The profile of cholesterol metabolism does not interfere with the cholesterol-lowering efficacy of phytostanol esters.

BACKGROUND & AIMS: Increasing evidence suggests that high cholesterol absorption efficiency enhances the risk of atherosclerotic cardiovascular diseases. It is not known whether inhibiting cholesterol absorption has different metabolic effects in high- vs. low cholesterol absorbers. We evaluated the effects of phytostanol esters on serum lipids and cholesterol metabolism in a post hoc study of three randomized, double-blind, controlled trials. The participants were classified into low (n = 20) and high (n = 21) cholesterol absorbers by median cholesterol absorption efficiency based on the plasma cholesterol absorption marker cholestanol at baseline. METHODS: The participants consumed mayonnaise or margarine without or with phytostanol esters for six to nine weeks without other changes in the diet or lifestyle. Serum cholesterol, cholestanol, lathosterol, and faecal neutral sterols and bile acids were analysed by gas-liquid chromatography. According to power calculations, the size of the study population (n = 41) was appropriate. RESULTS: During the control period, cholesterol synthesis, and faecal neutral sterols and bile acids were lower in high- vs. low absorbers (p < 0.05 for all). Phytostanol esters reduced low-density lipoprotein cholesterol by 10-13% in both groups, and directly measured cholesterol absorption efficiency by 41 ± 7% in low- and 47 ± 5% in high absorbers (p < 0.001 for all) without side effects. Cholesterol synthesis and faecal neutral sterols (p < 0.01) increased in both groups, more markedly in the high vs. low absorbers (p < 0.01). CONCLUSIONS: Low cholesterol absorption combined with high faecal neutral sterol excretion are components of reverse cholesterol transport. Thus, high- vs. low absorbers had a more disadvantageous metabolic profile at baseline. In both groups, phytostanol esters induced favourable changes in serum, lipoprotein, and metabolic variables known to help in prevention of the development of atherosclerotic cardiovascular diseases.

Impact of RYGB surgery on plasma immunoglobulins: association between blood pressure and glucose levels six months after surgery.

We aimed to study levels of natural antibodies in plasma, and their associations to clinical and fecal biomarkers, before and 6 months after Roux-en-Y gastric bypass (RYGB) surgery. Thirty individuals with obesity [16 type 2 diabetic, 14 non-diabetic (ND)] had RYGB surgery. Total plasma IgA, IgG and IgM antibody levels and specific antibodies to oxidized low-density lipoprotein (oxLDL), malondialdehyde-acetaldehyde adducts, Porphyromonas gingivalis gingipain A hemagglutinin domain (Rgp44), and phosphocholine were measured using chemiluminescence immunoassay. Associations between plasma and fecal antibodies as well as clinical markers were analyzed. RYGB surgery reduced blood pressure, and the glycemic state was improved. A higher level of diastolic blood pressure was associated with lower plasma antibodies to oxLDL after surgery. Also, lower level of glucose markers associated with lower level of plasma antibodies to bacterial virulence factors. Antibodies to oxLDL decreased after surgery, and positive association between active serum lipopolysaccharide and specific oxLDL antibodies was detected. Total IgG levels decreased after surgery, but only in ND individuals. Reduced level of total plasma IgG, improved state of hypertension and hyperglycemia and their associations with decreased levels of specific antibodies in plasma, suggest an improved state of systemic inflammation after RYGB surgery.

Immunoliposomes for detection of rupture-prone intracranial aneurysms.

