Rnd3 as a Novel Target to Ameliorate Microvascular Leakage

Background -Microvascular leakage of plasma proteins is a hallmark of inflammation that leads to tissue dysfunction. There are no current therapeutic strategies to reduce microvascular permeability. The purpose of this study was to identify the role of Rnd3, an atypical Rho family GTPase, in the control of endothelial barrier integrity. The potential therapeutic benefit of Rnd3 protein delivery to ameliorate microvascular leakage was also investigated. Methods and Results-Using immunofluorescence microscopy, Rnd3 was observed primarily in cytoplasmic areas around the nuclei of human umbilical vein endothelial cells. Permeability to fluorescein isothiocyanate-albumin and transendothelial electrical resistance of human umbilical vein endothelial cell monolayers served as indices of barrier function, and RhoA, Rac1, and Cdc42 activities were determined using G-LISA assays. Overexpression of Rnd3 significantly reduced the magnitude of thrombin-induced barrier dysfunction, and abolished thrombin-induced Racl inactivation. Depleting Rnd3 expression with siRNA significantly extended the time course of thrombin-induced barrier dysfunction and Racl inactivation. Time-lapse microscopy of human umbilical vein endothelial cells expressing GFP-actin showed that co-expression of mCherry-Rnd3 attenuated thrombin-induced reductions in local lamellipodia that accompany endothelial barrier dysfunction. Lastly, a novel Rnd3 protein delivery method reduced microvascular leakage in a rat model of hemorrhagic shock and resuscitation, assessed by both intravital microscopic observation of extravasation of fluorescein isothiocyanate-albumin from the mesenteric microcirculation, and direct determination of solute permeability in intact isolated venules. Conclusions-The data suggest that Rnd3 can shift the balance of RhoA and Racl signaling in endothelial cells. In addition, our findings suggest the therapeutic, anti-inflammatory potential of delivering Rnd3 to promote endothelial barrier recovery during inflammatory challenge

Identifying chemokines as therapeutic targets in renal disease: Lessons from antagonist studies and knockout mice

Chemokines, in concert with cytokines and adhesion molecules, play multiple roles in local and systemic immune responses. In the kidney, the temporal and spatial expression of chemokines correlates with local renal damage and accumulation of chemokine receptor-bearing leukocytes. Chemokines play important roles in leukocyte trafficking and blocking chemokines can effectively reduce renal leukocyte recruitment and subsequent renal damage. However, recent data indicate that blocking chemokine or chemokine receptor activity in renal disease may also exacerbate renal inflammation under certain conditions. An increasing amount of data indicates additional roles of chemokines in the regulation of innate and adaptive immune responses, which may adversively affect the outcome of interventional studies. This review summarizes available in vivo studies on the blockade of chemokines and chemokine receptors in kidney diseases, with a special focus on the therapeutic potential of anti-chemokine strategies, including potential side effects, in renal disease. Copyright (C) 2004 S. Karger AG, Basel

Black hole masses and starbursts in X-shaped radio sources

It has been suggested that the X-shaped morphology observed in some radio sources can reflect either a recent merger of two supermassive black holes (SMBHs) or the presence of a second active black hole in the galactic nucleus. These scenarios are tested by studying the relationship between the black hole mass, radio and optical luminosity, starburst history, and dynamic age of radio lobes in a sample of 29 X-shaped radio galaxies drawn from a list of 100 X-shaped radio source candidates identified from the FIRST survey. The same relationships are also studied in a control sample consisting of 36 radio-loud active nuclei with similar redshifts and optical and radio luminosities. The X-shaped objects are found to have statistically higher black hole masses and older starburst activity compared to the objects from the control sample. Implications of these findings are discussed for the black hole merger scenario and for the potential presence of active secondary black holes in post-merger galaxies.Comment: 11 pages, 8 figure

