Thirty-six C57BL/6J male mice were randomly divided into six groups (n = 6 per group). Obesity-related nephropathy was induced in mice by continuous feeding of high-fat diet (HFD) for 12 weeks. After 12 weeks, liraglutide (0.6 mg/kg) and AMP-activated protein kinase (AMPK) agonists bortezomib (200 μg/kg) were injected for 12 weeks, respectively. Enzyme-linked immunosorbent assay was employed to detect the levels of total cholesterol, triglycerides, low-density lipoprotein cholesterol, blood urea nitrogen, creatinine in serum, as well as urinary protein in urine. Besides, hematoxylin–eosin staining and periodic acid-Schiff staining were used to observe the pathological changes of kidney tissue; immunohistochemistry, western blot, and real-time quantitative PCR to assess the calmodulin-dependent protein kinase kinase beta (CaMKKβ)/AMPK signaling pathway activation. Results Liraglutide significantly reduced serum lipid loading, improved kidney function, and relieved kidney histopathological damage and glycogen deposition in the mouse model of obesity-related kidney disease induced by HFD. In addition, liraglutide also significantly inhibited the CaMKKβ/AMPK signaling pathway in kidney tissue of HFD-induced mice. However, bortezomib partially reversed the therapeutic effect of liraglutide on HDF-induced nephropathy in mice. Conclusions Liraglutide has a therapeutic effect on obesity-related kidney disease, and such an effect may be achieved by inhibiting the CaMKKβ/AMPK signaling pathway in kidney tissue.

Adoption of the cardiopulmonary exercise test in the exercise ability and cardiopulmonary function rehabilitation of coronary artery disease (CAD) patients

BackgroundThis study aimed to explore the application of cardiopulmonary exercise testing in coronary artery disease (CAD) patients, evaluate its impact on exercise ability and cardiopulmonary function in patients with coronary heart disease (CHD), and promote the application of cardiopulmonary exercise testing in CAD management.MethodsFifty CHD patients after percutaneous coronary intervention (PCI) were recruited and randomly enrolled into the control (Ctrl) group and intervention (Int) group. Routine health education and health education combined with RT training were carried out for the two groups. Blood lipid levels and lung function were compared between the two groups after intervention. Cardiac function was evaluated by Doppler ultrasonography, and cardiopulmonary fitness and exercise ability were evaluated by a cardiopulmonary exercise test (CPET). The self-rating anxiety scale (SAS) and self-rating depression scale (SDS) were employed to evaluate negative emotions. The 36-item short-form (SF-36) was adopted to evaluate quality of life.Result: Compared with those in the Ctrl group, the levels of serum total cholesterol (TC), triglycerides (TGs), high-density lipoprotein (HDL), and low-density lipoprotein (LDL) decreased in the Int group, while the levels of high-density lipoprotein increased (P < 0.05). The quantitative load results showed that compared with the Ctrl group, the heart rate (HR) and self-perceived fatigue degree of the Int group decreased, and the ST segment increased (P < 0.05). Compared with the Ctrl group, the left ventricular ejection fraction (LVEF), forced expiratory volume at 1 s (FEV1), ratio of forced expiratory volume to forced vital volume (FEV1/FVC%), and maximum chase volume (MVV) increased in the Int group, while the left ventricular end diastolic diameter and left ventricular end contractile diameter decreased (P < 0.05). The results of the CPET showed that compared with the Ctrl group, minute ventilation/carbon dioxide production slope, VE/VCO2 − Peak, anaerobic threshold (AT), peak oxygen pulse (VO2/HR peak), oxygen uptake efficiency platform (OUEP), increasing power exercise time (IPEt), HR recovery 1 min after exercise, peak load power (Watt peak), and value metabolic equivalent (Watt peak) increased in the Int group (P < 0.05). Compared with the Ctrl group, the SAS and SDS scores in the Int group decreased (P < 0.05). The results of the quality of life evaluation showed that compared with the Ctrl group, the score of the SF-36 dimensions increased in the Int group (P < 0.05).ConclusionRT training can reduce postoperative blood lipid and quantitative load levels in CAD patients and improve adverse mood. Furthermore, it can improve patients’ cardiopulmonary function, cardiopulmonary fitness, exercise ability, and quality of life.

