Dose-dependent von Willebrand Factor inhibition by aptamer BB-031 correlates with thrombolysis in a microfluidic model of arterial occlusion

Von Willebrand Factor (VWF) plays a critical role in thrombus formation, stabilization, and propagation. Previous studies have demonstrated that targeted inhibition of VWF induces thrombolysis when administered in vivo in animal models of ischemic stroke. The study objective was to quantify dose-dependent inhibition of VWF-platelet function and its relationship with thrombolysis using BB-031, an aptamer that binds VWF and inhibits its function. VWF:Ac, VWF:RCo, T-TAS, and ristocetin-induced impedance aggregometry were used to assess BB-031-mediated inhibition of VWF. Reductions in original thrombus surface area and new deposition during administration of treatment were measured in a microfluidic model of arterial thrombolysis. Rotational thromboelastometry was used to assess changes in hemostasis. BB-031 induced maximal inhibition at the highest dose (3384 nM) in VWF:Ac, and demonstrated dose-dependent responses in all other assays. BB-031, but not vehicle, induced recanalization in the microfluidic model. Maximal lytic efficacy in the microfluidic model was seen at 1692 nM and not 3384 nM BB-031 when assessed by surface area. Minor changes in ROTEM parameters were seen at 3384 nM BB-031. Targeted VWF inhibition by BB-031 results in clinically measurable impairment of VWF function, and specifically VWF-GPIb function as measured by VWF:Ac. BB-031 also induced thrombolysis as measured in a microfluidic model of occlusion and reperfusion. Moderate correlation between inhibition and lysis was observed. Additional studies are required to further examine off-target effects of BB-031 at high doses, however, these are expected to be above the range of clinical targeted dosing

レーザースペックルフローグラフィーを用いて測定した硝子体手術の術前、術中、術後の眼血流

BACKGROUND AND OBJECTIVE: Vitrectomy markedly alters the intraocular milieu, which can then affect the physiology of the retina and choroid. This study investigates whether vitrectomy also alters ocular blood flow as determined by laser speckle flowgraphy (LSFG). PATIENTS AND METHODS: Twenty eyes of 20 patients that underwent vitrectomy for idiopathic macular hole or epiretinal membrane were studied. Standard 23-gauge microincision vitreous surgery was performed. Ocular blood flow of the optic nerve head, retinal vessels, and choroid was determined by LSGF before, during, and 2 weeks and 1 month after vitrectomy. RESULTS: Postoperative blood flow of the optic nerve head, retinal vessels, and choroid did not differ significantly from preoperative values. Intraoperative blood flow of the optic nerve head and retinal vessels decreased significantly from baseline with increasing infusion pressure from 20 mm Hg to 40 mm Hg (P < .01), and choroidal blood flow decreased significantly when the infusion pressure increased from 8 mm Hg to 20 mm Hg and from 20 mm Hg to 40 mm Hg (both P < .01). CONCLUSION: Blood flow did not differ significantly postoperatively versus preoperatively, but it was significantly reduced during vitrectomy with increasing infusion pressure. Careful attention should be paid to infusion pressure during vitrectomy.博士（医学）・乙第1368号・平成27年11月27日Copyright ©2014, SLACK Incorporated. All Rights Reserved.The definitive version is available at " http://dx.doi.org/10.3928/23258160-20140306-04

Effect of structured use of preoperative portal vein embolization on outcomes after liver resection of perihilar cholangiocarcinoma

BACKGROUND: Portal vein embolization (PVE) is performed to reduce the risk of liver failure and subsequent mortality after major liver resection. Although a cut-off value of 2·7 per cent per min per m2 has been used with hepatobiliary scintigraphy (HBS) for future remnant liver function (FRLF), patients with perihilar cholangiocarcinoma (PHC) potentially benefit from an additional cut-off of 8·5 per cent/min (not corrected for body surface area). Since January 2016 a more liberal approach to PVE has been adopted, including this additional cut-off for HBS of 8·5 per cent/min. The aim of this study was to assess the effect of this approach on liver failure and mortality. METHODS: This was a single-centre retrospective study in which consecutive patients undergoing liver resection under suspicion of PHC in 2000-20

QTL and candidate gene mapping for polyphenolic composition in apple fruit

<p>Abstract</p> <p>Background</p> <p>The polyphenolic products of the phenylpropanoid pathway, including proanthocyanidins, anthocyanins and flavonols, possess antioxidant properties that may provide health benefits. To investigate the genetic architecture of control of their biosynthesis in apple fruit, various polyphenolic compounds were quantified in progeny from a 'Royal Gala' × 'Braeburn' apple population segregating for antioxidant content, using ultra high performance liquid chromatography of extracts derived from fruit cortex and skin.</p> <p>Results</p> <p>Construction of genetic maps for 'Royal Gala' and 'Braeburn' enabled detection of 79 quantitative trait loci (QTL) for content of 17 fruit polyphenolic compounds. Seven QTL clusters were stable across two years of harvest and included QTLs for content of flavanols, flavonols, anthocyanins and hydroxycinnamic acids. Alignment of the parental genetic maps with the apple whole genome sequence <it>in silico </it>enabled screening for co-segregation with the QTLs of a range of candidate genes coding for enzymes in the polyphenolic biosynthetic pathway. This co-location was confirmed by genetic mapping of markers derived from the gene sequences. <it>Leucoanthocyanidin reductase </it>(<it>LAR1</it>) co-located with a QTL cluster for the fruit flavanols catechin, epicatechin, procyanidin dimer and five unknown procyanidin oligomers identified near the top of linkage group (LG) 16, while <it>hydroxy cinnamate/quinate transferase </it>(<it>HCT</it>/<it>HQT</it>) co-located with a QTL for chlorogenic acid concentration mapping near the bottom of LG 17.</p> <p>Conclusion</p> <p>We conclude that <it>LAR1 </it>and <it>HCT</it>/<it>HQT </it>are likely to influence the concentration of these compounds in apple fruit and provide useful allele-specific markers for marker assisted selection of trees bearing fruit with healthy attributes.</p

