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Introduction HGF/c-MET signaling is a significant driver of glioblastoma (GBM) growth and disease progression. Unfortunately, c-MET targeted therapies have been found to be largely ineffective suggesting additional redundant mechanisms of c-MET activation. Methods Utilizing RNA-sequencing (RNA-seq) and ribosome profiling analyses of circular RNAs, circ-HGF (hsa_circ_0080914) was identified as markedly upregulated in primary GBM and found to potentially encode an HGF protein variant (C-HGF) 119 amino acids in length. This candidate HGF variant was characterized and evaluated for its ability to mediate c-MET activation and regulate PDX GBM cell growth, motility and invasive potential in vitro and tumor burden in intracranial xenografts in mice. Results An internal ribosome entry site (IRES) was identified within the circ-HGF RNA which mediated translation of the cross-junctional ORF encoding C-HGF and was observed to be highly expressed in GBM relative to normal brain tissue. C-HGF was also found to be secreted from GBM cells and concentrated cell culture supernatants or recombinant C-HGF activated known signaling cascades downstream of c-MET. C-HGF was shown to interact directly with the c-MET receptor resulting in its autophosphorylation and activation in PDX GBM lines. Knockdown of C-HGF resulted in suppression of c-MET signaling and marked inhibition of cell growth, motility and invasiveness, whereas overexpression of C-HGF displayed the opposite effects. Additionally, modulation of C-HGF expression regulated tumor growth in intracranial xenografted PDX GBM models. Conclusions These results reveal an alternative mechanism of c-MET activation via a circular RNA encoded HGF protein variant which is relevant in GBM biology. Targeting C-HGF may offer a promising approach for GBM clinical management.

Heat shock proteins (HSPs), especially Hsp70 (HSPA1), have been associated with cellular protection from various cellular stresses including heat, hypoxia-ischemia, neurodegeneration, toxins, and trauma. Endogenous HSPs are often synthesized in direct response to these stresses but in many situations are inadequate in protecting cells. The present study addresses the transduction of Hsp70 into cells providing protection from acute oxidative stress by H2O2. The recombinant Fv-Hsp70 protein and two mutant Fv-Hsp70 proteins minus the ATPase domain and minus the ATPase and terminal lid domains were tested at 0.5 and 1.0 μM concentrations after two different concentrations of H2O2 treatment. All three recombinant proteins protected SH-SY5Y cells from acute H2O2 toxicity. This data indicated that the protein binding domain was responsible for cellular protection. In addition, experiments pretreating cells with inhibitors of antioxidant proteins catalase and gamma-glutamylcysteine synthase (GGCS) before H2O2 resulted in cell death despite treatment with Fv-Hsp70, implying that both enzymes were protected from acute oxidative stress after treatment with Fv-Hsp70. This study demonstrates that Fv-Hsp70 is protective in our experiments primarily by the protein-binding domain. The Hsp70 terminal lid domain was also not necessary for protection.

Ξ− atomic X-ray spectroscopy is one of the most useful methods for investigation of the Ξ–nucleus strong interaction. Since the X-ray energy is shifted and/or broadened due to the Ξ–nucleus strong interaction compared to those calculated from electromagnetic interaction alone, the measurement of the energy shift, ΔE, and the width, Γ, give us information on the Ξ–nucleus potential. A serious problem in the measurement is the significant background derived from in-flight Ξ− decay. A novel method of identifying stopped Ξ− events using the nuclear emulsion was developed to realize the first Ξ− atomic X-ray spectroscopy experiment as the J-PARC E07 experiment, which also aimed at searching for ΛΛ and Ξ− hypernuclei in the emulsion. The X-rays emitted from Ξ− Br and Ξ− Ag atoms were measured using germanium detectors. No clear peaks were observed in the obtained spectra. However, we succeeded in reducing the background to 1/170 by this method employing coincidence measurements using nuclear emulsion and X-ray detectors.

