Identification of a variant hotspot in MYBPC3 and of a novel CSRP3 autosomal recessive alteration in a cohort of Polish patients with hypertrophic cardiomyopathy

INTRODUCTION Hypertrophic cardiomyopathy (HCM) is a heart disorder caused by autosomal dominant alterations affecting both sarcomeric genes and other nonsarcomeric loci in a minority of cases. However, in some patients, the occurrence of the causal pathogenic variant or variants in homozygosity, compound heterozygosity, or double heterozygosity has also been described. Most of the HCM pathogenic variants are missense and unique, but truncating mutations of the MYBPC3 gene have been reported as founder pathogenic variants in populations from Finland, France, Japan, Iceland, Italy, and the Netherlands. OBJECTIVES This study aimed to assess the genetic background of HCM in a cohort of Polish patients. PATIENTS AND METHODS Twenty–nine Polish patients were analyzed by a next–generation sequencing panel including 404 cardiovascular genes. RESULTS Pathogenic variants were found in 41% of the patients, with ultra–rare MYBPC3 c.2541C>G (p.Tyr847Ter) mutation standing for a variant hotspot and correlating with a lower age at HCM diagnosis. Among the nonsarcomeric genes, the CSRP3 mutation was found in a single case carrying the novel c.364C>T (p.Arg122Ter) variant in homozygosity. With this finding, the total number of known HCM cases with human CSRP3 knockout cases has reached 3

Defects of splicing in antithrombin deficiency

Background: There is increasing evidence supporting the relevance of aberrant splicing in multiple disorders. In antithrombin deficiency only 22 intronic mutations affecting splicing sites (7% of SERPINC1 mutations) are considered as splicing mutations. Methods: SERPINC1 was analyzed by Sanger sequencing and MLPA in 141 unrelated cases with antithrombin deficiency. Plasma antithrombin was studied by functional and western blot assays, purified by FPLC and characterized by proteomic analysis. In silico predictions on splicing was done with the Human Splicing Finder software. Results: We detected 89 different SERPINC1 defects, 13 with potential effect on splicing. Ten cases presented 9 mutations disturbing splicing sites, 5 new. Three gross or small gene defects also disturbed a correct splicing. Interestingly, the first duplication of a single exon ever described (c.1154-13_1218+115dup), caused mild deficiency (75%). A deeper intronic mutation (c.1154-14G>A), identified in three unrelated patients with traces of disulphide dimers of antithrombin in plasma, created a cryptic splicing site that might generate a variant with 4 additional in frame residues according to in silico predictions. This aberrant splicing was confirmed by proteomic analysis of the dimer purified from plasma. Conclusions: A high proportion of cases with antithrombin deficiency (up to 13%) may be explained by an aberrant splicing. Up to 15% of mutations in SERPINC1: splicing site variations, gross gene defects and deep intronic mutations, may affect a correct splicing with three potential consequences type I, type II, and even moderate antithrombin deficiency

Does Diabetes Accelerate the Progression of Aortic Stenosis through Enhanced Inflammatory Response within Aortic valves?

Diabetes predisposes to aortic stenosis (AS). We aimed to investigate if diabetes affects the expression of selected coagulation proteins and inflammatory markers in AS valves. Twenty patients with severe AS and concomitant type 2 diabetes mellitus (DM) and 40 well-matched patients without DM scheduled for valve replacement were recruited. Valvular tissue factor (TF), TF pathway inhibitor (TFPI), prothrombin, C-reactive protein (CRP) expression were evaluated by immunostaining and TF, prothrombin, and CRP transcripts were analyzed by real-time PCR. DM patients had elevated plasma CRP (9.2 [0.74–51.9] mg/l vs. 4.7 [0.59–23.14] mg/l, p = 0.009) and TF (293.06 [192.32–386.12] pg/ml vs. 140 [104.17–177.76] pg/ml, p = 0.003) compared to non-DM patients. In DM group, TF−, TFPI−, and prothrombin expression within valves was not related to demographics, body mass index, and concomitant diseases, whereas increased expression related to DM was found for CRP on both protein (2.87 [0.5–9]% vs. 0.94 [0–4]%, p = 0.01) and transcript levels (1.3 ± 0.61 vs. 0.22 ± 0.43, p = 0.009). CRP-positive areas were positively correlated with mRNA TF (r = 0.84, p = 0.036). Diabetes mellitus is associated with enhanced inflammation within AS valves, measured by CRP expression, which may contribute to faster AS progression

Association between the Interleukin-6 Promoter Polymorphism −174G/C and Serum Lipoprotein(a) Concentrations in Humans

