Oxidized low-density lipoproteins impair the pro-atherosclerotic effect of granulocyte-macrophage-colony-stimulating factor-producing T helper cells on macrophages.

T cells contribute to the pathogenesis of atherosclerosis. However, the presence and function of granulocyte-macrophage-colony-stimulating factor (GM-CSF)-producing T helper (ThGM) cells in atherosclerosis development is unknown. This study aims to characterize the phenotype and function of ThGM cells in experimental atherosclerosis. Atherosclerosis was induced by feeding apolipoprotein E knockout (ApoE-/-) mice with a high-fat diet. Aortic ThGM cells were detected and sorted by flow cytometry. The effect of oxidized low-density lipoprotein (oxLDL) on ThGM cells and the impact of ThGM cells on macrophages were evaluated by flow cytometry, quantitative RT-PCR, oxLDL binding/uptake assay, immunoblotting and foam cell formation assay. We found that GM-CSF+IFN-γ- ThGM cells existed in atherosclerotic aortas. Live ThGM cells were enriched in aortic CD4+CCR6-CCR8-CXCR3-CCR10+ T cells. Aortic ThGM cells triggered the expression of interleukin-1ß (IL-1ß), tumour necrosis factor (TNF), interleukin-6 (IL-6) and C-C motif chemokine ligand 2 (CCL2) in macrophages. Besides, aortic ThGM cells expressed higher CD69 than other T cells and bound to oxLDL. oxLDL suppressed the cytokine expression in ThGM cells probably via inhibiting the signal transducer and activator of transcription 5 (STAT5) signalling. Furthermore, oxLDL alleviated the effect of ThGM cells on inducing macrophages to produce pro-inflammatory cytokines and generate foam cells. The nuclear receptor subfamily 4 group A (NR4A) members NR4A1 and NR4A2 were involved in the suppressive effect of oxLDL on ThGM cells. Collectively, oxLDL suppressed the supportive effect of ThGM cells on pro-atherosclerotic macrophages.

Effects of portable pedal machines at work on lipoprotein subfraction profile in sedentary workers - the REMOVE study.

BACKGROUND: Sedentary behaviour at work is a major cause of atherosclerosis, particularly in tertiary workers. However, no studies have ever assessed the effect of active workstation on lipoprotein subfraction profile. This study aimed to evaluate the effect of 12-week portable pedal machines (PPMs) on lipoprotein subfraction profile among healthy sedentary workers. METHODS: Healthy administrative workers were randomized into an intervention group using PPMs for 12 weeks or a control group using normal-desk. Lipoprotein subfractions were assessed using Lipoprint® electrophoresis. Main outcomes were explored using mixed models with sensitivity analyses (four models). RESULTS: We included 40 participants (43.7 ± 8.6 years old, 100% women, BMI 23.8 ± 3.4 kg/m2; sedentary time at work 7.7 ± 1.8 h/day). Groups did not differ at baseline in any outcomes. 32 participants finished the trial. Changes in lipoprotein subfractions were especially marked for LDL profile. There was an interaction time x group for all parameters related to LDL and their subfractions: total LDL-cholesterol (p = 0.012), LDL particle size (p = 0.027), large LDL subfractions 1 and 2 (p = 0.001), and small dense LDL subfractions 3 to 7 (p = 0.046), using the crude model. The interaction reflects difference in the direction of changes between groups. The LDL particle size significantly increased in the intervention group (from 271.9 ± 2.5 at t0 to 272.8 ± 1.9 Ångström at t1, p = 0.037) while it did not change in the control group (272.5 ± 1.7 at t0 to 271.8 ± 1.5Å at t1, p = 0.52). All interactions were constantly significant whatever the models. Influencing variables were mainly stress at work that was associated with an increase in total LDL-cholesterol (coefficient 3.15, 95CI 0.20 to 6.11 mg/dl, p = 0.038), and BMI that was associated with Large-LDL, Large-HDL, IDL-C and triglycerides. CONCLUSIONS: Lipoprotein profile was improved after a 12-week PPMs intervention at work in healthy administrative workers. Changes were mainly showed for LDL and LDL subfractions. Lipoprotein profile was worsened by stress at work, BMI and age. TRIAL REGISTRATION: NCT04153214.

Targeting PCSK9 to upregulate MHC-II on the surface of tumor cells in tumor immunotherapy.

