Reovirus activates human dendritic cells to promote innate antitumor immunity

Oncolytic viruses can exert their antitumor activity via direct oncolysis or activation of antitumor immunity. Although reovirus is currently under clinical investigation for the treatment of localized or disseminated cancer, any potential immune contribution to its efficacy has not been addressed. This is the first study to investigate the ability of reovirus to activate human dendritic cells (DC), key regulators of both innate and adaptive immune responses. Reovirus induced DC maturation and stimulated the production of the proinflammatory cytokines IFN-alpha, TNF-alpha, IL-12p70, and IL-6. Activation of DC by reovirus was not dependent on viral replication, while cytokine production (but not phenotypic maturation) was inhibited by blockade of PKR and NF-kappaB signaling. Upon coculture with autologous NK cells, reovirus-activated DC up-regulated IFN-gamma production and increased NK cytolytic activity. Moreover, short-term coculture of reovirus-activated DC with autologous T cells also enhanced T cell cytokine secretion (IL-2 and IFN-gamma) and induced non-Ag restricted tumor cell killing. These data demonstrate for the first time that reovirus directly activates human DC and that reovirus-activated DC stimulate innate killing by not only NK cells, but also T cells, suggesting a novel potential role for T cells in oncolytic virus-induced local tumor cell death. Hence reovirus recognition by DC may trigger innate effector mechanisms to complement the virus's direct cytotoxicity, potentially enhancing the efficacy of reovirus as a therapeutic agent.     

A method for aligning RNA secondary structures and its application to RNA motif detection

Background  

Alignment of RNA secondary structures is important in studying functional RNA motifs. In recent years, much progress has been made in RNA motif finding and structure alignment. However, existing tools either require a large number of prealigned structures or suffer from high time complexities. This makes it difficult for the tools to process RNAs whose prealigned structures are unavailable or process very large RNA structure databases.     

Experimental flow studies in exact-replica phantoms of atherosclerotic carotid bifurcations under steady input conditions

Extensive flow studies are conducted in two carotid bifurcation flow phantoms. These phantoms exactly replicate the lumen of the plaque excised intact from two patients with severe carotid atherosclerosis. The input flow into the phantom's common carotid artery is steady. Novel scanning techniques for flow visualization and particle image velocimetry are used. In addition, a novel boundary treatment technique is employed in velocimetry to extract first order accurate velocity gradients at walls. The data show that the flow fields are highly three-dimensional. Numerous separation and recirculation zones dominate the flow domain, except at the lowest Reynolds numbers. The separation regions are often so severe that highly directed internal jets form. At high Reynolds numbers, the flows become unsteady and chaotic, even though the input flow is steady. Flow fields have large regions of energetic flow and almost stagnant recirculation zones. These recirculation zones range in size from the full size of the arteries to zones within crevasses smaller than 1 mm. Velocity field and streamline patterns conform well to the lumen geometry. The streamlines are highly tortuous. Stagnation points correlate well with the topological features of the stenosis. Vorticity maps confirm the highly complex and three dimensional nature of the flow. Wall shear stresses at the stenoses are estimated to be on the order of 10 Pa. These studies conclusively show that the nature of the flow in the diseased bifurcation is primarily dictated by the lumen geometry.     

Use of a sensitive EnVision +-based detection system for Western blotting: avoidance of streptavidin binding to endogenous biotin and biotin-containing proteins in kidney and other tissues

Western blotting remains a central technique in confirming identities of proteins, their quantitation and analysis of various isoforms. The biotin-avidin/streptavidin system is often used as an amplification step to increase sensitivity but in some tissues such as kidney, "nonspecific" interactions may be a problem due to high levels of endogenous biotin-containing proteins. The EnVision system, developed for immunohistochemical applications, relies on binding of a polymeric conjugate consisting of up to 100 peroxidase molecules and 20 secondary antibody molecules linked directly to an activated dextran backbone, to the primary antibody. This study demonstrates that it is also a viable and sensitive alternative detection system in Western blotting applications.     

