c-Src drives intestinal regeneration and transformation

The non‐receptor tyrosine kinase c‐Src, hereafter referred to as Src, is overexpressed or activated in multiple human malignancies. There has been much speculation about the functional role of Src in colorectal cancer (CRC), with Src amplification and potential activating mutations in up to 20% of the human tumours, although this has never been addressed due to multiple redundant family members. Here, we have used the adult <i>Drosophila</i> and mouse intestinal epithelium as paradigms to define a role for Src during tissue homeostasis, damage‐induced regeneration and hyperplasia. Through genetic gain and loss of function experiments, we demonstrate that Src is necessary and sufficient to drive intestinal stem cell (ISC) proliferation during tissue self‐renewal, regeneration and tumourigenesis. Surprisingly, Src plays a non‐redundant role in the mouse intestine, which cannot be substituted by the other family kinases Fyn and Yes. Mechanistically, we show that Src drives ISC proliferation through upregulation of EGFR and activation of Ras/MAPK and Stat3 signalling. Therefore, we demonstrate a novel essential role for Src in intestinal stem/progenitor cell proliferation and tumourigenesis initiation <i>in vivo.</i&gt

Herpes Simplex Virus-2 Glycoprotein Interaction with HVEM Influences Virus-Specific Recall Cellular Responses at the Mucosa

Infection of susceptible cells by herpes simplex virus (HSV) requires the interaction of the HSV gD glycoprotein with one of two principal entry receptors, herpes virus entry mediator (HVEM) or nectins. HVEM naturally functions in immune signaling, and the gD-HVEM interaction alters innate signaling early after mucosal infection. We investigated whether the gD-HVEM interaction during priming changes lymphocyte recall responses in the murine intravaginal model. Mice were primed with attenuated HSV-2 expressing wild-type gD or mutant gD unable to engage HVEM and challenged 32 days later with virulent HSV-2 expressing wild-type gD. HSV-specific CD8+ T cells were decreased at the genital mucosa during the recall response after priming with virus unable to engage HVEM but did not differ in draining lymph nodes. CD4+ T cells, which are critical for entry of HSV-specific CD8+ T cells into mucosa in acute infection, did not differ between the two groups in either tissue. An inverse association between Foxp3+ CD4+ regulatory T cells and CD8+ infiltration into the mucosa was not statistically significant. CXCR3 surface expression was not significantly different among different lymphocyte subsets. We conclude that engagement of HVEM during the acute phase of HSV infection influences the antiviral CD8+ recall response by an unexplained mechanism

False positive dobutamine stress echocardiograms: Characterization of clinical, echocardiographic and angiographic findings

AbstractObjectives. This study was designed to characterize the clinical, echocardiographic and angiographic findings in patients who have regional wall motion abnormalities predictive of coronary artery disease on dobutamine stress echocardiograms, although coronary angiography reveals no critical stenoses.Background. The specificity of dobutamine stress echocardiography has been reported to be lower than its sensitivity; the sources of false positive findings on dobutamine stress echocardiograms have not been previously defined.Methods. Clinical and echocardiographic characteristics were retrospectively reviewed for patients who had both a dobutamine stress echocardiogram indicate of coronary artery disease on the basis of wall motion abnormalities and <50% stenoses reported on coronary angiography performed within 6 weeks of the echocardiogram. A 16-segment model was used to perform wall motion scoring. Angiograms were independently reviewed, and stenosis severity was quantified with the use of digital calipers.Results. Thirty-nine (11.4%) of 342 studies met criteria for false positive test results, which occurred predominantly in women (72%, p < 0.601). Regional wall motion abnormalities were evident more often in the posterior circulation (62%), and 65% of them were limited to the basal segments. Twelve (28%) of 43 wall motion abnormalities were associated with coronary stenoses of at least intermediate grade (lumen diameter 40.3% to 68.1%). Abnormalities confined to basal segments of the posterior circulation were unlikely to have associated coronary lesions (p = 0.03).Conclusions. False positive findings on dobutamine stress echocardiograms tend to involve small wall motion abnormalities that are frequently located in basal segments of the posterior myocardial circulation. Approximately one third of false positive results occurred in patients with intermediate-grade coronary stenoses, and these studies may reflect true inducible ischemia. Additional sources of false positive study results may include poor endocardial visualization and abnormal motion dye to tethering to the fibrous skeleton of the heart. Altered echocardiographic diagnostic criteria may be appropriate for small wall motion abnormalities confined to basal segments of the posterior circulation