BACKGROUND: Proprotein convertase subtilisin/kexin type 9 (PCSK9), the last member of the proprotein convertase family, functions as a classic regulator of low-density lipoprotein (LDL) by interacting with low-density lipoprotein receptor (LDLR). Recent studies have shown that PCSK9 can affect the occurrence and development of tumors and can be used as a novel therapeutic target. However, a comprehensive pan-cancer analysis of PCSK9 has yet to be conducted. METHODS: The potential oncogenic effects of PCSK9 in 33 types of tumors were explored based on the datasets of The Cancer Genome Atlas (TCGA) dataset. In addition, the immune regulatory role of PCSK9 inhibition was evaluated via in vitro cell coculture and the tumor-bearing mouse model. Finally, the antitumor efficacy of targeted PCSK9 combined with OVA-II vaccines was verified. RESULTS: Our results indicated that PCSK9 was highly expressed in most tumor types and was significantly correlated with late disease stage and poor prognosis. Additionally, PCSK9 may regulate the tumor immune matrix score, immune cell infiltration, immune checkpoint expression, and major histocompatibility complex expression. Notably, we first found that dendritic cell (DC) infiltration and major histocompatibility complex-II (MHC-II) expression could be upregulated by PCSK9 inhibition and improve CD8+ T cell activation in the tumor immune microenvironment, thereby achieving potent tumor control. Combining PCSK9 inhibitors could enhance the efficacies of OVA-II tumor vaccine monotherapy. CONCLUSIONS: Conclusively, our pan-cancer analysis provided a more comprehensive understanding of the oncogenic and immunoregulatory roles of PCSK9 and demonstrated that targeting PCSK9 could increase the efficacy of long peptide vaccines by upregulating DC infiltration and MHC-II expression on the surface of tumor cells. This study reveals the critical oncogenic and immunoregulatory roles of PCSK9 in various tumors and shows the promise of PCSK9 as a potent immunotherapy target.

Cardiovascular risk and inflammation in a population with autoimmune diseases: a narrative review.

Juvenile Systemic Connective Tissue Diseases (JSCTD) are a heterogeneous group of chronic autoimmune diseases, associated with dyslipidemia and increased cardiovascular risk are related. Studies from the last 10 years, from 2013 to 2022, on lipid profiles in JSCTD were collected. Different studies on lipid profiles in children affected by JSCTD were selected, because the aim is to analyze the cardiovascular risk and the possibility of atherosclerosis in these patients in whom, sometimes, corticosteroid therapies and immunosuppressants increase the state of dyslipidemia. Several studies have shown that autoimmune diseases with an inflammatory substrate also share abnormalities in lipid profile and increased cardiovascular risk. Specifically, associations have been found between Juvenile Systemic Connective Tissue Diseases and elevated triglycerides, TC-C (Total Cholesterol), LDL-C (Low-Density Lipoprotein), low HDL-C (High-Density Lipoprotein), and increased risk of developing diseases such as myocardial infarction, peripheral vascular disease, pulmonary and arterial hypertension, and atrial fibrillation. Supplementation with alpha-linolenic acid (ALA) on the other hand has also been analyzed with positive results in reducing inflammatory parameters, such as IL-6 (Interleukin-6), CRP (C-reactive protein), and fasting glucose, in subjects with dyslipidemia. These observations suggest that supplementation with ALA, an omega-3 precursor, may positively modulate both the inflammatory status and dyslipidemic conditions in patients with autoimmune disorders.

A high-cholesterol diet promotes the intravasation of breast tumor cells through an LDL-LDLR axis.

Most metastases in breast cancer occur via the dissemination of tumor cells through the bloodstream. How tumor cells enter the blood (intravasation) is, however, a poorly understood mechanism at the cellular and molecular levels. Particularly uncharacterized is how intravasation is affected by systemic nutrients. High levels of systemic LDL-cholesterol have been shown to contribute to breast cancer progression and metastasis in various models, but the cellular and molecular mechanisms involved are still undisclosed. Here we show that a high- cholesterol diet promotes intravasation in two mouse models of breast cancer and that this could be reverted by blocking LDL binding to LDLR in tumor cells. Moreover, we show that LDL promotes vascular invasion in vitro and the intercalation of tumor cells with endothelial cells, a phenotypic change resembling vascular mimicry (VM). At the molecular level, LDL increases the expression of SERPINE2, previously shown to be required for both VM and intravasation. Overall, our manuscript unravels novel mechanisms by which systemic hypercholesterolemia may affect the onset of metastatic breast cancer by favouring phenotypic changes in breast cancer cells and increasing intravasation.

Knockdown of LINC01128 Downregulates FUT8 to Inhibit OxLDL-Induced Vascular Smooth Muscle Cell Apoptosis in Atherosclerosis.

