Hyperlipidemic Mouse Model Research Articles

Chronic exposure to E-cigarette aerosols potentiates atherosclerosis in a sex-dependent manner

Despite being designed for smoking cessation, e-cigarettes and their variety of flavors have become increasingly attractive to teens and young adults. This trend has fueled concerns regarding the potential role of e-cigarettes in advancing chronic diseases, notably those affecting the cardiovascular system. E-cigarettes contain a mixture of metals and chemical compounds, some of which have been implicated in cardiovascular diseases like atherosclerosis. Our laboratory has optimized in vivo exposure regimens to mimic human vaping patterns. Using these established protocols in an inducible (AAV-PCSK9) hyperlipidemic mouse model, this study tests the hypothesis that a chronic exposure to e-cigarette aerosols will increase atherosclerotic plaques. The exposures were conducted using the SCIREQ InExpose™ nose-only inhalation system and STLTH or Vuse products for 16 weeks. We observed that only male mice exposed to STLTH or Vuse aerosols had significantly increased plasma total cholesterol, triglycerides, and LDL cholesterol levels compared to mice exposed to system air. Moreover, these male mice also had a significant increase in aortic and sinus plaque area. Male mice exposed to e-cigarette aerosol had a significant reduction in weight gain over the exposure period. Our data indicate that e-cigarette use in young hyperlipidemic male mice increases atherosclerosis in the absence of significant pulmonary and systemic inflammation. These results underscore the need for extensive research to unravel the long-term health effects of e-cigarettes.

Widely Targeted Lipidomics and Microbiomics Perspectives Reveal the Mechanism of Auricularia auricula Polysaccharide's Effect of Regulating Glucolipid Metabolism in High-Fat-Diet Mice.

The role of Auricularia auricula polysaccharide (AP) in the regulation of glycolipid metabolism was investigated using a high-fat-diet-induced hyperlipidemic mouse model. In a further step, its potential mechanism of action was investigated using microbiome analysis and widely targeted lipidomics. Compared to high-fat mice, dietary AP supplementation reduced body weight by 13.44%, liver index by 21.30%, epididymal fat index by 50.68%, fasting blood glucose (FBG) by 14.27%, serum total cholesterol (TC) by 20.30%, serum total triglycerides (TGs) by 23.81%, liver non-esterified fatty acid (NEFA) by 20.83%, liver TGs by 20.00%, and liver malondialdehyde (MDA) by 21.05%, and increased liver glutathione oxidase (GSH-PX) activity by 52.24%, total fecal bile acid (TBA) by 46.21%, and fecal TG by 27.16%, which significantly regulated glucose and lipid metabolism. Microbiome analysis showed that AP significantly downregulated the abundance of the Desulfobacterota phylum, as well as the genii Desulfovibrio, Bilophila, and Oscillbacter in the cecum of hyperlipidemic mice, which are positively correlated with high lipid indexes, while it upregulated the abundance of the families Eubacterium_coprostanoligenes_group and Ruminococcaceae, as well as the genii Eubacterum_xylanophilum_group, Lachnospiraceae_NK4A136_group, Eubacterium_siraeum_group, and Parasutterella, which were negatively correlated with high lipid indexes. In addition, AP promoted the formation of SCFAs by 119.38%. Widely targeted lipidomics analysis showed that AP intervention regulated 44 biomarkers in metabolic pathways such as sphingolipid metabolism and the AGE-RAGE signaling pathway in the hyperlipidemic mice (of which 15 metabolites such as unsaturated fatty acids, phosphatidylserine, and phosphatidylethanolamine were upregulated, and 29 metabolites such as phosphatidylcholine, ceramide, carnitine, and phosphatidylinositol were downregulated), thereby correcting glucose and lipid metabolism disorders.

A novel lncRNA GM47544 modulates triglyceride metabolism by inducing ubiquitination-dependent protein degradation of APOC3

ObjectiveEmerging evidence highlights the pivotal roles of long non-coding RNAs (lncRNAs) in lipid metabolism. Apoprotein C3 (ApoC3) is a well-established therapeutic target for hypertriglyceridemia and exhibits a strong association with cardiovascular disease. However, the exact mechanisms via which the lncRNAs control ApoC3 expression remain unclear. MethodsWe identified a novel long noncoding RNA (lncRNA), GM47544, within the ApoA1/C3/A4/A5 gene cluster. Subsequently, the effect of GM47544 on intracellular triglyceride metabolism was analyzed. The diet-induced mouse models of hyperlipidemia and atherosclerosis were established to explore the effect of GM47544 on dyslipidemia and plaque formation in vivo. The molecular mechanism was explored through RNA sequencing, immunoprecipitation, RNA pull-down assay, and RNA immunoprecipitation. ResultsGM47544 was overexpressed under high-fat stimulation. GM47544 effectively improved hepatic steatosis, reduced blood lipid levels, and alleviated atherosclerosis in vitro and in vivo. Mechanistically, GM47544 directly bound to ApoC3 and facilitated the ubiquitination at lysine 79 in ApoC3, thereby facilitating ApoC3 degradation via the ubiquitin-proteasome pathway. Moreover, we identified AP006216.5 as the human GM47544 transcript, which fulfills a comparable function in human hepatocytes. ConclusionsThe identification of GM47544 as a lncRNA modulator of ApoC3 reveals a novel mechanism of post-translational modification, with significant clinical implications for the treatment of hypertriglyceridemia and atherosclerosis.