Coordination of bilayer properties by an inward-rectifier K+ channel is a cooperative process driven by protein-lipid interaction

Physical properties of biological membranes directly or indirectly govern biological processes. Yet, the interplay between membrane and integral membrane proteins is difficult to assess due to reciprocal effects between membrane proteins, individual lipids, and membrane architecture. Using solid-state NMR (SSNMR) we previously showed that KirBac1.1, a bacterial Inward-Rectifier K+ channel, nucleates bilayer ordering and microdomain formation through tethering anionic lipids. Conversely, these lipids cooperatively bind cationic residues to activate the channel and initiate K+ flux. The mechanistic details governing the relationship between cooperative lipid loading and bilayer ordering are, however, unknown. To investigate, we generated KirBac1.1 samples with different concentrations of 13C-lableded phosphatidyl glycerol (PG) lipids and acquired a full suite of SSNMR 1D temperature series experiments using the ordered all-trans (AT) and disordered trans-gauche (TG) acyl conformations as markers of bilayer dynamics. We observed increased AT ordered signal, decreased TG disordered signal, and increased bilayer melting temperature with increased PG concentration. Further, we identified cooperativity between ordering and direct binding of PG lipids, indicating KirBac1.1-driven bilayer ordering and microdomain formation is a classically cooperative Hill-type process driven by and predicated upon direct binding of PG lipids. Our results provide unique mechanistic insight into how proteins and lipids in tandem contribute to supramolecular bilayer heterogeneity in the lipid membrane.

Mitochondrial antiviral signaling protein enhances MASLD progression via the ERK/TNFα/NFκβ pathway.

Background and Aims: Mitochondrial antiviral signaling protein (MAVS) is a critical regulator that activates the host’s innate immunity against RNA viruses, and its signaling pathway has been linked to the secretion of proinflammatory cytokines. However, the actions of MAVS on inflammatory pathways during the development of metabolic dysfunction–associated steatotic liver disease (MASLD) have been little studied. Approach and Results: Liver proteomic analysis of mice with genetically manipulated hepatic p63, a transcription factor that induces liver steatosis, revealed MAVS as a target downstream of p63. MAVS was thus further evaluated in liver samples from patients and in animal models with MASLD. Genetic inhibition of MAVS was performed in hepatocyte cell lines, primary hepatocytes, spheroids, and mice. MAVS expression is induced in the liver of both animal models and people with MASLD as compared with those without liver disease. Using genetic knockdown of MAVS in adult mice ameliorates diet-induced MASLD. In vitro, silencing MAVS blunts oleic and palmitic acid–induced lipid content, while its overexpression increases the lipid load in hepatocytes. Inhibiting hepatic MAVS reduces circulating levels of the proinflammatory cytokine TNFα and the hepatic expression of both TNFα and NFκβ. Moreover, the inhibition of ERK abolished the activation of TNFα induced by MAVS. The posttranslational modification O-GlcNAcylation of MAVS is required to activate inflammation and to promote the high lipid content in hepatocytes. Conclusions: MAVS is involved in the development of steatosis, and its inhibition in previously damaged hepatocytes can ameliorate MASLD.

Cholesterol accumulation impairs HIF-1α-dependent immunometabolic reprogramming of LPS-stimulated macrophages by upregulating the NRF2 pathway

Lipid accumulation in macrophages (Mφs) is a hallmark of atherosclerosis. Yet, how lipid loading modulates Mφ inflammatory responses remains unclear. We endeavored to gain mechanistic insights into how pre-loading with free cholesterol modulates Mφ metabolism upon LPS-induced TLR4 signaling. We found that activities of prolyl hydroxylases (PHDs) and factor inhibiting HIF (FIH) are higher in cholesterol loaded Mφs post-LPS stimulation, resulting in impaired HIF-1α stability, transactivation capacity and glycolysis. In RAW264.7 cells expressing mutated HIF-1α proteins resistant to PHDs and FIH activities, cholesterol loading failed to suppress HIF-1α function. Cholesterol accumulation induced oxidative stress that enhanced NRF2 protein stability and triggered a NRF2-mediated antioxidative response prior to and in conjunction with LPS stimulation. LPS stimulation increased NRF2 mRNA and protein expression, but it did not enhance NRF2 protein stability further. NRF2 deficiency in Mφs alleviated the inhibitory effects of cholesterol loading on HIF-1α function. Mutated KEAP1 proteins defective in redox sensing expressed in RAW264.7 cells partially reversed the effects of cholesterol loading on NRF2 activation. Collectively, we showed that cholesterol accumulation in Mφs induces oxidative stress and NRF2 stabilization, which when combined with LPS-induced NRF2 expression leads to enhanced NRF2-mediated transcription that ultimately impairs HIF-1α-dependent glycolytic and inflammatory responses.