An apicobasal gradient of Rac activity determines protrusion form and position

Each cell within a polarised epithelial sheet must align and correctly position a wide range of subcellular structures, including actin-based dynamic protrusions. Using in vivo inducible transgenes that can sense or modify Rac activity, we demonstrate an apicobasal gradient of Rac activity that is required to correctly form and position distinct classes of dynamic protrusion along the apicobasal axis of the cell. We show that we can modify the Rac activity gradient in genetic mutants for specific polarity proteins, with consequent changes in protrusion form and position and additionally show, using photoactivatable Rac transgenes, that it is the level of Rac activity that determines protrusion form. Thus, we demonstrate a mechanism by which polarity proteins can spatially regulate Rac activity and the actin cytoskeleton to ensure correct epithelial cell shape and prevent epithelial-to-mesenchymal transitions

Myeloid cells expressing VEGF and arginase-1 following uptake of damaged retinal pigment epithelium suggests potential mechanism that drives the onset of choroidal angiogenesis in mice

Whilst data recognise both myeloid cell accumulation during choroidal neovascularisation (CNV) as well as complement activation, none of the data has presented a clear explanation for the angiogenic drive that promotes pathological angiogenesis. One possibility that is a pre-eminent drive is a specific and early conditioning and activation of the myeloid cell infiltrate. Using a laser-induced CNV murine model, we have identified that disruption of retinal pigment epithelium (RPE) and Bruch's membrane resulted in an early recruitment of macrophages derived from monocytes and microglia, prior to angiogenesis and contemporaneous with lesional complement activation. Early recruited CD11b(+) cells expressed a definitive gene signature of selective inflammatory mediators particularly a pronounced Arg-1 expression. Accumulating macrophages from retina and peripheral blood were activated at the site of injury, displaying enhanced VEGF expression, and notably prior to exaggerated VEGF expression from RPE, or earliest stages of angiogenesis. All of these initial events, including distinct VEGF (+) Arg-1(+) myeloid cells, subsided when CNV was established and at the time RPE-VEGF expression was maximal. Depletion of inflammatory CCR2-positive monocytes confirmed origin of infiltrating monocyte Arg-1 expression, as following depletion Arg-1 signal was lost and CNV suppressed. Furthermore, our in vitro data supported a myeloid cell uptake of damaged RPE or its derivatives as a mechanism generating VEGF (+) Arg-1(+) phenotype in vivo. Our results reveal a potential early driver initiating angiogenesis via myeloid-derived VEGF drive following uptake of damaged RPE and deliver an explanation of why CNV develops during any of the stages of macular degeneration and can be explored further for therapeutic gain

Monoclonal Antibodies against Accumulation-Associated Protein Affect EPS Biosynthesis and Enhance Bacterial Accumulation of Staphylococcus epidermidis

Because there is no effective antibiotic to eradicate Staphylococcus epidermidis biofilm infections that lead to the failure of medical device implantations, the development of anti-biofilm vaccines is necessary. Biofilm formation by S. epidermidis requires accumulation-associated protein (Aap) that contains sequence repeats known as G5 domains, which are responsible for the Zn2+-dependent dimerization of Aap to mediate intercellular adhesion. Antibodies against Aap have been reported to inhibit biofilm accumulation. In the present study, three monoclonal antibodies (MAbs) against the Aap C-terminal single B-repeat construct followed by the 79-aa half repeat (AapBrpt1.5) were generated. MAb18B6 inhibited biofilm formation by S. epidermidis RP62A to 60% of the maximum, while MAb25C11 and MAb20B9 enhanced biofilm accumulation. All three MAbs aggregated the planktonic bacteria to form visible cell clusters. Epitope mapping revealed that the epitope of MAb18B6, which recognizes an identical area within AapBrpt constructs from S. epidermidis RP62A, was not shared by MAb25C11 and MAb20B9. Furthermore, all three MAbs were found to affect both Aap expression and extracellular polymeric substance (EPS, including extracellular DNA and PIA) biosynthesis in S. epidermidis and enhance the cell accumulation. These findings contribute to a better understanding of staphylococcal biofilm formation and will help to develop epitope-peptide vaccines against staphylococcal infections

Evidence for the exclusive decay Bc+- to J/psi pi+- and measurement of the mass of the Bc meson