Combining bioinformatics and multiomics strategies to investigate the key microbiota and active components of Liupao tea ameliorating hyperlipidemia

Ethnopharmacological relevanceHyperlipidemia as a major health issue has attracted much public attention. As a geographical indication product of China, Liupao tea (LPT) is a typical representative of traditional Chinese dark tea that has shown good potential in regulating glucose and lipid metabolism. LPT has important medicinal value in hyperlipidemia prevention. However, the active ingredients and metabolic mechanisms by which LPT alleviates hyperlipidemia remain unclear. Aim of the studyThis study aimed to systematically investigate the metabolic mechanisms and active ingredients of LPT extract in alleviating hyperlipidemia. Materials and methodsFirstly, we developed a mouse model of hyperlipidemia to study the pharmacodynamics of LPT. Subsequently, network pharmacology and molecular docking were performed to predict the potential key active ingredients and core targets of LPT against hyperlipidemia. LC-MS/MS was used to validate the identity of key active ingredients in LPT with chemical standards. Finally, the effect and metabolic mechanisms of LPT extract in alleviating hyperlipidemia were investigated by integrating metabolomic, lipidomic, and gut microbiome analyses. ResultsResults showed that LPT extract effectively improved hyperlipidemia by suppressing weight gain, remedying dysregulation of glucose and lipid metabolism, and reducing hepatic damage. Network pharmacology analysis and molecular docking suggested that four potential active ingredients and seven potential core targets were closely associated with roles for hyperlipidemia treatment. Ellagic acid, catechin, and naringenin were considered to be the key active ingredients of LPT alleviating hyperlipidemia. Additionally, LPT extract modulated the mRNA expression levels of Fxr, Cyp7a1, Cyp8b1, and Cyp27a1 associated with bile acid (BA) metabolism, mitigated the disturbances of BA and glycerophospholipid (GP) metabolism in hyperlipidemia mice. Combining fecal microbiota transplantation and correlation analysis, LPT extract effectively improved species diversity and abundance of gut microbiota, particularly the BA and GP metabolism-related gut microbiota, in the hyperlipidemia mice. ConclusionsLPT extract ameliorated hyperlipidemia by modulating GP and BA metabolism by regulating Lactobacillus and Dubosiella, thereby alleviating hyperlipidemia. Three active ingredients of LPT served as the key factors in exerting an improvement on hyperlipidemia. These findings provide new insights into the active ingredients and metabolic mechanisms of LPT in improving hyperlipidemia, suggesting that LPT can be used to prevent and therapeutic hyperlipidemia.

Apolipoprotein O modulates cholesterol metabolism via NRF2/CYB5R3 independent of LDL receptor

Apolipoprotein O (APOO) plays a critical intracellular role in regulating lipid metabolism. Here, we investigated the roles of APOO in metabolism and atherogenesis in mice. Hepatic APOO expression was increased in response to hyperlipidemia but was inhibited after simvastatin treatment. Using a novel APOO global knockout (Apoo−/−) model, it was found that APOO depletion aggravated diet-induced obesity and elevated plasma cholesterol levels. Upon crossing with low-density lipoprotein receptor (LDLR) and apolipoprotein E (APOE) knockout hyperlipidemic mouse models, Apoo−/−Apoe−/− and Apoo−/−Ldlr−/− mice exhibited elevated plasma cholesterol levels, with more severe atherosclerotic lesions than littermate controls. This indicated the effects of APOO on cholesterol metabolism independent of LDLR and APOE. Moreover, APOO deficiency reduced cholesterol excretion through bile and feces while decreasing phospholipid unsaturation by inhibiting NRF2 and CYB5R3. Restoration of CYB5R3 expression in vivo by adeno-associated virus (AAV) injection reversed the reduced degree of phospholipid unsaturation while decreasing blood cholesterol levels. This represents the first in vivo experimental validation of the role of APOO in plasma cholesterol metabolism independent of LDLR and elucidates a previously unrecognized cholesterol metabolism pathway involving NRF2/CYB5R3. APOO may be a metabolic regulator of total-body cholesterol homeostasis and a target for atherosclerosis management.Apolipoprotein O (APOO) regulates plasma cholesterol levels and atherosclerosis through a pathway involving CYB5R3 that regulates biliary and fecal cholesterol excretion, independently of the LDL receptor. In addition, down-regulation of APOO may lead to impaired mitochondrial function, which in turn aggravates diet-induced obesity and fat accumulation.