BACKGROUND: The apoptosis of vascular smooth muscle cells (VSMCs) contributes to the progression of atherosclerosis (AS). Long intergenic non-protein coding RNA 1128 (LINC01128) has been implicated in AS, and this study aims to uncover the role and mechanism of LINC01128 in regulating oxidized low-density lipoprotein (oxLDL)-induced VSMCs. METHODS: The position of LINC01128 in cells and its target genes were predicted using bioinformatics. The localization of LINC01128 in human VSMCs was determined through fluorescence in situ hybridization. VSMCs were transfected, and the interaction between LINC01128 and fucosyltransferase 8 (FUT8) was validated by chromatin immunoprecipitation assay. The apoptotic VSMC model was established using oxLDL. LINC01128 expression in VSMCs was analyzed by quantitative real-time polymerase chain reaction (qRT-PCR), and FUT8 expression was detected by qRT-PCR and western blot. VSMC viability, migration, invasion abilities, and apoptosis were assessed using cell counting kit-8, transwell assay, and flow cytometry, respectively. RESULTS: OxLDL (200 µg/mL) upregulated the expression of LINC01128 and FUT8 mRNA, as well as FUT8 protein, in VSMCs. LINC01128 was expressed in the nucleus of VSMCs and bound to FUT8. Knockdown of LINC01128 alleviated the inhibitory effects of oxLDL (200 µg/mL) on viability, migration, and invasion, and mitigated the promotion of apoptosis and FUT8 expression in VSMCs. On the other hand, FUT8 overexpression enhanced the suppressive effects of oxLDL (200 µg/mL) on viability, migration, and invasion activities, and amplified the facilitating effect of oxLDL on apoptosis in VSMCs. Moreover, FUT8 overexpression reversed the impact of LINC01128 silencing on viability, migration, invasion, and apoptosis in oxLDL-stimulated VSMCs. CONCLUSION: The knockdown of LINC01128 downregulates FUT8, inhibiting the progression of VSMCs in AS.

Avocado-derived extracellular vesicles loaded with ginkgetin and berberine prevent inflammation and macrophage foam cell formation.

Atherosclerosis, a chronic inflammatory disease of aorta, remains the major cause of morbidity and mortality among cardiovascular disease patients. Macrophage foam cell formation and inflammation are critically involved in early stages of atherosclerosis, hence chemopreventive targeting of foam cell formation by nutraceuticals may be a promising approach to curbing the progression of atherosclerosis. However, many nutraceuticals including berberine and ginkgetin have low stability, tissue/cell penetration and bioavailability resulting in inadequate chemotherapeutic effects of these nutraceuticals. We have used avocado-derived extracellular vesicles (EV) isolated from avocado (EVAvo ) as a novel carrier of nutraceuticals, in a strategy to alleviate the build-up of macrophage foam cells and expression of inflammatory genes. Our key findings are: (i) Avocado is a natural source of plant-derived EVs as shown by the results from transmission electron microscopy, dynamic light scattering and NanoBrook Omni analysis and atomic force microscopy; (ii) EVAvo are taken up by macrophages, a critical cell type in atherosclerosis; (iii) EVAvo can be loaded with high amounts of ginkgetin and berberine; (iv) ginkgetin plus berberine-loaded EVAvo (EVAvo(B+G) ) suppress activation of NFκB and NLRP3, and inhibit expression of pro-inflammatory and atherogenic genes, specifically Cd36, Tnfα, Il1ß and Il6; (v) EVAvo(B+G) attenuate oxidized low-density lipoprotein (oxLDL)-induced macrophage foam cell formation and (vi) EVAvo(B+G) inhibit oxLDL uptake but not its cell surface binding during foam cell formation. Overall, our results suggest that using EVAvo as a natural carrier of nutraceuticals may improve strategies to curb the progression of atherosclerosis by limiting inflammation and pro-atherogenic responses.

Low-density lipoprotein receptor-related protein 5/6 promotes endometrial cancer progression and cancer cell immune escape.

The study investigated the potential association of the low-density lipoprotein (LDL) genome with endometrial cancer progression based on the Gene Expression Omnibus data set and The Cancer Genome Atlas data set. Differential and weighted gene coexpression network analysis was performed on endometrial cancer transcriptome datasets GSE9750 and GSE106191. The protein-protein interaction network was built using LDL-receptor proteins and the top 50 tumor-associated genes. Low-density lipoprotein-related receptors 5/6 (LRP5/6) in endometrial cancer tissues were correlated with oncogenes, cell cycle-related genes, and immunological checkpoints using Spearman correlation. MethPrimer predicted the LRP5/6 promoter CpG island. LRP2, LRP6, LRP8, LRP12, low-density lipoprotein receptor-related protein-associated protein, and LRP5 were major LDL-receptor-related genes associated with endometrial cancer. LRP5/6 was enriched in various cancer-related pathways and may be a key LDL-receptor-related gene in cancer progression. LRP5/6 may be involved in the proliferation process of endometrial cancer cells by promoting the expression of cell cycle-related genes. LRP5/6 may be involved in the proliferation of endometrial cancer cells by promoting the expression of cell cycle-related genes. LRP5/6 may promote the immune escape of cancer cells by promoting the expression of immune checkpoints, promoting endometrial cancer progression. The MethPrimer database predicted that the LRP5/6 promoter region contained many CpG islands, suggesting that DNA methylation can occur in the LRP5/6 promoter region. LRP5/6 may aggravate endometrial cancer by activating the phosphoinositide 3-kinase/protein kinase B pathway.