INTRODUCTION: Recent studies have suggested that dysregulated Hippo pathway signaling may contribute to glioblastoma proliferation and invasive characteristics. The downstream effector of the pathway, the Yes-associated protein (YAP) oncoprotein, has emerged as a promising target in glioblastoma multiforme (GBM). METHODS: Utilizing a high-throughput yeast two-hybrid based screen, a small molecule was identified which inhibits the association of the co-transcriptional activator YAP1 and the TEA domain family member 1 (TEAD1) transcription factor protein–protein interaction interface. This candidate inhibitor, NSC682769, a novel benzazepine compound, was evaluated for its ability to affect Hippo/YAP axis signaling and potential anti-glioblastoma properties. RESULTS: NSC682769 potently blocked association of YAP and TEAD in vitro and in GBM cells treated with submicromolar concentrations. Moreover, inhibitor-coupled bead pull down and surface plasmon resonance analyses demonstrate that NSC682769 binds to YAP. NSC682769 treatment of GBM lines and patient derived cells resulted in downregulation of YAP expression levels resulting in curtailed YAP-TEAD transcriptional activity. In GBM cell models, NSC682769 inhibited proliferation, colony formation, migration, invasiveness and enhanced apoptosis. In tumor xenograft and genetically engineered mouse models, NSC682769 exhibited marked anti-tumor responses and resulted in increased overall survival and displayed significant blood-brain barrier penetration. CONCLUSIONS: These results demonstrate that blockade of YAP-TEAD association is a viable therapeutic strategy for glioblastoma. On the basis of these favorable preclinical studies further clinical studies are warranted.

We investigate the impacts of neutrino cooling mechanism inside the neutron star (NS) core on the light curves of type-I X-ray bursts and X-ray superbursts. From several observations of NS thermal evolution, physical processes of fast neutrino cooling, such as the direct Urca (DU) process, are indicated. They significantly decrease the surface temperature of NSs, though the cooling effect could be suppressed by nucleon superfluidity. In the present study, focusing on the DU process and nucleon superfluidity, we investigate the effects of NS cooling on the X-ray bursts using a general-relativistic stellar-evolution code. We find that the DU process leads find the longer recurrence time and the higher peak luminosity, which could be obstructed by the neutrons superfluidity. We also apply our burst models to the comparison with {\it Clocked burster} GS 1826$-$24, and to the recurrence time of superburst triggered by carbon ignition. These effects are significant within a certain range of binary parameters and uncertainty of the NS equation of state., Comment: 12 pages, 8 figures, 3 Tables

INTRODUCTION: Clinical trials directed at mechanistic target of rapamycin (mTOR) inhibition have yielded disappointing results in glioblastoma (GBM). A major mechanism of resistance involves the activation of a salvage pathway stimulating internal ribosome entry site (IRES)-mediated protein synthesis. PRMT5 activity has been implicated in the enhancement of IRES activity. METHODS: We analyzed the expression and activity of PRMT5 in response to mTOR inhibition in GBM cell lines and short-term patient cultures. To determine whether PRMT5 conferred resistance we used genetic and pharmacological approaches to ablate PRMT5 activity and assessed the effects on in vitro and in vivo sensitivity. Mutational analyses of the requisite IRES-trans-acting factor (ITAF), hnRNP A1 determined whether PRMT5-mediated methylation was necessary for ITAF RNA binding and IRES activity. RESULTS: PRMT5 activity is stimulated in response to mTOR inhibitors. Knockdown or treatment with a PRMT5 inhibitor blocked IRES activity and sensitizes GBM cells. Ectopic expression of non-methylatable hnRNP A1 mutants demonstrated that methylation of either arginine residues 218 or 225 was sufficient to maintain IRES binding and hnRNP A1-dependent cyclin D1 or c-MYC IRES activity, however a double R218K/R225K mutant was unable to do so. The PRMT5 inhibitor EPZ015666 displayed synergistic anti-GBM effects in vitro and in a xenograft mouse model in combination with PP242. CONCLUSIONS: These results demonstrate that PRMT5 activity is stimulated upon mTOR inhibition in GBM. Our data further support a signaling cascade in which PRMT5-mediated methylation of hnRNP A1 promotes IRES RNA binding and activation of IRES-mediated protein synthesis and resultant mTOR inhibitor resistance.

