Apolipoprotein B Research Articles

Role of interleukin-6 and interleukin-10 in morphological and functional changes of the blood–brain barrier in hypertriglyceridemia

BackgroundHypertriglyceridemia is closely linked to atherosclerosis related inflammatory processes and blood–brain barrier (BBB) dysfunction. Using apolipoprotein B-100 (APOB-100) transgenic mice, an animal model of chronic hypertriglyceridemia, we analyzed BBB function and morphology in vitro and ex vivo. Our objective was to determine which BBB characteristics are produced mainly by interleukin (IL)-6, an atherosclerosis promoting cytokine, and whether these actions can be antagonized by IL-10, an anti-inflammatory cytokine.MethodsBrain endothelial and glial cell cultures and brain microvessels were isolated from wild type (WT) and APOB-100 transgenic mice and were treated with IL-6, IL-10 and their combination. First, IL-6 and IL-10 production was measured in WT and APOB-100 microvessels using qPCR. Then functional parameters of endothelial cell cultures were analyzed and immunocytochemistry for key BBB proteins was performed.ResultsIL-6 mRNA levels were higher in brain microvessels than in brain parenchyma of APOB-100 transgenic mice. Transendothelial electric resistance and P-glycoprotein activity were lower, and paracellular permeability was higher in cultured APOB-100 brain endothelial cells. These features were sensitive to both IL-6 and IL-10 treatments. A decreased P-glycoprotein immunostaining was measured in transgenic endothelial cells under control conditions and in WT cells after treating them with IL-6. This effect was antagonized by IL-10. Changes in immunostaining for tight junction proteins were observed after IL-6 exposure, which were in part antagonized by IL-10. In glial cell cultures an increase in aquaporin-4 immunolabeling in the transgenic group and an increase in microglia cell density in WT glia cultures was detected after IL-6 treatment, which was antagonized by IL-10. In isolated brain microvessels a decrease in P-glycoprotein immunolabeled area fraction was measured in APOB-100 microvessels under control conditions and in WT microvessels after every cytokine treatment. ZO-1 immunolabeling showed characteristics similar to that of P-glycoprotein. No change was seen in claudin-5 and occludin immunoreactive area fractions in microvessels. A decrease in aquaporin-4 immunoreactivity was measured in WT microvessels treated by IL-6, which was antagonized by IL-10.ConclusionIL-6 produced in microvessels contributes to BBB impairment observed in the APOB-100 mice. We showed that IL-10 partly antagonizes the effects of IL-6 at the BBB.

How Genetic Variants in Children with Familial Hypercholesterolemia Not Only Guide Detection, but Also Treatment.

Familial hypercholesterolemia (FH) is a hereditary disorder that causes severely elevated low-density lipoprotein (LDL-C) levels, which leads to an increased risk for premature cardiovascular disease. A variety of genetic variants can cause FH, namely variants in the genes for the LDL receptor (LDLR), apolipoprotein B (APOB), proprotein convertase subtilisin/kexin type 9 (PCSK9), and/or LDL-receptor adaptor protein 1 (LDLRAP1). Variants can exist in a heterozygous form (HeFH) or the more severe homozygous form (HoFH). If affected individuals are diagnosed early (through screening), they benefit tremendously from early initiation of lipid-lowering therapy, such as statins, and cardiovascular imaging to detect possible atherosclerosis. Over the last years, due to intensive research on the genetic basis of LDL-C metabolism, novel, promising therapies have been developed to reduce LDL-C levels and subsequently reduce cardiovascular risk. Results from studies on therapies focused on inhibiting PCSK9, a protein responsible for degradation of the LDLR, are impressive. As the effect of PCSK9 inhibitors (PCSK9-i) is dependent of residual LDLR activity, this medication is less potent in patients without functional LDLR (e.g., null/null variant). Novel therapies that are expected to become available in the near future focused on inhibition of another major regulatory protein in lipid metabolism (angiopoietin-like 3 (ANGPTL3)) might dramatically reduce the frequency of apheresis in children with HoFH, independently of their residual LDLR. At present, another independent risk factor for premature cardiovascular disease, elevated levels of lipoprotein(a) (Lp(a)), cannot be effectively treated with medication. Further understanding of the genetic basis of Lp(a) metabolism, however, offers a possibility for the development of novel therapies.

Plasma Apolipoprotein Concentrations Are Highly Altered in Severe Intensive Care Unit COVID-19 Patients: Preliminary Results from the LIPICOR Cohort Study.