Prior Exposure to Experimental Preeclampsia Increases Atherosclerotic Plaque Inflammation in Atherogenic Mice-Brief Report.

BACKGROUND: Women with a history of preeclampsia have evidence of premature atherosclerosis and increased risk of myocardial infarction and stroke compared with women who had a normotensive pregnancy. Whether this is due to common risk factors or a direct impact of prior preeclampsia exposure has never been tested in a mouse atherosclerosis model. METHODS: Pregnant LDLR-KO (low-density lipoprotein receptor knockout; n=35) female mice were randomized in midgestation to sFlt1 (soluble fms-like tyrosine kinase 1)-expressing adenovirus or identical control adenovirus. Postpartum, mice were fed high-fat diet for 8 weeks to induce atherogenesis. Comparison between the control and preeclampsia models was made for metabolic parameters, atherosclerosis burden and composition by histology, plaque inflammation by flow cytometry, and aortic cytokines and inflammatory markers using a cytokine array. RESULTS: In pregnant LDLR-KO mice, sFlt1 adenovirus significantly induced serum sFlt1, blood pressure, renal endotheliosis, and decreased pup viability. After 8 weeks of postpartum high fat feeding, body weight, fasting glucose, plasma cholesterol, HDL (high-density lipoprotein), and LDL (low-density lipoprotein) were not significantly different between groups with no change in aortic root plaque size, lipid content, or necrotic core area. Flow cytometry demonstrated significantly increased CD45+ aortic arch leukocytes and CD3+T cells and aortic lysate contained more CCL (CC motif chemokine ligand) 22 and fetuin A and decreased expression of IGFBP6 (insulin-like growth factor-binding protein 6) and CCL21 in preeclampsia-exposed mice compared with controls. CONCLUSIONS: In atherogenic LDLR-KO mice, exposure to sFlt1-induced preeclampsia during pregnancy increases future atherosclerotic plaque inflammation, supporting the concept that preeclampsia directly exacerbates atherosclerotic inflammation independent of preexisting risk factors. This mechanism may contribute to ischemic vascular disease in women after preeclampsia pregnancy.

Circ-ABCA1 promotes oxidized low-density lipoprotein-induced inflammation and phenotypic switch in vascular smooth muscle cells.

OBJECTIVE: Atherosclerosis (AS) is a chronic inflammatory disease of the arterial wall, in which Human Vascular Smooth Muscle Cells (HVSMCs) are involved. Nevertheless, the functions and mechanisms of circRNAs in oxidized Low-Density Lipoprotein (ox-LDL)-induced vascular smooth muscle cells remain unclear. METHODS: Circ-ABCA1 expression was measured in the models of AS. Then, in the vitro model, oligonucleotide transfection was performed, followed by an analysis of VSMC proliferation, migration, inflammation, and phenotypic switch. Also, in the in vivo model, mice were injected with shRNA lentivirus, followed by histological examination of aortic tissues. Finally, the interaction of circ-ABCA1, miR-885-5p, and ROCK2 was identified. RESULTS: Circ-ABCA1, was confirmed to be overexpressed in ox-LDL-induced VSMCs and mouse models of AS. Functionally, silencing circ-ABCA1 via oligonucleotide transfection suppressed VSMC proliferation, migration, inflammation, and phenotypic switch in vitro and prevented AS development in mice in vivo. Mechanistically, circ-ABCA1 absorbed miR-885-5p, which targeted ROCK2. CONCLUSION: Taken together, the data from this study suggest that circ-ABCA1 mediates cellular inflammation and phenotype switching through the miR-885-5p/ROCK2 axis in ox-LDL-induced VSMCs, and the circ-ABCA1/miR-885-5p/ROCK2 axis is a new potential biomarker for the treatment of AS.

The long coiled-coil protein NECC2 regulates oxLDL-induced endothelial oxidative damage and exacerbates atherosclerosis development in apolipoprotein E -/- mice.