Exploitation of an iron transporter for bacterial protein antibiotic import

Unlike their descendants, mitochondria and plastids, bacteria do not have dedicated protein import systems. However, paradoxically, import of protein bacteriocins, the mechanisms of which are poorly understood, underpins competition among pathogenic and commensal bacteria alike. Here, using X-ray crystallography, isothermal titration calorimetry, confocal fluorescence microscopy, and in vivo photoactivatable cross-linking of stalled translocation intermediates, we demonstrate how the iron transporter FpvAI in the opportunistic pathogen Pseudomonas aeruginosa is hijacked to translocate the bacteriocin pyocin S2 (pyoS2) across the outer membrane (OM). FpvAI is a TonB-dependent transporter (TBDT) that actively imports the small siderophore ferripyoverdine (Fe-Pvd) by coupling to the proton motive force (PMF) via the inner membrane (IM) protein TonB1. The crystal structure of the N-terminal domain of pyoS2 (pyoS2NTD) bound to FpvAI (Kd = 240 pM) reveals that the pyocin mimics Fe-Pvd, inducing the same conformational changes in the receptor. Mimicry leads to fluorescently labeled pyoS2NTD being imported into FpvAI-expressing P. aeruginosa cells by a process analogous to that used by bona fide TBDT ligands. PyoS2NTD induces unfolding by TonB1 of a force-labile portion of the plug domain that normally occludes the central channel of FpvAI. The pyocin is then dragged through this narrow channel following delivery of its own TonB1-binding epitope to the periplasm. Hence, energized nutrient transporters in bacteria also serve as rudimentary protein import systems, which, in the case of FpvAI, results in a protein antibiotic 60-fold bigger than the transporter’s natural substrate being translocated across the OM

Recent Developments and Applications of the HYDRUS Computer Software Packages

The HYDRUS-1D and HYDRUS (2D/3D) computer software packages are widely used finite-element models for simulating the one- and two- or three-dimensional movement of water, heat, and multiple solutes in variably saturated media, respectively. In 2008, Šimůnek et al. (2008b) described the entire history of the development of the various HYDRUS programs and related models and tools such as STANMOD, RETC, ROSETTA, UNSODA, UNSATCHEM, HP1, and others. The objective of this manuscript is to review selected capabilities of HYDRUS that have been implemented since 2008. Our review is not limited to listing additional processes that were implemented in the standard computational modules, but also describes many new standard and nonstandard specialized add-on modules that significantly expanded the capabilities of the two software packages. We also review additional capabilities that have been incorporated into the graphical user interface (GUI) that supports the use of HYDRUS (2D/3D). Another objective of this manuscript is to review selected applications of the HYDRUS models such as evaluation of various irrigation schemes, evaluation of the effects of plant water uptake on groundwater recharge, assessing the transport of particle-like substances in the subsurface, and using the models in conjunction with various geophysical methods

M19 Modulates Skeletal Muscle Differentiation and Insulin Secretion in Pancreatic β-Cells through Modulation of Respiratory Chain Activity

Mitochondrial dysfunction due to nuclear or mitochondrial DNA alterations contributes to multiple diseases such as metabolic myopathies, neurodegenerative disorders, diabetes and cancer. Nevertheless, to date, only half of the estimated 1,500 mitochondrial proteins has been identified, and the function of most of these proteins remains to be determined. Here, we characterize the function of M19, a novel mitochondrial nucleoid protein, in muscle and pancreatic β-cells. We have identified a 13-long amino acid sequence located at the N-terminus of M19 that targets the protein to mitochondria. Furthermore, using RNA interference and over-expression strategies, we demonstrate that M19 modulates mitochondrial oxygen consumption and ATP production, and could therefore regulate the respiratory chain activity. In an effort to determine whether M19 could play a role in the regulation of various cell activities, we show that this nucleoid protein, probably through its modulation of mitochondrial ATP production, acts on late muscle differentiation in myogenic C2C12 cells, and plays a permissive role on insulin secretion under basal glucose conditions in INS-1 pancreatic β-cells. Our results are therefore establishing a functional link between a mitochondrial nucleoid protein and the modulation of respiratory chain activities leading to the regulation of major cellular processes such as myogenesis and insulin secretion

Outer membrane protein size and LPS O-antigen define protective antibody targeting to the Salmonella surface

Lipopolysaccharide (LPS) O-antigen (O-Ag) is known to limit antibody binding to surface antigens, although the relationship between antibody, O-Ag and other outer-membrane antigens is poorly understood. Here we report, immunization with the trimeric porin OmpD from Salmonella Typhimurium (STmOmpD) protects against infection. Atomistic molecular dynamics simulations indicate this is because OmpD trimers generate footprints within the O-Ag layer sufficiently sized for a single IgG Fab to access. While STmOmpD differs from its orthologue in S. Enteritidis (SEn) by a single amino-acid residue, immunization with STmOmpD confers minimal protection to SEn. This is due to the OmpD-O-Ag interplay restricting IgG binding, with the pairing of OmpD with its native O-Ag being essential for optimal protection after immunization. Thus, both the chemical and physical structure of O-Ag are key for the presentation of specific epitopes within proteinaceous surface-antigens. This enhances combinatorial antigenic diversity in Gram-negative bacteria, while reducing associated fitness costs