Apolipoprotein B Research Articles

Abstract 13909: Use of Advanced Lipoprotein Testing in Real-World Data: A Cross Sectional Study Across Multiple Health Systems

Introduction: Per AHA guidelines, adults between 40-75 with borderline (5-7.5%) 10-year ASCVD risk (approximately 2.5% of this population in prior studies) should have the decision to start statin therapy personalized. Per AHA guidelines, elevated apolipoprotein B-100 (apoB) and elevated lipoprotein(a) (Lp(a)) are risk enhancing factors that may be considered in this decision. Hypothesis: We hypothesize that <10% of the estimated borderline 10-year ASCVD risk population between 40-75 years of age are tested for apo(B) or Lp(a). Methods: We performed a hypothesis-driven secondary analysis of a cross-sectional dataset of adults with LDL-c assessed in the most recent year of available data from 7 health systems participating in PaTH, a Partner Network in PCORnet. Participating sites reported summaries of demographics, diagnoses and lab testing for their health system populations as well as sub-cohorts of patients with prior ASCVD events, diabetes (DM), and a history of LDL > 190mg/dl. We evaluated the lifetime testing for apo(B) and Lp(a) in the whole population and subtracted out the testing in the high-risk cohorts. Results: From a population of 784,406 adults, we identified a cohort of 301,235 aged 40-75 without prior ASCVD event, DM, or LDL > 190. We estimated that 2.5% of this population (7,531) had borderline 10-year ASCVD risk. A total of 730 individuals were tested for apoB and 892 for Lp(a) across the entire non-high-risk cohort (0.2% and 0.3%). Assuming only the borderline risk group was tested, this represents testing rates of 9.7% for apoB and 11.8% for Lp(a), ranging across healthcare systems from 0-27% for apoB and 0-42% for Lp(a). Conclusions: Within the constraints of our population-based cross-sectional study, we found it plausible that >10% of the borderline-risk population utilized either apoB or Lp(a) testing for further risk stratification. Future work should be done to directly evaluate the use of these parameters in the borderline-risk population.

Abstract 14235: Hepatocyte Tissue Plasminogen Activator Limits Very-Low-Density Lipoprotein Production via Inhibiting ApoB-Microsomal Transfer Protein Interaction and ApoB Lipidation

Lower plasma tissue plasminogen activator (tPA), a key fibrinolytic enzyme, is associated with higher atherogenic cholesterol levels in humans, but the mechanism is unknown. Hepatocyte (HC) is the factory producing atherogenic apoB lipoproteins, also a major source of basal plasma tPA. Our hypothesis is that tPA limits apoB-VLDL production in HCs.Silencing tPA in HCs using AAV8-H1-sh-tPA in Ldlr -/- mice leads to increased plasma cholesterol, triglyceride, apoB, and exacerbated atherosclerosis compared with scramble-silenced controls. Fractionation of plasma lipoproteins by FPLC or ultracentrifugation shows increased cholesterol and apoB in VLDL and LDL fractions. Inhibiting VLDL hydrolysis by P407 leads to a faster plasma triglyceride-rising rate in HC-tPA-silenced wild type mice, suggesting higher hepatic VLDL production. In a pulse-chase assay, tPA-silenced human primary HC has higher apoB-associated radioactivity in cell medium and lysate, suggesting increased apoB production.Adequate apoB lipidation is required for its secretion. By electronic microscopy, the distribution of VLDL shifts to a larger diameter in plasma VLDL isolated from HC-tPA-silenced Ldlr -/- mice, suggesting higher lipid contents in VLDL. The endoplasmic reticulum (ER)-associated apoB is higher in tPA-silenced vs. control HCs, more prominent in the less denser density fractions. As density and lipidation are inversely related, these findings are consistent with the hypothesis that tPA limits apoB lipidation in ER. MTP is an ER chaperone incorporating neutral lipids onto apoB in HC. The tPA-silenced human primary HC has higher neutral lipid transfer activity than controls, despite similar MTP protein levels. Moreover, higher apoB is in the anti-MTP-precipitations from tPA-silenced vs. control HCs, suggesting that silencing tPA increases apoB-MTP interaction. Proximity ligation and immunoprecipitation assays show tPA interacts with apoB in HC. Solid-phase binding assay and native gel reveal purified human tPA binds to LDL, but not to MTP. tPA competes with MTP in binding to LDL. Moreover, tPA inhibits MTP-mediated neutral lipid transfer. These findings suggested tPA binds to apoB and inhibits MTP-dependent apoB lipidation and VLDL secretion.