Type I X-ray bursts are thermonuclear explosions on the neutron star (NS) surface caused by mass accretion from a companion star. Observations of X-ray bursts provide valuable information on X-ray binary systems, e.g., binary parameters, the chemical composition of accreted matter, and the nuclear equation of state (EOS) of NSs. There have been several theoretical studies to constrain the physics of X-ray bursters. However, they have mainly focused on the burning layers above the solid crust of the NS, which brings up issues of the treatment of NS gravitation and internal energy. In this study, focusing on the microphysics inside NSs, we calculate a series of X-ray bursts using a general-relativistic stellar-evolution code with several NS EOSs. We compare the X-ray-burst models with the burst parameters of a clocked burster associated with GS 1826-24. We find a monotonic correlation between the NS radius and the light-curve profile. A larger radius shows a higher recurrence time and a large peak luminosity. In contrast, the dependence of light curves on the NS mass becomes more complicated, where the neutrino cooling suppresses the efficiency of nuclear ignition. We also constrain the EOS and mass of GS 1826-24, i.e., stiffer EOSs, corresponding to larger NS radii, are not preferred due to too-high peak luminosity. The EOS and the cooling and heating of NSs are important to discuss the theoretical and observational properties of X-ray bursts., 19 pages, 16 figures, 3 tables, updated to match published version (ApJ)

We have performed the first direct measurement of the 83Rb(p,g) radiative capture reaction cross section in inverse kinematics using a radioactive beam of 83Rb at incident energies of 2.4 and 2.7 A MeV. The measured cross section at an effective relative kinetic energy of Ecm = 2.393 MeV, which lies within the relevant energy window for core collapse supernovae, is smaller than the prediction of statistical model calculations. This leads to the abundance of 84Sr produced in the astrophysical p process being higher than previously calculated. Moreover, the discrepancy of the present data with theoretical predictions indicates that further experimental investigation of p-process reactions involving unstable projectiles is clearly warranted., 6 pages, 4 figures, published in Physical Review Letters

Mosquitoes have developed specialized oviposition strategies that allow them to develop in a wide variety of aquatic habitats. Environmentally cued hatching traits may also play an important role in the successful colonization of some larval habitats, but this subject has remained largely unexplored in Culicidae. Aedes atropalpus (Coquillett) is an autogenous rock pool specialist that may maintain unique adaptations for oviposition and egg hatching. We investigated the egg-laying strategies of Ae. atropalpus exposed to standard (non-diapausing) rearing conditions and diapause-inducing conditions and tested the impact of physical agitation on egg hatch rates by exposing floating and submerged eggs to physical agitation treatments. The results of the oviposition experiment indicate that Ae. atropalpus females primarily lay non-diapausing eggs directly onto the water surface and lay diapausing eggs directly on solid surfaces. The egg-hatching experiment demonstrated that physical agitation significantly increases Ae. atropalpus hatch rates. Floating and submerged eggs responded similarly to the agitation treatment. These data suggest that oviposition behaviors based on both egg diapause status and environmentally-cued hatching strategies may be important adaptations for Ae. atropalpus in riverine rock pools.

The $\nu p$ process appears in proton-rich, hot matter which is expanding in a neutrino wind and may be realised in explosive environments such as core-collapse supernovae or in outflows from accretion disks. The impact of uncertainties in nuclear reaction cross sections on the finally produced abundances has been studied by applying Monte Carlo variation of all astrophysical reaction rates in a large reaction network. As the detailed astrophysical conditions of the $\nu p$ process still are unknown, a parameter study was performed, with 23 trajectories covering a large range of entropies and $Y_\mathrm{e}$. The resulting abundance uncertainties are given for each trajectory. The $\nu p$ process has been speculated to contribute to the light $p$ nuclides but it was not possible so far to reproduce the solar isotope ratios. It is found that it is possible to reproduce the solar $^{92}$Mo/$^{94}$Mo abundance ratio within nuclear uncertainties, even within a single trajectory. The solar values of the abundances in the Kr-Sr region relative to the Mo region, however, cannot be achieved within a single trajectory. They may still be obtained from a weighted superposition of different trajectories, though, depending on the actual conditions in the production site. For a stronger constraint of the required conditions, it would be necessary to reduce the uncertainties in the 3$\alpha$ and $^{56}$Ni(n,p)$^{56}$Co rates at temperatures $T>3$ GK., Comment: 6 pages, 1 figure, 2 tables; to appear in JPS Conference Proceedings (OMEG15, 2019); summarizes and expands on arXiv:1907.13129 with an additional discussion of isotopic ratios of p-nuclides; v2: typos fixed

Ξ− atomic X-ray spectroscopy is one of the most useful methods for investigation of the Ξ-nucleus strong interaction. A serious problem in the measurement is the significant background coming from in-flight Ξ− decay. For the first Ξ− atomic X-ray spectroscopy experiment, a novel method of identifying stopped Ξ− events using nuclear emulsion was developed to reject background photons from in-flight Ξ− decay. We succeeded in reducing the background to 1/170 by this method employing coincidence measurements using the nuclear emulsion and X-ray detectors.