Reactive oxygen species-responsive nano-platform with dual-targeting and fluorescent lipid-specific imaging capabilities for the management of atherosclerotic plaques

Atherosclerosis (AS), a pathological cause of cardiovascular disease, results from endothelial injury, local progressive inflammation, and excessive lipid accumulation. AS plaques rich in foam cells are prone to rupture and form thrombus, which can cause life-threatening complications. Therefore, the assessment of atherosclerotic plaque vulnerability and early intervention are crucial in reducing the mortality rates associated with cardiovascular disease. In this work, A fluorescent probe FC-TPA was synthesized, which switches the fluorescence state between protonated and non-protonated, reducing background fluorescence and enhancing imaging signal-to-noise ratio. On this basis, FC-TPA is loaded into cyclodextrin (CD) modified with phosphatidylserine targeting peptide (PTP) and coated with hyaluronic acid (HA) to construct the intelligent responsive diagnostic nanoplatform (HA@PCFT). HA@PCFT effectively targets atherosclerotic plaques, utilizing dual targeting mechanisms. HA binds strongly to CD44, while PTP binds to phosphatidylserine, enabling nanoparticle aggregation at the lesion site. ROS acts as a smart release switch for probes. Both in vitro and in vivo evaluations confirm impressive lipid-specific fluorescence imaging capabilities of HA@PCFT nanoparticles (NPs). The detection of lipid load in atherosclerotic plaque by fluorescence imaging will aid in assessing the vulnerability of atherosclerotic plaque. Statement of significanceCurrently, numerous fluorescent probes have been developed for lipid imaging. However, some challenges including inadequate water solubility, nonspecific distribution patterns, and fluorescence background interference, have greatly limited their further applications in vivo. To overcome these limitations, a fluorescent molecule has been designed and synthesized, thoroughly investigating its photophysical properties through both theoretical and experimental approaches. Interestingly, this fluorescent molecule exhibits the reversible fluorescence switching capabilities, mediated by hydrogen bonds, which effectively mitigate background fluorescence interference. Additionally, the fluorescent molecules has been successfully loaded into nanocarriers functionalized with the active targeting abilities, which has significantly improved the solubility of the fluorescent molecules and reduced their nonspecific distribution in vivo for an efficient target imaging in atherosclerosis. This study provides a valuable reference for evaluating the performance of such fluorescent dyes, and offers a promising perspective on the design of the target delivery systems for atherosclerosis.

Abstract 3083: Macrophage Klf15 Prevents Foam Cell Formation And Atherosclerosis Via Transcriptional Suppression Of Olr-1

Background: Macrophage-derived foam cells are a hallmark of atherosclerosis. Scavenger receptors, including lectin-like oxidized low-density lipoprotein (LDL) receptor-1 (OLR-1), are the principal receptors responsible for the uptake and modification of LDL, facilitating macrophage lipid load and the uptake of oxidized LDL by arterial wall cells. Krüppel-like factor 15 (KLF15) is a transcription factor that regulates the expression of genes by binding to the promoter during transcription. Therefore, this study aimed to investigate the precise role of macrophage KLF15 in atherogenesis. Methods: We used two murine models of atherosclerosis: mice injected with an adeno-associated virus (AAV) encoding the Asp374-to-Tyr mutant version of human PCSK9, followed by 12 weeks on a high-fat diet (HFD), and ApoE -/ - mice on a HFD. We subsequently injected mice with AAV- KLF15 and AAV- LacZ to assess the role of KLF15 in the development of atherosclerosis in vivo . Oil Red O, H&amp;E, and Masson’s trichome staining were used to evaluate atherosclerotic lesions. Western blots and RT-qPCR were used to assess protein and mRNA levels, respectively. Results: We determined that KLF15 expression was downregulated during atherosclerosis formation, and KLF15 overexpression prevented atherosclerosis progression. KLF15 expression levels did not affect body weight or serum lipid levels in mice. However, KLF15 overexpression in macrophages prevented foam cell formation by reducing OLR-1-meditated lipid uptake. KLF15 directly targeted and transcriptionally downregulated OLR-1 levels. Restoration of OLR-1 reversed the beneficial effects of KLF15 in atherosclerosis. Conclusion: Macrophage KLF15 transcriptionally downregulated OLR-1 expression to reduce lipid uptake, thereby preventing foam cell formation and atherosclerosis. Thus, our results suggest that KLF15 is a potential therapeutic target for atherosclerosis.