Indoleamine 2,3-Dioxygenase and Its Therapeutic Inhibition in Cancer

The tryptophan catabolic enzyme indoleamine 2,3-dioxygenase-1 (IDO1) has attracted enormous attention in driving cancer immunosuppression, neovascularization, and metastasis. IDO1 suppresses local CD8 + T effector cells and natural killer cells and induces CD4 + T regulatory cells (iTreg) and myeloid-derived suppressor cells (MDSC). The structurally distinct enzyme tryptophan dioxygenase (TDO) also has been implicated recently in immune escape and metastatic progression. Lastly, emerging evidence suggests that the IDO1-related enzyme IDO2 may support IDO1-mediated iTreg and contribute to B-cell inflammed states in certain cancers. IDO1 and TDO are upregulated widely in neoplastic cells but also variably in stromal, endothelial, and innate immune cells of the tumor microenviroment and in tumor-draining lymph nodes. Pharmacological and genetic proofs in preclinical models of cancer have validated IDO1 as a cancer therapeutic target. IDO1 inhibitors have limited activity on their own but greatly enhance “immunogenic” chemotherapy or immune checkpoint drugs. IDO/TDO function is rooted in inflammatory programming, thereby influencing tumor neovascularization, MDSC generation, and metastasis beyond effects on adaptive immune tolerance. Discovery and development of two small molecule enzyme inhibitors of IDO1 have advanced furthest to date in Phase II/III human trials (epacadostat and navoximod, respectively). Indoximod, a tryptophan mimetic compound with a different mechanism of action in the IDO pathway has also advanced in multiple Phase II trials. Second generation combined IDO/TDO inhibitors may broaden impact in cancer treatment, for example, in addressing IDO1 bypass (inherent resistance) or acquired resistance to IDO1 inhibitors. This review surveys knowledge about IDO1 function and how IDO1 inhibitors reprogram inflammation to heighten therapeutic responses in cancer

Indoleamine 2,3-Dioxygenase and Its Therapeutic Inhibition in Cancer

The tryptophan catabolic enzyme indoleamine 2,3-dioxygenase-1 (IDO1) has attracted enormous attention in driving cancer immunosuppression, neovascularization, and metastasis. IDO1 suppresses local CD8 + T effector cells and natural killer cells and induces CD4 + T regulatory cells (iTreg) and myeloid-derived suppressor cells (MDSC). The structurally distinct enzyme tryptophan dioxygenase (TDO) also has been implicated recently in immune escape and metastatic progression. Lastly, emerging evidence suggests that the IDO1-related enzyme IDO2 may support IDO1-mediated iTreg and contribute to B-cell inflammed states in certain cancers. IDO1 and TDO are upregulated widely in neoplastic cells but also variably in stromal, endothelial, and innate immune cells of the tumor microenviroment and in tumor-draining lymph nodes. Pharmacological and genetic proofs in preclinical models of cancer have validated IDO1 as a cancer therapeutic target. IDO1 inhibitors have limited activity on their own but greatly enhance “immunogenic” chemotherapy or immune checkpoint drugs. IDO/TDO function is rooted in inflammatory programming, thereby influencing tumor neovascularization, MDSC generation, and metastasis beyond effects on adaptive immune tolerance. Discovery and development of two small molecule enzyme inhibitors of IDO1 have advanced furthest to date in Phase II/III human trials (epacadostat and navoximod, respectively). Indoximod, a tryptophan mimetic compound with a different mechanism of action in the IDO pathway has also advanced in multiple Phase II trials. Second generation combined IDO/TDO inhibitors may broaden impact in cancer treatment, for example, in addressing IDO1 bypass (inherent resistance) or acquired resistance to IDO1 inhibitors. This review surveys knowledge about IDO1 function and how IDO1 inhibitors reprogram inflammation to heighten therapeutic responses in cancer

Visualizing fusion of pseudotyped HIV-1 particles in real time by live cell microscopy

RIGHTS : This article is licensed under the BioMed Central licence at http://www.biomedcentral.com/about/license which is similar to the 'Creative Commons Attribution Licence'. In brief you may : copy, distribute, and display the work; make derivative works; or make commercial use of the work - under the following conditions: the original author must be given credit; for any reuse or distribution, it must be made clear to others what the license terms of this work are.Abstract Background Most retroviruses enter their host cells by fusing the viral envelope with the plasma membrane. Although the protein machinery promoting fusion has been characterized extensively, the dynamics of the process are largely unknown. Results We generated human immunodeficiency virus-1 (HIV-1) particles pseudotyped with the envelope (Env) protein of ecotropic murine leukemia virus eMLV to study retrovirus entry at the plasma membrane using live-cell microscopy. This Env protein mediates highly efficient pH independent fusion at the cell surface and can be functionally tagged with a fluorescent protein. To detect fusion events, double labeled particles carrying one fluorophor in Env and the other in the matrix (MA) domain of Gag were generated and characterized. Fusion events were defined as loss of Env signal after virus-cell contact. Single particle tracking of >20,000 individual traces in two color channels recorded 28 events of color separation, where particles lost the Env protein, with the MA layer remaining stable at least for a short period. Fourty-five events were detected where both colors were lost simultaneously. Importantly, the first type of event was never observed when particles were pseudotyped with a non-fusogenic Env. Conclusion These results reveal rapid retroviral fusion at the plasma membrane and permit studies of the immediate post-fusion events.Published versio

