Recurrent F8 and F9 gene variants result from a founder effect in two large French haemophilia cohorts.

INTRODUCTION: Haemophilia A (HA) and haemophilia B (HB) are X-linked recessive diseases, caused by a large number of pathogenic variants in the F8 and F9 genes. With the exception of introns 22 and 1 inversions which are frequent in severe HA cases, about 2000 unique variants in F8 and 1000 in F9 have been described in databases and their recurrence remains limited. AIM AND METHODS: During routine analysis, we identified two recurrent missense variants, the F8 gene c.1244C>T, p.Ala415Val variant in 27 HA patients and the F9 gene c.835G>A, p.Ala279Thr variant in 34 HB patients, in two groups of haemophiliac patients from two different regions of France. We aimed to identify whether these variants result from a founder effect. We performed haplotype reconstruction after analysis of extragenic and intragenic polymorphic markers. The ESTIAGE programme was used to estimate the age of the variant. RESULTS: We identified a common ancestral haplotype HA1, in all the HA patients sharing the p.Ala415Val variant, and HB1 for 22 of 34 HB patients sharing the p.Ala279Thr variant. The estimated time of occurrence of the founder variant was between the 13th and 17th century (95% CI: 16 to 29 generations) for the F8 variant and between the 3rd and the 11th century for the F9 variant (95% CI: 44 to 72 generations). CONCLUSION: This study supports a founder effect for these two variants in the two largest reported cohorts of haemophilia patients with an identical variant. These pathogenic variants are among the three most early reported variants in haemophilia.

On-chip free beam optics on a polymer-based photonic integration platform.

This paper presents on-chip free beam optics on polymer-based photonic components. Due to the circumstance that waveguide-based optics allows no direct beam access we use Gradient index (GRIN) lenses assembled into the chip to collimate the beam from the waveguides. This enables low loss power transmission over a length of 1432 µm. Even though the beam propagates through air it is possible to create a resonator with a wavelength shift of 0.002 nm/°C, hence the allowed deviations from the ITU-T grid (100 GHz) are met for ± 20 °C. In order to guarantee reliable laser stability, it is necessary to implement optical isolators at the output of the laser. This requires the insertion of bulk material into the chip and is realized by a 1050 µm thick coated glass. Due to the large gap of the free-space section, it is possible to combine different resonators together. This demonstrates the feasibility of an integrated wavelength-meter.

Hypostomus sertanejo (Siluriformes: Loricariidae), new armoured catfish species from north-eastern Brazil.

A re-evaluation of the armoured catfish species of Hypostomus in the Rio Jaguaribe, north-eastern Brazil, was prompted by the discovery of specimens with pale spots on a dark background collected from that basin c. 1936 and deposited at the Academy of Natural Sciences of Philadelphia. Recent field collections in the Rio Jarguaribe basin confirmed the presence of the pale-spotted specimens and its distinctiveness as a new species. Hypostomus sertanejo n. sp. is diagnosed from other species of Hypostomus by having fins and dermal-plated regions of head and body with pale spots or vermiculations on darker background, teeth slender, asymmetrically bicuspid and numerous (34-75) on dentary and premaxilla, depressed dorsal-fin spine not reaching adipose spine, unbranched pectoral-fin spine longer than unbranched pelvic-fin ray, seven branched dorsal-fin rays and one (rarely two) predorsal plate(s) bordering supraoccipital. Ancistrus salgadae Fowler 1941 is hypothesized to be a junior synonym of Hypostomus carvalhoi (Miranda-Ribeiro, 1937), a dark-spotted Hypostomus described from the Rio Granjeiro, a tributary to the upper Rio Salgado.

Comparison of different blood compartments for the detection of circulating DNA using a rat model of Pneumocystis pneumonia.