Overexpression of Rictor has been demonstrated to result in increased mechanistic target of rapamycin C2 (mTORC2) nucleation and activity leading to tumor growth and increased invasive characteristics in glioblastoma multiforme (GBM). However, the mechanisms regulating Rictor expression in these tumors is not clearly understood. In this report, we demonstrate that Rictor is regulated at the level of mRNA translation via heat-shock transcription factor 1 (HSF1)-induced HuR activity. HuR is shown to directly bind the 3' untranslated region of the Rictor transcript and enhance translational efficiency. Moreover, we demonstrate that mTORC2/AKT signaling activates HSF1 resulting in a feed-forward cascade in which continued mTORC2 activity is able to drive Rictor expression. RNAi-mediated blockade of AKT, HSF1 or HuR is sufficient to downregulate Rictor and inhibit GBM growth and invasive characteristics in vitro and suppress xenograft growth in mice. Modulation of AKT or HSF1 activity via the ectopic expression of mutant alleles support the ability of AKT to activate HSF1 and demonstrate continued HSF1/HuR/Rictor signaling in the context of AKT knockdown. We further show that constitutive overexpression of HuR is able to maintain Rictor expression under conditions of AKT or HSF1 loss. The expression of these components is also examined in patient GBM samples and correlative associations between the relative expression of these factors support the presence of these signaling relationships in GBM. These data support a role for a feed-forward loop mechanism by which mTORC2 activity stimulates Rictor translational efficiency via an AKT/HSF1/HuR signaling cascade resulting in enhanced mTORC2 activity in these tumors.

Internal ribosome entry site (IRES)-mediated protein synthesis has been demonstrated to play an important role in resistance to mechanistic target of rapamycin (mTOR) targeted therapies. Previously, we have demonstrated that the IRES trans-acting factor (ITAF), hnRNP A1 is required to promote IRES activity and small molecule inhibitors which bind specifically to this ITAF and curtail IRES activity, leading to mTOR inhibitor sensitivity. Here we report the identification of riluzole (Rilutek&reg, ), an FDA-approved drug for amyotrophic lateral sclerosis (ALS), via an in silico docking analysis of FDA-approved compounds, as an inhibitor of hnRNP A1. In a riluzole-bead coupled binding assay and in surface plasmon resonance imaging analyses, riluzole was found to directly bind to hnRNP A1 and inhibited IRES activity via effects on ITAF/RNA-binding. Riluzole also demonstrated synergistic anti-glioblastoma (GBM) affects with mTOR inhibitors in vitro and in GBM xenografts in mice. These data suggest that repurposing riluzole, used in conjunction with mTOR inhibitors, may serve as an effective therapeutic option in glioblastoma.

Tissue reactions and stochastic effects after exposure to ionising radiation are variable between individuals but the factors and mechanisms governing individual responses are not well understood. Individual responses can be measured at different levels of biological organization and using different endpoints following varying doses of radiation, including: cancers, non-cancer diseases and mortality in the whole organism; normal tissue reactions after exposures; and, cellular endpoints such as chromosomal damage and molecular alterations. There is no doubt that many factors influence the responses of people to radiation to different degrees. In addition to the obvious general factors of radiation quality, dose, dose rate and the tissue (sub)volume irradiated, recognized and potential determining factors include age, sex, life style (e.g., smoking, diet, possibly body mass index), environmental factors, genetics and epigenetics, stochastic distribution of cellular events, and systemic comorbidities such as diabetes or viral infections. Genetic factors are commonly thought to be a substantial contributor to individual response to radiation. Apart from a small number of rare monogenic diseases such as ataxia telangiectasia, the inheritance of an abnormally responsive phenotype among a population of healthy individuals does not follow a classical Mendelian inheritance pattern. Rather it is considered to be a multi-factorial, complex trait.