Abstract 3073: Gene Regulation By RNP Granule Formation In Atherosclerosis

Background: Atherosclerosis, the leading cause of cardiovascular disease related death, is a chronic inflammatory condition incited by the accumulation of low-density lipoproteins (LDL) and recruitment of macrophages in the arterial intima. Macrophages become lipid laden foam cells that persist in the arterial vasculature, inciting chronic inflammation and plaque formation. A better understanding of the mechanisms underlying this maladaptive inflammatory response may reveal novel therapeutic strategies for intervention. Hypothesis: We hypothesize that the transformation into a foam cell state, a process that requires transcriptional rewiring, is facilitated by stress granules (SG). SGs are cytoplasmic membrane-less organelles that sequester translation initiation factors, mRNAs, and noncoding RNAs due to stress. We previously reported that SG formation is increased in plaque macrophages of atherosclerotic mice and can be induced in vitro during cellular cholesterol loading. However, the consequences of SG formation and the transcripts sequestered by these organelles is unknown. Methods and Results: To investigate whether SGs influence macrophage phenotype, we used siRNA to knockdown G3bp1 and treated with Dil-oxLDL. We found that G3bp1 knockdown reduced macrophage lipid loading and induced transcriptional changes associated with canonical M2 anti-inflammatory macrophage polarization. Furthermore, RNA-seq of macrophages treated with si G3bp1 and oxLDL showed downregulation of pathways involved in phagosome formation and S100 family signaling. RNA immunoprecipitation of G3BP1 from macrophages treated with an ER stressor revealed preferential binding of G3BP1 to transcripts implicated in cholesterol metabolism. Altogether, these data support a role for SGs in maintaining a pro-inflammatory state and regulating cholesterol pathways in plaque macrophages. Conclusion: Our data support a role for SGs in repressing the translation of RNA transcripts in lipid-laden foam cells that accumulate in atherosclerotic plaque. Identification of the specific proteins and RNAs sequestered in SGs will provide insight into how they regulate gene expression and cellular phenotypic switching that prevents the resolution of inflammation.

Obesity and polycystic ovary syndrome influence on intestinal permeability at fasting, and modify the effect of diverse macronutrients on the gut barrier

Women with the extremely prevalent polycystic ovary syndromegather multiple cardiovascular risk factors and chronic subclinical inflammation. Interactions between diet, adiposity, and gut microbiota modulate intestinal permeabilityand bacterial product translocation, and may contribute to the chronic inflammation process associated with the polycystic ovary syndrome. In the present study, we aimed to address the effects of obesity, functional hyperandrogenism, and diverse oral macronutrients on intestinal permeabilityby measuring circulating markers of gut barrier dysfunction and endotoxemia. Participants included 17 non-hyperandrogenic control women, 17 women with polycystic ovary syndrome, and 19 men that were submitted to glucose, lipid, and protein oral loads. Lipopolysaccharide-binding protein, plasma soluble CD14, succinate, zonulin family peptide, and glucagon-like peptide-2 were determined at fasting and after oral challenges. Macronutrient challenges induced diverse changes on circulating intestinal permeabilitybiomarkers in the acute postprancial period, with lipids and proteins showing the most unfavorable and favorable effects, respectively. Particularly, lipopolysaccharide-binding protein, zonulin family peptide, and glucagon-like peptide-2 responses were deregulated by the presence of obesity after glucose and lipid challenges. Obese subjects showed higher fasting intestinal permeabilitybiomarkers levels than non-obese individuals, except for plasma soluble CD14. The polycystic ovary syndromeexacerbated the effect of obesity further increasing fasting glucagon-like peptide-2, lipopolysaccharide-binding protein, and succinate concentrations. We observed specific interactions of the polycystic ovary syndromewith obesity in the postprandial response of succinate, zonulin family peptide, and glucagon-like peptide-2. In summary, obesity and polycystic ovary syndromemodify the effect of diverse macronutrients on the gut barrier, and alsoinfluence intestinal permeabilityat fasting,contributing to the morbidity of functional hyperandrogenism by inducing endotoxemia and subclinical chronic inflammation.