Pneumocystis is mostly found in the alveolar spaces, but circulation of viable organisms also occurs and suggests that the detection of DNA in blood could be used as a noninvasive procedure to improve the diagnosis of Pneumocystis pneumonia (PcP). In order to determine the optimal compartment for Pneumocystis DNA detection, we used a rat model of PcP and tested the presence of Pneumocystis with a quantitative mtLSU targeting real-time PCR in four blood compartments: whole blood, clot, serum and Platelet-Rich-Plasma (PRP). All samples from 4 Pneumocystis-free control rats were negative. Pneumocystis was detected in 79, 64, 57, and 57% of samples from 14 PcP rats, respectively, but DNA release was not related to pulmonary loads. These data confirm the potential usefulness of Pneumocystis DNA detection in the blood for PcP diagnosis and suggest that whole blood could be the most appropriate compartment for Pneumocystis detection.

Karyotype structure of Hypostomus cf. plecostomus (Linnaeus, 1758) from Tapajós River basin, Southern Amazon: occurrence of sex chromosomes (ZZ/ZW) and their evolutionary implications.

Hypostomus is a group of fish with numerical and struc-tural karyotypic variability. Among them, only six species, three of which belong to the Amazon basin, show a sex chromosome. In this study, we present the karyotype structure of Hypostomus cf. plecos-tomus from the Teles Pires river basin in the municipality of Alta Flo-resta, MT. The species has 2n = 68 and the karyotype formula 14m+ 24sm+ 14st+ 16a [fundamental number (FN) = 120] in males and 15m+ 24sm+14st+15a (FN = 121) in females and sex chromosomes ZZ/ZW. Argyrophilic nucleolar organizer regions (AgNORs) were identified in two pairs of chromosomes at different positions: short arm of the pair 21and long arm of the pair 27, matching the signals displayed by 18S FISH and indicating multiple NORs. Analysis of band C detected few blocks of constitutive heterochromatin in the pericentromeric regions of most chromosomes and the telomeric regions of some pairs, includ-ing the nucleolar pair 21. However, large blocks on the long arm of the nucleolar pair 27 still stood out. GC-rich heterochromatin (CMA3) was visualized only coincidently with nucleolar sites. Mapping of 5S rDNA sites with FISH revealed markings in eight chromosomes, demonstrat-ing synteny between the 18S and 5S sites. The data obtained for H. cf. plecostomus are important for taxonomic studies of this Amazon com-plex "H. plecostomus group". The occurrence of sex chromosomes in Amazon species of Hypostomus suggests an evolutionary event that is independent of other species in the group.

Cytogenetic description of Ancistrus abilhoai (Siluriformes: Loricariidae) from Iguaçu River basin, southern Brazil.

The Iguaçu River basin is a tributary to the upper Paraná River in southern Brazil, and is considered an important aquatic ecoregion that, although having few species of fish, 51-71% of these are apparently endemic. Ancistrus abilhoai is one of three recently described species for this basin and is currently considered endemic to the basin. In this study, we present the chromosomal structure of two populations of Ancistrus abilhoai one collected in the Iguaçu River, in Paraná State, and another collected in the Timbó River, a tributary of the Iguaçu River, in the State of Santa Catarina. Karyotype analyzes were performed in 11 specimens from the Iguaçu River (four females and seven males) and 12 specimens (all males) from Timbó River, revealing 2n = 48 chromosomes with a karyotype formula of 22m + 14sm + 6st + 6a in both populations. Analysis of active nucleolar organizer regions (Ag-NORs) and fluorescent in situ hybridization (FISH) with 18S rDNA probes revealed the submetacentric pair 13 bearing marks at terminal positions on the short arms. Considered as plesiomorphic chromosomal markers in Loricariidae, asynteny 18S and 5S rDNA, and small amounts of heterochromatin were observed. In this study, the first chromosomal data of A. abilhoai are presented with comments on karyotypic characteristics of the genus.

Karyotypic diversity of the armoured catfish genus Hypostomus (Siluriformes: Loricariidae) in the context of its occurrence and distribution.

