AAV2-Mediated Combined Subretinal Delivery of IFN-α and IL-4 Reduces the Severity of Experimental Autoimmune Uveoretinitis

We previously showed that adeno-associated virus 2 (AAV2) mediated subretinal delivery of human interferon-alpha (IFN-α) could effectively inhibit experimental autoimmune uveoretinitis (EAU). In this study we investigated whether subretinal injection of both AVV2.IFN-α and AAV2.IL-4 had a stronger inhibition on EAU activity. B10RIII mice were subretinally injected with AAV2.IFN-α alone (1.5×107 vg), AAV2.IL-4 alone (3.55×107 vg), and AAV2.IFN-α combined with AAV2.IL-4. PBS, AAV2 vector encoding green fluorescent protein (AAV2.GFP) (5×107 vg) was subretinally injected as a control. IFN-α and IL-4 were effectively expressed in the eyes from three weeks to three months following subretinal injection of AAV2 vectors either alone or following combined administration and significantly attenuated EAU activity clinically and histopathologically. AAV2.IL-4 showed a better therapeutic effect as compared to AAV2.IFN-α. The combination of AAV2.IL-4 and AAV2.IFN-α was not significantly different as compared to AAV2.IL-4 alone. There was no difference concerning DTH (delayed-type hypersensitivity) reaction, lymphocyte proliferation and IL-17 production among the investigated treatment groups, suggesting that local retinal gene delivery did not affect the systemic immune response

AAV2-Mediated Subretinal Gene Transfer of hIFN-α Attenuates Experimental Autoimmune Uveoretinitis in Mice

BACKGROUND: Recent reports show that gene therapy may provide a long-term, safe and effective intervention for human diseases. In this study, we investigated the effectiveness of adeno-associated virus 2 (AAV2) based human interferon-alpha (hIFN-α) gene therapy in experimental autoimmune uveoretinitis (EAU), a classic model for human uveitis. METHODOLOGY/PRINCIPAL FINDINGS: An AAV2 vector harboring the hIFN-α gene (AAV2.hIFN-α) was subretinally injected into B10RIII mice at two doses (1.5×10(6) vg, 1.5×10(8) vg). AAV2 vector encoding green fluorescent protein (AAV2.GFP) was used as a control (5×10(8) vg). The expression of hIFN-α in homogenized eyes and serum was detected by ELISA three weeks after injection. The biodistribution of vector DNA in the injected eyes, contralateral eyes and distant organs was determined by PCR. EAU was induced by immunization with IRBP(161-180) three weeks following vector injections, and evaluated clinically and pathologically. IRBP-specific proliferation and IL-17 expression of lymphocytes from the spleen and lymph nodes were assayed to test the influence of the subretinal delivery of AAV2.hIFN-α on the systemic immune response. hIFN-α was effectively expressed in the eyes from three weeks to three months following subretinal injection of AAV2.hIFN-α vector. DNA of AAV2.GFP was observed only in the injected eyes, but not in the distant organs or contralateral eyes. Subretinal injection of both doses significantly attenuated EAU activity clinically and histologically. For the lower dose, there was no difference concerning lymphocyte proliferation and IL-17 production among the AAV2.hIFN-α, AAV2.GFP and PBS injected mice. However, the higher dose of AAV2.hIFN-α significantly suppressed lymphocyte proliferation and IL-17 production. CONCLUSIONS/SIGNIFICANCE: Subretinal delivery of AAV2.hIFN-α lead to an effective expression within the eye for at least three months and significantly attenuated EAU activity. AAV2.hIFN-α was shown to inhibit the systemic IRBP-specific immune response

A Novel Function of Apolipoprotein E: Upregulation of ATP-Binding Cassette Transporter A1 Expression

Despite the well known importance of apolipoprotein (Apo) E in cholesterol efflux, the effect of ApoE on the expression of ATP-binding cassette transporter A1 (ABCA1) has never been investigated. The objective of this study was to determine the effect of ApoE on ApoB-carrying lipoprotein-induced expression of ABCA1, a protein that mediates cholesterol efflux. Our data demonstrate that ApoB-carrying lipoproteins obtained from both wild-type and ApoE knockout mice induced ApoAI-mediated cholesterol efflux in mouse macrophages, which was associated with an enhanced ABCA1 promoter activity, and an increased ABCA1 mRNA and protein expression. In addition, these lipoproteins increased the level of phosphorylated specificity protein 1 (Sp1) and the amount of Sp1 bound to the ABCA1 promoter. However, all these inductions were significantly diminished in cells treated with ApoE-free lipoproteins, when compared to those treated with wild-type lipoproteins. Enrichment with human ApoE3 reversed the reduced inducibility of ApoE-free lipoproteins. Moreover, we observed that inhibition of Sp1 DNA-binding by mithramycin A diminished ABCA1 expression and ApoAI-mediated cholesterol efflux induced by ApoB-carrying lipoproteins, and that mutation of the Sp1-binding motif in the ABCA1 promoter region diminished ApoB-carrying lipoprotein-induced ABCA1 promoter activity. Collectively, these data suggest that ApoE associated with ApoB-carrying lipoproteins has an upregulatory role on ABCA1 expression, and that induction of Sp1 phosphorylation is a mechanism by which ApoE upregulates ABCA1 expression