SARS-CoV-2 infection goes beyond acute pneumonia, as it also impacts lipid metabolism. Decreased HDL-C and LDL-C levels have been reported in patients with COVID-19. The lipid profile is a less robust biochemical marker than apolipoproteins, components of lipoproteins. However, the association of apolipoprotein levels during COVID-19 is not well described and understood. The objective of our study is to measure plasma levels of 14 apolipoproteins in patients with COVID-19 and to evaluate the relationships between apolipoprotein levels, severity factors and patient outcomes. From November to March 2021, 44 patients were recruited on admission to the intensive care unit because of COVID-19. Fourteen apolipoproteins and LCAT were measured by LC-MS/MS in plasma of 44 COVID-19 patients on admission to the ICU and 44 healthy control subjects. Absolute apolipoprotein concentrations were compared between COVID-19 patients and controls. Plasma apolipoproteins (Apo) A (I, II, IV), C(I, II), D, H, J and M and LCAT were lower in COVID-19 patients, whereas Apo E was higher. COVID-19 severity factors such as PaO2/FiO2 ratio, SO-FA score and CRP were correlated with certain apolipoproteins. Lower Apo B100 and LCAT levels were observed in non-survivors of COVID-19 versus survivors. To conclude, in this study, lipid and apolipoprotein profiles are altered in COVID-19 patients. Low Apo B100 and LCAT levels may be predictive of non-survival in COVID-19 patients.

Remnant Indices for Estimating Postprandial Lipidemia in Young Women

Postprandial hyperglycemia is well established as a major risk factor for prediabetes and type 2 diabetes. However, for postprandial hyperlipidemia, no definitive criteria exist, and fasting values of lipids and lipoproteins had limited efficacy for estimating postprandial lipidemia. In the present study, we performed a fat-ingestion test and aimed to propose indices for estimating postprandial lipoprotein metabolism. Methods: Healthy young Japanese women (n=54, age 21.1 ± 1.0 y) with apolipoprotein E phenotype 3/3 were enrolled. They ingested fat cream (OFTT cream™, Jomo, Japan; 1 g/kg as cream, 0.35 g/kg as fat). Venous blood samples were taken before (0 h) and at 0.5, 1, 2, 4, and 6 h after ingestion. Results: The serum triglyceride (TG) level peaked at 2 h and returned to below baseline at 6 h. The remnant-like particle-TG (RP-TG) level increased at 1 h, peaked at 2 h, and returned to baseline at 6 h. The remnant lipoprotein-cholesterol (RLP-C) level increased at 2 h, peaked at 4 h, and returned to baseline at 6 h. The apolipoprotein B48 level increased at 1 h, peaked at 4 h, and did not return to baseline at 6 h. The apolipoprotein B100 concentration slightly decreased at 2 h and increased at 6 h. TG−RP-TG did not change during 6 h, but RP-TG/TG rose at 2–6 h compared to the fasting value. RP-TG/RLP-C increased at 2–4 h and returned to baseline at 6 h. Conclusion: After fat ingestion, while the concentration of non-remnant TG was stable, remnant TG increased. The content of TG per remnant particle increased up to 2 h and decreased from 2 h to 6 h, and the size became smaller. The remnant indices, RP-TG/TG and RP-TG/RLP-C, may be useful for estimating postprandial lipidemia.

Targeted proteomics using stable isotope labeled protein fragments enables precise and robust determination of total apolipoprotein(a) in human plasma.

Lipoprotein(a), also known as Lp(a), is an LDL-like particle composed of apolipoprotein(a) (apo(a)) bound covalently to apolipoprotein B100. Plasma concentrations of Lp(a) are highly heritable and vary widely between individuals. Elevated plasma concentration of Lp(a) is considered as an independent, causal risk factor of cardiovascular disease (CVD). Targeted mass spectrometry (LC-SRM/MS) combined with stable isotope-labeled recombinant proteins provides robust and precise quantification of proteins in the blood, making LC-SRM/MS assays appealing for monitoring plasma proteins for clinical implications. This study presents a novel quantitative approach, based on proteotypic peptides, to determine the absolute concentration of apo(a) from two microliters of plasma and qualified according to guideline requirements for targeted proteomics assays. After optimization, assay parameters such as linearity, lower limits of quantification (LLOQ), intra-assay variability (CV: 4.7%) and inter-assay repeatability (CV: 7.8%) were determined and the LC-SRM/MS results were benchmarked against a commercially available immunoassay. In summary, the measurements of an apo(a) single copy specific peptide and a kringle 4 specific peptide allow for the determination of molar concentration and relative size of apo(a) in individuals.