This study taxonomically reviewed the specimens studied by Artoni & Bertollo (1996) and assimilated species of Hypostomus into three groups according to their cytogenetic characteristics, vagility and occurrence environments.

The occurrence of Austrodiplostomum compactum (Lutz, 1928) (Digenea: Diplostomidae) metacercariae in the eyes of loricariid fish (Siluriformes: Osteichthyes: Loricariidae) from Brazil.

The aim of this study was to report the occurrence of Austrodiplostomum compactum metacercariae in the eyes of 98 specimens of loricariid fish (Hypostomus ancistroides, H. hermanni, H. iheringii, H. margaritifer, H. regani, H. strigaticeps, Hypostomus sp. and Megalancistrus parananus) from the Chavantes reservoir (23°07'36″S and 49°37'35″W) located in the rio Paranapanema, upper Paraná river basin, municipality of Ipaussu, São Paulo State, Brazil. Fish were collected from October 2007 to February 2009 using nylon monofilament gill nets and transported to the field laboratory where they were euthanized and the eyes were taken and examined under a stereomicroscope. Hypostomus ancistroides and M. parananus were not infected by this diplostomid. Hypostomus hermanni and H. margaritifer were represented by only one specimen but both had a high intensity of A. compactum metacercarie (27 and 35, respectively). Hypostomus strigaticeps (n = 45) and H. iheringii (n = 28) were the most representative specimens and the prevalence, mean intensity of infection and mean abundance were 24.4%, 10.3 and 2.7, and 64.2%, 13.1 and 8.4, respectively. No correlation was observed between the intensity of infection and the standard length (r = - 0.223; P = 0.827) and weight (r = 0.03; P = 0.779) of studied fish. Similarly, linear regression among these variables showed a poor correlation and indicated that the infection by A. compactum metacercariae occurs similarly in small and large fish specimens. A seasonal pattern of infection was not observed. Hypostomus hermanni, H. iheringii, H. margaritifer and H. strigaticeps were new hosts recorded for A. compactum metacercariae. A review of morphometric data of A. compactum metacercariae is presented.

Karyotype description of two species of Hypostomus (Siluriformes, Loricariidae) of the Planalto da Bodoquena, Brazil.

Hypostomus sp 3-Córrego Salobrinha NUP 4247 and Hypostomus sp 2-Rio Perdido NUP 4249, collected in the Planalto da Bodoquena, Paraguay River basin, Brazil, were characterized cytogenetically. Hypostomus sp 3-Córrego Salobrinha showed two modal numbers. This polymorphism consists of the presence of two extrachromosomes. It was not possible to define the diploid number in four specimens, where cell lineages had 2n = 83 and 2n = 84 chromosomes in one individual, and 2n = 82, 2n = 83 and 2n = 84 chromosomes in the others. These results reveal the existence of a genetic mosaic due to the occurrence of one or two extrachromosomes in this species. Hypostomus sp 2-Rio Perdido NUP 4249 showed a 2n = 84, FN = 106 with size heteromorphism in one pair of chromosomes stained with AgNO3. In both species, C banding showed a pattern of heterochromatin distribution with a few small bands in the centromeric and pericentromeric regions coinciding with chromomycin A3 staining. Until now, the major diploid number for the genus Hypostomus was 2n = 80, but the species studied here had chromosomes that in creased this number and the variation for this genus. Our results are also the first cytogenetic data on Hypostomus from the Paraguay River basin.

Study of six patients with complete F9 deletion characterized by cytogenetic microarray: role of the SOX3 gene in intellectual disability.

