Tissue Localization and Extracellular Matrix Degradation by PI, PII and PIII Snake Venom Metalloproteinases: Clues on the Mechanisms of Venom-Induced Hemorrhage

20 páginas, 4 figuras, 3 tablas y 7 tablas en material suplementario.Snake venom hemorrhagic metalloproteinases (SVMPs) of the PI, PII and PIII classes were compared in terms of tissue localization and their ability to hydrolyze basement membrane components in vivo, as well as by a proteomics analysis of exudates collected in tissue injected with these enzymes. Immunohistochemical analyses of co-localization of these SVMPs with type IV collagen revealed that PII and PIII enzymes co-localized with type IV collagen in capillaries, arterioles and post-capillary venules to a higher extent than PI SVMP, which showed a more widespread distribution in the tissue. The patterns of hydrolysis by these three SVMPs of laminin, type VI collagen and nidogen in vivo greatly differ, whereas the three enzymes showed a similar pattern of degradation of type IV collagen, supporting the concept that hydrolysis of this component is critical for the destabilization of microvessel structure leading to hemorrhage. Proteomic analysis of wound exudate revealed similarities and differences between the action of the three SVMPs. Higher extent of proteolysis was observed for the PI enzyme regarding several extracellular matrix components and fibrinogen, whereas exudates from mice injected with PII and PIII SVMPs had higher amounts of some intracellular proteins. Our results provide novel clues for understanding the mechanisms by which SVMPs induce damage to the microvasculature and generate hemorrhage.This work was performed in partial fulfillment of the requirements for the PhD degree for Cristina Herrera at Universidad de Costa Rica.Peer reviewe

Adaptive Evolution of the Myo6 Gene in Old World Fruit Bats (Family: Pteropodidae)

PMCID: PMC3631194This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited

Methylation of class II transactivator gene promoter IV is not associated with susceptibility to Multiple Sclerosis

<p>Abstract</p> <p>Background</p> <p>Multiple sclerosis (MS) is a complex trait in which alleles at or near the class II loci <it>HLA-DRB1 </it>and <it>HLA-DQB1 </it>contribute significantly to genetic risk. The MHC class II transactivator (<it>MHC2TA</it>) is the master controller of expression of class II genes, and methylation of the promoter of this gene has been previously been shown to alter its function. In this study we sought to assess whether or not methylation of the <it>MHC2TA </it>promoter pIV could contribute to MS disease aetiology.</p> <p>Methods</p> <p>In DNA from peripheral blood mononuclear cells from a sample of 50 monozygotic disease discordant MS twins the <it>MHC2TA </it>promoter IV was sequenced and analysed by methylation specific PCR.</p> <p>Results</p> <p>No methylation or sequence variation of the <it>MHC2TA </it>promoter pIV was found.</p> <p>Conclusion</p> <p>The results of this study cannot support the notion that methylation of the pIV promoter of <it>MHC2TA </it>contributes to MS disease risk, although tissue and timing specific epigenetic modifications cannot be ruled out.</p

Expression of the Multiple Sclerosis-Associated MHC Class II Allele HLA-DRB1*1501 Is Regulated by Vitamin D

Multiple sclerosis (MS) is a complex trait in which allelic variation in the MHC class II region exerts the single strongest effect on genetic risk. Epidemiological data in MS provide strong evidence that environmental factors act at a population level to influence the unusual geographical distribution of this disease. Growing evidence implicates sunlight or vitamin D as a key environmental factor in aetiology. We hypothesised that this environmental candidate might interact with inherited factors and sought responsive regulatory elements in the MHC class II region. Sequence analysis localised a single MHC vitamin D response element (VDRE) to the promoter region of HLA-DRB1. Sequencing of this promoter in greater than 1,000 chromosomes from HLA-DRB1 homozygotes showed absolute conservation of this putative VDRE on HLA-DRB1*15 haplotypes. In contrast, there was striking variation among non–MS-associated haplotypes. Electrophoretic mobility shift assays showed specific recruitment of vitamin D receptor to the VDRE in the HLA-DRB1*15 promoter, confirmed by chromatin immunoprecipitation experiments using lymphoblastoid cells homozygous for HLA-DRB1*15. Transient transfection using a luciferase reporter assay showed a functional role for this VDRE. B cells transiently transfected with the HLA-DRB1*15 gene promoter showed increased expression on stimulation with 1,25-dihydroxyvitamin D3 (P = 0.002) that was lost both on deletion of the VDRE or with the homologous “VDRE” sequence found in non–MS-associated HLA-DRB1 haplotypes. Flow cytometric analysis showed a specific increase in the cell surface expression of HLA-DRB1 upon addition of vitamin D only in HLA-DRB1*15 bearing lymphoblastoid cells. This study further implicates vitamin D as a strong environmental candidate in MS by demonstrating direct functional interaction with the major locus determining genetic susceptibility. These findings support a connection between the main epidemiological and genetic features of this disease with major practical implications for studies of disease mechanism and prevention