Dawn-to-dusk dry fasting induces anti-atherosclerotic, anti-inflammatory, and anti-tumorigenic proteome in peripheral blood mononuclear cells in subjects with metabolic syndrome

BackgroundMetabolic syndrome characterized by abdominal obesity, high blood pressure, elevated fasting glucose and triglyceride levels and low high-density lipoprotein cholesterol level is associated with pro-inflammatory state, increased risk for atherosclerosis, and multiple cancers. Our previous results on subjects with metabolic syndrome showed that 4-week dawn-to-dusk (sunset) dry fasting resulted in significant changes in the serum proteome and improvement in several metabolic risk factors. Peripheral blood mononuclear cells (PBMC) proteomics is a powerful tool that can provide mechanistic insights into how dawn-to-dusk dry fasting affects protein expression in metabolic pathways at cellular level. In this study, we determined whether dawn-to-dusk dry fasting would induce favorable changes in PBMC proteome in subjects with metabolic syndrome, similar to the changes induced by dawn-to-dusk dry fasting in the same subjects' serum proteome. MethodsWe conducted a prospective study on subjects with metabolic syndrome and collected blood specimens before 4-week dawn-to-dusk dry fasting, at the end of 4-week dawn-to-dusk dry fasting, and one week after 4-week dawn-to-dusk dry fasting. We performed untargeted proteomics using nano ultra-high performance liquid chromatography-tandem mass spectrometry to assess the impact of 4-week dawn-to-dusk dry fasting on PBMC proteome. ResultsThere were 14 subjects with metabolic syndrome with a mean age of 59 who fasted from dawn to dusk (strict dry fasting without any liquid or food intake) for more than 14 h daily for 29 days. The quantitative proteome analysis showed that apolipoprotein B (APOB) gene protein products (GP) levels were downregulated and had the most statistical significance of the observed difference at the end of 4-week dawn-to-dusk dry fasting (P = 0.008) and one week after 4-week dawn-to-dusk dry fasting (P = 0.0004) compared with the levels before 4-week dawn-to-dusk dry fasting. The comparison between GP levels before and at the end of 4-week dawn-to-dusk dry fasting showed an alteration in the expression of genes associated with lipid and atherosclerosis pathway (P = 6.014e-4) and C-type lectin receptor signaling pathway (P = 1.064e-5). The genes that were differentially expressed in the lipid and atherosclerosis pathway were APOB (P = 0.008), CD36 (P = 0.040), CALM1, CALM2, CALM3 (P = 0.015), and HSPA8 (P = 0.047). One of the differentially expressed genes in the C-type lectin receptor signaling pathway was lymphocyte-specific protein 1 (LSP1), which showed an average of 19-fold increase at the end of 4-week dawn-to-dusk dry fasting compared with the GP levels before fasting (P = 0.004). Several GPs associated with tumor-suppressor effect (TUBB4B, LSP1, ACTR3B) were upregulated, and GPs associated with tumor-promoter effect (CD36, CALM1, CALM2, CALM3, FLOT2, PPIF) were downregulated at the end of 4-week dawn-to-dusk dry fasting or one week after 4-week dawn-to-dusk dry fasting compared with the GP levels before 4-week dawn-to-dusk dry fasting. ConclusionBased on our results, we conclude that in subjects with metabolic syndrome, 4-week dawn-to-dusk dry fasting induced anti-atherosclerotic, anti-inflammatory, and anti-tumorigenic PMBC proteome. Randomized, controlled clinical trials are needed to further investigate the effect of dawn-to-dusk dry fasting on subjects with chronic metabolic diseases and metabolic syndrome-induced cancers.

Lp(a), oxidized phospholipids and oxidation-specific epitopes are increased in subjects with keloid formation

BackgroundKeloid formation following trauma or surgery is common among darkly pigmented individuals. Since lipoprotein(a) [Lp(a)] has been postulated to have a putative role in wound healing, and also mediates atherosclerotic cardiovascular disease, it was assessed whether Lp(a), its associated oxidized phospholipids and other oxidation-specific biomarkers were associated with keloid formation.MethodsThis case-control study included darkly pigmented individuals of African ancestry, 100 with keloid scarring and 100 non-keloid controls. The lipid panel, hsCRP, Lp(a), oxidized phospholipids on apolipoprotein B-100 (OxPL-apoB), IgG and IgM apoB-immune complexes and IgG and IgM autoantibodies to a malondialdehyde mimotope (MDA-mimotope) were measured. Immunohistochemistry of keloid specimens was performed for both Lp(a) and OxPL staining.ResultsCases and controls were well matched for age, sex and lipid profile. Mean Lp(a) (57.8 vs. 44.2 mg/dL; P = 0.01, OxPL-apoB 17.4 vs. 15.7 nmol/L; P = 0.009) and IgG and IgM apoB-immune complexes and IgG and IgM MDA-mimotope levels were significantly higher in keloid cases. Keloid tissue stained strongly for OxPL.ConclusionDarkly pigmented individuals of African ancestry with keloids have higher plasma levels of Lp(a), OxPL-apoB and oxidation-specific epitopes. The commonality of excessive wound healing in keloids and chronic complications from coronary revascularization suggests avenues of investigation to define a common mechanism driven by Lp(a) and the innate response to oxidized lipids.

