The proangiogenic capacity of polymorphonuclear neutrophils delineated by microarray technique and by measurement of neovascularization in wounded skin of CD18-deficient mice

Growing evidence supports the concept that polymorphonuclear neutrophils (PMN) are critically involved in inflammation-mediated angiogenesis which is important for wound healing and repair. We employed an oligonucleotide microarray technique to gain further insight into the molecular mechanisms underlying the proangiogenic potential of human PMN. In addition to 18 known angiogenesis-relevant genes, we detected the expression of 10 novel genes, namely midkine, erb-B2, ets-1, transforming growth factor receptor-beta(2) and -beta(3), thrombospondin, tissue inhibitor of metalloproteinase 2, ephrin A2, ephrin B2 and restin in human PMN freshly isolated from the circulation. Gene expression was confi rmed by the RT-PCR technique. In vivo evidence for the role of PMN in neovascularization was provided by studying neovascularization in a skin model of wound healing using CD18-deficient mice which lack PMN infi ltration to sites of lesion. In CD18-deficient animals, neo- vascularization was found to be signifi cantly compromised when compared with wild- type control animals which showed profound neovascularization within the granulation tissue during the wound healing process. Thus, PMN infiltration seems to facilitate inflammation mediated angiogenesis which may be a consequence of the broad spectrum of proangiogenic factors expressed by these cells. Copyright (c) 2006 S. Karger AG, Basel

LPS unmasking of Shigella flexneri reveals preferential localisation of tagged outer membrane protease IcsP to septa and new poles

The Shigella flexneri outer membrane (OM) protease IcsP (SopA) is a member of the enterobacterial Omptin family of proteases which cleaves the polarly localised OM protein IcsA that is essential for Shigella virulence. Unlike IcsA however, the specific localisation of IcsP on the cell surface is unknown. To determine the distribution of IcsP, a haemagglutinin (HA) epitope was inserted into the non-essential IcsP OM loop 5 using Splicing by Overlap Extension (SOE) PCR, and IcsP(HA) was characterised. Quantum Dot (QD) immunofluorescence (IF) surface labelling of IcsP(HA) was then undertaken. Quantitative fluorescence analysis of S. flexneri 2a 2457T treated with and without tunicaymcin to deplete lipopolysaccharide (LPS) O antigen (Oag) showed that IcsP(HA) was asymmetrically distributed on the surface of septating and non-septating cells, and that this distribution was masked by LPS Oag in untreated cells. Double QD IF labelling of IcsP(HA) and IcsA showed that IcsP(HA) preferentially localised to the new pole of non-septating cells and to the septum of septating cells. The localisation of IcsP(HA) in a rough LPS S. flexneri 2457T strain (with no Oag) was also investigated and a similar distribution of IcsP(HA) was observed. Complementation of the rough LPS strain with rmlD resulted in restored LPS Oag chain expression and loss of IcsP(HA) detection, providing further support for LPS Oag masking of surface proteins. Our data presents for the first time the distribution for the Omptin OM protease IcsP, relative to IcsA, and the effect of LPS Oag masking on its detection.Elizabeth Ngoc Hoa Tran, Matthew Thomas Doyle, Renato Moron

Proteome Based Construction of the Lymphocyte Function-Associated Antigen 1 (LFA-1) Interactome in Human Dendritic Cells.

The β2-integrin lymphocyte function-associated antigen 1 (LFA-1) plays an important role in the migration, adhesion and intercellular communication of dendritic cells (DCs). During the differentiation of human DCs from monocyte precursors, LFA-1 ligand binding capacity is completely lost, even though its expression levels were remained constant. Yet LFA-1-mediated adhesive capacity on DCs can be regained by exposing DCs to the chemokine CCL21, suggesting a high degree of regulation of LFA-1 activity during the course of DC differentiation. The molecular mechanisms underlying this regulation of LFA-1 function in DCs, however, remain elusive. To get more insight we attempted to identify specific LFA-1 binding partners that may play a role in regulating LFA-1 activity in DCs. We used highly sensitive label free quantitative mass-spectrometry to identify proteins co-immunoprecipitated (co-IP) with LFA-1 from ex vivo generated DCs. Among the potential binding partners we identified not only established components of integrin signalling pathways and cytoskeletal proteins, but also several novel LFA-1 binding partners including CD13, galectin-3, thrombospondin-1 and CD44. Further comparison to the LFA-1 interaction partners in monocytes indicated that DC differentiation was accompanied by an overall increase in LFA-1 associated proteins, in particular cytoskeletal, signalling and plasma membrane (PM) proteins. The here presented LFA-1 interactome composed of 78 proteins thus represents a valuable resource of potential regulators of LFA-1 function during the DC lifecycle