Abstract 2069: Impact Of Endothelial Stiffening And CD36-mediated Lipid Uptake On Atherosclerotic Lesion Formation

Objective: To determine the impact of endothelial stiffening induced by CD36 (cluster of differentiation 36) lipid uptake in the development of atherosclerosis. Approach and Results: Endothelial stiffening of intact en face monolayers, as assessed by atomic force microscopy, reveal the hyperlipidemic mouse model of atherosclerosis, LDLR -/- (low density lipoprotein receptor knockout) is significantly stiffer, especially in the atheroprone aortic arch, as compared to wild type controls (n=4 male mice with 8-15 tissue measurement sites per condition per mouse, p<0.05). Furthermore, a 4-5 month high fat diet (HFD) feeding in LDLR -/- mice resulted in significantly enhanced stiffening compared to regular chow feeding, an effect that was observed in males but not females (n=6-8 mice, p<0.05). There was a significantly greater plaque development, as analyzed using Oil O Red staining for lipids, in male HFD-fed LDLR -/- mice, as compared to age-matched females (n=5-6, p<0.05). No significant difference between males and females was found in lipid panel parameters (n=4-6) or weight gain (n=10-14). Specific downregulation of CD36 in male hypercholesterolemic Cdh5.CreERT2CD36fl/fl LDLR -/- mice led to both a significant decrease in endothelial stiffening (n=5 mice, p<0.05) and in the areas of atherosclerotic lesion formation (n=5-6 mice, p<0.05). Mechanistically, we show that the saturated fatty acid, palmitic acid, increased endothelial stiffening in the en face monolayers of wild type mice, an effect which is prevented by the pre-treatment of the CD36 irreversible inhibitor, SSO (sulfo-N-succinimidyl oleate, n=5-6 male mice, p<0.05). Conclusions: These results highlight that endothelial stiffness from intact aortas and atherosclerotic plaque formation following HFD feeding is strongly dependent on endothelial CD36.

Untargeted metabolomics based on ultra-high performance liquid chromatography-mass spectrometry/MS reveals the lipid-lowering mechanism of taurine in hyperlipidemia mice.

Taurine has a prominent lipid-lowering effect on hyperlipidemia. However, a comprehensive analysis of the effects of taurine on endogenous metabolites in hyperlipidemia has not been documented. This study aimed to explore the impact of taurine on multiple metabolites associated with hyperlipidemia. The hyperlipidemic mouse model was induced by high-fat diet (HFD). Taurine was administered via oral gavage at doses of 700 mg/kg/day for 14 weeks. Evaluation of body weight, serum lipid levels, and histopathology of the liver and adipose tissue was performed to confirm the lipid-lowering effect of taurine. Ultra-high performance liquid chromatography-mass spectrometry (UPLC-MS)-based metabonomics analyses of serum, urine, feces, and liver, coupled with multivariate data analysis, were conducted to assess changes in the endogenous metabolites. Biochemical and histological examinations demonstrated that taurine administration prevented weight gain and dyslipidemia, and alleviated lipid deposition in the liver and adipose tissue in hyperlipidemic mice. A total of 76 differential metabolites were identified by UPLC-MS-based metabolomics approach, mainly involving BAs, GPs, SMs, DGs, TGs, PUFAs and amino acids. Taurine was found to partially prevent HFDinduced abnormalities in the aforementioned metabolites. Using KEGG database and MetaboAnalyst software, it was determined that taurine effectively alleviates metabolic abnormalities caused by HFD, including fatty acid metabolism, sphingolipid metabolism, glycerophospholipid metabolism, diacylglycerol metabolism, amino acid metabolism, bile acid and taurine metabolism, taurine and hypotaurine metabolism. Moreover, DGs, GPs and SMs, and taurine itself may serve as active metabolites in facilitating various anti-hyperlipidemia signal pathways associated with taurine. This study provides new evidence for taurine to prevent hyperlipidemia.

Disulfiram Reduces Atherosclerosis and Enhances Efferocytosis, Autophagy, and Atheroprotective Gut Microbiota in Hyperlipidemic Mice.

