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Congenital Factor XI (FXI) deficiency shows a high variability in clinical phenotype. To date, many allele variants have been shown to cause this bleeding disorder. However, the genotype-phenotype relationship is difficult to establish. This report provides insights into this bleeding disorder. Sixteen unrelated Italian index cases with congenital FXI deficiency and their relatives were investigated. After the identification of the deficiency, we obtained DNA from each subject and analyzed the FXI gene using direct sequencing. We identified 5 and 11 individuals with severe and moderate deficiency of FXI activity, respectively. Most patients (8/16) carried mutations in the Apple 2 domain and 4 patients showed c.403G>T (p.Glu135*; type II mutation). Four novel compound heterozygosities were identified. Bleeding symptoms were present in two severely deficient subjects carrying the combinations c.901T>C (p.Phe301Leu)/c.1556G>A (p.Trp519*) and c.943G>A (p.Glu315)/c.1556G>A (p.Trp519*), respectively. Bleeding episodes were also observed in the presence of a moderate deficiency in two individuals heterozygous for c.449C>T (p.Thr150Met) and c.1253G>T (p.Gly418Val), respectively. One novel mutation, c.1682C>A (p.Ala561Asp), was identified as potentially deleterious in an asymptomatic individual. We confirm an unclear prediction of phenotype from mutational data. The FXI levels should be coupled with FXI analysis for a more comprehensive prediction of the bleeding phenotype in FXI deficiency., Competing Interests: The authors declare no conflict of interest.

Background: The JAK2 V617F mutation is an independent risk factor for MPN and SVT. Gender-related differences in MPN distribution have been reported and, recently, variability in the JAK2 V617F allele burden between sexes has been suggested. We wondered whether gender would modulate the role of the JAK2 V617F mutation as susceptibility risk factor for SVT., Materials and Methods: In 180 patients presenting with SVT, medical history was collected. The presence of the JAK2 V617F mutation and 46/1 haplotype was determined by polymerase chain reaction followed by TaqMan SNP genotyping assays., Results: Among patients with SVT, 43 (23.9%; 95%-CI: 18.2-30.7) carried the JAK2 V617F mutation. The JAK2 V617F mutation was found more frequently in women (29/95: 30.5%; 95%-CI: 22.1-40.4) than in men (14/85: 16.5%; 95%-CI: 10.0-25.9; OR: 2.2; 95%-CI: 1.1-4.5). The distribution of 46/1 haplotype frequencies did not differ significantly between men and women. In women carrying the rs12343867 CC genotype, the frequency observed for the occurrence of the V617F mutation was significantly higher than that observed in those not carrying (60.0% [95% CI: 31.2-83.3] vs. 26.8% [95% CI: 18.4-37.4]; OR: 4.1; 95%-CI: 1.1-14.9). In men, a similar prevalence was found among carriers of the rs12343867 CC genotype (16.7% [95% CI: 3.5-46.0]) and in non carriers (16.4% [95% CI: 9.3-27.2]). The V617F allele burden was unrelated to clinical characteristics and significantly higher in carriers of the rs12343867 CC genotype., Conclusions: Present findings suggest that, in patients presenting with SVT, the JAK2 V617F mutation is frequently found in women and, possibly by interacting with the 46/1 haplotype, may represent a gender-related susceptibility allele for SVT., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

Background and Objectives: Although a large number of gene mutations have been characterized in patients with factor VII (FVII) deficiency, few naturally occurring mutations have been described in epidermal growth factor (EGF)-like domains. We investigated a 6-year old Italian girl who had low functional and antigenic FVII plasma levels., Design and Methods: Plasma levels of FVII activity and antigen were evaluated in the propositus and her relatives. Mutation screening was performed by sequencing the FVII gene. The effect of the identified FVII mutations was investigated by protein expression in transfected cells., Results: The propositus was shown to be a compound heterozygote for a known (Arg110Cys) and a novel (Asp123Tyr) missense mutation both occurring in the second EGF-like domain. In transfected cells, expression of the Arg110Cys mutation reduced the amount of intracellular and secreted FVII protein (48% and 18%, respectively). Likewise, cells transfected with the Asp123Tyr mutation gave rise to low intracellular (40%) and extracellular (4%) FVII antigen levels. In conditioned media, FVII procoagulant activity was reduced accordingly (10% and <1%, respectively)., Interpretation and Conclusions: Transient expression of the identified FVII mutations caused severely reduced but detectable FVII antigen and activity levels. The present findings suggest that the two naturally occurring missense mutations identified within the second EGF-like domain severely affect FVII protein handling, impairing the correct folding of FVII.

Congenital afibrinogenemia is a rare autosomal recessive disorder characterized by a hemorrhagic diathesis of variable severity. Although more than 100 families with this disorder have been described, genetic defects have been characterized in few cases. An investigation of a young propositus, offspring of a consanguineous marriage, with undetectable levels of functional and quantitative fibrinogen, was conducted. Sequence analysis of the fibrinogen genes showed a homozygous G-to-A mutation at the fifth nucleotide (nt 2395) of the third intervening sequence (IVS) of the gamma-chain gene. Her first-degree relatives, who had approximately half the normal fibrinogen values and showed concordance between functional and immunologic levels, were heterozygtes. The G-to-A change predicts the disappearance of a donor splice site. After transfection with a construct, containing either the wild-type or the mutated sequence, cells with the mutant construct showed an aberrant messenger RNA (mRNA), consistent with skipping of exon 3, but not the expected mRNA. Sequencing of the abnormal mRNA showed the complete absence of exon 3. Skipping of exon 3 predicts the deletion of amino acid sequence from residue 16 to residue 75 and shifting of reading frame at amino acid 76 with a premature stop codon within exon 4 at position 77. Thus, the truncated gamma-chain gene product would not interact with other chains to form the mature fibrinogen molecule. The current findings show that mutations within highly conserved IVS regions of fibrinogen genes could affect the efficiency of normal splicing, giving rise to congenital afibrinogenemia.