Oxidized low density lipoprotein (oxLDL)-induced endothelial oxidative damage promotes the development of atherosclerosis. Caveolae play an essential role in maintaining the survival and function of vascular endothelial cell (VEC). It is reported that the long coiled-coil protein NECC2 is localized in caveolae and is associated with neural cell differentiation and adipocyte formation, but its role in VECs needs to be clarified. Our results showed NECC2 expression increased in the endothelium of plaque-loaded aortas and oxLDL-treated HUVECs. Down-regulation of NECC2 by NECC2 siRNA or compound YF-307 significantly inhibited oxLDL-induced VEC apoptosis and the adhesion factors expression. Remarkably, inhibition of NECC2 expression in the endothelium of apoE-/- mice by adeno-associated virus (AAV)-carrying NECC2 shRNA or compound YF-307 alleviated endothelium injury and restricted atherosclerosis development. The immunoprecipitation results confirmed that NECC2 interacted with Tyk2 and caveolin-1(Cav-1) in VECs, and NECC2 further promoted the phosphorylation of Cav-1 at Tyr14 b y activating Tyk2 phosphorylation. On the other hand, inhibiting NECC2 levels suppressed oxLDL-induced phosphorylation of Cav-1, uptake of oxLDL by VECs, accumulation of intracellular reactive oxygen species and activation of NF-κB. Our findings suggest that NECC2 may contribute to oxLDL-induced VEC injury and atherosclerosis via modulating Cav-1 phosphorylation through Tyk2. This work provides a new concept and drug target for treating atherosclerosis.

Sinomenine protects against atherosclerosis in apolipoprotein E-knockout mice by inhibiting of inflammatory pathway.

Atherosclerosis, a multifaceted and persistent inflammatory condition, significantly contributes to the progression of cardiocerebrovascular disorders, such as myocardial infarctions and cerebrovascular accidents. It involves the accumulation of cholesterol, fatty deposits, calcium and cellular debris in the walls of arteries, leading to the formation of plaques. Our aim is to investigate the potential of sinomenine to counteract atherosclerosis in mice lacking Apolipoprotein E (ApoE-/-) Mice. We employed the high-fat diet-induced method to induce atherosclerosis in ApoE-/- mice, and the mice were treated with sinomenine (5, 10, and 15 mg/kg) and simvastatin (0.5 mg/kg) for 12 weeks. Body weight, water intake, and food intake were assessed. Lipid parameters, oxidative stress, inflammatory cytokines, and mRNA levels were estimated. Sinomenine treatment remarkably (P < 0.001) suppressed body weight, along with food and water intake. Sinomenine altered the levels of total cholesterol (TC), high-density lipoprotein (HDL), triglyceride (TG), low-density lipoprotein (LDL), and very low-density lipoprotein (VLDL), which were modulated in the atherosclerosis group. Sinomenine treatment also altered the levels of oxidative stress parameters such as glutathione peroxidase (GPx), catalase (CAT), malonaldehyde (MDA), superoxide dismutase (SOD) and glutathione (GSH). In addition, it modulated cardiac parameters like C-reactive protein (CRP), endothelin-1 (ET-1), thromboxane B2 (TXB2), nitric oxide (NO), cardiac troponin I (cTnI), lactate dehydrogenase (LDH), and creatinine kinase isoenzymes (CK-MB). Inflammatory cytokines interleukin (IL)-1α, IL-1ß, TNF-α, IL-6, and IL-10 were also affected. Sinomenine further suppressed the mRNA expression of IL-6, IL-17, IL-10, tumor necrosis factor-α (TNF-α), Il-1ß, monocyte chemoattractant protein-1 (MCP-1), MCP-2, MCP-3, transforming Growth Factor-1ß (TGF-1ß), vascular cell adhesion molecule 1 (VCAM-1), and intercellular adhesion molecule-1 (ICAM-1). The results suggest that sinomenine remarkably suppressed the development of atherosclerosis in the early stage.