BACKGROUND: It is estimated that significant (3.2%) of population carries intracranial aneurysm (IA). An increasing number of imaging studies have caused that the chance of finding an incidental aneurysm is becoming more common. Since IA rupture causes subarachnoidal hemorrhage (SAH) and have significant mortality and morbidity prophylactic treatment should be considered when IA is detected. The benefit and risk of treatment of IA is based on epidemiological estimate which takes account patient and aneurysm characteristics. However we know that aneurysm rupture is biological process where inflammation of aneurysm wall is actively leading to degeneration of aneurysm wall and finally weakens it until it bursts. Until now, there have not been imaging method to detect inflammatory process of aneurysm wall METHODS: We created targeting immunoliposome for use in the imaging of aneurysm. Immunoliposome comprises antibodies against at least one vascular inflammatory marker associated with aneurysm inflammation and a label and/or a contrast agent. RESULTS: Histological analysis of IAs where immunoliposome comprises antibodies against vascular inflammation with a label shows promising results for selectively detecting aneurysms inflammation. In magnetic resonance imaging (MRI) we were able to detect immunoliposomes carrying gadolinium. CONCLUSION: Our work opens a new avenue for using contrast labeled immunoliposomes for detecting rupture-prone aneurysms. Immunoliposomes can cary gadolinium and selectively bind to inflammatory section of aneurysm that can be detected with MRI. Further research is needed to develop immunoliposomes to be used with MRI in humans to target treatment to those patients who benefit from it the most.

The angiopoietin receptor Tie2 is atheroprotective in arterial endothelium.

Leukocytes and resident cells in the arterial wall contribute to atherosclerosis, especially at sites of disturbed blood flow. Expression of endothelial Tie1 receptor tyrosine kinase is enhanced at these sites, and attenuation of its expression reduces atherosclerotic burden and decreases inflammation. However, Tie2 tyrosine kinase function in atherosclerosis is unknown. Here we provide genetic evidence from humans and from an atherosclerotic mouse model to show that TIE2 is associated with protection from coronary artery disease. We show that deletion of Tie2, or both Tie2 and Tie1, in the arterial endothelium promotes atherosclerosis by increasing Foxo1 nuclear localization, endothelial adhesion molecule expression and accumulation of immune cells. We also show that Tie2 is expressed in a subset of aortic fibroblasts, and its silencing in these cells increases expression of inflammation-related genes. Our findings indicate that unlike Tie1, the Tie2 receptor functions as the dominant endothelial angiopoietin receptor that protects from atherosclerosis.

Lipoprotein(a) induces caspase-1 activation and IL-1 signaling in human macrophages.

Introduction: Lipoprotein(a) (Lp(a)) is an LDL-like particle with an additional apolipoprotein (apo)(a) covalently attached. Elevated levels of circulating Lp(a) are a risk factor for atherosclerosis. A proinflammatory role for Lp(a) has been proposed, but its molecular details are incompletely defined. Methods and results: To explore the effect of Lp(a) on human macrophages we performed RNA sequencing on THP-1 macrophages treated with Lp(a) or recombinant apo(a), which showed that especially Lp(a) induces potent inflammatory responses. Thus, we stimulated THP-1 macrophages with serum containing various Lp(a) levels to investigate their correlations with cytokines highlighted by the RNAseq, showing significant correlations with caspase-1 activity and secretion of IL-1ß and IL-18. We further isolated both Lp(a) and LDL particles from three donors and then compared their atheroinflammatory potentials together with recombinant apo(a) in primary and THP-1 derived macrophages. Compared with LDL, Lp(a) induced a robust and dose-dependent caspase-1 activation and release of IL-1ß and IL-18 in both macrophage types. Recombinant apo(a) strongly induced caspase-1 activation and IL-1ß release in THP-1 macrophages but yielded weak responses in primary macrophages. Structural analysis of these particles revealed that the Lp(a) proteome was enriched in proteins associated with complement activation and coagulation, and its lipidome was relatively deficient in polyunsaturated fatty acids and had a high n-6/n-3 ratio promoting inflammation. Discussion: Our data show that Lp(a) particles induce the expression of inflammatory genes, and Lp(a) and to a lesser extent apo(a) induce caspase-1 activation and IL-1 signaling. Major differences in the molecular profiles between Lp(a) and LDL contribute to Lp(a) being more atheroinflammatory.

High cholesterol absorption: A risk factor of atherosclerotic cardiovascular diseases?