The Search for Associations of Serum Proteins with the Presence of Unstable Atherosclerotic Plaque in Coronary Atherosclerosis.

To study the associations of blood proteins with the presence of unstable atherosclerotic plaques in the arteries of patients with coronary atherosclerosis using quantitative proteomics. The studies involved two groups of men with coronary atherosclerosis (group 1 (St) had only stable atherosclerotic plaques; group 2 (Ns) had only unstable atherosclerotic plaques, according to histological analysis of tissue samples); the average age of patients was 57.95 ± 7.22. Protein concentrations in serum samples were determined using the PeptiQuant Plus Proteomics Kit. The identification of protein fractions was carried out by monitoring multiple reactions on a Q-TRAP 6500 mass spectrometer combined with a liquid chromatograph. Mass spectrometric identification revealed in serum samples from patients with unstable atherosclerotic plaques a reduced concentration of proteins in the blood: α-1-acid glycoprotein, α-1-antichymotrypsin, α-1-antitrypsin, ceruloplasmin, hemopexin, haptoglobin, apolipoprotein B-100, apolipoprotein L1, afamin and complement component (C3, C7, C9). Moreover, at the same time a high concentration complements factor H and attractin. The differences were considered significant at p < 0.05. It was found that the instability of atherosclerotic plaques is associated with the concentration of proteins: afamin, attractin, components of the complement system, hemopexin and haptoglobin. The data of our study showed the association of some blood proteins with the instability of atherosclerotic plaques in coronary atherosclerosis. Their potential role in the development of this disease and the possibility of using the studied proteins as biomarkers requires further research.

Genetic and molecular architecture of familial hypercholesterolemia.

Atherosclerotic cardiovascular disease is the leading cause of death globally. Despite its important risk of premature atherosclerosis and cardiovascular disease, familial hypercholesterolemia (FH) is still largely underdiagnosed worldwide. It is one of the most frequently inherited diseases due to mutations, for autosomal dominant forms, in either of the LDLR, APOB, and PCSK9 genes or possibly a few mutations in the APOE gene and, for the rare autosomal forms, in the LDLRAP1 gene. The discovery of the genes implicated in the disease has largely helped to improve the diagnosis and treatment of FH from the LDLR by Brown and Goldstein, as well as the introduction of statins, to PCSK9 discovery in FH by Abifadel etal., and the very rapid availability of PCSK9 inhibitors. In the last two decades, major progress has been made in clinical and genetic diagnostic tools and the therapeutic arsenal against FH. Improving prevention, diagnosis, and treatment and making them more accessible to all patients will help reduce the lifelong burden of the disease.

LPA disruption with AAV-CRISPR potently lowers plasma apo(a) in transgenic mouse model: A proof-of-concept study

Lipoprotein(a) (Lp(a)) represents a unique subclass of circulating lipoprotein particles and consists of an apolipoprotein(a) (apo(a)) molecule covalently bound to apolipoprotein B-100. The metabolism of Lp(a) particles is distinct from that of low-density lipoprotein (LDL) cholesterol, and currently approved lipid-lowering drugs do not provide substantial reductions in Lp(a), a causal risk factor for cardiovascular disease. Somatic genome editing has the potential to be a one-time therapy for individuals with extremely high Lp(a). We generated an LPA transgenic mouse model expressing apo(a) of physiologically relevant size. Adeno-associated virus (AAV) vector delivery of CRISPR-Cas9 was used to disrupt the LPA transgene in the liver. AAV-CRISPR nearly completely eliminated apo(a) from the circulation within a week. We performed genome-wide off-target assays to determine the specificity of CRISPR-Cas9 editing within the context of the human genome. Interestingly, we identified intrachromosomal rearrangements within the LPA cDNA in the transgenic mice as well as in the LPA gene in HEK293T cells, due to the repetitive sequences within LPA itself and neighboring pseudogenes. This proof-of-concept study establishes the feasibility of using CRISPR-Cas9 to disrupt LPA invivo, and highlights the importance of examining the diverse consequences of CRISPR cutting within repetitive loci and in the genome globally.

Physiological Bases for the Superiority of Apolipoprotein B Over Low-Density Lipoprotein Cholesterol and Non-High-Density Lipoprotein Cholesterol as a Marker of Cardiovascular Risk.