Pyroptosis executor GsdmD (gasdermin D) promotes atherosclerosis in mice and humans. Disulfiram was recently shown to potently inhibit GsdmD, but the invivo efficacy and mechanism of disulfiram's antiatherosclerotic activity is yet to be explored. We used human/mouse macrophages, endothelial cells, and smooth muscle cells and a hyperlipidemic mouse model of atherosclerosis to determine disulfiram antiatherosclerotic efficacy and mechanism. The effects of disulfiram on several atheroprotective pathways such as autophagy, efferocytosis, phagocytosis, and gut microbiota were determined. Atomic force microscopy was used to determine the effects of disulfiram on the biophysical properties of the plasma membrane of macrophages. Disulfiram-fed hyperlipidemic apolipoprotein E-/- mice showed significantly reduced interleukin-1β release upon invivo Nlrp3 (NLR family pyrin domain containing 3) inflammasome activation. Disulfiram-fed mice showed smaller atherosclerotic lesions (~27% and 29% reduction in males and females, respectively) and necrotic core areas (~50% and 46% reduction in males and females, respectively). Disulfiram induced autophagy in macrophages, smooth muscle cells, endothelial cells, hepatocytes/liver, and atherosclerotic plaques. Disulfiram modulated other atheroprotective pathways (eg,efferocytosis, phagocytosis) and gut microbiota. Disulfiram-treated macrophages showed enhanced phagocytosis/efferocytosis, with the mechanism being a marked increase in cell-surface expression of efferocytic receptor MerTK. Atomic force microscopy analysis revealed altered biophysical properties of disulfiram-treated macrophages, showing increased order-state of plasma membrane and increased adhesion strength. Furthermore, 16sRNA sequencing of disulfiram-fed hyperlipidemic mice showed highly significant enrichment in atheroprotective gut microbiota Akkermansia and a reduction in atherogenic Romboutsia species. Taken together, our data show that disulfiram can simultaneously modulate several atheroprotective pathways in a GsdmD-dependent as well as GsdmD-independent manner.

Role of inflammatory signaling pathways involving the CD40–CD40L–TRAF cascade in diabetes and hypertension—insights from animal and human studies

CD40L–CD40–TRAF signaling plays a role in atherosclerosis progression and affects the pathogenesis of coronary heart disease (CHD). We tested the hypothesis that CD40L–CD40–TRAF signaling is a potential therapeutic target in hyperlipidemia, diabetes, and hypertension. In mouse models of hyperlipidemia plus diabetes (db/db mice) or hypertension (1 mg/kg/d angiotensin-II for 7 days), TRAF6 inhibitor treatment (2.5 mg/kg/d for 7 or 14 days) normalized markers of oxidative stress and inflammation. As diabetes and hypertension are important comorbidities aggravating CHD, we explored whether the CD40L–CD40–TRAF signaling cascade and their associated inflammatory pathways are expressed in CHD patients suffering from comorbidities. Therefore, we analyzed vascular bypass material (aorta or internal mammary artery) and plasma from patients with CHD with diabetes and/or hypertension. Our Olink targeted plasma proteomic analysis using the IMMUNO-ONCOLOGY panel revealed a pattern of step-wise increase for 13/92 markers of low-grade inflammation with significant changes. CD40L or CD40 significantly correlated with 38 or 56 other inflammatory targets. In addition, specific gene clusters that correlate with the comorbidities were identified in isolated aortic mRNA of CHD patients through RNA-sequencing. These signaling clusters comprised CD40L–CD40–TRAF, immune system, hemostasis, muscle contraction, metabolism of lipids, developmental biology, and apoptosis. Finally, immunological analysis revealed key markers correlated with comorbidities in CHD patients, such as CD40L, NOX2, CD68, and 3-nitrotyrosine. These data indicate that comorbidities increase inflammatory pathways in CHD, and targeting these pathways will be beneficial in reducing cardiovascular events in CHD patients with comorbidities.

A novel mouse model of familial combined hyperlipidemia and atherosclerosis.

