Significant associations of common genetic variants in <i>KNG1</i> (NM_001102416.2) and <i>F11</i> (NM_000128.3) with plasma FXI levels.

<p>Significant associations of common genetic variants in <i>KNG1</i> (NM_001102416.2) and <i>F11</i> (NM_000128.3) with plasma FXI levels.</p

Plot of the association between variants within the <i>F11</i> locus with plasma FXI levels.

<p>Markers represented common variants organized by genomic position. Tthe diamond-shaped marker represented the top SNP (rs56810541) with the lowest variant-plasma FXI level association p-value. The left axis shows the statistical significance expressed as -log₁₀ of the p-values and colour intensities show the level of linkage disequilibrium between all variants and the top SNP. The recombination rate in the HapMap II sample [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0176301#pone.0176301.ref033" target="_blank">33</a>] for this region is measured on the right axis.</p

Plot of the collapsing method association in the <i>KNG1</i> locus with plasma FXI levels.

<p>Markers represented the mean position of low-frequency variant sets. All of the markers located above the dotted horizontal line obtained a p-value <0.05 after the collapsing method association. Diamond-shaped markers represented the five significant low-frequency variant sets with controlling FWER = 0.05. The biggest diamond-shaped marker is the top low-frequency variant set.</p

Plot of the collapsing method association in the <i>F11</i> locus.

<p>Markers represented the mean position of low-frequency variant sets. The diamond-shaped marker represented the top low-frequency variant set and markers located above the dotted horizontal line are the most significant low-frequency variant sets (p-value <0.05). None of the low-frequency variant sets were significantly associated after controlling FWER = 0.05.</p

Role of multimeric analysis of von Willebrand factor (VWF) in von Willebrand disease (VWD) diagnosis: Lessons from the PCM-EVW-ES Spanish project

<div><p>The multimeric analysis (MA) of plasma von Willebrand factor (VWF) evaluates structural integrity and helps in the diagnosis of von Willebrand disease (VWD). This assay is a matter of controversy, being considered by some investigators cumbersome and only slightly informative. The centralised study ‘Molecular and Clinical Profile of von Willebrand Disease in Spain (PCM-EVW-ES)’ has been carried out by including the phenotypic assessment and the genetic analysis by next generation sequencing (NGS) of the VWF gene (VWF). The aim of the present study was to evaluate the role of MA to the diagnosis of these patients and their potential discrepancies. Two hundred and seventy out of 480 patients centrally diagnosed with VWD had normal multimers, 168 had abnormal multimers and 42 a total absence of multimers. VWF MA was of great significance in the diagnosis of 83 patients (17.3%), it was also of help in the diagnosis achieved in 365 additional patients (76%) and was not informative in 32 cases (6.7%). With regard to discrepancies, 110 out of 480 (23%) patients centrally diagnosed with VWD presented some kind of discordance between VWF:RCo/VWF:Ag and/or VWF:CB/VWF:Ag ratios, multimeric study and/or genetic results. The VWF MA was key in the presence of novel mutations as well as in cases with phenotypic discrepancies. A comparison between the contribution of MA and VWF:CB showed a clearly higher contribution of the former in the diagnostic process. These data seem to reinforce the relevance of the VWF MA in VWD diagnosis, despite all its limitations.</p></div

Demographic data of the cohort of patients.

<p>Demographic data of the cohort of patients.</p

Multimeric analysis of von Willebrand factor (VWF) in low-resolution SDS-agarose gels in patients with type 2A VWD and some discrepancy.

<p>VWF from platelet lysate (NPt), plasmas of a normal subject (NP), patients with type 2A VWD and a patient with VWD type 2A (IIA) used as a control 2A are shown. <b>(a-b):</b> Patients with discrepancy between ratios and multimeric analysis; <b>(c-d):</b> Patients re-classified as type 2A on the basis of the genetic study.</p

Multimeric analysis of von Willebrand factor (VWF) in low-resolution SDS-agarose gels in patients with type 2A/2M VWD and some discrepancy.

<p>VWF from platelet lysate (NPt), plasmas of a normal subject (NP), patients with type 2A/2M VWD and a patient with VWD type 2A (IIA) used as a control 2A are shown. <b>(a-c)</b> Patients that showed discrepancy between ratios and multimeric analysis. In the case of the patients C30P012F07, C30P013F08 and C37P003F03 the mutations had not been described previously and the multimeric analysis (smeary) was of great significance to establish the diagnosis.</p

Evaluation of the contribution of different laboratory steps in the diagnosis definition.

<p>Four different successive laboratory assessment steps are considered. The progress in the diagnostic definition of the patients according to each step is shown. After the first and second steps, the multimeric analysis (MA) was of great significance in the diagnosis definition of 83 additional patients. Moreover, MA was in agreement with the diagnostic definition accomplished in steps 1 and 2 (179 patients). Finally, MA was also in agreement with the diagnostic definition achieved by molecular analysis in 186 additional patients. Thus, MA contributed to the diagnosis definition in a total of 448 (93.3%) patients.</p

Distribution of patients according to their coincidence between ratios and multimeric analysis before genetic study and between multimeric analysis and mutation after genetic study.

<p>Of 110 patients with some type of discrepancy, in 76 (48 + 28) the MA was in line with the molecular study while in 18 (14+4) patients there was not concordance. In the remaining 16, the similarity could not be demonstrated because the mutation found has not been described previously.</p