Lower Richness of Small Wild Mammal Species and Chagas Disease Risk

A new epidemiological scenario involving the oral transmission of Chagas disease, mainly in the Amazon basin, requires innovative control measures. Geospatial analyses of the Trypanosoma cruzi transmission cycle in the wild mammals have been scarce. We applied interpolation and map algebra methods to evaluate mammalian fauna variables related to small wild mammals and the T. cruzi infection pattern in dogs to identify hotspot areas of transmission. We also evaluated the use of dogs as sentinels of epidemiological risk of Chagas disease. Dogs (n = 649) were examined by two parasitological and three distinct serological assays. kDNA amplification was performed in patent infections, although the infection was mainly sub-patent in dogs. The distribution of T. cruzi infection in dogs was not homogeneous, ranging from 11–89% in different localities. The interpolation method and map algebra were employed to test the associations between the lower richness in mammal species and the risk of exposure of dogs to T. cruzi infection. Geospatial analysis indicated that the reduction of the mammal fauna (richness and abundance) was associated with higher parasitemia in small wild mammals and higher exposure of dogs to infection. A Generalized Linear Model (GLM) demonstrated that species richness and positive hemocultures in wild mammals were associated with T. cruzi infection in dogs. Domestic canine infection rates differed significantly between areas with and without Chagas disease outbreaks (Chi-squared test). Geospatial analysis by interpolation and map algebra methods proved to be a powerful tool in the evaluation of areas of T. cruzi transmission. Dog infection was shown to not only be an efficient indicator of reduction of wild mammalian fauna richness but to also act as a signal for the presence of small wild mammals with high parasitemia. The lower richness of small mammal species is discussed as a risk factor for the re-emergence of Chagas disease

High-Throughput Construction of Intron-Containing Hairpin RNA Vectors for RNAi in Plants

With the wide use of double-stranded RNA interference (RNAi) for the analysis of gene function in plants, a high-throughput system for making hairpin RNA (hpRNA) constructs is in great demand. Here, we describe a novel restriction-ligation approach that provides a simple but efficient construction of intron-containing hpRNA (ihpRNA) vectors. The system takes advantage of the type IIs restriction enzyme BsaI and our new plant RNAi vector pRNAi-GG based on the Golden Gate (GG) cloning. This method requires only a single PCR product of the gene of interest flanked with BsaI recognition sequence, which can then be cloned into pRNAi-GG at both sense and antisense orientations simultaneously to form ihpRNA construct. The process, completed in one tube with one restriction-ligation step, produced a recombinant ihpRNA with high efficiency and zero background. We demonstrate the utility of the ihpRNA constructs generated with pRNAi-GG vector for the effective silencing of various individual endogenous and exogenous marker genes as well as two genes simultaneously. This method provides a novel and high-throughput platform for large-scale analysis of plant functional genomics

Jet energy measurement with the ATLAS detector in proton-proton collisions at root s=7 TeV