Essentials Some hemophilia B (HB) patients with complete F9 deletion present with intellectual disability (ID). We delineate six F9 complete deletions and investigate genotype/phenotype correlation. We identify SOX3 as a candidate gene for ID, acting through haploinsufficiency, in HB patients. All complete F9 deletions in ID patients should be explored with cytogenetic microarrays. SUMMARY: Background Large deletions encompassing both the complete F9 gene and contiguous genes have been detected in patients with severe hemophilia B (HB). Some of these patients present other clinical features, such as intellectual disability (ID). Objectives/Methods In this study, we characterized six unrelated large deletions encompassing F9, by cytogenetic microarray analysis (CMA), to investigate genotype/phenotype correlation. Results Five of the six patients included in this study presented with ID associated with HB. CMA showed that the six large deletions, ranging in size from approximately 933 kb to 9.19 Mb, were located within the Xq26.3 to Xq28 bands. In all cases, the complete deletion of F9 was associated with the loss of various neighboring genes (5-28 other genes). The smallest region of overlap for ID was a 1.26-Mb region encompassing seven OMIM genes (LOC389895, SOX3, LINC00632, CDR1, SPANXF1, LDOC1, SPANXC). SOX3, our candidate gene for ID, encodes an early transcription factor involved in pituitary development. All of the patients studied who had both HB and ID had deletion of the SOX3 gene. Conclusions All HB patients with an atypical phenotype, especially if complete deletion of F9 is suspected, should be referred to a geneticist for possible pangenomic assessment, because haploinsufficiency of genes flanking F9, such as SOX3 in particular, may result in a broader phenotype, including ID. Such assessment would be of particular value for the genetic counseling of female carriers with F9 deletions, as it would facilitate analysis of the risk of transmitting HB associated with ID.

Relevance of tissue factor in cardiovascular disease.

Overexpression and exposition of tissue factor (TF) in atherosclerotic plaques and/or arterial thrombi are critical events in atherothrombosis. TF, the receptor for factor VII (FVII) and activated factor VII (FVIIa), is the principal initiator of blood coagulation and induces thrombin generation leading to fibrin formation and platelet activation. TF also plays a major role in cell migration and angiogenesis. TF activity is downregulated by Tissue Factor Pathway Inhibitor (TFPI), a Kunitz-type inhibitor, which forms a neutralizing complex with TF, FVIIa and activated factor X. In physiological conditions, TF is absent from vascular cells which come into contact with flowing blood and is present as an inactive pool in fibroblasts and smooth muscle cells (SMC). In contrast, TF is widely expressed in atherosclerotic plaques and is found in macrophages, SMCs, and foam-cells and also in extracellular matrix and acellular lipid-rich core. TF expression is up-regulated by inflammatory cytokines and oxidized lipids. Plaque thrombogenicity is directly correlated to their TF content. After fibrous cap disruption, TF is exposed on plaque surface and triggers thrombus formation leading to arterial lumen occlusion and/or downstream embolization. In coronary and carotid plaques, TF content was found to be higher in plaques from symptomatic than asymptomatic patients. Soluble forms of TF and microparticles of monocyte and platelet origin, and bearing TF, constitute "blood-born TF". The contribution of this TF pool to arterial thrombosis is still under discussion. TF pathway is a target for new therapeutic agents that can decrease TF activity, such as active site-inactivated factor VIIa, recombinant TFPI and antibodies against TF or peptides interfering with TF-FVIIa complex activity.

The coronary artery disease-associated gene C6ORF105 is expressed in human macrophages under the transcriptional control of PPARγ.

Coronary artery disease (CAD) is a major cause of morbidity and mortality. Mutations in C6ORF105, associated with decreased gene expression, positively correlate with the risk of CAD in Chinese populations. Moreover, the C6ORF105-encoded protein may play a role in coagulation. Here, we report that C6ORF105 gene expression is lower in circulating mononuclear cells from obese diabetic than lean subjects. Moreover, C6ORF105 is expressed in human macrophages and atherosclerotic lesions, where its expression positively correlates with expression of the transcription factor Peroxisome Proliferator-Activated Receptor (PPAR)γ. Activation of PPARγ increases, in a PPARγ-dependent manner, the expression of C6ORF105 in human macrophages and atherosclerotic lesions.