Within the context of residual cardiovascular risk in post-statin era, emerging evidence from epidemiologic and human genetic studies have demonstratedthattriglyceride (TG)-rich lipoproteinsand their remnants are causally related to cardiovascular risk. While,carriers of loss-of-function mutations of ApoC3 havelow TG levelsand are protected from cardiovascular disease (CVD). Of translational significance,siRNAs/antisense oligonucleotide (ASO) targeting ApoC3 is beneficial for patients with atheroscleroticCVD. Therefore, animal models of atherosclerosiswith bothhypercholesterolemia and hypertriglyceridemia are important for the discovery of novel therapeutic strategies targeting TG-lowering on top of traditional cholesterol-lowering.In this study, we constructed a novel mouse model of familial combined hyperlipidemia through inserting a human ApoC3 transgene(hApoC3-Tg) into C57BL/6 J mice and injecting a gain-of-function variant of adeno-associated virus-proprotein convertase subtilisin/kexin type 9 (AAV-PCSK9)-D377Y concurrently with high cholesterol diet (HCD) feeding for 16 weeks. In the last 10 weeks, hApoC3-Tg mice were orally treated with a combination of atorvastatin (10 mg·kg-1·d-1) and fenofibrate (100 mg·kg-1·d-1). HCD-treated hApoC3-Tg mice demonstrated elevated levels of serum TG, total cholesterol (TC) and low density lipoprotein-cholesterol (LDL-C). Oral administration of atorvastatin and fenofibrate significantly decreased the plaque sizes of en face aorta, aortic sinus and innominate artery accompanied by improved lipid profileand distribution. In summary,this novel mouse model is of considerable clinical relevance for evaluation of anti-atherosclerotic drugsby targeting both hypercholesterolemia and hypertriglyceridemia.

Incidence of microvascular dysfunction is increased in hyperlipidemic mice, reducing cerebral blood flow and impairing remote memory.

The development of cognitive dysfunction is not necessarily associated with diet-induced obesity. We hypothesized that cognitive dysfunction might require additional vascular damage, for example, in atherosclerotic mice. We induced atherosclerosis in male C57BL/6N mice by injecting AAV-PCSK9DY (2x1011 VG) and feeding them a cholesterol-rich Western diet. After 3 months, mice were examined for cognition using Barnes maze procedure and for cerebral blood flow. Cerebral vascular morphology was examined by immunehistology. In AAV-PCSK9DY-treated mice, plaque burden, plasma cholesterol, and triglycerides are elevated. RNAseq analyses followed by KEGG annotation show increased expression of genes linked to inflammatory processes in the aortas of these mice. In AAV-PCSK9DY-treated mice learning was delayed and long-term memory impaired. Blood flow was reduced in the cingulate cortex (-17%), caudate putamen (-15%), and hippocampus (-10%). Immunohistological studies also show an increased incidence of string vessels and pericytes (CD31/Col IV staining) in the hippocampus accompanied by patchy blood-brain barrier leaks (IgG staining) and increased macrophage infiltrations (CD68 staining). We conclude that the hyperlipidemic PCSK9DY mouse model can serve as an appropriate approach to induce microvascular dysfunction that leads to reduced blood flow in the hippocampus, which could explain the cognitive dysfunction in these mice.

Subchronic inhalation exposure to ultrafine particulate matter alters the intestinal microbiome in various mouse models

Exposure to ultrafine particles (UFPs) has been associated with multiple adverse health effects. Inhaled UFPs could reach the gastrointestinal tract and influence the composition of the gut microbiome. We have previously shown that oral ingestion of UFPs alters the gut microbiome and promotes intestinal inflammation in hyperlipidemic Ldlr−/− mice. Particulate matter (PM)2.5 inhalation studies have also demonstrated microbiome shifts in normolipidemic C57BL/6 mice. However, it is not known whether changes in microbiome precede or follow inflammatory effects in the intestinal mucosa. We hypothesized that inhaled UFPs modulate the gut microbiome prior to the development of intestinal inflammation. We studied the effects of UFP inhalation on the gut microbiome and intestinal mucosa in two hyperlipidemic mouse models (ApoE−/− mice and Ldlr−/− mice) and normolipidemic C57BL/6 mice. Mice were exposed to PM in the ultrafine-size range by inhalation for 6 h a day, 3 times a week for 10 weeks at a concentration of 300–350 μg/m3.16S rRNA gene sequencing was performed to characterize sequential changes in the fecal microbiome during exposures, and changes in the intestinal microbiome at the end. PM exposure led to progressive differentiation of the microbiota over time, associated with increased fecal microbial richness and evenness, altered microbial composition, and differentially abundant microbes by week 10 depending on the mouse model. Cross-sectional analysis of the small intestinal microbiome at week 10 showed significant changes in α-diversity, β-diversity, and abundances of individual microbial taxa in the two hyperlipidemic models. These alterations of the intestinal microbiome were not accompanied, and therefore could not be caused, by increased intestinal inflammation as determined by histological analysis of small and large intestine, cytokine gene expression, and levels of fecal lipocalin. In conclusion, 10-week inhalation exposures to UFPs induced taxonomic changes in the microbiome of various animal models in the absence of intestinal inflammation.

Metabolomics and network pharmacology reveal partial insights into the hypolipidemic mechanisms of ferulic acid in a dyslipidemia mouse model.