In 2019, the European Society of Cardiology/European Atherosclerosis Society stated that apolipoprotein B (apoB) was a more accurate marker of cardiovascular risk than low-density lipoprotein cholesterol (LDL-C) and non-high-density lipoprotein cholesterol. Since then, the evidence has continued to mount in favor of apoB. This review explicates the physiological mechanisms responsible for the superiority of apoB as a marker of the cardiovascular risk attributable to the atherogenic apoB lipoprotein particles chylomicron remnants, very low-density lipoprotein, and low-density lipoprotein particles. First, the nature and relative numbers of these different apoB particles will be outlined. This will make clear why low-density lipoprotein particles are almost always the major determinants of cardiovascular risk and why the concentrations of triglycerides and LDL-C may obscure this relation. Next, the mechanisms that govern the number of very low-density lipoprotein and low-density lipoprotein particles will be outlined because, except for dysbetalipoproteinemia, the total number of apoB particles determines cardiovascular risk, Then, the mechanisms that govern the cholesterol mass within very low-density lipoprotein and low-density lipoprotein particles will be reviewed because these are responsible for the discordance between the mass of cholesterol within apoB particles, measured either as LDL-C or non-high-density lipoprotein cholesterol, and the number of apoB particles measured as apoB, which creates the superior predictive power of apoB over LDL-C and non-high-density lipoprotein cholesterol. Finally, the major apoB dyslipoproteinemias will be briefly outlined. Our objective is to provide a physiological framework for health care givers to understand why apoB is a more accurate marker of cardiovascular risk than LDL-C or non-high-density lipoprotein cholesterol.

Inhibition on XBP1s-driven lipogenesis by Qushi Huayu Decoction contributes to amelioration of hepatic steatosis induced by fructose

Ethnopharmacological relevanceQushi Huayu Decoction (QHD) is a traditional Chinese medicine formula consisting of five herbs, which has been used for non-alcoholic fatty liver disease (NAFLD) treatment in clinic for decades in China and validated in several NAFLD animal models. The hepatic de novo lipogenesis (DNL) is enhanced greatly to contribute to steatosis in NAFLD. The spliced form of X-box binding protein 1 (XBP1s) initiates DNL independently of sterol regulatory element-binding protein (SREBP) and carbohydrate-responsive element-binding protein (ChREBP). Aim of the studyTo disclose the mechanism of inhibition on hepatic DNL by QHD and the responsible compounds. MethodsThe effects of QHD on hepatic DNL were evaluated in mice induced by high-fructose diet (HFru). The effects of the serum-absorbed compounds of QHD on XBP1s were evaluated in HepG2 cells induced by tunicamycin. Hepatic histology, triglyceride (TG) and nonesterified fatty acids were observed. Hepatic apolipoprotein B100 and very low-density lipoprotein were measured to reflect lipid out-transport. The mRNA expression of XBP1s and its target genes were detected by real-time polymerase chain reaction. The protein expression of TG synthetases and DNL enzymes, and inositol requirement enzyme 1 alpha (IRE1α), phosphorylated IRE1α and XBP1s were detected in liver tissue and HepG2 cells by western-blot. The binding activity of SREBP1, protein expression of ChREBP and XBP1s were detected in the nuclear extracts of liver tissue. ResultsDynamical observing suggested feeding with HFru for 2 weeks was sufficient to induce hepatic lipogenesis and XBP1s. QHD ameliorated liver steatosis without enhancing out-transport of lipids, accompanied with more inhibitory effects on DNL enzymes than TG synthetases. QHD inhibits the nuclear XBP1s without affecting ChREBP and SREBP1. In QHD, chlorogenic acid, geniposide and polydatin inhibit lipogenesis initiated by XPB1s. ConclusionQHD probably decreases hepatic DNL by inhibiting XBP1s independent of SREBP1 and ChREBP. Chlorogenic acid, geniposide and polydatin are the potential responsible compounds.

Effect of synbiotic bread containing lactic acid on blood lipids and apolipoproteins in patients with type 2 diabetes: Arandomized controlled trial.

Recently, the use of synbiotics for managing various diseases has dramatically increased. Synbiotics have been shown to be a good approach to influence the composition of the gut microbiota with positive health effects. Management of type 2 diabetes mellitus (T2DM) complications is one of the reasons for the ingestion of synbiotics and so the aim of the current study was to determine the effects of synbiotic bread intake on markers of lipid profile in T2DM patients. One hundred T2DM patients (age between 20 and 60 years) were randomly assigned to four groups to consume different types of synbiotic bread, three times/day, for 8 weeks: "synbiotic + lactic acid" (n= 25; IV), "synbiotic" (n= 25; III), "lactic acid brad" (n= 25; II), or "control" (n= 25; I). The measured outcomes included anthropometric characteristics, glycemic control parameters, blood lipids, and apolipoproteins. The consumption of "synbiotic + lactic acid bread" (group IV) and "lactic acid bread" (group II) led to a significant decrease in total cholesterol (TC) and glycated hemoglobin (HbA1c) compared to the "control bread." The HbA1c levels were also significantly lower when compared to group II. Additionally, apolipoprotein A (Apo A1) levels were significantly decreased in group IV, compared to control and other groups (post hoc analysis). No significant differences between groups were observed for triglyceride (TG), high-density lipoprotein (HDL), low-density lipoprotein (LDL), and apolipoprotein B100 (Apo B100) levels. The observed results show that the synbiotic bread (with or without lactic acid) promoted a decrease in total cholesterol (TC) and Apo A1 in diabetic patients when consumed daily for 8 weeks.