Description of first germinal mosaic mutation identified in dominant skeletal mutation experiments and considerations about how to deal with this kind of spontaneous mutation in analyses

Germinal mosaicism is a well-established mechanism by which new spontaneous mutations enter the human population, but it is only rather recently that clusters of mutations arising in that way have been acknowledged and dealt with in specific-locus experiments on male mice. This paper reports the first cluster of germinal mosaic mutations to have been identified in experiments on the induction of dominant skeletal mutations. The mutation was detected in six offspring of a control male from the radiation part of an Assessment-of-Dominant-Damage (ADD) experiment. Reasons are provided to explain why this one litter of six mutants was excluded from the analysis of induction of dominant mutations causing the more common skeletal anomalies, which is reported in another paper. The effects of excluding this litter from that analysis are fully described. There is discussion of why such clusters should be included in some analyses but omitted in others. They should certainly always be reported because, in some cases, they can have a major impact on conclusions. Details on this one cluster of FCGM mutations provide numerous examples of how a dominant skeletal mutation that causes rare effects can also cause many of the more common anomalies.     

Using virally expressed melanoma cDNA libraries to identify tumor-associated antigens that cure melanoma

Multiple intravenous injections of a cDNA library, derived from human melanoma cell lines and expressed using the highly immunogenic vector vesicular stomatitis virus (VSV), cured mice with established melanoma tumors. Successful tumor eradication was associated with the ability of mouse lymphoid cells to mount a tumor-specific CD4(+) interleukin (IL)-17 recall response in vitro. We used this characteristic IL-17 response to screen the VSV-cDNA library and identified three different VSV-cDNA virus clones that, when used in combination but not alone, achieved the same efficacy against tumors as the complete parental virus library. VSV-expressed cDNA libraries can therefore be used to identify tumor rejection antigens that can cooperate to induce anti-tumor responses. This technology should be applicable to antigen discovery for other cancers, as well as for other diseases in which immune reactivity against more than one target antigen contributes to disease pathology.     

Redox heterogeneity of subsurface waters in the Mesoproterozoic ocean

A substantial body of evidence suggests that subsurface water masses in mid‐Proterozoic marine basins were commonly anoxic, either euxinic (sulfidic) or ferruginous (free ferrous iron). To further document redox variations during this interval, a multiproxy geochemical and paleobiological investigation was conducted on the approximately 1000‐m‐thick Mesoproterozoic (Lower Riphean) Arlan Member of the Kaltasy Formation, central Russia. Iron speciation geochemistry, supported by organic geochemistry, redox‐sensitive trace element abundances, and pyrite sulfur isotope values, indicates that basinal calcareous shales of the Arlan Member were deposited beneath an oxygenated water column, and consistent with this interpretation, eukaryotic microfossils are abundant in basinal facies. The Rhenium–Osmium (Re–Os) systematics of the Arlan shales yield depositional ages of 1414 ± 40 and 1427 ± 43 Ma for two horizons near the base of the succession, consistent with previously proposed correlations. The presence of free oxygen in a basinal environment adds an important end member to Proterozoic redox heterogeneity, requiring an explanation in light of previous data from time‐equivalent basins. Very low total organic carbon contents in the Arlan Member are perhaps the key—oxic deep waters are more likely (under any level of atmospheric O₂) in oligotrophic systems with low export production. Documentation of a full range of redox heterogeneity in subsurface waters and the existence of local redox controls indicate that no single stratigraphic section or basin can adequately capture both the mean redox profile of Proterozoic oceans and its variance at any given point in time.     

Are exhaled nitric oxide measurements using the portable NIOX MINO repeatable?

Background

Exhaled nitric oxide is a non-invasive marker of airway inflammation and a portable analyser, the NIOX MINO (Aerocrine AB, Solna, Sweden), is now available. This study aimed to assess the reproducibility of the NIOX MINO measurements across age, sex and lung function for both absolute and categorical exhaled nitric oxide values in two distinct groups of children and teenagers.

Methods

Paired exhaled nitric oxide readings were obtained from 494 teenagers, aged 16-18 years, enrolled in an unselected birth cohort and 65 young people, aged 6-17 years, with asthma enrolled in an interventional asthma management study.

Results

The birth cohort participants showed a high degree of variability between first and second exhaled nitric oxide readings (mean intra-participant difference 1.37 ppb, 95% limits of agreement -7.61 to 10.34 ppb), although there was very close agreement when values were categorised as low, normal, intermediate or high (kappa = 0.907, p < 0.001). Similar findings were seen in subgroup analyses by sex, lung function and asthma status. Similar findings were seen in the interventional study participants.

Conclusions

The reproducibility of exhaled nitric oxide is poor for absolute values but acceptable when values are categorised as low, normal, intermediate or high in children and teenagers. One measurement is therefore sufficient when using categorical exhaled nitric oxide values to direct asthma management but a mean of at least two measurements is required for absolute values.