Hawthorn leaf flavonoids alleviate the deterioration of atherosclerosis by inhibiting SCAP-SREBP2-LDLR pathway through sPLA2-â ¡A signaling in macrophages in mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Hawthorn leaves are a combination of the dried leaves of the Rosaceae plants, i.e., Crataegus pinnatifida Bge. or Crataegus pinnatifida Bge. var. major N. E. Br., is primarily cultivated in East Asia, North America, and Europe. hawthorn leaf flavonoids (HLF) are the main part of extraction. The HLF have demonstrated potential in preventing hypertension, inflammation, hyperlipidemia, and atherosclerosis. However, the potential pharmacological mechanism behind its anti-atherosclerotic effect has yet to be explored. AIM OF THE STUDY: The in vivo and in vitro effects of HLF on lipid-mediated foam cell formation were investigated, with a specific focus on the levels of secreted phospholipase A2 type IIA (sPLA2-II A) in macrophage cells. MATERIALS AND METHODS: The primary constituents of HLF were analyzed using ultra-high performance liquid chromatography and liquid chromatography-tandem mass spectrometry. In vivo, HLF, at concentrations of 5 mg/kg, 20 mg/kg, and 40 mg/kg, were administered to apolipoprotein E knockout mice (ApoE-/-) fed by high-fat diet (HFD) for 16 weeks. Aorta and serum samples were collected to identify lesion areas and lipids through mass spectrometry analysis to dissect the pathological process. RAW264.7 cells were incubated with oxidized low-density lipoprotein (ox-LDL) alone, or ox-LDL combined with different doses of HLF (100, 50, and 25 µg/ml), or ox-LDL plus 24-h sPLA2-IIA inhibitors, for cell biology analysis. Lipids and inflammatory cytokines were detected using biochemical analyzers and ELISA, while plaque size and collagen content of plaque were assessed by HE and the Masson staining of the aorta. The lipid deposition in macrophages was observed by Oil Red O staining. The expression of sPLA2-IIA and SCAP-SREBP2-LDLR was determined by RT-qPCR and Western blot analysis. RESULTS: The chemical profile of HLF was studied using UPLC-Q-TOF-MS/MS, allowing the tentative identification of 20 compounds, comprising 1 phenolic acid, 9 flavonols and 10 flavones, including isovitexin, vitexin-4″-O-glucoside, quercetin-3-O-robibioside, rutin, vitexin-2″-O-rhamnoside, quercetin, etc. HLF decreased total cholesterol (TC), triglycerides (TG), low-density lipoprotein cholesterol (LDL-C), and non-high-density lipoprotein cholesterol (non-HDL-C) levels in ApoE-/- mice (P < 0.05), reduced ox-LDL uptake, inhibited level of inflammatory factors, such as IL-6, IL-8, TNF-α, and IL-1ꞵ (P < 0.001), and alleviated aortic plaques with a thicker fibrous cap. HLF effectively attenuated foam cell formation in ox-LDL-treated RAW264.7 macrophages, and reduced levels of intracellular TC, free cholesterol (FC), cholesteryl ester (CE), IL-6, TNF-α, and IL-1ß (P < 0.001). In both in vivo and in vitro experiments, HLF significantly downregulated the expression of sPLA2-IIA, SCAP, SREBP2, LDLR, HMGCR, and LOX-1 (P < 0.05). Furthermore, sPLA2-IIA inhibitor effectively mitigated inflammatory release in RAW264.7 macrophages and regulated SCAP-SREBP2-LDLR signaling pathway by inhibiting sPLA2-IIA secretion (P < 0.05). CONCLUSION: HLF exerted a protective effect against atherosclerosis through inhibiting sPLA2-IIA to diminish SCAP-SREBP2-LDLR signaling pathway, to reduce LDL uptake caused foam cell formation, and to slow down the progression of atherosclerosis in mice.

Accurate and sensitive low-density lipoprotein (LDL) detection based on the proximity ligation assisted rolling circle amplification (RCA).

Metabolic-associated fatty liver disease (MAFLD) is one of the leading causes of mortality from chronic diseases worldwide, and it is strongly linked to dyslipidemia. Dyslipidemia typically presents as an elevated concentration of low density lipoprotein (LDL). Hence, accurate quantification of LDL particles is crucial for predicting the risks of cardiovascular illnesses. Nevertheless, conventional techniques can merely provide indirect measurements of LDL particle concentrations through the detection of cholesterol or proteins within LDL particles, and they often require significant effort and time. Therefore, an accurate and effective method for identifying intact LDL particles is highly desired. We have devised a method that allows for the measurement of LDL concentration without the need for isolation. This method relies on proximity ligation rolling circle amplification (RCA). This technique enables the direct and precise measurement of the concentration of "actual" LDL particles, rather than measuring the cholesterol content inside LDL. It has a detection limit of 7.3 µg dL-1, which also meets the requirements for analyzing lipoproteins in clinical samples. Hence, this platform exhibits immense potential in clinical applications and health management.

LDLR c.415G > A causes familial hypercholesterolemia by weakening LDLR binding to LDL.

BACKGROUND: Familial hypercholesterolemia (FH) is a prevalent hereditary disease that can cause aberrant cholesterol metabolism. In this study, we confirmed that c.415G > A in low-density lipoprotein receptor (LDLR), an FH-related gene, is a pathogenic variant in FH by in silico analysis and functional experiments. METHODS: The proband and his family were evaluated using the diagnostic criteria of the Dutch Lipid Clinic Network. Whole-exome and Sanger sequencing were used to explore and validate FH-related variants. In silico analyses were used to evaluate the pathogenicity of the candidate variant and its impact on protein stability. Molecular and biochemical methods were performed to examine the effects of the LDLR c.415G > A variant in vitro. RESULTS: Four of six participants had a diagnosis of FH. It was estimated that the LDLR c.415G > A variant in this family was likely pathogenic. Western blotting and qPCR suggested that LDLR c.415G > A does not affect protein expression. Functional studies showed that this variant may lead to dyslipidemia by impairing the binding and absorption of LDLR to low-density lipoprotein ( LDL). CONCLUSION: LDLR c.415G > A is a pathogenic variant in FH; it causes a significant reduction in LDLR's capacity to bind LDL, resulting in impaired LDL uptake. These findings expand the spectrum of variants associated with FH.

Porcine low-density lipoprotein receptor plays an important role in classical swine fever virus infection.