Associations of genetically determined lipid traits and lipid-modifying agents with the risk of diabetic retinopathy: A Mendelian randomization study

Background and aimsThe evidence that dyslipidemia is associated with hyperglycemia calls for an investigation of whether dyslipidemia, as well as lipid-modifying agents, could affect the subsequent development of diabetic retinopathy (DR). Therefore, we aimed to address these unanswered questions by utilizing Mendelian randomization (MR) analysis. MethodsGenetic variants were selected from the UK Biobank as instruments to serve as proxies for lipid traits [high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein cholesterol (LDL-C), triglyceride (TG), apolipoprotein A-I (APOA-I) and apolipoprotein B (APOB)]. Univariable and multivariable MR analyses were performed to examine the associations of these lipid traits with DR and different levels of severity of DR. Based on the evidence for the effects of lipids on outcomes, we estimated the causal relevance of cholesteryl ester transfer protein (CETP) inhibitors in severe nonproliferative and proliferative DR using protein quantitative trait loci (pQTLs) and expression quantitative trait loci (eQTLs) as instruments. ResultsGenetically determined HDL-C levels were inversely associated with the risk of severe nonproliferative DR (OR = 0.70, 95% CI = 0.52–0.94) and proliferative DR (OR = 0.90, 95% CI = 0.83–0.97) in the main analyses utilizing the inverse variance-weighted (IVW) MR method and a couple of sensitivity analyses. No association was noted between genetically proxied CETP inhibitors and DR. ConclusionsThis MR study suggests the casual protective roles of HDL-C in severe nonproliferative DR and proliferative DR, which calls for further studies to confirm these findings. Current lipid-modifying agents acting on HDL-C may not reduce the risk of DR and new treatments are required in the future.

Identification of genetic susceptibility in preterm newborns with bronchopulmonary dysplasia by whole-exome sequencing: BIVM gene may play a role

Bronchopulmonary dysplasia (BPD) is a common chronic respiratory diseasein preterm infants caused by multifactorial etiology. Genetic factors areinvolved in the occurrence of BPD, but studies have found that candidate geneshave poor reproducibility and are influenced by ethnic heterogeneity;therefore, more exploration is still needed. We performed whole-exon sequencingin 34 preterm infants with BPD and 32 non-BPD control neonates. The data wereanalyzed and interpreted by Fisher difference comparison, PLINK and eQTLassociation analysis, KEGG and GO enrichment analysis, STRING tool, Cytoscapesoftware, ProtParam tool, HOPE online software, and GEOR2 analysis on NCBI GEOdataset. BPD has a highly heterogeneity in different populations, and we found35 genes overlapped with previous whole-exon sequencing studies, such as APOBgene. Arterial and epithelial cell development and energy metabolism pathwaysaffect BPD. In this study, 24 key genes were identified, and BIVM rs3825519mutation leads to prolonged assisted ventilation in patients with BPD. A novelDDAH1 mutation site (NM_012137: exon1: c.89 T > G: p.L30R) was found in 9 BPD patients.Conclusion: BIVM gene rs3825519 mutation may play a role in the pathogenesis of BPD by affecting cilia movement, and the DDAH1 and APOB genes mutations may have a pathogenic role in BPD.What is Known:• Genetic factors are involved in the occurrence of bronchopulmonary dysplasia.• The candidate genes have poor reproducibility and are influenced by ethnic heterogeneity, therefore, more exploration is still needed.What is New:• We identified the role of susceptible SNPs in BPD in Shenzhen, China, and identified 24 key genes that influence the pathogenesis of BPD, and also found 35 genes overlapped with previous whole exon sequencing studies, such as APOB gene.• We found that BIVM and DDAH1 genes may play a pathogenic role in the pathogenesis of BPD.