The propagation of uncertainties in reaction cross sections and rates of neutron-, proton-, and alpha-induced reactions into the final isotopic abundances obtained in nucleosynthesis models is an important issue in studies of nucleosynthesis and Galactic Chemical Evolution. We developed a Monte Carlo method to allow large-scale postprocessing studies of the impact of nuclear uncertainties on nucleosynthesis. Temperature-dependent rate uncertainties combining realistic experimental and theoretical uncertainties are used. From detailed statistical analyses uncertainties in the final abundances are derived as probability density distributions. Furthermore, based on rate and abundance correlations an automated procedure identifies the most important reactions in complex flow patterns from superposition of many zones or tracers. The method so far was already applied to a number of nucleosynthesis processes. Here we focus on the production of p-nuclei in white dwarfs exploding as thermonuclear (type Ia) supernovae. We find generally small uncertainties in the final abundances despite of the dominance of theoretical nuclear uncertainties. A separate analysis of low- and high-density regions indicates that the total uncertainties are dominated by the high-density regions., 6 pages, 3 figures; invited talk at 13th Int. Conf. on Nucleus-Nucleus Collisions, Saitama, Japan, 2018. To be published in JPS Conf. Proceedinngs; summarizes results from arXiv:1807.11475; v2: updated reference and receipt date; as accepted for publication

Phosgene is classified as a chemical warfare agent, yet data on its short-duration high concentration toxicity in a nose-only exposure rat model is sparse and inconsistent. Hence, an exposure system for short term/high concentration exposure was developed and characterized. Herein, we report the median lethal concentration (LC(50)) for a 10-min nasal exposure of phosgene in a 24-h rat survival model. Male Wistar rats (Envigo) weighing 180-210 g on the day of exposure, were exposed to phosgene gas via nose-only inhalation using a system specifically designed to allow the simultaneous exposure and quantification of phosgene. After 24 h, the surviving rats were euthanized, the lung/body mass ratio determined, and lung tissues analyzed for histopathology. Increased terminal airway edema in the lungs located primarily at the alveoli (resulting in an increased lung/body mass ratio) coincided with the observed mortality. An LC(50) value of 129.2 mg/m(3) for a 10-min exposure was determined. Furthermore, in agreement with other highly toxic compounds, this study reveals a LC(50) concentration value supportive of a non-linear toxic load model, where the toxic load exponent is > 1 (n(e) = 1.17). Thus, in line with other chemical warfare agents, phosgene toxicity is predicted to be more severe with short-duration, high-concentration exposures than long-duration, low-concentration exposures. This model is anticipated to be refined and developed to screen novel therapeutics against relevant short-term high concentration phosgene exposures expected from a terrorist attack, battlefield deployment, or industrial accident.

Several thousand tracers from a 2D model of a thermonuclear supernova were used in a Monte Carlo post-processing approach to determine p-nuclide abundance uncertainties originating from nuclear physics uncertainties in the reaction rates.

TP53 is a tumor suppressor gene that is mutated in 50% of cancers, and its function is tightly regulated by the E3 ligase, Mdm2. Both p53 and Mdm2 are localized in the cell nucleus, a site that is impervious to therapeutic regulation by most antibodies. We identified a cell-penetrating lupus monoclonal anti-DNA antibody, mAb 3E10, that targets the nucleus, and we engineered mAb 3E10 to function as an intranuclear transport system to deliver therapeutic antibodies into the nucleus as bispecific single chain Fv (scFv) fragments. Bispecific scFvs composed of 3E10 include PAb421 (3E10-PAb421) that binds p53 and restores the function of mutated p53, and 3G5 (3E10-3G5) that binds Mdm2 and prevents destruction of p53 by Mdm2. We documented the therapeutic efficacy of these bispecific scFvs separately in previous studies. In this study, we show that combination therapy with 3E10-PAb421 and 3E10-3G5 augments growth inhibition of cells with p53 mutations compared to the effect of either antibody alone. By contrast, no enhanced response was observed in cells with wild-type p53 or in cells homozygous null for p53.