Lateral distribution of muons in IceCube cosmic ray events

In cosmic ray air showers, the muon lateral separation from the center of the shower is a measure of the transverse momentum that the muon parent acquired in the cosmic ray interaction. IceCube has observed cosmic ray interactions that produce muons laterally separated by up to 400 m from the shower core, a factor of 6 larger distance than previous measurements. These muons originate in high p(T) (>2 GeV/c) interactions from the incident cosmic ray, or high-energy secondary interactions. The separation distribution shows a transition to a power law at large values, indicating the presence of a hard p(T) component that can be described by perturbative quantum chromodynamics. However, the rates and the zenith angle distributions of these events are not well reproduced with the cosmic ray models tested here, even those that include charm interactions. This discrepancy may be explained by a larger fraction of kaons and charmed particles than is currently incorporated in the simulations.R. Abbasi ... G. C. Hill ... et al. (IceCube Collaboration

Multi-messenger observations of a binary neutron star merger

On 2017 August 17 a binary neutron star coalescence candidate (later designated GW170817) with merger time 12:41:04 UTC was observed through gravitational waves by the Advanced LIGO and Advanced Virgo detectors. The Fermi Gamma-ray Burst Monitor independently detected a gamma-ray burst (GRB 170817A) with a time delay of ~1.7 s with respect to the merger time. From the gravitational-wave signal, the source was initially localized to a sky region of 31 deg2 at a luminosity distance of 40+8-8 Mpc and with component masses consistent with neutron stars. The component masses were later measured to be in the range 0.86 to 2.26 Mo. An extensive observing campaign was launched across the electromagnetic spectrum leading to the discovery of a bright optical transient (SSS17a, now with the IAU identification of AT 2017gfo) in NGC 4993 (at ~40 Mpc) less than 11 hours after the merger by the One- Meter, Two Hemisphere (1M2H) team using the 1 m Swope Telescope. The optical transient was independently detected by multiple teams within an hour. Subsequent observations targeted the object and its environment. Early ultraviolet observations revealed a blue transient that faded within 48 hours. Optical and infrared observations showed a redward evolution over ~10 days. Following early non-detections, X-ray and radio emission were discovered at the transient’s position ~9 and ~16 days, respectively, after the merger. Both the X-ray and radio emission likely arise from a physical process that is distinct from the one that generates the UV/optical/near-infrared emission. No ultra-high-energy gamma-rays and no neutrino candidates consistent with the source were found in follow-up searches. These observations support the hypothesis that GW170817 was produced by the merger of two neutron stars in NGC4993 followed by a short gamma-ray burst (GRB 170817A) and a kilonova/macronova powered by the radioactive decay of r-process nuclei synthesized in the ejecta

Search for time-independent neutrino emission from astrophysical sources with 3 yr of IceCube data

We present the results of a search for neutrino point sources using the IceCube data collected between 2008 April and 2011 May with three partially completed configurations of the detector: the 40-, 59-, and 79-string configurations. The live-time of this data set is 1040 days. An unbinned maximum likelihood ratio test was used to search for an excess of neutrinos above the atmospheric background at any given direction in the sky. By adding two more years of data with improved event selection and reconstruction techniques, the sensitivity was improved by a factor of 3.5 or more with respect to the previously published results obtained with the 40-string configuration of IceCube. We performed an all-sky survey and a dedicated search using a catalog of a priori selected objects observed by other telescopes. In both searches, the data are compatible with the background-only hypothesis. In the absence of evidence for a signal, we set upper limits on the flux of muon neutrinos. For an E-2 neutrino spectrum, the observed limits are (0.9-5) x 10(-12) TeV-1 cm(-2) s(-1) for energies between 1 TeV and 1 PeV in the northern sky and (0.9-23.2) x 10(-12) TeV-1 cm(-2) s(-1) for energies between 10(2) TeV and 10(2) PeV in the southern sky. We also report upper limits for neutrino emission from groups of sources that were selected according to theoretical models or observational parameters and analyzed with a stacking approach. Some of the limits presented already reach the level necessary to quantitatively test current models of neutrino emission

Biphasic toxicodynamic features of some antimicrobial agents on microbial growth: a dynamic mathematical model and its implications on hormesis

<p>Abstract</p> <p>Background</p> <p>In the present work, we describe a group of anomalous dose-response (DR) profiles and develop a dynamic model that is able to explain them. Responses were obtained from conventional assays of three antimicrobial agents (nisin, pediocin and phenol) against two microorganisms (<it>Carnobacterium piscicola </it>and <it>Leuconostoc mesenteroides</it>).</p> <p>Results</p> <p>Some of these anomalous profiles show biphasic trends which are usually attributed to hormetic responses. But they can also be explained as the result of the time-course of the response from a microbial population with a bimodal distribution of sensitivity to an effector, and there is evidence suggesting this last origin. In light of interest in the hormetic phenomenology and the possibility of confusing it with other phenomena, especially in the bioassay of complex materials we try to define some criteria which allow us to distinguish between <it>sensu stricto </it>hormesis and biphasic responses due to other causes. Finally, we discuss some problems concerning the metric of the dose in connection with the exposure time, and we make a cautionary suggestion about the use of bacteriocins as antimicrobial agents.</p> <p>Conclusions</p> <p>The mathematical model proposed, which combines the basis of DR theory with microbial growth kinetics, can generate and explain all types of anomalous experimental profiles. These profiles could also be described in a simpler way by means of bisigmoidal equations. Such equations could be successfully used in a microbiology and toxicology context to discriminate between hormesis and other biphasic phenomena.</p