Functional Characterization of p.(Arg160Gln) PCSK9 Variant Accidentally Found in a Hypercholesterolemic Subject

Familial hypercholesterolaemia (FH) is an autosomal dominant dyslipidaemia, characterised by elevated LDL cholesterol (LDL-C) levels in the blood. Three main genes are involved in FH diagnosis: LDL receptor (LDLr), Apolipoprotein B (APOB) and Protein convertase subtilisin/kexin type 9 (PCSK9) with genetic mutations that led to reduced plasma LDL-C clearance. To date, several PCSK9 gain-of-function (GOF) variants causing FH have been described based on their increased ability to degrade LDLr. On the other hand, mutations that reduce the activity of PCSK9 on LDLr degradation have been described as loss-of-function (LOF) variants. It is therefore important to functionally characterise PCSK9 variants in order to support the genetic diagnosis of FH. The aim of this work is to functionally characterise the p.(Arg160Gln) PCSK9 variant found in a subject suspected to have FH. Different techniques have been combined to determine efficiency of the autocatalytic cleavage, protein expression, effect of the variant on LDLr activity and affinity of the PCSK9 variant for the LDLr. Expression and processing of the p.(Arg160Gln) variant had a result similar to that of WT PCSK9. The effect of p.(Arg160Gln) PCSK9 on LDLr activity is lower than WT PCSK9, with higher values of LDL internalisation (13%) and p.(Arg160Gln) PCSK9 affinity for the LDLr is lower than WT, EC50 8.6 ± 0.8 and 25.9 ± 0.7, respectively. The p.(Arg160Gln) PCSK9 variant is a LOF PCSK9 whose loss of activity is caused by a displacement of the PCSK9 P' helix, which reduces the stability of the LDLr-PCSK9 complex.

Genetic Heterogeneity of Familial Hypercholesterolemia: Repercussions for Molecular Diagnosis.

Genetics of Familial Hypercholesterolemia (FH) is ascribable to pathogenic variants in genes encoding proteins leading to an impaired LDL uptake by the LDL receptor (LDLR). Two forms of the disease are possible, heterozygous (HeFH) and homozygous (HoFH), caused by one or two pathogenic variants, respectively, in the three main genes that are responsible for the autosomal dominant disease: LDLR, APOB and PCSK9 genes. The HeFH is the most common genetic disease in humans, being the prevalence about 1:300. Variants in the LDLRAP1 gene causes FH with a recessive inheritance and a specific APOE variant was described as causative of FH, contributing to increase FH genetic heterogeneity. In addition, variants in genes causing other dyslipidemias showing phenotypes overlapping with FH may mimic FH in patients without causative variants (FH-phenocopies; ABCG5, ABCG8, CYP27A1 and LIPA genes) or act as phenotype modifiers in patients with a pathogenic variant in a causative gene. The presence of several common variants was also considered a genetic basis of FH and several polygenic risk scores (PRS) have been described. The presence of a variant in modifier genes or high PRS in HeFH further exacerbates the phenotype, partially justifying its variability among patients. This review aims to report the updates on the genetic and molecular bases of FH with their implication for molecular diagnosis.

Molecular Characteristics of Toxicity of Acrolein Produced from Spermine.

Acrolein (CH2=CH-CHO), an unsaturated aldehyde produced from spermine, is one of the major contributors to oxidative stress. Acrolein has been found to be more toxic than reactive oxygen species (H2O2 and •OH), and it can be easily conjugated with proteins, bringing about changes in nature of the proteins. Acrolein is detoxified by glutathione in cells and was found to be mainly produced from spermine through isolating two cell lines of acrolein-resistant Neuro2a cells. The molecular characteristics of acrolein toxicity and tissue damage elicited by acrolein were investigated. It was found that glyceraldehyde-3-phosphate dehydrogenase (GAPDH); cytoskeleton proteins such as vimentin, actin, α- and β-tubulin proteins; and apolipoprotein B-100 (ApoB100) in LDL are strongly damaged by acrolein conjugation. In contrast, activities of matrix metalloproteinase-9 (MMP-9) and proheparanase (proHPSE) are enhanced, and antibody-recognizing abilities of immunoglobulins are modified by acrolein conjugation, resulting in aggravation of diseases. The functional changes of these proteins by acrolein have been elucidated at the molecular level. The findings confirmed that acrolein is the major contributor causing tissue damage in the elderly.

Case report of familial hypobetalipoproteinemia: a novel APOB mutation and literature review.

Familial hypobetalipoproteinemia is an autosomal dominantly inherited disorder of lipid metabolism characterized by a <5th percentile plasma levels of low-density lipoprotein cholesterol or total apolipoprotein B, caused by mutations in the APOB gene. Patients with heterozygous APOB-related familial hypobetalipoproteinemia are usually asymptomatic or manifest with mild liver dysfunction or hepatic steatosis. A 14-year-old Korean boy was referred to our clinic because of low low-density lipoprotein cholesterol and elevated aminotransferase levels. He did not complain specific symptoms of hypobetalipoproteinemia, but his laboratory and radiologic tests showed low low-density lipoprotein cholesterol, low apolipoprotein B, high aminotransferases, severe fatty liver, and hepatosplenomegaly. A multigene panel test identified a heterozygous mutation for p.Lys3846Ter (c.11536A>T) in the APOB gene, which has not been reported in previous studies and was not detected in his parents with normal lipid profiles. Limiting intake of saturated fat, increased physical activity, and vitamin E supplementation were performed. Subsequent laboratory tests revealed normalized aminotransferase levels.