Hepatic inactivation of murine Surf4 results in marked reduction in plasma cholesterol.

PCSK9 negatively regulates low-density lipoprotein receptor (LDLR) abundance on the cell surface, leading to decreased hepatic clearance of LDL particles and increased levels of plasma cholesterol. We previously identified SURF4 as a cargo receptor that facilitates PCSK9 secretion in HEK293T cells (Emmer et al., 2018). Here, we generated hepatic SURF4-deficient mice (Surf4fl/fl Alb-Cre+) to investigate the physiologic role of SURF4 in vivo. Surf4fl/fl Alb-Cre+ mice exhibited normal viability, gross development, and fertility. Plasma PCSK9 levels were reduced by ~60% in Surf4fl/fl Alb-Cre+ mice, with a corresponding ~50% increase in steady state LDLR protein abundance in the liver, consistent with SURF4 functioning as a cargo receptor for PCSK9. Surprisingly, these mice exhibited a marked reduction in plasma cholesterol and triglyceride levels out of proportion to the partial increase in hepatic LDLR abundance. Detailed characterization of lipoprotein metabolism in these mice instead revealed a severe defect in hepatic lipoprotein secretion, consistent with prior reports of SURF4 also promoting the secretion of apolipoprotein B (APOB). Despite a small increase in liver mass and lipid content, histologic evaluation revealed no evidence of steatohepatitis or fibrosis in Surf4fl/fl Alb-Cre+ mice. Acute depletion of hepatic SURF4 by CRISPR/Cas9 or liver-targeted siRNA in adult mice confirms these findings. Together, these data support the physiologic significance of SURF4 in the hepatic secretion of PCSK9 and APOB-containing lipoproteins and its potential as a therapeutic target in atherosclerotic cardiovascular diseases.

P037: High breadth whole exome sequencing of circulating tumor DNA identifies novel recurrent genetic alterations in Hodgkin lymphoma

Hodgkin lymphoma (HL) is a B-cell lymphoma with a generally favorable prognosis. However, side effects of aggressive front-line treatment and difficult-to-treat relapsed/refractory disease are still a clinical challenge. A better understanding of the biological diversity of HL at baseline might help to further individualize the treatment of patients and thus ultimately improve outcome. A broad characterization of the genetic drivers of HL is highly important to improve our understanding of the biological diversity of HL. Here, we present the results of our study using whole-exome sequencing of circulating tumor (ct)DNA to broadly characterize the HL oncogenome in 165 patients. After extraction of germline and cell-free DNA, WES was performed, targeting 37 Mb of the human genome. An in-house customized bioinformatic pipeline including twofold error reduction using unique molecular identifiers (UMIs) and digital error suppression was used as previously described by us (Sobesky et. al., Med, 2021). The median ctDNA concentration per patient was 2.26 log haploid genome equivalents per milliliter (log hGE/ml) (range: 1.41–4.18 log hGE/ml) without significant correlation with clinical parameters such as sex, international prognostic score (IPS) or clinical stage. In addition to the detection of several genetic aberrations involved in the pathogenesis of HL (such as B2M, CSF2RB, GNA13, SOCS1), we were able to detect several novel, recurrently mutated potential oncogenes and tumor suppressor genes. Among the most frequently mutated genes not previously described in HL were olfactory receptor family 10 subfamily G member 7 (OR10G7) (11.5%), Apolipoprotein B (APOB) (10.3%) and Filaggrin 2 (FLG2) (9.7%). Furthermore, several previously undescribed copy number variations (CNV) were detected (losses: 13q13.1, 1p36.13a, 6p21.33; gains: 19q13.42, 9p24, 12q12). To decipher the detected genes’ and CNVs’ contribution to the biological diversity of HL, we performed non-negative matrix factorization. Furthermore, neoantigen load was assessed through human-leukocyte antigen (HLA) sequencing. Detailed results will be presented at the meeting. In conclusion, our ctDNA based WES approach provides sufficient sensitivity to both perform genotyping in HL and detecting novel genomic aberrations potentially involved in the pathogenesis of HL.

A Low-Frequency APOB p.(Pro955Ser) Variant Contributes to the Severity of/Variability in Familial Hypercholesterolemia.