Chandra Observations of NGC 4438: An Environmentally Damaged Galaxy in the Virgo Cluster

We present results from a 25 ksec CHANDRA ACIS-S observation of galaxies NGC4438 and NGC4435 in the Virgo Cluster. X-ray emission in NGC4438 is observed in a ~700 pc nuclear region, a 2.3 kpc spherical bulge, and a network of filaments extending 4-10 kpc to the W and SW of the galaxy. The X-ray emission in all 3 regions is highly correlated to similar features observed in Halpha. Spectra of the filaments and bulge are well represented by a 0.4 keV MEKAL model with combined 0.3-2 keV intrinsic luminosity of 1.24x10^{40}erg/s, electron densities ~ 0.02-0.04 cm^{-3}, cooling times of 400-700 Myr and X-ray gas mass <~ 3.7x10^8 Msolar. In the nuclear region of NGC4438 X-ray emission is seen from the nucleus and from two outflow bubbles extending 360(730) pc to the NW(SE) of the nucleus. The spectrum of the NW outflow bubble plus nucleus is well fitted by an absorbed (n_H=1.9x10^{21} cm^{-2}) 0.58 keV MEKAL plasma model plus a heavily absorbed (n_H = 2.9 x10^{22} cm^{-2}) Gamma = 2, power law component. The electron density, cooling time, and X-ray gas mass in the NW outflow are ~0.5 cm^{-3}, 30 Myr and 3.5x10^6 Msolar. Weak X-ray emission is observed in the central region of NGC4435 with the peak of the hard emission coincident with the galaxy's optical center; while the peak of the soft X-ray emission is displaced 316 pc to the NE. The spectrum of NGC 4435 is well fitted by a non-thermal power law plus a thermal component from 0.2-0.3 keV diffuse ISM gas. We argue that the X-ray properties of gas outside the nuclear region in NGC4438 and in NGC4435 favor a high velocity, off-center collision between these galaxies ~ 100 Myr ago; while the nuclear X-ray emitting outflow gas in NGC4438 has been heated only recently (within ~ 1-2 Myr) by shocks (v_s ~ 600 kms^{-1}) possibly powered by a central AGN.Comment: 40 pages, 7 figures; minor changes to conform to published version, improved spectral fits to NGC 4435, improved figures 3,5; new figures 6b,

Functional Interaction between Herpes Simplex Virus Type 2 gD and HVEM Transiently Dampens Local Chemokine Production after Murine Mucosal Infection

Herpes virus entry mediator (HVEM) is one of two principal receptors mediating herpes simplex virus (HSV) entry into murine and human cells. It functions naturally as an immune signaling co-receptor, and may participate in enhancing or repressing immune responses depending on the natural ligand used. To investigate whether engagement of HVEM by HSV affects the in vivo response to HSV infection, we generated recombinants of HSV-2(333) that expressed wild-type gD (HSV-2/gD) or mutant gD able to bind to nectin-1 (the other principal entry receptor) but not HVEM. Replication kinetics and yields of the recombinant strains on Vero cells were indistinguishable from those of wild-type HSV-2(333). After intravaginal inoculation with mutant or wild-type virus, adult female C57BL/6 mice developed vaginal lesions and mortality in similar proportions, and mucosal viral titers were similar or lower for mutant strains at different times. Relative to HSV-2/gD, percentages of HSV-specific CD8+ T-cells were similar or only slightly reduced after infection with the mutant strain HSV-2/gD-Δ7-15, in all tissues up to 9 days after infection. Levels of HSV-specific CD4+ T-cells five days after infection also did not differ after infection with either strain. Levels of the cytokine IL-6 and of the chemokines CXCL9, CXCL10, and CCL4 were significantly lower in vaginal washes one day after infection with HSV-2/gD compared with HSV-2/gD-Δ7-15. We conclude that the interaction of HSV gD with HVEM may alter early innate events in the murine immune response to infection, without significantly affecting acute mortality, morbidity, or initial T-cell responses after lethal challenge

Copolymerization Studies of Vinyl Chloride and Vinyl Acetate with Ethylene Using a Transition-Metal Catalyst

Since the advent of Ziegler−Natta polymerization of ethylene, attempts have been made to extend coordination polymerization to commercially important monomers with polar functionality. In this study we examined the copolymerization of perdeuterated vinyl chloride (VC) and perdeuterated vinyl acetate (VA) with ethylene using a tridentate Fe(II) dichloride pyridine diimine metal catalyst. The resulting ethylene oligomers were examined by GC/MS and ^2H NMR spectroscopy. It was shown that VC was inserted once for every ∼180 ethylene monomers and VA was inserted once for every ∼350 ethylene monomers. VC and VA behave as comonomers for coordination/insertion polymerizations with ethylene. However, we find that insertion with either monomer leads to termination of the growing chain via β-elimination processes. The deuterium atoms are exclusively located at the olefin terminus for each of the monomers