Genomic analysis of lean individuals with NAFLD identifies monogenic disorders in a prospective cohort study

Lean patients with non-alcoholic fatty liver disease (NAFLD) represent 10-20% of the affected population and may have heterogeneous drivers of disease. We have recently proposed the evaluation of patients with lean NAFLD without visceral adiposity for rare monogenic drivers of disease. Here, we aimed to validate this framework in a well-characterised cohort of patients with biopsy-proven NAFLD by performing whole exome sequencing. This prospective study included 124 patients with biopsy-proven NAFLD and paired liver biopsies who underwent standardised research visits including advanced magnetic resonance imaging (MRI) assessment of liver fat and stiffness. Six patients with lean NAFLD were identified and underwent whole exome sequencing. Two lean patients (33%) were identified to have monogenic disorders. The lean patients with monogenic disorders had similar age, and anthropometric and MRI characteristics to lean patients without a monogenic disorder. Patient 1 harbours a rare homozygous pathogenic mutation in ALDOB (aldolase B) and was diagnosed with hereditary fructose intolerance. Patient 2 harbours a rare heterozygous mutation in apolipoprotein B (APOB). The pathogenicity of this APOB variant (p.Val1856CysfsTer2) was further validated in the UK Biobank and associated with lower circulating APOB levels (beta= -0.51g/L, 95% CI -0.65 to -0.36g/L, p= 1.4×10-11) and higher liver fat on MRI (beta=+10.4%, 95% CI 4.3-16.5%, p= 8.8×10-4). Hence, patient 2 was diagnosed with heterozygous familial hypobetalipoproteinaemia. In this cohort of well-characterised patients with lean NAFLD without visceral adiposity, 33% (2/6) had rare monogenic drivers of disease, highlighting the importance of genomic analysis in this NAFLD subtype. Although most people with non-alcoholic fatty liver disease (NAFLD) are overweight or obese, a subset are lean and may have unique genetic mutations that cause their fatty liver disease. We show that 33% of study participants with NAFLD who were lean harboured unique mutations that cause their fatty liver, and that these mutations had effects beyond the liver. This study demonstrates the value of genetic assessment of NAFLD in lean individuals to identify distinct subtypes of disease.

Association of Rare Protein-Truncating DNA Variants in APOB or PCSK9 With Low-density Lipoprotein Cholesterol Level and Risk of Coronary Heart Disease

Protein-truncating variants (PTVs) in apolipoprotein B (APOB) and proprotein convertase subtilisin/kexin type 9 (PCSK9) are associated with significantly lower low-density lipoprotein (LDL) cholesterol concentrations. The association of these PTVs with coronary heart disease (CHD) warrants further characterization in large, multiracial prospective cohort studies. To evaluate the association of PTVs in APOB and PCSK9 with LDL cholesterol concentrations and CHD risk. This studied included participants from 5 National Heart, Lung, and Blood Institute (NHLBI) studies and the UK Biobank. NHLBI study participants aged 5 to 84 years were recruited between 1971 and 2002 across the US and underwent whole-genome sequencing. UK Biobank participants aged 40 to 69 years were recruited between 2006 and 2010 in the UK and underwent whole-exome sequencing. Data were analyzed from June 2021 to October 2022. PTVs in APOB and PCSK9. Estimated untreated LDL cholesterol levels and CHD. Among 19 073 NHLBI participants (10 598 [55.6%] female; mean [SD] age, 52 [17] years), 139 (0.7%) carried an APOB or PCSK9 PTV, which was associated with 49 mg/dL (95% CI, 43-56) lower estimated untreated LDL cholesterol level. Over a median (IQR) follow-up of 21.5 (13.9-29.4) years, incident CHD was observed in 12 of 139 carriers (8.6%) vs 3029 of 18 934 noncarriers (16.0%), corresponding to an adjusted hazard ratio of 0.51 (95% CI, 0.28-0.89; P = .02). Among 190 464 UK Biobank participants (104 831 [55.0%] female; mean [SD] age, 57 [8] years), 662 (0.4%) carried a PTV, which was associated with 45 mg/dL (95% CI, 42-47) lower estimated untreated LDL cholesterol level. Estimated CHD risk by age 75 years was 3.7% (95% CI, 2.0-5.3) in carriers vs 7.0% (95% CI, 6.9-7.2) in noncarriers, corresponding to an adjusted hazard ratio of 0.51 (95% CI, 0.32-0.81; P = .004). Among 209 537 individuals in this study, 0.4% carried an APOB or PCSK9 PTV that was associated with less exposure to LDL cholesterol and a 49% lower risk of CHD.