Lowering elevated low-density lipoprotein cholesterol (LDL-C) concentrations reduces the risk of atherosclerotic cardiovascular diseases (ASCVDs). However, increasing evidence suggests that cholesterol metabolism may also be involved in the risk reduction of ASCVD events. In this review, we discuss if the different profiles of cholesterol metabolism, with a focus on high cholesterol absorption, are atherogenic, and what could be the possible mechanisms. The potential associations of cholesterol metabolism and the risk of ASCVDs are evaluated from genetic, metabolic, and population-based studies and lipid-lowering interventions. According to these studies, loss-of-function genetic variations in the small intestinal sterol transporters ABCG5 and ABCG8 result in high cholesterol absorption associated with low cholesterol synthesis, low cholesterol elimination from the body, and a high risk of ASCVDs. In contrast, loss-of-function genetic variations in another intestinal sterol transporter, NPC1L1 result in low cholesterol absorption associated with high cholesterol synthesis, elevated cholesterol elimination from the body, and low risk of ASCVDs. Statin monotherapy is not sufficient to reduce the ASCVD risk in cases of high cholesterol absorption, and these individuals need combination therapy of statin with cholesterol absorption inhibition. High cholesterol absorption, i.e., >60%, is estimated to occur in approximately one third of a population, so taking it into consideration is important to optimise lipid-lowering therapy to prevent atherosclerosis and reduce the risk of ASCVD events.

Fatty fish consumption reduces lipophilic index in erythrocyte membranes and serum phospholipids.

BACKGROUND AND AIMS: Lipophilic index (LI) has been introduced to assess the overall fatty acid lipophilicity and as a simple estimate of membrane fluidity. However, little is known on effect of diet on LI. We tested if Camelina sativa oil (CSO) high in ALA, fatty fish (FF) or lean fish (LF) affect LI as compared to control diet and, secondarily, if the LI is associated with HDL lipids and functionality and LDL lipidome. METHODS AND RESULTS: We used data from two randomized clinical trials. The AlfaFish intervention lasted 12 weeks and 79 subjects with impaired glucose tolerance were randomized to FF, LF, CSO or control group. In the Fish trial, 33 subjects with myocardial infarction or unstable ischemic heart attack were randomized to FF, LF or control group for 8 weeks. LI was calculated from erythrocyte membrane fatty acids in AlfaFish and from serum phospholipids in Fish trial. HDL lipids were measured using high-throughput proton nuclear magnetic resonance spectroscopy. There was a significant decrease in LI in the FF group in the AlfaFish (fold change 0.98 ± 0.03) and in the Fish trial (0.95 ± 0.04) and the decrease differed from that of control group in both trials and from CSO group in the AlfaFish study. There were no significant changes in LI in LF or CSO groups. The mean diameter of HDL particles and concentration of large HDL particles were inversely associated with LI. CONCLUSION: FF consumption decreased LI indicating better membrane fluidity in subjects with impaired glucose tolerance or coronary heart disease.

OxLDL sensitizes platelets for increased formation of extracellular vesicles capable of finetuning macrophage gene expression.

Platelet extracellular vesicles (PEVs) generated upon platelet activation may play a role in inflammatory pathologies such as atherosclerosis. Oxidized low-density lipoprotein (oxLDL), a well-known contributor to atherogenesis, activates platelets and presensitizes them for activation by other agonists. We studied the effect of oxLDL on the secretion, composition, and inflammatory functions of PEVs using contemporary EV analytics. Platelets were activated by co-stimulation with thrombin (T) and collagen (C) ± oxLDL and characterized by high-resolution flow cytometry, nanoparticle tracking analysis, proximity extension assay, western blot, and electron microscopy. The effect of PEVs on macrophage differentiation and functionality was examined by analyzing macrophage surface markers, cytokine secretion, and transcriptome. OxLDL upregulated TC-induced formation of CD61+, P-selectin+ and phosphatidylserine+ PEVs. Blocking the scavenger receptor CD36 significantly suppressed the oxLDL+TC-induced PEV formation, and HDL caused a slight but detectable suppression. The inflammatory protein cargo differed between the PEVs from stimulated and unstimulated platelets. Both oxLDL+TC- and TC-induced PEVs enhanced macrophage HLA-DR and CD86 expression and decreased CD11c expression as well as secretion of several cytokines. Pathways related to cell cycle and regulation of gene expression, and immune system signaling were overrepresented in the differentially expressed genes between TC PEV -treated vs. control macrophages and oxLDL+TC PEV -treated vs. control macrophages, respectively. In conclusion, we speculate that oxLDL and activated platelets contribute to proatherogenic processes by increasing the number of PEVs that provide an adhesive and procoagulant surface, contain inflammatory mediators, and subtly finetune the macrophage gene expression.