The neuron-derived orphan receptor 1 (NOR1) is induced upon human alternative macrophage polarization and stimulates the expression of markers of the M2 phenotype.

BACKGROUND: Atherosclerosis is an inflammatory disease in which macrophages play a crucial role. Macrophages are present in different phenotypes, with at the extremes of the spectrum the classical M1 pro-inflammatory and the alternative M2 anti-inflammatory macrophages. The neuron-derived orphan receptor 1 (NOR1), together with Nur77 and Nurr1, are members of the NR4A orphan nuclear receptor family, expressed in human atherosclerotic lesion macrophages. However, the role of NOR1 in human macrophages has not been studied yet. OBJECTIVES: To determine the expression and the functions of NOR1 in human alternative macrophages. METHODS AND RESULTS: In vitro IL-4 polarization of primary monocytes into alternative M2 macrophages enhances NOR1 expression in human but not in mouse macrophages. Moreover, NOR1 expression is most abundant in CD68+MR+ alternative macrophage-enriched areas of human atherosclerotic plaques in vivo. Silencing NOR1 in human alternative macrophages decreases the expression of several M2 markers such as the Mannose Receptor (MR), Interleukin-1 Receptor antagonist (IL-1Ra), CD200 Receptor (CD200R), coagulation factor XIII A1 polypeptide (F13A1), Interleukin 10 (IL-10) and the Peroxisome Proliferator-Activated Receptor (PPAR)γ. Bioinformatical analysis identified F13A1, IL-1Ra, IL-10 and the Matrix Metalloproteinase-9 (MMP9) as potential target genes of NOR1 in human alternative macrophages. Moreover, expression and enzymatic activity of MMP9 are induced by silencing and repressed by NOR1 overexpression in M2 macrophages. CONCLUSIONS: These data identify NOR1 as a transcription factor induced during alternative differentiation of human macrophages and demonstrate that NOR1 modifies the alternative macrophage phenotype.

Pentoxifylline prevents upregulation of monocyte tissue factor in renal transplant recipients undergoing post-graft complications.

Pentoxifylline (PTX) has been demonstrated to improve graft survival in renal transplant recipients undergoing post graft complications. As activated monocytes are possible initiators of vascular damage through tissue factor (TF) expression, we evaluated the monocyte TF expression and endothelium activation markers in 140 consecutive patients receiving cadaveric kidney grafts, randomized in a double-blind study comparing PTX versus placebo. Monocyte TF expression and plasma von Willebrand factor, tissue plasminogen activator, thrombomodulin and tumor necrosis factor-alpha (TNF-alpha) levels were determined before transplantation and each month after. Additional samplings were realized in case of acute rejection. TF and TNF-alpha expression were significantly modified after graft. In patients with complications, PTX prevented the increase of TF expression at month one, and after rejection episodes. Endothelium activation markers were significantly modified after graft and in patients with complications but PTX had no significant effect on their plasma levels. These results suggest that the protective effect of PTX on graft survival could be related to the prevention of monocyte TF upregulation associated with complications.

Homozygous variant of antithrombin with lack of affinity for heparin: management of severe thrombotic complications associated with intrauterine fetal demise.