The correlation between blood-lipid ratio in the first trimester and large-for-gestational-age infants

BackgroundTo investigate the correlation between maternal glucose and lipid metabolism indexes and blood-lipid ratio in the first trimester and large-for- gestational-age (LGA) infants.MethodsWomen in the first trimester of pregnancy who underwent regular obstetric examination in the obstetric outpatient department of the Affiliated Hospital of Chengde Medical College from June 2018 to March 2019 were included according to the standard. Basic information were collected based on questionnaires at the first visit of pregnant women, including early fasting blood glucose (FBG), fasting insulin (FINS), glycated hemoglobin (HbA1c), high-density lipoprotein (HDL), low-density lipoprotein (LDL), triglyceride (TG), total cholesterol (TC), apolipoprotein A1 (APO-A1), apolipoprotein B (APO-B), lipoprotein a (LP(a)), LDL/HDL, TG/HDL, TC/HDL, APO-B/APO-A1 ratio, birth weight of newborns, gestational age at delivery etc.ResultsA total of 418 cases were included for analysis. The incidence rate of LGA infants was 13.88%, and that of small-for-gestational-age (SGA) infants was 4.78%. Univariate analysis revealed that the age, pre-pregnancy body mass index (BMI), weight gain during pregnancy, APO-B/APO-A1 between LGA group and appropriate-for-gestational-age (AGA) group were significantly different (P < 0.05); multivariate stepwise logistic regression analysis indicated that the correlation between maternal age, pre-pregnancy BMI, weight gain during pregnancy, APO-B/APO-A1 level and LGA were statistically significant (P < 0.05); compared with the reference range of APO-B/APO-A1 of 0.46–0.65, values < 0.46 and > 0.65 were protective factor of LGA (P < 0.05). The receiver operating curve(ROC) indicated that the area under the curve (AUC)s for predicting LGA using maternal age, pre-pregnancy BMI, weight gain during pregnancy, and early pregnancy APO-B/APO-A1 were 0.585, 0.606, 0.637, 0.531, respectively. The AUC for a combined prediction model was 0.742, showing greater predictive value than any other factors individually.ConclusionMaternal age, pre-pregnancy BMI, weight gain during pregnancy, and APO-B/APO-A1 levels in first trimester are significant factors influencing the occurrence of LGA infants, and the combination of the four factors would have certain predictive value for LGA.

Levels of pretreatment serum lipids predict responses to PD-1 inhibitor treatment in advanced intrahepatic cholangiocarcinoma.

It has been identified that serum lipids can be used as prognostic biomarkers in several types of cancer and are associated with patient survival. We aimed to clarify the prognostic value of the serum lipids and to establish a novel effective nomogram for overall survival (OS) in intrahepatic cholangiocarcinoma (iCCA) patients receiving anti-PD1 therapy. Pretreatment serum lipids were retrospectively analyzed for prognostic value, including apolipoprotein B (APOB), apolipoproteinA-1 (APOA1), total cholesterol (TC), high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein cholesterol (LDL-C) and triglycerides (TG), which were assessed for prediction accuracy using Kaplan-Meier survival curves and time-dependent receiver operating characteristic (ROC). Cox regression analysis with univariate and multivariate factors was used to identify prognostic factors predictive of OS, and prognostic nomograms were constructed. All the serum lipids showed good discriminatory ability in terms of OS (all P<0.05), the higher the lipid levels, the better the prognosis, while APOA1 and TG were remarkable independent predictors for OS in multivariate analysis (hazard ratio, 2.177,2.035; confidence interval, 1.393-3.402, 1.184-3.498; P=0.001, P=0.01). Four (CA19-9, APOA1, tumor number and TG) independent prognostic factors were chosen to generate the nomogram for OS. The area under the ROC curve at 1-year and 2-year consistently demonstrated that the predictive value of the nomogram was superior to serum lipids. In our study, serum lipid levels were used as a prognostic nomogram in the prediction of anti-PD-1 therapy efficacy in patients with iCCA.