We report first evidence for a fully reconstructed decay mode of the B_c^{\pm} meson in the channel B_c^{\pm} \to J/psi \pi^{\pm}, with J/psi \to mu^+mu^-. The analysis is based on an integrated luminosity of 360 pb$^{-1} in p\bar{p} collisions at 1.96 TeV center of mass energy collected by the Collider Detector at Fermilab. We observe 14.6 \pm 4.6 signal events with a background of 7.1 \pm 0.9 events, and a fit to the J/psi pi^{\pm} mass spectrum yields a B_c^{\pm} mass of 6285.7 \pm 5.3(stat) \pm 1.2(syst) MeV/c^2. The probability of a peak of this magnitude occurring by random fluctuation in the search region is estimated as 0.012%.Comment: 7 pages, 3 figures. Version 3, accepted by PR

Evolution of chloroplast retrograde signaling facilitates green plant adaptation to land

Chloroplast retrograde signaling networks are vital for chloroplast biogenesis, operation, and signaling, including excess light and drought stress signaling. To date, retrograde signaling has been considered in the context of land plant adaptation, but not regarding the origin and evolution of signaling cascades linking chloroplast function to stomatal regulation. We show that key elements of the chloroplast retrograde signaling process, the nucleotide phosphatase (SAL1) and 3'-phosphoadenosine-5'-phosphate (PAP) metabolism, evolved in streptophyte algae-the algal ancestors of land plants. We discover an early evolution of SAL1-PAP chloroplast retrograde signaling in stomatal regulation based on conserved gene and protein structure, function, and enzyme activity and transit peptides of SAL1s in species including flowering plants, the fern Ceratopteris richardii, and the moss Physcomitrella patens Moreover, we demonstrate that PAP regulates stomatal closure via secondary messengers and ion transport in guard cells of these diverse lineages. The origin of stomata facilitated gas exchange in the earliest land plants. Our findings suggest that the conquest of land by plants was enabled by rapid response to drought stress through the deployment of an ancestral SAL1-PAP signaling pathway, intersecting with the core abscisic acid signaling in stomatal guard cells

Top quark mass measurement using the template method at CDF

We present a measurement of the top quark mass in the lepton+jets and dilepton channels of $t\bar{t}$ decays using the template method. The data sample corresponds to an integrated luminosity of 5.6 fb$^{-1}$ of $p\bar{p}$ collisions at Tevatron with $\sqrt{s}=1.96$ TeV, collected with the CDF II detector. The measurement is performed by constructing templates of three kinematic variables in the lepton+jets and two kinematic variables in the dilepton channel. The variables are two reconstructed top quark masses from different jets-to-quarks combinations and the invariant mass of two jets from the $W$ decay in the lepton+jets channel, and a reconstructed top quark mass and $m_{T2}$, a variable related to the transverse mass in events with two missing particles, in the dilepton channel. The simultaneous fit of the templates from signal and background events in the lepton+jets and dilepton channels to the data yields a measured top quark mass of $M_{top} = 172.1 \pm 1.1(stat) \pm 0.9(syst).$Comment: submitted to Phys. Rev.

Measurement of the ttbar Production Cross Section in ppbar Collisions at sqrt(s) = 1.96 TeV

We present a measurement of the top quark pair production cross section in ppbar collisions at sqrt(s)=1.96 TeV using 318 pb^{-1} of data collected with the Collider Detector at Fermilab. We select ttbar decays into the final states e nu + jets and mu nu + jets, in which at least one b quark from the t-quark decays is identified using a secondary vertex-finding algorithm. Assuming a top quark mass of 178 GeV/c^2, we measure a cross section of 8.7 +-0.9 (stat) +1.1-0.9 (syst) pb. We also report the first observation of ttbar with significance greater than 5 sigma in the subsample in which both b quarks are identified, corresponding to a cross section of 10.1 +1.6-1.4(stat)+2.0-1.3 (syst) pb.Comment: Accepted for publication in Physics Review Letters, 7 page