Several cellular factors have been reported to be required for replication of classical swine fever virus (CSFV), a member of the genus Pestivirus within the family Flaviviridae. However, many steps of its replication cycle are still poorly understood. The low-density lipoprotein receptor (LDLR) is involved in cell entry and post-entry processes of different viruses including other members of the Flaviviridae. In this study, the relevance of LDLR in replication of CSFV and another porcine pestivirus, Bungowannah pestivirus (BuPV), was investigated by antibody-mediated blocking of LDLR and genetically engineered porcine cell lines providing altered LDLR expression levels. An LDLR-specific antibody largely blocked infection with CSFV, but had only a minor impact on BuPV. Infections of the genetically modified cells confirmed an LDLR-dependent replication of CSFV. Compared to wild type cells, lower and higher expression of LDLR resulted in a 3.5-fold decrease or increase in viral titers already 20 h post infection. Viral titers were 25-fold increased in LDLR-overexpressing cells compared to cells with reduced LDLR expression at 72 h post infection. The varying LDLR expression levels had no clear effect on permissivity to BuPV. A decoy receptor assay using recombinant soluble LDLR provided no evidence that LDLR may function as a receptor for CSFV or BuPV. Differences in their dependency on LDLR suggest that CSFV and BuPV likely use different mechanisms to interact with their host cells. Moreover, this study reveals similarities in the replication cycles of CSFV and other members of the family Flaviviridae that are dependent on LDLR.

Feather meal processing methods impact the production parameters, blood biochemical indices, gut function, and hepatic enzyme activity in broilers.

This study investigated the effects of feather meal (FM) processing methods on production parameters, blood biochemical indices, intestinal morphology, digestive and hepatic enzyme activities, and gastrointestinal tract pH and microflora of broilers. A total of 480-d-old male broilers were used for 42 d in a completely randomized design with eight treatments and five replicates (12 chicks/replicate). Treatments were 1) a control diet (without FM), 2) a diet containing 4% raw FM (RFM), 3) a diet containing 4% processed FM (PFM) by autoclave (Au-PFM), 4) a diet containing 4% fermented FM (FFM) by Bacillus licheniformis (Bl-FFM), 5) a diet containing 4% FFM by Bacillus subtilis (Bs-FFM), 6) a diet containing 4% FFM by Aspergillus niger (An-FFM), 7) a diet containing 4% FFM by B. licheniformis + B. subtilis + A. niger (Co-FFM), and 8) a diet containing 4% PFM by an enzyme (En-PFM). Results showed that in the FFMs the contents of ash, ether extract, total volatile nitrogen, and amino acids including Lys, Met, Thr, Trp, His, Leu, Gly, Ile, Phe, and Tyr increased (P < 0.05), while crude fiber, crude protein, and dry matter content decreased (P < 0.05). Compared with the control, the Co-FFM diet had no significant differences (P > 0.05) in total body weight gain (2,827 vs. 2,791 g/chick), total feed intake (5,018 vs. 4,991 g/chick), European production efficiency factor (375 vs. 377), European Broiler Index (371 vs. 371), and feed conversion ratio (1.77 vs. 1.78 g/g). Feeding FFM decreased (P < 0.05) serum total cholesterol (1.46-fold), triglyceride (1.61-fold), very low-density lipoprotein cholesterol (1.61-fold), and low-density lipoprotein cholesterol (2.27-fold) compared to the control. Also, FFM increased (P < 0.05) villus height (1,045 to 1,351, 661 to 854, and 523 to 620 µm), and villus height to crypt depth ratio (6.15 to 8.45, 4.55 to 7.04, and 4.27 to 5.45), in the duodenum, jejunum, and ileum, respectively, compared to the control. Compared to the control, the Co-FFM diet increased (P < 0.05) protease (34, 39, and 45 %) in the pancreas, duodenum, and jejunum, as well as amylase (73, and 97 %) activities in the duodenum, and jejunum, respectively. Diets containing FFM reduced (P < 0.05) pH in the crop, gizzard, and ileum, and decreased (P < 0.05) Escherichia coli (6.12 to 5.70) count in ileum compared to the control. The Co-FFM diet increased (P < 0.05) lactic acid bacteria count in crop (6.77 to 7.50) and ileum (6.94 to 7.73), also decreased (P < 0.05) coliforms (6.31 to 5.75) count in ileum compared to the control. In conclusion, FM fermentation, particularly Co-FFM, improves the nutritional value of FM, converting it into a decent source of dietary protein for broilers.

