PDB ID : 3UX1 Title : Structural Characterization of Adeno-Associated Virus Serotype 9 Authors : DiMattia, M.A.; Nam, H.-J.; Van Vliet, K.; Mitchell, M.; McCall, A.; Bennett, A.; Gurda, B.; McKenna, R.; Potter, M.; Sakai, Y.; Byrne, B.J.; Muzyczka, N.; Aslanidi, G.; Zolotukhin, S.; Olson, N.; Sinkovitis, R.; Baker, T.S.; AgbandjeMcKenna, M. Deposited on : 2011-12-03 Resolution : 2.80 Å(reported)

Non-return-valves are commonly used across the range of industries for prevention of reverse flow conditions where they often exhibit premature failure due to self-sustained oscillations that are a result of fluid structure interactions. This paper details the design of a proof of concept hardware-in-the-loop (HWIL) simulator and its controller for simulating fluid-structure interactions of a non-return-valve with an arbitrary air system geometry. A numerical model of an airbitrary air system is coupled to the physical non-return-valve via a purpose built fast acting control valve and associated instrumentation to simulate both stable and unstable vibrations. The vibrations of the non-return-valve in the HWIL simulator are compared against its vibrations in the real air system and are found to have an excellent agreement for a range of massflows and air system volumes. The simulator described in this paper can be used for simulating other types of passive flow control devices with arbitrary air systems thereby offering potential for significantly reducing the cost of full system tests.

This paper presents a theoretical model for predicting the behavior of electro-active polymer actuators. This model takes the form of an array of masses with the interconnecting forces derived using Hooke's law coupled with an electrostatic force, and is solved using time-marching integration. Forming the model in this way allows the response of actuators of various sizes and pre-strains to be simulated with a range of electrode designs and mechanically stiffened sections. To validate the response of the model a series of Nusil MED4905 polydimethylsiloxane actuators were built and tested with differing electrode designs and with additional mechanical stiffening of the membrane. Through a comparison of maximum electrode displacement, with applied electrode voltage, tracking of a grid on the membrane surface and dynamic testing, the model was found to show excellent agreement with the experimental results.

Practical observations of the nonreturn valve wear in aero-engine cabin-bleed systems suggest that such valves are subject to unstable behavior. A theoretical model for the prediction of nonreturn valve instabilities in air systems is proposed and a nonlinear state-space model of the nonreturn valve and air volume interaction is derived from first principles. Experimental work is used to identify both the dynamic characteristics and the flow properties of the valve, which are used to identify the coefficients within the model. Through frequency analysis of valve oscillatory behavior, the levels of damping within the system are identified. Finally, using a local linearization of the state-space model an explicit mathematical prediction of valve stability is derived based on system parameters. These predictions are used to generate a map of the transition from stable to unstable system behavior for low-speed air flow, which is in excellent agreement with experimental data.

TLR-induced innate immunity and inflammation are mediated by signaling cascades leading to activation of the MAPK family of Ser/Thr protein kinases, including p38 MAPK, which controls cytokine release during innate and adoptive immune responses. Failure to terminate such inflammatory reactions may lead to detrimental systemic effects, including septic shock and autoimmunity. In this study, we provide genetic evidence of a critical and nonredundant role of MAPK phosphatase (MKP)-1 in the negative control of MAPK-regulated inflammatory reactions in vivo. MKP-1-/- mice are hyperresponsive to low-dose LPS-induced toxicity and exhibit significantly increased serum TNF-alpha, IL-6, IL-12, MCP-1, IFN-gamma, and IL-10 levels after systemic administration of LPS. Furthermore, absence of MKP-1 increases systemic levels of proinflammatory cytokines and exacerbates disease development in a mouse model of rheumatoid arthritis. When activated through TLR2, TLR3, TLR4, TLR5, and TLR9, bone marrow-derived MKP-1-/- macrophages exhibit increased cytokine production and elevated expression of the differentiation markers B7.2 (CD86) and CD40. MKP-1-deficient macrophages also show enhanced constitutive and TLR-induced activation of p38 MAPK. Based on these findings, we propose that MKP-1 is an essential component of the intracellular homeostasis that controls the threshold and magnitude of p38 MAPK activation in macrophages, and inflammatory conditions accentuate the significance of this regulatory function.