Background: Lipoprotein(a) [Lp(a)] is an independent risk factor for cardiovascular disease. The interleukin-6 (IL-6) receptor antagonist tocilizumab has been shown to lower serum Lp(a) concentrations. We investigated whether the IL-6 single nucleotide polymorphism 2174G/C is associated with baseline serum Lp(a) concentrations. Methodology/Principal Findings: We divided 2321 subjects from the Lipid Analytic Cologne (LIANCO) cohort into 2 groups, the ones with substantially elevated Lp(a), defined as concentrations $60 mg/dl (n = 510), and the ones with Lp(a),60 mg/ dl (n = 1811). The association with the genotypes GG (33.7%), GC (50.75%) and CC (15.55%) was investigated. The GC and the CC genotype were associated with a significantly increased odds ratio of having substantially elevated Lp(a) concentrations (OR = 1.3, 95 % CI 1.04 to 1.63, P = 0.02 and OR = 1.44, 95 % CI 1.06 to 1.93, P = 0.018). These associations remained significant after adjusting for age, sex, smoking behavior, body mass index, serum lipoproteins, hypertension and diabetes. Of these covariates, only LDL cholesterol was significantly and independently associated with elevated Lp(a) concentrations. Conclusions/Significance: The IL-6 single nucleotide polymorphism 2174G/C is associated with increased odds of having elevated Lp(a). Whether this association plays a role in the Lp(a)-lowering effects of IL-6 receptor antagonists remains to b

Mast Cells in Human Stenotic Aortic Valves Are Associated with the Severity of Stenosis

Aortic valve stenosis (AS) is characterized by extensive calcification of the aortic valve leaflets and infiltration of inflammatory cells. Activated mast cells (MCs) may participate in the induction of fibrosis and calcification with ensuing valve stiffening. We sought to investigate whether the number of MCs within stenotic aortic valves is associated with the severity of AS. We studied 43 patients (19 men, 24 women) with dominant AS (age, 64.2±5.9 years; mean transvalvular pressure gradient, 62.11±24.47 mmHg) without atherosclerotic vascular disease, undergoing elective aortic valve replacement. MCs were detected in the excised valves by immunostaining. Aortic valves from five healthy subjects obtained on autopsy served as negative controls. The number of tryptase- and chymase-positive MCs was increased in AS valves compared with the control valves (6.9 [2.3-18.9]/$mm^{2}$ vs. 0.7 [0-2.2]/$mm^{2}$, P00.0001 and 3.2 [2.1-9.4]/$mm^{2}$ vs. 0.3 [0-1.9]/$mm^{2}$, P00.002, respectively). MCs that colocalized with macrophages and neovessels were detected mainly in the calcified regions of the leaflets. The number of MCs positively correlated with maximal (r00.73, P< 0.0001) and mean (r00.78, P<0.0001) gradients and maximal aortic jet velocity (r00.68, P00.0005). An inverse correlation with aortic valve area (r0-0.71, P00.0001) was also observed. Multivariate regression analysis revealed that MC number and valve thickness were significantly associated with mean transvalvular gradient ($R^{2}$00.74, P<0.000001) in AS patients. Increased MC number within human stenotic aortic valves is associated with the severity of AS

Hypoglycosylation is a common finding in antithrombin deficiency in the absence of a SERPINC1 gene defect

Essentials We investigated the molecular base of antithrombin deficiency in cases without SERPINC1 defects. 27% of cases presented hypoglycosylation, transient in 62% and not restricted to antithrombin. Variations in genes involved in N-glycosylation underline this phenotype. These results support a new form of thrombophilia. Click here to listen to Dr Huntington's perspective on thrombin inhibition by the serpins SUMMARY: Background Since the discovery of antithrombin deficiency, 50 years ago, few new thrombophilic defects have been identified, all with weaker risk of thrombosis than antithrombin deficiency. Objective To identify new thrombophilic mechanisms. Patients/methods We studied 30 patients with antithrombin deficiency but no defects in the gene encoding this key anticoagulant (SERPINC1). Results A high proportion of these patients (8/30: 27%) had increased hypoglycosylated forms of antithrombin. All N-glycoproteins tested in these patients (alpha1-antitrypsin, FXI and transferrin) had electrophoretic, HPLC and Q-TOF patterns indistinguishable from those of the congenital disorders of glycosylation (rare recessive multisystem disorders). However, all except one had no mental disability. Moreover, intermittent antithrombin deficiency and hypoglycosylation was recorded in five out of these eight patients, all associated with moderate alcohol intake. Genetic analysis, including whole exome sequencing, revealed mutations in different genes involved in the N-glycosylation pathway. Conclusions Our study provides substantial and novel mechanistic insights into two disease processes, with potential implications for diagnosis and clinical care. An aberrant N-glycosylation causing a recessive or transient antithrombin deficiency is a new form of thrombophilia. Our data suggest that congenital disorders of glycosylation are probably underestimated, especially in cases with thrombosis as the main or only clinical manifestation