Uncertainties in nucleosynthesis models originating from uncertainties in astrophysical reaction rates were estimated in a Monte Carlo variation procedure. Thousands of rates were simultaneously varied within individual, temperature-dependent errors to calculate their combined effect on final abundances. After a presentation of the method, results from application to three different nucleosynthesis processes are shown: the $\gamma$-process and the s-process in massive stars, and the main s-process in AGB stars (preliminary results). Thermal excitation of nuclei in the stellar plasma and the combined action of several reactions increase the final uncertainties above the level of the experimental errors. The total uncertainty, on the other hand, remains within a factor of two even in processes involving a large number of unmeasured rates, with some notable exceptions for nuclides whose production is spread over several stellar layers and for s-process branchings., Comment: 8 pages, 4 figures; Proceedings of OMEG 2017, Daejeon, Korea, June 27-30, 2017; to appear in AIP Conf. Proc

We have investigated the revival of a shock wave by nuclear burning reactions at the central region of core-collapse supernovae. For this purpose, we performed hydrodynamic simulations of core collapse and bounce for 15 M⊙ progenitor model, using ZEUS-MP code in axi-symmetric coordinates. Our numerical code is equipped with a simple nuclear reaction network including 13 α nuclei form 4He to 56Ni, and accounting for energy feedback from nuclear reactions as well as neutrino heating and cooling. We found that the energy released by nuclear reactions is significantly helpful in accelerating shock waves and is able to produce energetic explosion even if the input neutrino luminosity is low.

Background Although early reperfusion is the most desirable intervention after ischemic myocardial insult, it may add to damage through oxidative stress. Objectives This study investigated the cardioprotective effects of a single intravenous dose of heat shock protein-72 (HSP72) coupled to a single-chain variable fragment (Fv) of monoclonal antibody 3E10 (3E10Fv) in a rabbit ischemia-reperfusion model. The Fv facilitates rapid transport of HSP72 into cells, even with intact membranes. Methods A left coronary artery occlusion (40 min) reperfusion (3 h) model was used in 31 rabbits. Of these, 12 rabbits received the fusion protein (Fv-HSP72) intravenously. The remaining 19 control rabbits received a molar equivalent of 3E10Fv alone (n = 6), HSP72 alone (n = 6), or phosphate-buffered saline (n = 7). Serial echocardiographic examinations were performed to assess left ventricular function before and after reperfusion. Micro–single-photon emission computed tomography imaging of 99mTc-labeled annexin-V was performed with micro–computed tomography scanning to characterize apoptotic damage in vivo, followed by gamma counting of the excised myocardial specimens to quantify cell death. Histopathological characterization of the myocardial tissue and sequential cardiac troponin I measurements were also undertaken. Results Myocardial annexin-V uptake was 43% lower in the area at risk (p = 0.0003) in Fv-HSP72–treated rabbits compared with control animals receiving HSP72 or 3E10Fv alone. During reperfusion, troponin I release was 42% lower and the echocardiographic left ventricular ejection fraction 27% higher in the Fv-HSP72–treated group compared with control animals. Histopathological analyses confirmed penetration of 3E10Fv-containing molecules into cardiomyocytes in vivo, and treatment with Fv-HSP72 showed fewer apoptotic nuclei compared with control rabbits. Conclusions Single-dose administration of Fv-HSP72 fusion protein at the time of reperfusion reduced myocardial apoptosis by almost one-half and improved left ventricular functional recovery after myocardial ischemia-reperfusion injury in rabbits. It might have potential to serve as an adjunct to early reperfusion in the management of myocardial infarction.

Insects with complex life cycles show a variety of developmental strategies when faced with low nutrient conditions requiring trade-offs in timing and nutrient storage. Previously documented strategies among insects include plasticity or canalization (fixation) of the pupation threshold, postthreshold developmental time, and adult size or stored resources. Using four models previously developed by Juliano et al. (2004), we tested the plasticity of these traits in the aquatic larvae of the frog-biting midge, Corethrella appendiculata Grabham. Rates of prey consumption and timing of pupation of fourth instar midges were compared across a range of prey densities. Model comparisons revealed that final instar C. appendiculata larvae exhibit a canalized pupation threshold followed by a canalized lengthy postthreshold period. Males entered the final instar earlier and reached a significantly lower threshold while females entered later and averaged more prey consumed daily, suggesting differing developmental strategies between sexes. Plasticity in body and egg clutch sizes of females from differing nutrient treatments was observed, but all females produced eggs autogenously. The canalized developmental pattern and long postthreshold period displayed in C. appendiculata is hypothesized to be related to the midge's preference for relatively permanent aquatic habitats.