Hyperlipidemia is a condition characterized by abnormal levels of lipids and lipoproteins in the plasma, posing significant health risks. Ferulic acid (FA) is an organic acid with therapeutic properties for diabetes and hyperlipidemia. To explore biomarkers for FA treatment of hyperlipidemia and elucidate the mechanisms of lipid-lowering-related changes in metabolic pathways by metabolomics and network pharmacology. Initially, a hyperlipidemic mouse model induced by triton WR-1339 was established to evaluate the therapeutic effects of FA. Subsequently, serum metabolomics was utilized to identify differential metabolites, and metabolic pathway analysis was performed using MetaboAnalyst 6.0. Thirdly, network pharmacology was employed to identify potential targets of FA for hyperlipidemia. Finally, the compound-target-metabolite (C-T-M) network obtained core targets and validated them with molecular docking. Biochemical analysis and histological examination showed that FA had lipid-lowering effects on hyperlipidemic mice. It identified 31 potential biomarkers for FA against hyperlipidemia by metabolomics involving lipid and amino acid metabolism. Lipid and atherosclerosis signaling pathways were identified as the key signaling pathways of FA against hyperlipidemia by KEGG analysis. Conjoint analysis showed that FA against hyperlipidemia was associated with 18 core targets and six biomarkers. Molecular docking results showed that FA has a high binding affinity to these core targets. Through the synergy of network pharmacology and metabolomics, this study provides insights into how FA regulates endogenous metabolites, underscoring its promise as a treatment for hyperlipidemia.

Expression and Role of PDK4 on Childhood Dyslipidemia and Lipid Metabolism in Hyperlipidemic Mice.

Hyperlipidemia is a common metabolic disorder that can lead to cardiovascular disease. PDK4 is a key enzyme that regulates glucose and fatty acid metabolism and homeostasis. The aim of this study is to explore the correlation between PDK4 expression and dyslipidemia in obese children, and to find new therapeutic targets for hyperlipidemia in children. The expression of PDK4 in serum was detected by qRT-PCR. Receiver operating characteristic curve was used to analyze the relationship between PDK4 and dyslipidemia. Upstream miRNAs of PDK4 were predicted by the database and verified by dual luciferase reporter gene assay and detected by qRT-PCR. The hyperlipidemia mouse model was established by high-fat diet (HFD) feeding, and the metabolic disorders of mice were detected. PDK4 is poorly expressed in the serum of obese children. The upstream of PDK4 may be inhibited by miR-107, miR-27a-3p, and miR-106b-5p, which are highly expressed in the serum of obese children. Overexpression of PDK4 improves lipid metabolism in HFD mice. miR-27a-3p silencing upregulates PDK4 to improve lipid metabolism. In conclusion, PDK4 has a diagnostic effect on dyslipidemia in children, while lipid metabolism in hyperlipidemic mice could be mitigated by upregulation of PDK4, which was inhibited by miR-107, miR-27a-3p and miR-106b-5p on upstream.

Danlou Recipe promotes cholesterol efflux in macrophages RAW264.7 and reverses cholesterol transport in mice with hyperlipidemia induced by P407

IntroductionLiver X Receptor (LXR) agonists could attenuate the development of atherosclerosis but bring excess lipid accumulation in the liver. Danlou Recipe was believed to be a benefit for improving the lipid profile. Thus, it is unclear whether Danlou Recipe could attenuate hyperlipidemia without excess lipid accumulated in the liver of mice. This study aimed to clarify if Danlou Recipe could alleviate the progression of hyperlipidemia in mice without extra lipids accumulated in the liver.MethodsMale murine macrophage RAW264.7 cells and murine peritoneal macrophages were used for the in vitro experiments. Cellular cholesterol efflux was determined using the fluorescent cholesterol labeling method. Those genes involved in lipid metabolism were evaluated by qRT‐PCR and western blotting respectively. In vivo, a mouse model of hyperlipidemia induced by P407 was used to figure out the effect of Danlou Recipe on reverse cholesterol transport (RCT) and hyperlipidemia. Ethanol extract of Danlou tablet (EEDL) was prepared by extracting the whole powder of Danlou Prescription from ethanol, and the chemical composition was analyzed by ultra-performance liquid chromatography (UPLC).ResultsEEDL inhibits the formation of RAW264.7 macrophage-derived foam cells, and promotes ABCA1/apoA1 conducted cholesterol efflux in RAW264.7 macrophages and mouse peritoneal macrophages. In the P407-induced hyperlipidemia mouse model, oral administration of EEDL can promote RCT in vivo and improve fatty liver induced by a high-fat diet. Consistent with the findings in vitro, EEDL promotes RCT by upregulating the LXR activities.ConclusionOur results demonstrate that EEDL has the potential for targeting RCT/LXR in the treatment of lipid metabolism disorders to be developed as a safe and effective therapy.