Microstructural characterization of corticospinal tract in subacute and chronic stroke patients with distal lesions by means of advanced diffusion MRI

PURPOSE: The aim of the paper is to evaluate if advanced dMRI techniques, including diffusion kurtosis imaging (DKI) and neurite orientation dispersion and density imaging (NODDI), could provide novel insights into the subtle microarchitectural modifications occurring in the corticospinal tract (CST) of stroke patients in subacute and chronic phases. METHODS: Seventeen subjects (age 68 ± 11 years) in the subacute phase (14 ± 3 days post-stroke), 10 of whom rescanned in the chronic phase (231 ± 36 days post-stroke), were enrolled. Images were acquired using a 3-T MRI scanner with a two-shell EPI protocol (20 gradient directions, b = 700 s/mm2, 3 b = 0; 64 gradient directions, b = 2000 s/mm2, 9 b = 0). DTI-, DKI-, and NODDI-derived parameters were calculated in the posterior limb of the internal capsule (PLIC) and in the cerebral peduncle (CP). RESULTS: In the subacute phase, a reduction of FA, AD, and KA values was correlated with an increase of ODI, RD, and AK parameters, in both the ipsilesional PLIC and CP, suggesting that increased fiber dispersion can be the main structural factor. In the chronic phase, a reduction of FA and an increase of ODI persisted in the ipsilesional areas. This was associated with reduced Fic and increased MD, with a concomitant reduction of MK and increase of RD, suggesting that fiber reduction, possibly due to nerve degeneration, could play an important role. CONCLUSIONS: This study shows that advanced dMRI approaches can help elucidate the underpinning architectural modifications occurring in the CST after stroke. Further follow-up studies on bigger cohorts are needed to evaluate if DKI- and NODDI-derived parameters might be proposed as complementary biomarkers of brain microstructural alterations

Prostaglandin F2-alpha receptor (FPr) expression on porcine corpus luteum microvascular endothelial cells (pCL-MVECs)

<p>Abstract</p> <p>Background</p> <p>The corpus luteum (CL) is a transient endocrine gland and prostaglandin F2-alpha is considered to be the principal luteolysin in pigs. In this species, the in vivo administration of prostaglandin F2-alpha induces apoptosis in large vessels as early as 6 hours after administration. The presence of the prostaglandin F2-alpha receptor (FPr) on the microvascular endothelial cells (pCL-MVECs) of the porcine corpus luteum has not yet been defined. The aim of the study was to assess FPr expression in pCL-MVECs in the early and mid-luteal phases (EL-p, ML-p), and during pregnancy (P-p). Moreover, the effectiveness of prostaglandin F2-alpha treatment in inducing pCL-MVEC apoptosis was tested.</p> <p>Methods</p> <p>Porcine CLs were collected in the EL and ML phases and during P-p. All CLs from each animal were minced together and the homogenates underwent enzymatic digestion. The pCL-MVECs were then positively selected by an immunomagnetic separation protocol using Dynabeads coated with anti-CD31 monoclonal antibody and seeded in flasks in the presence of EGM 2-MV (Microvascular Endothelial Cell Medium-2). After 4 days of culture, the cells underwent additional immunomagnetic selection and were seeded in flasks until the confluent stage.</p> <p>PCR Real time, western blot and immunodetection assays were utilized to assess the presence of FPr on pCL-MVEC primary cultures. Furthermore, the influence of culture time (freshly isolated, cultured overnight and at confluence) and hormonal treatment (P4 and E2) on FPr expression in pCL-MVECs was also investigated. Apoptosis was detected by TUNEL assay of pCL-MVECs exposed to prostaglandin F2-alpha.</p> <p>Results</p> <p>We obtained primary cultures of pCL-MVECs from all animals. FPr mRNA and protein levels showed the highest value (ANOVA) in CL-MVECs derived from the early-luteal phase. Moreover, freshly isolated MVECs showed a higher FPr mRNA value than those cultured overnight and confluent cells (ANOVA). prostaglandin F2-alpha treatment failed to induce an apoptotic response in all the pCL-MVEC cultures.</p> <p>Conclusion</p> <p>Our data showing the presence of FPr on MVECs and the inability of prostaglandin F2-alpha to evoke an in vitro apoptotic response suggest that other molecules or mechanisms must be considered in order to explain the in vivo direct pro-apoptotic effect of prostaglandin F2-alpha at the endothelial level.</p