The success of an invasive species in a new region depends on its interactions with ecologically similar resident species. Invasions by disease vector mosquitoes are important as they may have ecological and epidemiological consequences. Potential interactions of a recent invasive mosquito, Aedes japonicus Theobald, with resident species in Virginia were evaluated by sampling larvae from containers and trapping adults. Distinct species compositions were observed for artificial containers and rock pools, with Ae. albopictus most abundant in the former and Ae. japonicus in the latter. However, these two species were found to co-occur in 21.2% of containers sampled. Among the six mosquito species most common in containers from May through September, 2006, only interspecific associations of Ae. japonicus with Aedes albopictus (Skuse) and Aedes triseriatus (Say) were significant, and both were negative. In addition to differences in habitat preference, mean crowding estimates suggest that interspecific repulsion may contribute to the significant negative associations observed between these species. High relative abundances of late instars and pupae of Ae. japonicus seem to provide this species with a mechanism of evading competition with Ae. albopictus, facilitating their coexistence in artificial containers. Although annual fluctuations were observed, trends in adult populations over a 6-yr period provide no evidence of declines. In summary, this survey of diverse container types and all life stages provided only limited evidence for competitive displacements or reductions of resident container species by Ae. japonicus, as observed elsewhere in its invasive range.

The therapeutic use of antibodies is restricted by the limited access of antibodies to intracellular compartments. To overcome this limitation, we developed a cell-penetrating monoclonal antibody, mAb 3E10, as an intracellular delivery vehicle for the intracellular and intranuclear delivery of antibodies constructed as bispecific single-chain Fv fragments. Because MDM2 is an important target in cancer therapy, we selected monoclonal antibody (mAb) 3G5 for intracellular transport. mAb 3G5 binds MDM2 and blocks binding of MDM2 to p53. Here, we show that the resulting 3E10-3G5 bispecific antibody retains cell-penetrating and MDM2-binding activity, increases tumor p53 levels, and inhibits growth of MDM2-addicted tumors. The use of cell-penetrating bispecific antibodies in targeted molecular therapy will significantly broaden the spectrum of accessible intracellular targets and may have a profound impact in cancer therapy. Mol Cancer Ther; 11(10); 2169–73. ©2012 AACR.

Upon mating, male mosquitoes transfer accessory gland proteins (Acps) that induce refractoriness to further mating in females. This can also occur because of cross-insemination by males of related species, a process known as mating interference (satyrization). This mechanism could explain the competitive displacement of resident Aedes aegypti by the invasive Aedes albopictus where they co-occur. We tested this hypothesis in mosquito populations in Florida. A new polymerase chain reaction species diagnostic applied to sperm dissected from 304 field-collected females revealed bidirectional cross-mating in five (1.6%) individuals. Cross-injections of females with Acps showed that Ae. albopictus males induced monogamy in heterospecific females but not Ae. aegypti males. Despite its low frequency in the areas under study, the first evidence of cross-mating in nature and the asymmetric effect of Acps on mating suggest that satyrization may have initially contributed to the observed competitive reduction of Ae. aegypti by invasive Ae. albopictus in many areas.

Foxp3 is a nuclear transcription factor that is both a tumor suppressor factor and regulator of T-regulatory cell (Treg) function, and is a potential therapeutic target in both autoimmunity and cancer. In order to distinguish molecular pathways responsible for these separate Foxp3 functions, deletion mutants of Foxp3 proteins were transduced and analyzed for cytotoxic activity in human cancer cell lines Skov3, MDA-MB-231, MCF-7 and Jurkat. Human Foxp3 cDNA mutants were amplified and ligated to produce plasmids for direct cell transfection. Constructs were produced and confirmed by DNA sequencing. Lipofectamine 2000 was used for plasmid transfection. Foxp3 cells were then examined. The results of our experiments reveal retention of tumor suppressor function in the absence of NFAT binding and transcriptional activation required for Treg function. Our results have significant implications for the design of autoimmune and cancer therapies that target Foxp3 and Treg cells.