Heterozygous familial hypercholesterolemia (HeFH) is caused by a rare pathogenic variant in the LDLR, APOB, and PCSK9 genes. However, the causative variants in these genes have not been identified in approximately 40% of HeFH patients. Our aim was to identify novel (or additional) genes/variants that contribute to HeFH. Whole-exome sequencing was performed for 215 family members from 122 families with HeFH without pathogenic variants in the LDLR or PCSK9 genes. We could not find novel causative familial hypercholesterolemia (FH) genes/variants by family analysis. Next, we examined all APOB variants. Twenty-four nonsynonymous APOB variants were identified. The allele frequencies of the c.2863C > T:p.(Pro955Ser) variant in the HeFH probands and the general Japanese population were 0.15 and 0.034, respectively [odds ratio 4.9 (95% CI 3.4-7.1); P = 6.9 × 10-13]. The patients harboring the c.2863C > T:p.(Pro955Ser) variant accounted for 9.8% (n = 63) of unrelated patients with HeFH (n = 645). The penetrance of the c.2863C > T:p.(Pro955Ser) variant was low in the pedigree-based genetic analysis. In an in vitro assay, low-density lipoprotein (LDL) uptake from patients with the homozygous c.2863C > T:p.(Pro955Ser) variant was 44% of the LDL uptake from control subjects, and it was similar to that of the LDL uptake from patients with the known pathogenic heterozygous p.(Arg3527Gln) variant. The low-frequency APOB c.2863C > T:p.(Pro955Ser) variant is not an FH-causative variant, but it has a moderate effect size in HeFH. These findings suggest that the combination of the APOB c.2863C > T:p.(Pro955Ser) variant and age, environmental factors, or other genetic factors contributes to the severity of or variability in the HeFH phenotype.

Lipoprotein(a) and cardiovascular outcomes in patients with coronary artery disease and impaired glucose metabolism

Abstract Introduction Lipoprotein(a) [Lp(a)] is an LDL-like molecule composed of a part of apolipoprotein(a) bounding covalently to apolipoprotein B-100. High plasma Lp(a) levels were associated with MACE in stable CAD patients. Recent research shows contradictory results in stable CAD patients with high Lp(a) plasmatic levels and impaired glucose metabolism in MACE occurrence. Purpose Investigate whether high Lp(a) levels were associated with MACE in CAD patients with impaired glucose metabolism, at an extended follow-up. Methods A prospective cohort of 1,127 CAD patients with impaired glucose metabolism (pre-diabetes and diabetes) was observed during 4.9±3.4 years. Pre-diabetes was considered when fasting plasma glucose ranged from 5.6 to 6.9 mmol/L, or hemoglobin A1c levels ranging from 5.7 to 6.4%. Lp(a) levels ≥30 mg/dL were considered high. Bivariate and multivariate Cox regression analysis evaluated the risk of Lp(a) ≥30 mg/dL for MACE occurrence. Kaplan-Meier curves estimated the survival probability for high and low Lp(a) levels. Results Of the patients with Lp(a) levels ≥30, 44.4% presented MACE and 32.0% had no MACE (p<0.0001). Cox regression analysis with smoking, hypertension, dyslipidemia, physical inactivity and kidney failure (creatinine clearance <60 mL/min) showed that high Lp(a) remained in the equation as an independent risk factor for MACE (HR=1.24; p=0.031). The Kaplan-Meier showed, at 10-year' follow-up, a better survival in the group with lower Lp(a) levels (p=0.023). Conclusion Our study demonstrated that high Lp(a) levels were an independent predictor of MACE and cardiovascular mortality in a CAD population with impaired glucose metabolism. Lp(a) measurement may help further risk stratification for diabetes and pre-diabetes patients suffering CAD. With the recent development of drugs that selectively lower Lp(a) levels, this marker can become a clinical target for reducing CVD risk. Funding Acknowledgement Type of funding sources: Public hospital(s). Main funding source(s): SESARAM EPERAM

Association of Lipoprotein A, a Predisposing Risk Factor for In-stent Restenosis

Abstract
 Elevated plasma lipoprotein (A) concentration is considered as a predisposing risk factor for cardiovascular disease and in-stent restenosis. Lipoprotein (A) is a lipoprotein with an LDL-like particle that contains Apo-lipoprotein B100 and is associated with cardiovascular complications. The most prevalent complication associated with percutaneous coronary intervention is in-stent restenosis.In-stent Restenosis affected 40% of all angioplasties in the pre-stent era, then 28% in the (BMS) era. Furthermore, the introduction of (DES) second generation and drug-coated balloons has further reduced restenosis rates to less than 10%. The aim of this review paper is to investigate the clinical association of Lipoprotein (A) concentration in the occurrence of in-stent restenosis. Our paper possesses some limitations Firstly, it does not explain how lipoprotein concentrations are established. Secondly, the mechanism underlying the lipoprotein (A) lowering effect of current therapy.
 Keywords: Lipoprotein A, in-stent restenosis, predisposing factor, Diabetes mellitus, Cardio-vascular disease.

Identification of Genomic Regions and Candidate Genes Associated with Body Weight and Body Conformation Traits in Karachai Goats.