Fermentation represents an attractive alternative method for feather meal (FM) efficient bioconversion and its nutritional value enhancement. This study investigated the effects of FM processing methods on broilers. Experimental diets were 1) a control diet (without FM), 2) a diet containing 4% raw FM (RFM), 3) a diet containing 4% processed FM (PFM) by autoclave (Au-PFM), 4) a diet containing 4% fermented FM (FFM) by Bacillus licheniformis (Bl-FFM), 5) a diet containing 4% FFM by Bacillus subtilis (Bs-FFM), 6) a diet containing 4% FFM by Aspergillus niger (An-FFM), 7) a diet containing 4% FFM by B. licheniformis + B. subtilis + A. niger (Co-FFM), and 8) a diet containing 4% PFM by an enzyme (En-PFM). Results showed that FFMs increased the contents of ash, ether extract, total volatile nitrogen, and amino acids including Lys, Met, Thr, Trp, His, Leu, Gly, Ile, Phe, and Tyr, while decreased crude fiber, crude protein, and dry matter content. The production parameters of birds fed Co-FFM were similar to the control group. In addition, FFMs decreased serum total cholesterol (1.46-fold), triglyceride (1.61-fold), very low-density lipoprotein cholesterol (1.61-fold), and low-density lipoprotein cholesterol (2.27-fold). Furthermore, Co-FFM improved intestinal morphology, enzyme activities, and beneficial bacterial populations. In conclusion, Co-FFM, improves the nutritional value of FM, converting it into a decent source of dietary protein for broilers.

Oxidized-LDL Induces Metabolic Dysfunction in Retinal Pigment Epithelial Cells.

Recently, mitochondrial dysfunction has gained attention as a causative factor in the pathogenesis and progression of age-related macular degeneration (AMD). Mitochondrial damage plays a key role in metabolism and disrupts the balance of intracellular metabolic pathways, such as oxidative phosphorylation (OXPHOS) and glycolysis. In this study, we focused on oxidized low-density lipoprotein (ox-LDL), a major constituent of drusen that accumulates in the retina of patients with AMD, and investigated whether it could be a causative factor for metabolic alterations in retinal pigment epithelial (RPE) cells. We found that prolonged exposure to ox-LDL induced changes in fatty acid ß-oxidation (FAO), OXPHOS, and glycolytic activity and increased the mitochondrial reactive oxygen species production in RPE cells. Notably, the effects on metabolic alterations varied with the concentration and duration of ox-LDL treatment. In addition, we addressed the limitations of using ARPE-19 cells for retinal disease research by highlighting their lower barrier function and FAO activity compared to those of induced pluripotent stem cell-derived RPE cells. Our findings can aid in the elucidation of mechanisms underlying the metabolic alterations in AMD.

The association between inflammation, arterial stiffness, oxidized LDL and cardiovascular disease in Finnish men with metabolic syndrome - a 15-year follow-up study.

BACKGROUND: All-cause mortality and cardiovascular disease are increased in subjects with metabolic syndrome (MetS). Risk scores are used to predict individual risk of heart disease. We performed a long-term follow-up study to investigate whether risk scores and cardiovascular risk factors such as arterial stiffness, high-sensitive C-reactive protein (hs-CRP) and oxidized LDL (OxLDL) can be used to predict cardiovascular events in Finnish men with MetS. METHODS: After baseline measurements we followed 105 Finnish men aged 30 to 65 years with MetS for a mean period of 16.4 years. The primary outcome of the study was a composite of myocardial infarction, stroke, symptomatic vascular disease diagnosed with invasive angiography, coronary or peripheral revascularization, amputation due to peripheral vascular disease, cardiovascular death and non-cardiovascular death. The endpoints were retrieved from electronic medical records. RESULTS: The number of acute myocardial infarctions and strokes during the first 10 years was lower than estimated by FINRISK score but SCORE predicted cardiovascular death correctly. During the whole follow-up period, 27 of 105 participants (25.8%) had 30 endpoint events. The incidence of the primary composite outcome was significantly lower in subjects with hs-CRP < 1.0 mg/L than in subjects with hs-CRP ≥ 1.0 mg/L (6 of 41 subjects [14.6%] vs. 21 of 64 subjects [32.8%]; p = 0.036). The incidence of the primary composite outcome was higher among subjects with large artery elasticity classified as borderline compared to subjects with normal large artery elasticity (5 of 10 subjects [50%] vs. 22 of 93 subjects [24%]; p = 0.05). There was no difference in the incidence of primary composite outcome in groups with different degrees of small artery elasticity or different level of oxLDL. CONCLUSIONS: Men with MetS who had hs-CRP ≥ 1.0 mg/L had higher risk for CVD and all-cause mortality than those with hs-CRP of < 1.0 mg/L. This also applies to subjects with borderline decreased large artery elasticity. The amount of OxLDL had no predictive value on the incidence of CVD and all-cause mortality. Men with MetS participating in the Hämeenlinna Metabolic Syndrome Research Program without lifestyle or drug intervention had better outcome for myocardial infarction or stroke than estimated by the FINRISK score. TRIAL REGISTRATION: ClinicalTrials.gov NCT01119404 retrospectively registered 07/05/2010.