Within 2 yr of the arrival of the invasive container mosquito Aedes albopictus (Skuse), the previously dominant invasive mosquito Aedes aegypti (L.) disappeared from many Florida cemeteries. At some cemeteries, however, Ae. aegypti populations seem stable despite Ae. albopictus invasion. We sought to understand this variation in the outcome (exclusion, coexistence) of this invasion, given that previous experiments show that Ae. albopictus is the superior larval competitor. We tested experimentally the hypothesis that climate-dependent egg survivorship differs between exclusion and coexistence cemeteries and that differences in invasion outcome are associated with microclimate. Viability of eggs oviposited in the laboratory and suspended in vases at six cemeteries was significantly greater for Ae. aegypti than for Ae. albopictus, and greater in 2001 than in 2006. Cemeteries differed significantly in egg survivorship of Ae. albopictus, but not of Ae. aegypti, which is consistent with the hypothesis that Ae. albopictus suffers site-specific, climate-driven egg mortality that mitigates the competitive superiority of larval Ae. albopictus. Principal component (PC) analysis of microclimate records from vases during the experiments yielded three PCs accounting for >96% of the variance in both years of experiments. Multivariate analysis of variance of the three PCs revealed significant microclimate differences among the six cemeteries and between exclusion versus coexistence cemeteries. Stepwise logistic regression of egg survivorship versus microclimate PCs yielded significant fits for both species, and twice as much variance explained for Ae. albopictus as for Ae. aegypti in both years, Higher mortalities in 2006 were associated with high average daily maximum temperatures in vases, with lethal thresholds for both species at ≈40°C. From 1990 to 2007, vase occupancy by Ae. albopictus increased and that by Ae. aegypti decreased, with increasing seasonal precipitation at one well-sampled cemetery. Results support the hypothesis that locally variable climate-driven mortality of Ae. albopictus eggs contributes to patterns of exclusion of, or coexistence with, Ae. aegypti.

Background and Purpose— This study investigated the effects of intravenous recombinant Fv-Hsp70 protein on infarction volume and behavior after experimental ischemic stroke. Methods— Focal cerebral ischemia was produced by occluding the middle cerebral artery using the intraluminal suture technique. Rats subjected to 2 hours of focal ischemia were allowed to survive 24 hours. At 2¼ hours and 3 hours after onset of ischemia, Fv-Hsp70 recombinant protein (0.5 mg/kg) or saline was injected through the tail vein. Sensorimotor function and infarction volume were assessed at 24 hours after ischemia. Results— Administration of Fv-Hsp70 after focal cerebral ischemia significantly decreased infarct volume by 68% and significantly improved sensorimotor function compared with the saline-treated control group. Western blots showed Fv-Hsp70 in ischemic but not in control brain; and Fv-Hsp70 suppressed endogenous Hsp70. Conclusion— Fv-Hsp70 protected the ischemic brain in this experimental stroke model.

Native predators are postulated to have an important role in biotic resistance of communities to invasion and community resilience. Effects of predators can be complex, and mechanisms by which predators affect invasion success and impact are understood for only a few well-studied communities. We tested experimentally whether a native predator limits an invasive species' success and impact on a native competitor for a community of aquatic insect larvae in water-filled containers. The native mosquito Aedes triseriatus alone had no significant effect on abundance of the invasive mosquito Aedes albopictus. The native predatory midge Corethrella appendiculata, at low or high density, significantly reduced A. albopictus abundance. This effect was not caused by trait-mediated oviposition avoidance of containers with predators, but instead was a density-mediated effect caused by predator-induced mortality. The presence of this predator significantly reduced survivorship of the native species, but high predator density also significantly increased development rate of the native species when the invader was present, consistent with predator-mediated release from interspecific competition with the invader. Thus, a native predator can indirectly benefit its native prey when a superior competitor invades. This shows the importance of native predators as a component of biodiversity for both biotic resistance to invasion and resilience of a community perturbed by successful invasion.

mTORC2 is a multiprotein kinase composed of mTOR, mLST8, PRR5, mSIN1 and Rictor. The complex is insensitive to rapamycin and has demonstrated functions controlling cell growth, motility, invasion and cytoskeletal assembly. mTORC2 is the major hydrophobic domain kinase which renders Akt fully active via phosphorylation on serine 473. We isolated Hsp70 as a putative Rictor interacting protein in a yeast two-hybrid assay and confirmed this interaction via co-immunoprecipitation and colocalization experiments. In cells expressing an antisense RNA targeting Hsp70, mTORC2 formation and activity were impaired. Moreover, in cells lacking Hsp70 expression, mTORC2 activity was inhibited following heat shock while controls demonstrated increased mTORC2 activity. These differential effects on mTORC2 activity were specific, in that mTORC1 did not demonstrate Hsp70-dependent alterations under these conditions. These data suggest that Hsp70 is a component of mTORC2 and is required for proper assembly and activity of the kinase both constitutively and following heat shock.