Murine leukemia virus (MLV) replication monitored with fluorescent proteins

Background: Cancer gene therapy will benefit from vectors that are able to replicate in tumor tissue and cause a bystander effect. Replication-competent murine leukemia virus (MLV) has been described to have potential as cancer therapeutics, however, MLV infection does not cause a cytopathic effect in the infected cell and viral replication can only be studied by immunostaining or measurement of reverse transcriptase activity. Results: We inserted the coding sequences for green fluorescent protein (GFP) into the proline-rich region (PRR) of the ecotropic envelope protein (Env) and were able to fluorescently label MLV. This allowed us to directly monitor viral replication and attachment to target cells by flow cytometry. We used this method to study viral replication of recombinant MLVs and split viral genomes, which were generated by replacement of the MLV env gene with the red fluorescent protein (RFP) and separately cloning GFP-Env into a retroviral vector. Co-transfection of both plasmids into target cells resulted in the generation of semi-replicative vectors, and the two color labeling allowed to determine the distribution of the individual genomes in the target cells and was indicative for the occurrence of recombination events. Conclusions: Fluorescently labeled MLVs are excellent tools for the study of factors that influence viral replication and can be used to optimize MLV-based replication-competent viruses or vectors for gene therapy

Addressing Language and Culture Differences in Health Care Settings

Health care encounters can be stressful, compromising a patient’s ability to fully engage and understand. This is particularly challenging for patients who are limited-English proficient (LEP) and not able to speak, read, write or understand English at levels appropriate for successful encounters in healthcare settings. To provide quality care to the increasing number of LEP patients from different cultural backgrounds, health care practices have to bridge that communication gap with linguistically and culturally appropriate services

Vaccination directed against the human endogenous retrovirus-K envelope protein inhibits tumor growth in a murine model system

Human endogenous retrovirus (HERV) genomes are chromosomally integrated in all cells of an individual. They are normally transcriptionally silenced and transmitted only vertically. Enhanced expression of HERV-K accompanied by the emergence of anti-HERV-K-directed immune responses has been observed in tumor patients and HIV-infected individuals. As HERV-K is usually not expressed and immunological tolerance development is unlikely, it is an appropriate target for the development of immunotherapies. We generated a recombinant vaccinia virus (MVA-HKenv) expressing the HERV-K envelope glycoprotein (ENV), based on the modified vaccinia virus Ankara (MVA), and established an animal model to test its vaccination efficacy. Murine renal carcinoma cells (Renca) were genetically altered to express E. coli beta-galactosidase (RLZ cells) or the HERV-K ENV gene (RLZ-HKenv cells). Intravenous injection of RLZ-HKenv cells into syngenic BALB/c mice led to the formation of pulmonary metastases, which were detectable by X-gal staining. A single vaccination of tumor-bearing mice with MVA-HKenv drastically reduced the number of pulmonary RLZ-HKenv tumor nodules compared to vaccination with wild-type MVA. Prophylactic vaccination of mice with MVA-HKenv precluded the formation of RLZ-HKenv tumor nodules, whereas wild-type MVA-vaccinated animals succumbed to metastasis. Protection from tumor formation correlated with enhanced HERV-K ENV-specific killing activity of splenocytes. These data demonstrate for the first time that HERV-K ENV is a useful target for vaccine development and might offer new treatment opportunities for diverse types of cancer

Abduction in Annotated Probabilistic Temporal Logic

Annotated Probabilistic Temporal (APT) logic programs are a form of logic programs that allow users to state (or systems to automatically learn)rules of the form ``formula G becomes true K time units after formula F became true with L to U% probability.\u27\u27 In this paper, we develop a theory of abduction for APT logic programs. Specifically, given an APT logic program Pi, a set of formulas H that can be ``added\u27\u27 to Pi, and a goal G, is there a subset S of H such that Pi cup S is consistent and entails the goal G? In this paper, we study the complexity of the Basic APT Abduction Problem (BAAP). We then leverage a geometric characterization of BAAP to suggest a set of pruning strategies when solving BAAP and use these intuitions to develop a sound and complete algorithm

Apurinic/Apyrimidinic Endonuclease 1 Regulates Inflammatory Response in Macrophages

The multi-functional apyrimidinic endonuclease 1/redox factor-1 (APE1/Ref-1) DNA repair and redox signaling protein has been shown to have a role in cancer growth and survival, however, little has been investigated concerning its role in inflammation. In this study, an APE1 redox-specific inhibitor (E3330) was used in lypopolysaccharide (LPS)-stimulated macrophages (RAW264.7). E3330 clearly suppressed secretion of inflammatory cytokines including tumor necrosis factor-α (TNF-α), interleukin (IL-6) and IL-12 and inflammatory mediators nitric oxide (NO) as well as prostaglandin E2 (PGE2) from the LPS-stimulated RAW264.7 cells. These data were supported by the down-regulation of the LPS-dependent expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) genes in the RAW264.7 cells. The effects of E3330 were mediated by the inhibition of transcription factors nuclear factor-κB (NF-κB) and activator protein 1 (AP-1) in the LPS-stimulated macrophages, both known targets of APE1. In conclusion, pharmacological inhibition of APE1 by E3330 suppresses inflammatory response in activated macrophages and can be considered as a novel therapeutic strategy for the inhibition of tumor-associated macrophages

Honokiol suppresses metastasis of renal cell carcinoma by targeting KISS1/KISS1R signaling

Renal cell carcinoma (RCC) is a common urological cancer worldwide and is known to have a high risk of metastasis, which is considered responsible for more than 90% of cancer associated deaths. Honokiol is a small-molecule biphenol isolated from Magnolia spp. bark and has been shown to be a potential anticancer agent involved in multiple facets of signal transduction. In this study, we demonstrated that honokiol inhibited the invasion and colony formation of highly metastatic RCC cell line 786-0 in a dose-dependent manner. DNA-microarray data showed the significant upregulation of metastasis-suppressor gene KISS1 and its receptor, KISS1R. The upregulation was confirmed by qRT-PCR analysis. Overexpression of KISS1 and KISS1R was detected by western blotting at the translation level as well. Of note, the decreased invasive and colonized capacities were reversed by KISS1 knockdown. Taken together, the results first indicate that activation of KISS1/KISS1R signaling by honokiol suppresses multistep process of metastasis, including invasion and colony formation, in RCC cells 786-0. Honokiol may be considered as a natural agent against RCC metastasis

How much does skin barrier count for allergic dermatitis improvement?