Analysis of Systemic and Serum Risk Factors in Patients with Age-Related Macular Degeneration and Polypoidal Choroidal Vasculopathy.

It remains controversial whether polypoidal choroidal vasculopathy (PCV) represents a subtype of neovascular age-related macular degeneration (nAMD) or is a distinct disease entity. This study aimed to compare and analyze systemic and serum risk factors for nAMD and PCV in an aging Chinese population. A retrospective study was performed on 108 patients with nAMD, 131 patients with PCV, and 219 control subjects. Serum levels of total cholesterol (TC), triglycerides (TG), low-density lipoprotein (LDL), high-density lipoprotein (HDL), apolipoprotein A1 (APOA1), apolipoprotein B (APOB), complement 3 (C3), and complement 4 (C4) together with data on systemic risk factors, including hyperlipidemia, hypertension, diabetes mellitus (DM), coronary artery disease (CAD), and asthma, were collected. Chi-square tests, independent-samples t tests, and binary logistic regression analyses were performed to evaluate the associations ofrisk factors with nAMD and PCV. Patients with PCV and those with nAMD were likely to have hyperlipidemia (P < 0.001). CAD (P = 0.020) and hypertension (P = 0.006) correlated significantly with nAMD and PCV, respectively. Although no association of age and asthma with PCV or nAMD was found (P > 0.05), DM was associated with PCV development (OR = 0.535, P = 0.044). Regarding serum risk factors, HDL, LDL, TG, APOB, and C3 were significantly associated with nAMD (OR < 0.001, P < 0.001; OR = 0.028, P < 0.001; OR = 0.175, P < 0.001; OR = 0.922, P = 0.022; OR < 0.001, P < 0.001) and PCV (OR = 0.001, P = 0.001; OR = 0.097, P = 0.003; OR = 0.410, P = 0.037; OR = 0.895, P = 0.001; OR = 0.001, P < 0.001). Compared with nAMD, higher levels of HDL (P = 0.003) and LDL (P = 0.016) and lower levels of TG (P = 0.039) were found in patients with PCV, but the association of systemic risk factors between the two diseases was not significant(P > 0.05). Our findings indicate that hyperlipidemia is significantly associated with both nAMD and PCV. Serum lipid and complement levels have an effect on the pathogenesis of nAMD and PCV, and consideration of the differences between systemic and serum risk factors should be taken into account in clinical management.

Targeted sequencing of a gene panel in patients with familial hypercholesterolemia from Southern Poland.

Familial hypercholesterolemia (FH) is an autosomal dominant monogenic lipid metabolism disorder characterized by a significantly elevated level of low‑density lipoprotein (LDL) cholesterol and leading to premature ischemic heart disease. FH is caused by mutations in the LDLR, APOB, and PCSK9 genes; however, these mutations account for only about 40% of FH cases. In order to obtain a genetic diagnosis of FH, sequencing of other genes involved in the lipid metabolism might be useful. This study aimed to describe genetic variants in genes associated with FH in a group of patients from the Małopolska province in Southern Poland, using the targeted next generation sequencing (NGS) technology. The study involved 90 unrelated adults (age range, 18-70 years) with FH diagnosed clinically according to the Simon Broome Register criteria. A custom‑designed capture assay and the Illumina MiSeq platform were used. The panel included exons and exon / intron boundaries of known FH‑causing genes: LDLR, APOB, and PCSK9, as well as genes previously associated with high cholesterol levels: APOE, ABCG5, ABCG8, LPL, NPC1, LDLRAP1, LIPC, STAP1, and CELSR2. Genetic variants were classified based on in silico predictions and ClinVar reports. We detected 4 patients with variants in the LDLR and APOB genes that had not been previously linked to FH in ClinVar. We also found APOB mutations outside the common LDL receptor-binding region, in exons 26 and 29. Interestingly, we observed a high frequency of pathogenic variants in exon 4 of the APOE gene: rs7412, probably damaging (4 patients) and rs429358, benign (16 patients). NGS is a useful and reliable method to detect new variants in genes related to FH. In addition, the results enable the detection of FH phenocopies and introduction of appropriate treatment.