The role of rock joint frictional strength in the containment of fracture propagation

The fracturing phenomenon within the reservoir environment is a complex process that is controlled by several factors and may occur either naturally or by artificial drivers. Even when deliberately induced, the fracturing behaviour is greatly influenced by the subsurface architecture and existing features. The presence of discontinuities such as joints, artificial and naturally occurring faults and interfaces between rock layers and microfractures plays an important role in the fracturing process and has been known to significantly alter the course of fracture growth. In this paper, an important property (joint friction) that governs the shear behaviour of discontinuities is considered. The applied numerical procedure entails the implementation of the discrete element method to enable a more dynamic monitoring of the fracturing process, where the joint frictional property is considered in isolation. Whereas fracture propagation is constrained by joints of low frictional resistance, in non-frictional joints, the unrestricted sliding of the joint plane increases the tendency for reinitiation and proliferation of fractures at other locations. The ability of a frictional joint to suppress fracture growth decreases as the frictional resistance increases; however, this phenomenon exacerbates the influence of other factors including in situ stresses and overburden conditions. The effect of the joint frictional property is not limited to the strength of rock formations; it also impacts on fracturing processes, which could be particularly evident in jointed rock masses or formations with prominent faults and/or discontinuities

Network Modeling Identifies Molecular Functions Targeted by miR-204 to Suppress Head and Neck Tumor Metastasis

Due to the large number of putative microRNA gene targets predicted by sequence-alignment databases and the relative low accuracy of such predictions which are conducted independently of biological context by design, systematic experimental identification and validation of every functional microRNA target is currently challenging. Consequently, biological studies have yet to identify, on a genome scale, key regulatory networks perturbed by altered microRNA functions in the context of cancer. In this report, we demonstrate for the first time how phenotypic knowledge of inheritable cancer traits and of risk factor loci can be utilized jointly with gene expression analysis to efficiently prioritize deregulated microRNAs for biological characterization. Using this approach we characterize miR-204 as a tumor suppressor microRNA and uncover previously unknown connections between microRNA regulation, network topology, and expression dynamics. Specifically, we validate 18 gene targets of miR-204 that show elevated mRNA expression and are enriched in biological processes associated with tumor progression in squamous cell carcinoma of the head and neck (HNSCC). We further demonstrate the enrichment of bottleneckness, a key molecular network topology, among miR-204 gene targets. Restoration of miR-204 function in HNSCC cell lines inhibits the expression of its functionally related gene targets, leads to the reduced adhesion, migration and invasion in vitro and attenuates experimental lung metastasis in vivo. As importantly, our investigation also provides experimental evidence linking the function of microRNAs that are located in the cancer-associated genomic regions (CAGRs) to the observed predisposition to human cancers. Specifically, we show miR-204 may serve as a tumor suppressor gene at the 9q21.1–22.3 CAGR locus, a well established risk factor locus in head and neck cancers for which tumor suppressor genes have not been identified. This new strategy that integrates expression profiling, genetics and novel computational biology approaches provides for improved efficiency in characterization and modeling of microRNA functions in cancer as compared to the state of art and is applicable to the investigation of microRNA functions in other biological processes and diseases

Improving Quality of Life in Nursing Homes: The Structured Resident Interview Approach

The quality of life (QOL) of the approximately 1.5 million nursing facility (NF) residents in the US is undoubtedly lower than desired by residents, families, providers, and policy makers. Although there have been important advances in defining and measuring QOL for this population, there is a need for interventions that are tied to standardized measurement and quality improvement programs. This paper describes the development and testing of a structured, tailored assessment and care planning process for improving the QOL of nursing home residents. The Quality of Life Structured Resident Interview and Care Plan (QOL.SRI/CP) builds on a decade of research on measuring QOL and is designed to be easily implemented in any US nursing home. The approach was developed through extensive and iterative pilot testing and then tested in a randomized controlled trial in three nursing homes. Residents were randomly assigned to receive the assessment alone or both the assessment and an individualized QOL care plan task. The results show that residents assigned to the intervention group experienced improved QOL at 90-and 180-day follow-up, while QOL of residents in the control group was unchanged