Patients with homozygous heparin-binding-site (HBS) qualitative antithrombin deficiencies are at significant risk of venous and arterial thrombosis. We report on the eighth case of homozygous HBS deficiency, and the fourth case concerning the Arg 47-Cys mutation. The proposita is a 25 year old, without known thrombotic antecedent, despite an oral contraceptive therapy for 7 years. After 25 weeks of a first pregnancy, she presented an intrauterine fetal demise complicated with deep vein thrombosis and pulmonary embolism. Heparin therapy was inefficient (no clinical nor angiographic improvement, no biological hypocoagulability). Heparin cofactor activity was < 10%, antigen concentration was normal. The crossed immunoelectrophoresis of patient's plasma, with and without heparin, showed a typical profile of qualitative HBS antithrombin deficiency. The molecular analysis revealed an homozygous Arg 4-Cys mutation. Antithrombotic therapy was achieved with continuous infusion of antithrombin concentrates (80 IU/kg/day) and unfractionated heparin (500 IU/kg/day) during 12 days, leading to clinical improvement, and followed by treatment with vitamin K antagonists. This observation emphasizes the risk of intrauterine fetal demise and the inefficiency of heparin therapy without antithrombin infusion in type II HBS homozygous deficiency. The management of a future pregnancy will probably require repeated infusions of antithrombin.

Differential expression for tissue-specific isozymes in three species of Hypostomus Lacépède, 1803 (Teleostei: Loricariidae).

The expression patterns of 14 enzymatic systems in skeletal muscle, liver and heart tissues of three species of Hypostomus from the Iguaçu River basin (Brazil) were investigated. Although the patterns were similar for the three species, different tissues showed differential expressions, and the data showed that differential tissue expressions of isoperoxidases may be due to preferential combination or association of polypeptide subunits. The detected patterns for SOD isozymes showed that the quaternary structures of these enzymes were in disagreement with the subunit number reported for the majority of other vertebrate groups. Tissue-specific restriction on heterotetramer formation also were described in LDH and MDHP isozymes. Thus, these tissue-specific gene expression character in the species of Hypostomus have the greatest potential to be recognized and applied in systematic studies among species of Hypostomus.

[G20210A transition in the prothrombin gene and venous thromboembolic disease]. / La transition G20210A dans le gène de la prothrombine et la maladie thromboembolique veineuse.

INTRODUCTION: Genetic predisposition to venous thrombosis can be due to coagulation inhibitor deficiencies (antithrombin, protein C or protein S) or to activated protein C resistance resulting from factor V Leiden mutation (FV Leiden). Poort et al. recently identified a new polymorphism in the 3'-untranslated region of the prothrombin gene, the G20210A transition (FII G20210A), which was found to be associated with an increased risk of venous thrombosis. EXEGESIS: The prevalence of the A allele is approximately 1 to 4% in the general population, and 5 to 7% in patients with venous thrombosis. Heterozygous carriers have a three to five times increased risk of thrombosis. The diagnosis is based on a polymerase chain reaction technique and restriction enzyme digestion from genomic DNA. Recent studies aim to determine the relative risk of thrombosis and the clinical features which are associated with the mutation (age of first thrombosis, recurrence). The thrombotic risk seems to be higher when FII G20210A transition is associated with the FV Leiden mutation. CONCLUSION: The presence of heterozygous FII G20210A transition does not modify the management of acute thrombotic events but can lead to an increase in the duration of the anticoagulant treatment. When such a genetic abnormality is identified, thorough information of the patient is needed, including on the prophylactic heparin in high-risk situations and caution on the prescription of oral contraceptives containing estrogens.

Ichthyofauna from the Emas National Park region: composition and structure.

The relationship between habitats and the ichthyofauna composition in the Parque Nacional das Emas (PNE) and adjacent areas (the Araguaia and Sucuriú rivers) are provided and could be applied in determining the Park's future zoning. Samples of the ichthyofauna and limnological parameters were obtained during both dry (September 1999) and wet (December 1999) seasons. Ichthyofauna collections resulted in the capture of 4,740 specimens of 22 species. The most abundant species in the Araguaia River during the two sampling seasons were Astyanax sp. 2 and Hasemania sp. In the Sucuriú River and PNE, Astyanax scabripinnis cf. paranae and Hoplias aff. malabaricus were the most frequent species. The largest number of species and diversity index were recorded for the Araguaia River. However, sound management policies require more detailed studies on the fish communities of the Cerrado biome.