Interactions of fentanyl with blood platelets and plasma proteins: platelet sensitivity to prasugrel metabolite is not affected by fentanyl under in vitro conditions

BackgroundClinical trials indicate that fentanyl, like morphine, may impair intestinal absorption and thus decrease the efficacy of oral P2Y12 inhibitors, such as clopidogrel, ticagrelor, and prasugrel. However, the ability of fentanyl to directly negate or reduce the inhibitory effect of P2Y12 receptor antagonists on platelet function has not been established. A series of in vitro experiments was performed to investigate the ability of fentanyl to activate platelets, potentiate platelet response to ADP, and/or diminish platelet sensitivity to prasugrel metabolite (R-138727) in agonist-stimulated platelets. The selectivity and specificity of fentanyl toward major carrier proteins has been also studied.MethodsBlood was obtained from healthy volunteers (19 women and 12 men; mean age 40 ± 13 years). Platelet function was measured in whole blood, platelet-rich plasma and in suspensions of isolated platelets by flow cytometry, impedance and optical aggregometry. Surface plasmon resonance and molecular docking were employed to determine the binding kinetics of fentanyl to human albumin, α1-acid glycoprotein, apolipoprotein A-1 and apolipoprotein B-100.ResultsWhen applied at therapeutic and supratherapeutic concentrations under various experimental conditions, fentanyl had no potential to stimulate platelet activation and aggregation, or potentiate platelet response to ADP, nor did it affect platelet susceptibility to prasugrel metabolite in ADP-stimulated platelets. In addition, fentanyl was found to interact with all the examined carrier proteins with dissociation constants in the order of 10–4 to 10–9 M.ConclusionsIt does not seem that the delayed platelet responsiveness to oral P2Y12 inhibitors, such as prasugrel, in patients undergoing percutaneous coronary intervention, results from direct interactions between fentanyl and blood platelets. Apolipoproteins, similarly to albumin and α1-acid glycoprotein, appear to be important carriers of fentanyl in blood.

New Genetically Determined Markers of the Functional State of the Cardiovascular System.

Nowadays, cardiovascular diseases (CVDs) occupy a leading position in population mortality. Since it is known that the development of cardiovascular pathologies is determined mainly by the human genetic burden, an urgent task of primary prevention of CVDs is to assess the contribution of gene polymorphism to the formation of cardiovascular risk. The material for the study was the blood of volunteers aged 21 to 102 years. Polymorphisms were determined by real-time PCR. Multichannel volumetric sphygmography was performed to analyze the functional state of the vascular wall. The study revealed that the rs5742904 polymorphism of the ApoB gene was found to be absent in the studied groups of long-livers and descendants of long-livers. Results indicated that the carriage of the heterozygous variant of the MMP9 polymorphism is associated with a favorable prognosis for cardiovascular system functioning. A tendency towards an increase in the rate of biological age acceleration among subgroups with AA and GG genotypes of the MMP9 gene and a negative value of biological age acceleration among heterozygous carriers of this polymorphism allele were found. The conducted studies make it possible to identify new associations of the studied polymorphisms with the functional state of the cardiovascular system, which is of great clinical importance and requires further study.

Bayesian network analysis of panomic biological big data identifies the importance of triglyceride-rich LDL in atherosclerosis development.

We sought to explore biomarkers of coronary atherosclerosis in an unbiased fashion. We analyzed 665 patients (mean ± SD age, 56 ± 11 years; 47% male) from the GLOBAL clinical study (NCT01738828). Cases were defined by the presence of any discernable atherosclerotic plaque based on comprehensive cardiac computed tomography (CT). De novo Bayesian networks built out of 37,000 molecular measurements and 99 conventional biomarkers per patient examined the potential causality of specific biomarkers. Most highly ranked biomarkers by gradient boosting were interleukin-6, symmetric dimethylarginine, LDL-triglycerides [LDL-TG], apolipoprotein B48, palmitoleic acid, small dense LDL, alkaline phosphatase, and asymmetric dimethylarginine. In Bayesian analysis, LDL-TG was directly linked to atherosclerosis in over 95% of the ensembles. Genetic variants in the genomic region encoding hepatic lipase (LIPC) were associated with LIPC gene expression, LDL-TG levels and with atherosclerosis. Triglyceride-rich LDL particles, which can now be routinely measured with a direct homogenous assay, may play an important role in atherosclerosis development. GLOBAL clinical study (Genetic Loci and the Burden of Atherosclerotic Lesions); [https://clinicaltrials.gov/ct2/show/NCT01738828?term=NCT01738828&rank=1], identifier [NCT01738828].