ABSTRACT Coccidioidomycosis is a common fungal infection acquired in the southwestern United States. This is the first study in over 2 decades to determine the seroincidence of Coccidioides immitis infections among U.S. military members performing training exercises in an area of endemicity. Only 8% of participants were aware of coccidioidomycosis, despite the majority having visited or lived previously in an area of endemicity. One (0.6%) of the 178 participants developed “definite” serologic evidence of infection over a 5-week training period; four (2.3%) additional patients developed “possible” coccidioidomycosis infections. None had complicated disease. The calculated annual incidence ranged from 6 to 32%. This study suggests that the risk of serious coccidioidomycosis is low among military personnel during desert training exercises; however, disease incidence may vary depending on specific activities and geographic factors. Due to the potential morbidity and mortality of this infection, preventative strategies, including vaccine development, are advocated.

Conventional methods for rAAV purification that are based on cesium chloride ultracentrifugation have often produced vector preparations of variable quality and resulted in significant loss of particle infectivity. We report here several novel purification strategies that involve the use of non-ionic iodixanol gradients followed by ion exchange or heparin affinity chromatography by either conventional or HPLC columns. These methods result in more than 50% recovery of rAAV from a crude lysate and routinely produce vector that is more than 99% pure. More importantly, the new purification procedures consistently produce rAAV stocks with particle-to-infectivity ratios of less than 100, which is significantly better than conventional methods. The new protocol increases the overall yield of infectious rAAV by at least 10-fold and allows for the complete purification of rAAV in 1 working day. Several of these methods should also be useful for large-scale production.

The green fluorescent protein (GFP) has previously been adapted as a reporter for gene transfer and expression in mammalian cells in culture and in tissue sections. Herein is described a new method for detecting GFP in situ within epithelia accessible to fiberoptic endoscopy by incorporating fluorescent filters for detection of GFP into an existing fiberoptic endoscopy system. This device was used to detect expression of GFP from adeno-associated virus (AAV; dose of 3 × 107 IU) and adenovirus (Ad; dose of 1 × 109 to 1 × 1010 p.f.u.) vectors within the bronchial epithelium of New Zealand white rabbits. GFP expression was confirmed by fluorescence-activated cell sorting (FACS), direct fluorescence microscopy of cytospin preparations of brushed cells, and by fluorescence microscopy of fixed tissue sections. This reporter gene/detection system was then used to track the time course of expression of the AAV vector in the bronchial epithelium over the first 30 days after administration. The transduction frequency in the treated region of the epithelium peaked at around 50% at 21 days after transduction. Vector expression was still present at around 20% efficiency at 30 days after administration. This example indicates how this method could be used to reliably track gene transfer in living animals or patients.

We constructed gfph, a synthetic version of the jellyfish Aequorea victoria green fluorescent protein (gfp) cDNA that is adapted for high-level expression in mammalian cells, especially those of human origin. A total of 92 base substitutions were made in 88 codons in order to change the codon usage within the gfp10 coding sequence so that it was more appropriate for expression in mammalian cells. We also describe a series of versatile recombinant adeno-associated virus and adenovirus vectors for delivery and expression of genes into mammalian cells and, using these vectors, demonstrate the efficient transduction and expression of the gfph gene in the human cell line 293 and also in vivo, within neurosensory cells of guinea pig eye. Cells infected with recombinant adeno-associated virus-GFPH can be readily sorted by fluorescence-activated cell sorting, suggesting that the newly designed gfph gene could be widely used as a reporter in many gene delivery technologies, including human gene therapy.

Schizophrenia is a common disorder with a life time prevalence of approximately 1 per cent. The illness often develops in young adults, who were previously normal, and is characterized by a constellation of symptoms including hallucinations and delusions (psychotic symptoms) and symptoms such as severely inappropriate emotional responses, a disorder of thinking and concentration, erratic behaviour as well as social and occupational deterioration. A considerable proportion of the variance in the liability to develop schizophrenia may be genetic, but segregation analysis, to establish a mode of transmission, has not produced a consistent result. One of these studies was carried out in Iceland and made use of the large family size and extensive geneaological information present in that country. Here we demonstrate genetic linkage of two DNA polymorphisms on the long arm of human chromosome 5 to schizophrenia in seven British and Icelandic families with multiple affected members. The results indicate the existence of a gene locus with a dominant schizophrenia-susceptibility allele. Inheritance of the allele in the families studied suggests that it may also predispose to psychiatric conditions such as schizophrenia spectrum disorders and a variety of other disorders. This report provides the first strong evidence for the involvement of a single gene in the causation of schizophrenia.