IL-1 alpha is myristoylated for monocyte surface translocation and promotes atherosclerosclerosis in mice

Abstract Background The role of Interleukin-1 alpha (IL-1α) and beta (IL-1β) in the pathogenesis of atherosclerosis in vivo has been controversial for years. In contrast to IL-1β, which is dependent on the NLR family pyrin domain containing 3 (NLRP3) inflammasome, IL-1α can be tethered to the plasma membrane (csIL-1α) where it exerts pro-inflammatory effects. We characterised the role of IL-1α and the Nlrp3 inflammasome in a nongenetic hyperlipidemic mouse model and examined the function of csIL-1α in human monocytes. Methods Atherosclerosis was induced by a single injection of mutant PCSK9-AAV8 virus in wild-type (WT, referred to as PCSK9-AAV), Il1a-/-, Nlrp3-/-, and Il1b-/- mice, in combination with high-fat western diet (21% fat, 0.2% cholesterol) for 12 weeks. Plaque size and lipid content, were histologically analysed. The serum proteome was studied using a targeted multiplex PEA (Olink Proteomics). Human primary monocytes and human umbilical vein endothelial cells (HUVECs) were used to examine csIL-1α-IL1R interaction by analyzing VCAM1 expression and monocyte adhesion. LPS was used to induce IL-1α protein synthesis and membrane translocation in vitro. Myristoylation was measured via flow cytometry. Results Il1a-/- animals showed significantly reduced atherosclerotic lesion size (-62% vs. PCSK9-AAV) and fat accumulation within the plaque (-64% vs. PCSK9-AAV). Knockout of Nlrp3 and IL-1β did not reduce plaque size or lipid content, indicating an Nlrp3-independent mechanism of the protective effect of Il1a-/-. Circulating pro-inflammatory cytokines such as IL-1α, IL-1β, and IL-6 were downregulated in serum of Il1a-/-, Nlrp3-/-, and Il1b-/- mice compared to PCSK9-AAV mice suggesting that the development of atherosclerotic plaques is independent of circulating inflammatory cytokines at an early stage. Therefore, we investigated the role of csIL-1α on human monocytes. Stimulation with LPS showed enrichment of csIL-1α by 13.2% (p<0.05) on monocytes without releasing IL-1α. Treatment of HUVECs with LPS-stimulated monocytes resulted in increased VCAM1 expression (+22.2%, p<0.05) and monocyte adhesion (1.9 fold, p<0.05) through a csIL-1α-IL1R interaction which was abolished by the addition of neutralising antibodies for IL1R1 and IL-1α (p<0.05, vs. LPS-treated monocytes). To investigate whether the IL-1α translocation is mediated by protein myristoylation, human monocytes were pre-incubated with the n-myristoyl-transferase inhibitor IMP-1088 before LPS stimulation, resulting in a reduction of csIL-1α expression (p<0.05, vs. LPS-treated monocytes). Conclusion This study demonstrated a protective effect of Il1a deficiency on the development of atherosclerosis. CsIL-1α mediates monocyte-to-endothelial adhesion by increasing VCAM1 expression through endothelial IL1R1 signaling. The data underscore the importance of the juxtacrine signaling of IL-1α and its role in the development of atherosclerosis independent of circulating cytokine levels.

Chemical Profiling of Nitraria roborowskii Kom. by UPLC-Q-Orbitrap-MS and Their Hypolipidemic Effects in Vivo.

The Nitraria roborowskii Kom. (NRK) berries, as fruits of the genus Nitraria of the Zygophyllceae family, have been widely used as folk medicine. Modern pharmacological research has demonstrated that Nitraria berries had hypolipidemic, hypoglycemic, and immunomodulatory effects. However, more research needs to be reported on the chemical composition and biological activity of NRK. Hence, the phenolic compounds in the NRK berries were comprehensively analyzed and characterized by Ultra Performance Liquid Chromatography-Quadruple-Orbitrap MS system (UPLC-Q-Orbitrap MS) in this study. In total, 52 phenolics were identified, and all were reported for the first time. In addition, the hypolipidemic efficacy of NRK berries extract was studied in the hyperlipidemic mouse model. After treatment, the high dose group of NRK substantially reversed total cholesterol, low-density lipoprotein cholesterol, and high-density lipoprotein cholesterol levels. Through lipidomics technology, 27 potential biomarkers were characterized. And there was a significant callback at 25 of them after NRK treatment by using statistical analysis methods. Pathway analysis results demonstrated that NRK might exert therapeutic effects by regulating glycerophospholipid and glycerolipid metabolism pathways. This study could provide firsthand information on NRK berries for their phenolic compounds and potential application in preventing and treating hyperlipidemia.