Resolvin T4 enhances macrophage cholesterol efflux to reduce vascular disease

While cardiovascular disease (CVD) is one of the major co-morbidities in patients with rheumatoid arthritis (RA), the mechanism(s) that contribute to CVD in patients with RA remain to be fully elucidated. Herein, we observe that plasma concentrations of 13-series resolvin (RvT)4 negatively correlate with vascular lipid load in mouse inflammatory arthritis. Administration of RvT4 to male arthritic mice fed an atherogenic diet significantly reduces atherosclerosis. Assessment of the mechanisms elicited by this mediator demonstrates that RvT4 activates cholesterol efflux in lipid laden macrophages via a Scavenger Receptor class B type 1 (SR-BI)-Neutral Cholesterol Ester Hydrolase-dependent pathway. This leads to the reprogramming of lipid laden macrophages yielding tissue protection. Pharmacological inhibition or knockdown of macrophage SR-BI reverses the vasculo-protective activities of RvT4 in vitro and in male mice in vivo. Together these findings elucidate a RvT4-SR-BI centered mechanism that orchestrates macrophage responses to limit atherosclerosis during inflammatory arthritis.

Influence of coffee roasting degree on antioxidant and metabolic parameters: Comprehensive in vitro and in vivo analysis

This study aimed to assess the impact of roasting degree on antioxidant and metabolic parameters in vitro and in vivo. In vitro, we evaluated radical scavenging, lipid peroxidation, and the activity of digestive enzymes (α-glucosidase, α-amylase, and lipase). In vivo, we first examined coffee's effect on carbohydrate and lipid absorption in healthy rats, followed by a chronic evaluation of metabolic disorders and antioxidant markers using a diet-induced obesity model. In vitro results revealed that increased roasting degree reduced the antioxidant capacity of coffee brews. All brews showed lower inhibition of α-glucosidase and α-amylase, and lipase inhibition compared to the positive control (acarbose or orlistat). In vivo, all roasting degrees consistently reduced postprandial glucose levels by 20%. Notably, coffee with a high roasting degree (HRD) decreased serum triglycerides (TG) by ∼44% after a lipid load, while other roasts did not. Chronic administration of unroasted (UN) or HRD coffee significantly reduced weight gain compared to the obese control (∼15% and ∼10%, respectively). Notably, all coffee samples improved lipid metabolism parameters. UN and HRD coffee significantly decreased adipocyte volume by 58% and 48%, respectively, compared to the obese control. Additionally, all groups exhibited less than 30% hepatic lipid droplets independent of roasting degree. HRD treatment notably increased liver catalase (CAT) activity and reduced lipid peroxidation in serum (∼90%), liver (∼59%), and adipose tissue (∼37%) compared to the obese control group. These findings suggest that HRD in coffee may confer certain biological advantages.

Plasticizer acetyl triethyl citrate (ATEC) induces lipogenesis and obesity

Environmental chemicals, such as plasticizers, have been linked to increased rates of obesity, according to epidemiological studies. Acetyl triethyl citrate (ATEC) is a plasticizer that is commonly utilized in pharmaceutical products and food packaging as a non-phthalate alternative. Due to its direct contact with the human body and high leakage rate from the polymers, assessment of the potential risk of ATEC exposure at environmentally relevant low doses to human health is needed. Male C57BL/6 J mice were fed diets containing ATEC at doses of either 0.1 or 10 μg/kg per day in a period of 12 weeks to mimic the real exposure environment. The findings suggest that in C57BL/6 J mice, ATEC exposure resulted in increased body weight gain, body fat percentage, and benign hepatocytes, as well as adipocyte size. Consistent with in vivo models, ATEC treatment obviously stimulated the increase of intracellular lipid load in both mouse and human hepatocytes. Mechanically, ATEC induced the transcriptional expression of genes involved in de novo lipogenesis and lipid uptake. Using both enzyme inhibitor and small interfering RNA (siRNA) transfection, we found that stearoyl-coenzyme A desaturase 1 (SCD1) played a significant role in ATEC-induced intracellular lipid accumulation. This study for the first time provided initial evidence suggesting the obesogenic and fatty liver-inducing effect of ATEC at low doses near human exposure levels, and ATEC might be a potential environmental obesogen and its effect on human health need to be further evaluated.