Background Allergic dermatitis is a genetic-based skin condition affecting an increasing number of dogs. A more efficient treatment approach is often multimodal, including a special focus on skin barrier condition, from which many pruritic triggers depend as well as common complications. Aim: To evaluate the specific role of skin barrier-directed reestablishing treatment in allergic dermatitis clinical improvement. Method Five dogs with allergic dermatitis were selected from an allergy outpatient consultation. Owners’ informed consent was obtained upon approval by an ethics committee. Patients were subjected to clinical work up including Canine Atopic Dermatitis Extent and Severity Index 4 (CADESI-4) and specific diagnosis for atopy and/or food allergy, including intradermal testing (IDT). Skin biopsies were performed from a non-infected inflamed and adjacent non-affected area, and conserved in 4% buffered formaldehyde. Follow up lasted for 2-3 months, applying avoidance measures directed to the implicated allergen sources and treating with a phytosphingosine-containing shampoo/lotion, according to the manufacturer’s directions. Reassessment was done each 10-15 days with CADESI-4 and further IDT and skin biopsies performed at the end. Skin samples were histologically processed including staining for mastocytes and collagen fibers. Thickness of the non-keratinized epidermal layer and stratum corneum as well this lamina organization (scored 1-5), mast cell density and integrity, and collagen density in the dermal layer were assessed. Normal (N) and lesion (L) data, before and after treatment, were compared. Results Three patients showed evident clinical improvement, 1 moderately and 1 not (10-19, 3 and 1 respectively decrease in CADESI-4). IDT results did not differ. Non-keratinized epidermal layer and stratum corneum thickness ranged respectively 12/4 (N) to 330/300 μm (L) before treatment and 10/4 (N) to 120/120 μm (L) after treatment. Lamina cornea organization tended to increase with treatment, with no clear change in collagen fibers. Mast cell density improved in the recovered patients with 15.2% drop and integrity increased 22.8%. Conclusion Skin barrier-directed reestablishing treatment in allergic dermatitis allows for effective and significant clinical improvement, even as single choice treatment, and therefore should be considered as a non-side-effect treatment, especially when a curative immunological approach is not possible or ineffective.CEVA Animal Healt

BreastDefend enhances effect of tamoxifen in estrogen receptor-positive human breast cancer in vitro and in vivo

BACKGROUND: Tamoxifen (TAM) has been widely used for the treatment of estrogen receptor (ER)-positive breast cancer and its combination with other therapies is being actively investigated as a way to increase efficacy and decrease side effects. Here, we evaluate the therapeutic potential of co-treatment with TAM and BreastDefend (BD), a dietary supplement formula, in ER-positive human breast cancer. METHODS: Cell proliferation and apoptosis were determined in ER-positive human breast cancer cells MCF-7 by MTT assay, quantitation of cytoplasmic histone-associated DNA fragments and expression of cleaved PARP, respectively. The molecular mechanism was identified using RNA microarray analysis and western blotting. Tumor tissues from xenograft mouse model were analyzed by immunohistochemistry. RESULTS: Our data clearly demonstrate that a combination of 4-hydroxytamoxifen (4-OHT) with BD lead to profound inhibition of cell proliferation and induction of apoptosis in MCF-7 cells. This effect is consistent with the regulation of apoptotic and TAM resistant genes at the transcription and translation levels. Importantly, TAM and BD co-treatment significantly enhanced apoptosis, suppressed tumor growth and reduced tumor weight in a xenograft model of human ER-positive breast cancer. CONCLUSION: BD sensitized ER-positive human breast cancer cells to 4-OHT/TAM treatment in vitro and in vivo. BreastDefend can be used in an adjuvant therapy to increase the therapeutic effect of tamoxifen in patients with ER-positive breast cancer

Novel approach to plasma facing materials in nuclear fusion reactors

A novel material design in nuclear fusion reactors is proposed based on W-nDiamond nanostructured composites. Generally, a microstructure refined to the nanometer scale improves the mechanical strength due to modification of plasticity mechanisms. Moreover, highly specific grainboundary area raises the number of sites for annihilation of radiation induced defects. However, the low thermal stability of fine-grained and nanostructured materials demands the presence of particles at the grain boundaries that can delay coarsening by a pinning effect. As a result, the concept of a composite is promising in the field of nanostructured materials. The hardness of diamond renders nanodiamond dispersions excellent reinforcing and stabilization candidates and, in addition, diamond has extremely high thermal conductivity. Consequently, W-nDiamond nanocomposites are promising candidates for thermally stable first-wall materials. The proposed design involves the production of WAV-nDiamondAV-Cu/Cu layered castellations. The W, W-nDiamond and W-Cu layers are produced by mechanical alloying followed by a consolidation route that combines hot rolling with spark plasma sintering (SPS). Layer welding is achieved by spark plasma sintering. The present work describes the mechanical alloying processsing and consolidation route used to produce W-nDiamond composites, as well as microstructural features and mechanical properties of the material produced Long term plasma exposure experiments are planned at ISTTOK and at FTU (Frascati)