Mechanisms of weight-loss effect in obese mice by the endogenous cannabinoid receptor 2 agonist beta-caryophyllene

Background and aimsThe endogenous cannabinoid system (ECS) is involved in the regulation of a variety of physiological activities in the body, such as metabolism and energy uptake, and cannabinoid receptor 2 (CNR2) is one of these receptors that is predominantly distributed in the periphery. β-caryophyllene (BCP) is an agonist of CNR2 which is known to possess pharmacological activities such as anti-inflammatory and antioxidant properties. In this study, we wanted to investigate whether BCP possesses pharmacological effects on obese mice and its mechanism. MethodsReversed feeding rhythm, propylthiouracil was delivered intraperitoneally, and BCP was gavaged once daily for four weeks to establish a hyperlipidemic obese mouse model. A glucose tolerance test, lipid level measurements, liver, peritoneal, and subcutaneous fat removal, HE and Oil Red O staining of the liver, and immunohistochemistry (IHC) staining with an anti-CNR2 antibody were all carried out. The liver was examined using tools like GO and KEGG databases for differentially expressed genes and signaling pathways linked to medication effectiveness. ResultsBCP had significant effects on weight reduction and improvement of dyslipidemia. What’s more, it significantly reduced body fat percentage, improved steatosis and ballooning of liver cells, and reduced fat accumulation, while inhibiting the proliferation of peri-abdominal adipocytes. BCP exerted its effects to improve dyslipidemia and reduce body weight probably through circadian regulation and cholesterol metabolic pathways. Finally, and its efficacy in improving dyslipidemia and reducing body weight may be mainly through activating CNR2, activating SIRT1/PGC-1α/PPARγ and SIRT1/AMPK pathways. ConclusionBCP activates the CNR2, SIRT1/PGC-1α/PPARγ signaling pathway, and SIRT1/AMPK signaling pathway to exert dyslipidemia-improving and weight-loss effects.

Human circulating CD24hi marginal zone B cells produce IgM targeting atherogenic antigens and confer protection from vascular disease.

IgMs that inactivate oxidation-specific epitopes (IgMOSE), which are secondary products of lipid peroxidization, protect against inflammatory diseases, including diet-induced atherosclerosis. However, the human B cell subtype that produces IgMOSE remains unknown. In this study, we used single-cell mass cytometry and adoptive transfer of B cell subtypes to NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ (NSG) mice to identify B27+IgM+CD24hi cells as the major producers of IgMOSE in humans. Notably, these cells have characteristics of human circulatory marginal zone B (MZB) cells, which are known to be atheoroprotective IgM producers in mice. CD24 antibody treatment to reduce MZB cells and IgM in a hyperlipidemic humanized mouse model provides the evidence that MZB cells protect against vascular inflammation. Consistent with these findings, the frequency of B27+IgM+CD24hi cells (MZB) in patients inversely correlates with coronary artery disease severity.

Anti-Hyperlipidemic Effect of Fucoidan Fractions Prepared from Iceland Brown Algae Ascophyllum nodosum in an Hyperlipidemic Mice Model.

Ascophyllum nodosum, a brown algae abundantly found along the North Atlantic coast, is recognized for its high polysaccharide content. In this study, we investigated the anti-hyperlipidemic effect of fucoidans derived from A. nodosum, aiming to provide information for their potential application in anti-hyperlipidemic therapies and to explore comprehensive utilization of this Iceland brown seaweed. The crude fucoidan prepared from A. nodosum was separated using a diethylethanolamine column, resulting in two fucoidan fractions, AFC-1 and AFC-2. Both fractions were predominantly composed of fucose and xylose. AFC-1 exhibited a higher sulfate content of 27.8% compared to AFC-2 with 17.0%. AFC-2 was primarily sulfated at the hydroxy group of C2, whereas AFC-1 was sulfated at both the hydroxy groups of C2 and C4. To evaluate the anti-hyperlipidemic effect, a hyperlipidemia mouse model was established by feeding mice a high-fat diet. The effects of AFC-1, AFC-2, and the crude extract were investigated, with the drug atorvastatin used as a positive comparison. Among the different fucoidan fractions and doses, the high dose of AFC-2 administration demonstrated the most significant anti-hyperlipidemic effect across various aspects, including physiological parameters, blood glucose levels, lipid profile, histological analysis, and the activities of oxidative stress-related enzymes and lipoprotein-metabolism-related enzymes (p < 0.05 for the final body weight and p < 0.01 for the rest indicators, compared with the model group), and its effect is comparable to the atorvastatin administration. Furthermore, fucoidan administration resulted in a lower degree of loss in gut flora diversity compared to atorvastatin administration. These findings highlight the significant biomedical potential of fucoidans derived from A. nodosum as a promising therapeutic solution for hypolipidemia.