The jet energy scale and its systematic uncertainty are determined for jets measured with the ATLAS detector at the LHC in proton-proton collision data at a centre-of-mass energy of √s = 7TeV corresponding to an integrated luminosity of 38 pb-1. Jets are reconstructed with the anti-kt algorithm with distance parameters R=0. 4 or R=0. 6. Jet energy and angle corrections are determined from Monte Carlo simulations to calibrate jets with transverse momenta pT≥20 GeV and pseudorapidities {pipe}η{pipe}<4. 5. The jet energy systematic uncertainty is estimated using the single isolated hadron response measured in situ and in test-beams, exploiting the transverse momentum balance between central and forward jets in events with dijet topologies and studying systematic variations in Monte Carlo simulations. The jet energy uncertainty is less than 2. 5 % in the central calorimeter region ({pipe}η{pipe}<0. 8) for jets with 60≤pT<800 GeV, and is maximally 14 % for pT<30 GeV in the most forward region 3. 2≤{pipe}η{pipe}<4. 5. The jet energy is validated for jet transverse momenta up to 1 TeV to the level of a few percent using several in situ techniques by comparing a well-known reference such as the recoiling photon pT, the sum of the transverse momenta of tracks associated to the jet, or a system of low-pT jets recoiling against a high-pT jet. More sophisticated jet calibration schemes are presented based on calorimeter cell energy density weighting or hadronic properties of jets, aiming for an improved jet energy resolution and a reduced flavour dependence of the jet response. The systematic uncertainty of the jet energy determined from a combination of in situ techniques is consistent with the one derived from single hadron response measurements over a wide kinematic range. The nominal corrections and uncertainties are derived for isolated jets in an inclusive sample of high-pT jets. Special cases such as event topologies with close-by jets, or selections of samples with an enhanced content of jets originating from light quarks, heavy quarks or gluons are also discussed and the corresponding uncertainties are determined. © 2013 CERN for the benefit of the ATLAS collaboration

Variants of ST8SIA1 Are Associated with Risk of Developing Multiple Sclerosis

Multiple sclerosis (MS) is an inflammatory demyelinating disease of the central nervous system of unknown etiology with both genetic and environmental factors playing a role in susceptibility. To date, the HLA DR15/DQ6 haplotype within the major histocompatibility complex on chromosome 6p, is the strongest genetic risk factor associated with MS susceptibility. Additional alleles of IL7 and IL2 have been identified as risk factors for MS with small effect. Here we present two independent studies supporting an allelic association of MS with polymorphisms in the ST8SIA1 gene, located on chromosome 12p12 and encoding ST8 alpha-N-acetyl-neuraminide alpha-2,8-sialyltransferase 1. The initial association was made in a single three-generation family where a single-nucleotide polymorphism (SNP) rs4762896, was segregating together with HLA DR15/DQ6 in MS patients. A study of 274 family trios ( affected child and both unaffected parents) from Australia validated the association of ST8SIA1 in individuals with MS, showing transmission disequilibrium of the paternal alleles for three additional SNPs, namely rs704219, rs2041906, and rs1558793, with p = 0.001, p = 0.01 and p = 0.01 respectively. These findings implicate ST8SIA1 as a possible novel susceptibility gene for MS

Measurement of the inclusive and dijet cross-sections of b-jets in pp collisions at sqrt(s) = 7 TeV with the ATLAS detector

The inclusive and dijet production cross-sections have been measured for jets containing b-hadrons (b-jets) in proton-proton collisions at a centre-of-mass energy of sqrt(s) = 7 TeV, using the ATLAS detector at the LHC. The measurements use data corresponding to an integrated luminosity of 34 pb^-1. The b-jets are identified using either a lifetime-based method, where secondary decay vertices of b-hadrons in jets are reconstructed using information from the tracking detectors, or a muon-based method where the presence of a muon is used to identify semileptonic decays of b-hadrons inside jets. The inclusive b-jet cross-section is measured as a function of transverse momentum in the range 20 < pT < 400 GeV and rapidity in the range |y| < 2.1. The bbbar-dijet cross-section is measured as a function of the dijet invariant mass in the range 110 < m_jj < 760 GeV, the azimuthal angle difference between the two jets and the angular variable chi in two dijet mass regions. The results are compared with next-to-leading-order QCD predictions. Good agreement is observed between the measured cross-sections and the predictions obtained using POWHEG + Pythia. MC@NLO + Herwig shows good agreement with the measured bbbar-dijet cross-section. However, it does not reproduce the measured inclusive cross-section well, particularly for central b-jets with large transverse momenta.Comment: 10 pages plus author list (21 pages total), 8 figures, 1 table, final version published in European Physical Journal