The human liver lipidome is significantly related to the lipid composition and aggregation susceptibility of low-density lipoprotein (LDL) particles.

BACKGROUND AND AIMS: The susceptibility of low-density lipoprotein (LDL) to aggregation predicts atherosclerotic cardiovascular disease. However, causes of interindividual variation in LDL lipid composition and aggregation susceptibility remain unclear. We examined whether the lipid composition and aggregation susceptibility of LDL reflect the lipid composition of the human liver. METHODS: Liver biopsies and blood samples for isolation of LDL particles were obtained from 40 obese subjects (BMI 45.9 ± 6.1 kg/m2, age 43 ± 8 years). LDL was isolated using sequential ultracentrifugation and lipidomic analyses of liver and LDL samples were determined using ultra-high performance liquid chromatography-mass spectrometry. LDL aggregation susceptibility ex vivo was analyzed by inducing aggregation by human recombinant secretory sphingomyelinase and following aggregate formation. RESULTS: The composition (acyl carbon number and double bond count) of hepatic triglycerides, phosphatidylcholines, and sphingomyelins (SMs) was closely associated with that of LDL particles. Hepatic dihydroceramides and ceramides were positively correlated with concentrations of the corresponding SM species in LDL as well with LDL aggregation. These relationships remained statistically significant after adjustment for age, sex, and body mass index. CONCLUSIONS: Lipid composition of LDL reflects that of the human liver in obese patients. Changes in hepatic sphingolipid metabolism may contribute to interindividual variation of LDL lipid composition and susceptibility to aggregation.

The spoils of war and the long-term spoiling of health conditions of entire nations.

The healthcare system of Ukraine was already suffering from several shortfalls before February 2022, but the war of aggression started by the Russian leadership is poised to inflict a further severe blow that will have long-lasting consequences for the health of all Ukrainians. In pre-war Ukraine, noncommunicable diseases (NCDs) contributed to 91% of deaths, especially cardiovascular diseases (67%). Ukrainians have a high prevalence of risk factors for NCDs ranking among the highest levels reported by the World Health Organization (WHO) in the European (EU) Region. Cardiovascular disease is one of the key health risks for the conflict-affected Ukrainian population due to significant limitations in access to health care and interruptions in the supply of medicines and resources. The excess mortality observed during the COVID-19 pandemic, due to a combination of viral illness and chronic disease states, is bound to increase exponentially from poorly treated NCDs. In this report, we discuss the impact of the war on the public health of Ukraine and potential interventions to provide remote health assistance to the Ukrainian population.

Modified Lipoproteins Induce Arterial Wall Inflammation During Atherogenesis.

Circulating apolipoprotein B-containing lipoproteins, notably the low-density lipoproteins, enter the inner layer of the arterial wall, the intima, where a fraction of them is retained and modified by proteases, lipases, and oxidizing agents and enzymes. The modified lipoproteins and various modification products, such as fatty acids, ceramides, lysophospholipids, and oxidized lipids induce inflammatory reactions in the macrophages and the covering endothelial cells, initiating an increased leukocyte diapedesis. Lipolysis of the lipoproteins also induces the formation of cholesterol crystals with strong proinflammatory properties. Modified and aggregated lipoproteins, cholesterol crystals, and lipoproteins isolated from human atherosclerotic lesions, all can activate macrophages and thereby induce the secretion of proinflammatory cytokines, chemokines, and enzymes. The extent of lipoprotein retention, modification, and aggregation have been shown to depend largely on differences in the composition of the circulating lipoprotein particles. These properties can be modified by pharmacological means, and thereby provide opportunities for clinical interventions regarding the prevention and treatment of atherosclerotic vascular diseases.