The objective of this study was to identify the SNPs and candidate genes related to body weight and seven body conformation traits at the age of 8 months in the Russian aboriginal Karachai goats (n = 269) by conducting genome-wide association studies (GWAS), using genotypes generated by Goat SNP BeadChip (Illumina Inc., USA). We identified 241 SNPs, which were significantly associated with the studied traits, including 47 genome-wide SNPs (p < 10−5) and 194 suggestive SNPs (p < 10−4), distributed among all goat autosomes except for autosome 23. Fifty-six SNPs were common for two and more traits (1 SNP for six traits, 2 SNPs for five traits, 12 SNPs for four traits, 20 SNPs for three traits, and 21 SNPs for two traits), while 185 SNPs were associated with single traits. Structural annotation within a window of 0.4 Mb (±0.2 Mb from causal SNPs) revealed 238 candidate genes. The largest number of candidate genes was identified at Chr13 (33 candidate genes for the five traits). The genes identified in our study were previously reported to be associated with growth-related traits in different livestock species. The most significant genes for body weight were CRADD, HMGA2, MSRB3, MAX, HACL1 and RAB15, which regulate growth processes, body sizes, fat deposition, and average daily gains. Among them, the HMGA2 gene is a well-known candidate for prenatal and early postnatal development, and the MSRB3 gene is proposed as a candidate gene affecting the growth performance. APOB, PTPRK, BCAR1, AOAH and ASAH1 genes associated with withers height, rump height and body length, are involved in various metabolic processes, including fatty acid metabolism and lipopolysaccharide catabolism. In addition, WDR70, ZBTB24, ADIPOQ, and SORCS3 genes were linked to chest width. KCNG4 was associated with rump height, body length and chest perimeter. The identified candidate genes can be proposed as molecular markers for growth trait selection for genetic improvement in Karachai goats.

A New Structural Model of Apolipoprotein B100 Based on Computational Modeling and Cross Linking.

ApoB-100 is a member of a large lipid transfer protein superfamily and is one of the main apolipoproteins found on low-density lipoprotein (LDL) and very low-density lipoprotein (VLDL) particles. Despite its clinical significance for the development of cardiovascular disease, there is limited information on apoB-100 structure. We have developed a novel method based on the “divide and conquer” algorithm, using PSIPRED software, by dividing apoB-100 into five subunits and 11 domains. Models of each domain were prepared using I-TASSER, DEMO, RoseTTAFold, Phyre2, and MODELLER. Subsequently, we used disuccinimidyl sulfoxide (DSSO), a new mass spectrometry cleavable cross-linker, and the known position of disulfide bonds to experimentally validate each model. We obtained 65 unique DSSO cross-links, of which 87.5% were within a 26 Å threshold in the final model. We also evaluated the positions of cysteine residues involved in the eight known disulfide bonds in apoB-100, and each pair was measured within the expected 5.6 Å constraint. Finally, multiple domains were combined by applying constraints based on detected long-range DSSO cross-links to generate five subunits, which were subsequently merged to achieve an uninterrupted architecture for apoB-100 around a lipoprotein particle. Moreover, the dynamics of apoB-100 during particle size transitions was examined by comparing VLDL and LDL computational models and using experimental cross-linking data. In addition, the proposed model of receptor ligand binding of apoB-100 provides new insights into some of its functions.

Effect of APOB gene polymorphisms on body mass index, blood pressure, and total cholesterol levels: A cross-sectional study in Mexican population.

APOB gene polymorphisms are considered risk factors for the development of dyslipidemia, hypertension, and cardiovascular disease (CVD) in several populations. In Mexico, these pathologies are frequent and studies regarding this gene are scarce. The aim of this cross-sectional study was to determined genotype, allele, and haplotype frequencies of APOB polymorphisms and performed analyses of association among the biochemical, hemodynamic, anthropometrical, and genetic variables. Blood samples were taken from 361 subjects from unselected Mexican population for biochemical analysis and for deoxyribonucleic acid extraction; besides blood pressure and body mass index (BMI) were measured. APOB polymorphisms rs934197, rs533617, rs693, and rs1042031 were genotyped by polymerase chain reaction (PCR)-restriction fragment length polymorphism; whereas, rs17240441 and c.66_67insCTGCTG were genotyped by PCR followed by electrophoresis. Genotype and allele frequencies were obtained by simple counting and deviations from Hardy-Weinberg equilibrium (HWE) were calculated by chi-square test. The effect of the polymorphisms on the quantitative variables was determined using analysis of variance, Student’s t test, Pearson’s and Spearman’s correlations and multiple linear regression models. All the polymorphisms were within HWE. Frequencies of mutated alleles were highly heterogeneous: rs934197-T 33.6%, rs17240441-D 39.3%, c.66_67insCTGCTG-I 3.9%, rs533617-G 0.9%, rs693-T 40.5%, and rs1042031-G 17.3%. Chronic degenerative diseases were frequent in the studied population: overweight-obesity 55.1%, dyslipidemia 45.8%, and hypertension 23.5%. The association analyses showed that despite adjustments for age and sex the mutated alleles rs934197-T, rs1042031G, c.66_67-insCTGCTG-I, and rs533617-G, were related to lower values of BMI, total cholesterol (TC), systolic blood pressure, and diastolic blood pressure, respectively. All polymorphisms analyzed except rs533517 and c.66_67insCTGCTG showed high frequencies of the mutated allele, making them useful for association studies. Our results revealed that, APOB gene polymorphisms could be contributing to the development of several chronic diseases, such as essential hypertension, dyslipidemias, obesity, among others. However, specific studies with each pathology are needed to know the possible implications of the polymorphisms.