Children with familial hypercholesterolemia display changes in LDL and HDL function: A cross-sectional study.

BACKGROUND: The functional status of lipoprotein particles contributes to atherogenesis. The tendency of plasma low-density lipoprotein (LDL) particles to aggregate and the ability of igh-density lipoprotein (HDL) particles to induce and mediate reverse cholesterol transport associate with high and low risk for cardiovascular disease in adult patients, respectively. However, it is unknown whether children with familial hypercholesterolemia (FH) display lipoprotein function alterations. HYPOTHESIS: We hypothesized that FH children had disrupted lipoprotein functions. METHODS: We analyzed LDL aggregation susceptibility and HDL-apoA-I exchange (HAE), and activity of four proteins that regulate lipoprotein metabolism (cholesteryl ester transfer protein, lecithin-cholesterol acyltransferase, phospholipid transfer protein, and paraoxonase-1) in plasma samples derived from children with FH (n = 47) and from normocholesterolemic children (n = 56). Variation in lipoprotein functions was further explored using an nuclear magnetic resonance-based metabolomics profiling approach. RESULTS: LDL aggregation was higher, and HAE was lower in FH children than in normocholesterolemic children. LDL aggregation associated positively with LDL cholesterol (LDL-C) and negatively with triglycerides, and HAE/apoA-I associated negatively with LDL-C. Generally, the metabolomic profile for LDL aggregation was opposite of that of HAE/apoA-I. CONCLUSIONS: FH children displayed increased atherogenicity of LDL and disrupted HDL function. These newly observed functional alterations in LDL and HDL add further understanding of the risk for atherosclerotic cardiovascular disease in FH children.

Lipidomic changes of LDL after consumption of Camelina sativa oil, fatty fish and lean fish in subjects with impaired glucose metabolism-A randomized controlled trial.

BACKGROUND: There is little knowledge on the effects of alpha-linolenic acid (ALA) and n-3 long-chain polyunsaturated fatty acids (n-3 LCPUFA) on the LDL lipidome and aggregation of LDL particles. OBJECTIVE: We examined if consumption of Camelina sativa oil (CSO) as a source of ALA, fatty fish (FF) as a source of n-3 LCPUFA and lean fish (LF) as a source of fish protein affect the lipidome of LDL as compared to a control diet. METHODS: Participants with impaired glucose tolerance (39 women and 40 men) were randomized to 4 study groups (CSO providing 10 g/d ALA, FF and LF [both 4 fish meals/wk] and control limiting their fish and ALA intake) in a 12-week, parallel trial. Diets were instructed and dietary fats were provided to the participants. The lipidome of LDL particles isolated from samples collected at baseline and after intervention was analyzed with electrospray ionization-tandem mass spectrometry. RESULTS: In the CSO group, the relative concentrations of saturated and monounsaturated cholesteryl ester species in LDL decreased and the species with ALA increased. In the FF group, LDL phosphatidylcholine (PC) species containing n-3 LCPUFA increased. There was a significant positive correlation between the change in total sphingomyelin and change in LDL aggregation, while total PC and triunsaturated PC species were inversely associated with LDL aggregation when all the study participants were included in the analysis. CONCLUSION: Dietary intake of CSO and FF modifies the LDL lipidome to contain more polyunsaturated and less saturated lipid species. The LDL surface lipids are associated with LDL aggregation.

Overfeeding Saturated Fat Increases LDL (Low-Density Lipoprotein) Aggregation Susceptibility While Overfeeding Unsaturated Fat Decreases Proteoglycan-Binding of Lipoproteins.

Objective: We recently showed that measurement of the susceptibility of LDL (low-density lipoprotein) to aggregation is an independent predictor of cardiovascular events. We now wished to compare effects of overfeeding different dietary macronutrients on LDL aggregation, proteoglycan-binding of plasma lipoproteins, and on the concentration of oxidized LDL in plasma, 3 in vitro parameters consistent with increased atherogenicity. Approach and Results: The participants (36 subjects; age, 48+/-10 years; body mass index, 30.9+/-6.2 kg/m2) were randomized to consume an extra 1000 kcal/day of either unsaturated fat, saturated fat, or simple sugars (CARB) for 3 weeks. We measured plasma proatherogenic properties (susceptibility of LDL to aggregation, proteoglycan-binding, oxidized LDL) and concentrations and composition of plasma lipoproteins using nuclear magnetic resonance spectroscopy, and in LDL using liquid chromatography mass spectrometry, before and after the overfeeding diets. LDL aggregation increased in the saturated fat but not the other groups. This change was associated with increased sphingolipid and saturated triacylglycerols in LDL and in plasma and reduction of clusterin on LDL particles. Proteoglycan binding of plasma lipoproteins decreased in the unsaturated fat group relative to the baseline diet. Lipoprotein properties remained unchanged in the CARB group. Conclusions: The type of fat during 3 weeks of overfeeding is an important determinant of the characteristics and functional properties of plasma lipoproteins in humans.

Native and oxidised lipoproteins negatively regulate the serum amyloid A-induced NLRP3 inflammasome activation in human macrophages.

OBJECTIVES: The NLRP3 inflammasome plays a key role in arterial wall inflammation. In this study, we elucidated the role of serum lipoproteins in the regulation of NLRP3 inflammasome activation by serum amyloid A (SAA) and other inflammasome activators. METHODS: The effect of lipoproteins on the NLRP3 inflammasome activation was studied in primary human macrophages and THP-1 macrophages. The effect of oxidised low-density lipoprotein (LDL) was examined in an in vivo mouse model of SAA-induced peritoneal inflammation. RESULTS: Native and oxidised high-density lipoproteins (HDL3) and LDLs inhibited the interaction of SAA with TLR4. HDL3 and LDL inhibited the secretion of interleukin (IL)-1ß and tumor necrosis factor by reducing their transcription. Oxidised forms of these lipoproteins reduced the secretion of mature IL-1ß also by inhibiting the activation of NLRP3 inflammasome induced by SAA, ATP, nigericin and monosodium urate crystals. Specifically, oxidised LDL was found to inhibit the inflammasome complex formation. No cellular uptake of lipoproteins was required, nor intact lipoprotein particles for the inhibitory effect, as the lipid fraction of oxidised LDL was sufficient. The inhibition of NLRP3 inflammasome activation by oxidised LDL was partially dependent on autophagy. Finally, oxidised LDL inhibited the SAA-induced peritoneal inflammation and IL-1ß secretion in vivo. CONCLUSIONS: These findings reveal that both HDL3 and LDL inhibit the proinflammatory activity of SAA and this inhibition is further enhanced by lipoprotein oxidation. Thus, lipoproteins possess major anti-inflammatory functions that hinder the NLRP3 inflammasome-activating signals, particularly those exerted by SAA, which has important implications in the pathogenesis of cardiovascular diseases.

Streptococcus pneumoniae pneumolysin and neuraminidase A convert high-density lipoproteins into pro-atherogenic particles.

High-density lipoproteins (HDLs) are a group of different subpopulations of sialylated particles that have an essential role in the reverse cholesterol transport (RCT) pathway. Importantly, changes in the protein and lipid composition of HDLs may lead to the formation of particles with reduced atheroprotective properties. Here, we show that Streptococcus pneumoniae pneumolysin (PLY) and neuraminidase A (NanA) impair HDL function by causing chemical and structural modifications of HDLs. The proteomic, lipidomic, cellular, and biochemical analysis revealed that PLY and NanA induce significant changes in sialic acid, protein, and lipid compositions of HDL. The modified HDL particles have reduced cholesterol acceptor potential from activated macrophages, elevated levels of malondialdehyde adducts, and show significantly increased complement activating capacity. These results suggest that accumulation of these modified HDL particles in the arterial intima may present a trigger for complement activation, inflammatory response, and thereby promote atherogenic disease progression.