Association between Genetic Polymorphisms and Bleeding in Patients on Direct Oral Anticoagulants

Objectives: The purpose of our study is to investigate the effects of apolipoprotein B (APOB) and APOE gene polymorphisms on bleeding complications in patients receiving direct oral anticoagulants (DOACs). Methods: A total of 16 single nucleotide polymorphisms (SNPs) in 468 patients were genotyped. Six SNPs of ABCB1 (rs3842, rs1045642, rs2032582, rs1128503, rs3213619, and rs3747802), one SNP of CYP3A5 (rs776746), seven SNPs of APOB (rs1042034, rs2163204, rs693, rs679899, rs13306194, rs13306198, and rs1367117), and two SNPs of APOE (rs429358 and rs7412) were analyzed by a TaqMan genotyping assay. Multivariable logistic regression analysis with selected variables was performed for the construction of a risk scoring system. Two risk scoring systems were compared (demographic factors only vs. demographic factors and genetic factors). Results: In the multivariable analyses, two models were constructed; only demographic factors were included in Model I and both demographic factors and genetic factors in Model II. Rivaroxaban and anemia showed significant association with bleeding in both models. Additionally, ABCB1 rs3842 variant homozygote carriers (CC) and APOB rs13306198 variant allele carriers (AG, AA) had a higher risk of bleeding risk compared with that of wild-type allele carriers (TT, TC) and wild-type homozygote carriers (GG), respectively. Whereas the area under the receiver operating characteristic curve (AUROC) value using demographic factors only was 0.65 (95% confidence interval (CI): 0.56–0.74), the AUROC increased to 0.72 by adding genetic factors (95% CI: 0.65–0.80). The predicted bleeding risks of bleeding in patients with 0, 1, 2, 3, 4, 5, 6, 7 and 8 points from the logistic regression curve were 0.8%, 2.0%, 5.4%, 5.2%, 12.5%, 26.9%, 47.0%, 64.3% and 82.3%, respectively. Conclusions: The study results can be used for enhancing individualized treatment strategies in patients taking DOACs, helping clinicians predict the bleeding risk.

Physiological interindividual variability in endogenous estradiol concentration does not influence adipose tissue and hepatic lipid kinetics in women.

Increased triglyceride (TG) and apolipoprotein B-100 (apoB-100) concentrations in plasma are important risk factors for cardiovascular disease in women. Administration of some estrogen preparations raises plasma TG and apoB-100 concentrations by increasing hepatic very low-density lipoprotein (VLDL) TG and apoB-100 secretion rates. However, the influence of physiological variation in endogenous estradiol on VLDL-TG and VLDL-apoB-100 metabolism and on free fatty acid (FFA) release into plasma (the major source of fatty acids for VLDL-TG production) is not known. We measured basal VLDL-TG, VLDL-apoB-100, and plasma FFA kinetics by using stable isotopically labeled tracers in 36 eumenorrheic, premenopausal women (age: 33 ± 2 years, BMI: 31 ± 1 kg/m2; mean ± s.e.m.) during the follicular phase of the menstrual cycle; participants were divided into two groups based on low (n = 18) or high (n = 18) plasma estradiol concentrations (defined as below or above the median value of 140 pmol/L in the whole group). Mean plasma estradiol concentration was >3-fold higher in the high-estradiol than in the low-estradiol group (299 ± 37 and 96 ± 7 pmol/L, P < 0.001); there was no difference in plasma progesterone concentrations between the two groups (P = 0.976). There were no significant differences in plasma FFA concentration, FFA rate of appearance in plasma, VLDL-TG and VLDL-apoB-100 concentrations, hepatic VLDL-TG and VLDL-apoB-100 secretion rates, VLDL-TG and VLDL-apoB-100 plasma clearance rates, and mean residence times (all P ≥ 0.45). No significant associations were found between plasma estradiol concentration and FFA, VLDL-TG, and VLDL-apoB-100 concentrations and kinetics (all P > 0.19). Plasma estradiol concentration is not an important correlate of basal plasma FFA, VLDL-TG, and VLDL-apoB-100 kinetics in premenopausal women.