Accounting Problems Under the Excess Profits Tax

DNA vaccines based on subunits from pathogens have several advantages over other vaccine strategies. DNA vaccines can easily be modified, they show good safety profiles, are stable and inexpensive to produce, and the immune response can be focused to the antigen of interest. However, the immunogenicity of DNA vaccines which is generally quite low needs to be improved. Electroporation and co-delivery of genetically encoded immune adjuvants are two strategies aiming at increasing the efficacy of DNA vaccines. Here, we have examined whether targeting to antigen-presenting cells (APC) could increase the immune response to surface envelope glycoprotein (Env) gp120 from Human Immunodeficiency Virus type 1 (HIV- 1). To target APC, we utilized a homodimeric vaccine format denoted vaccibody, which enables covalent fusion of gp120 to molecules that can target APC. Two molecules were tested for their efficiency as targeting units: the antibody-derived single chain Fragment variable (scFv) specific for the major histocompatilibility complex (MHC) class II I-E molecules, and the CC chemokine ligand 3 (CCL3). The vaccines were delivered as DNA into muscle of mice with or without electroporation. Targeting of gp120 to MHC class II molecules induced antibodies that neutralized HIV-1 and that persisted for more than a year after one single immunization with electroporation. Targeting by CCL3 significantly increased the number of HIV-1 gp120-reactive CD8(+) T cells compared to non-targeted vaccines and gp120 delivered alone in the absence of electroporation. The data suggest that chemokines are promising molecular adjuvants because small amounts can attract immune cells and promote immune responses without advanced equipment such as electroporation.Funding Agencies|Research Council of Norway; Odd Fellow</p

Dendritic cell vaccination and immune monitoring

We exploited dendritic cells (DC) to vaccinate melanoma patients. We recently demonstrated a statistical significant correlation between favorable clinical outcome and the presence of vaccine-related tumor antigen-specific T cells in delayed type hypersensitivity (DTH) skin biopsies. However, favorable clinical outcome is only observed in a minority of the treated patients. Therefore, it is obvious that current DC-based protocols need to be improved. For this reason, we study in small proof of principle trials the fate, interactions and effectiveness of the injected DC

Adipose Tissue Gene Expression of Factors Related to Lipid Processing in Obesity

BACKGROUND: Adipose tissue lipid storage and processing capacity can be a key factor for obesity-related metabolic disorders such as insulin resistance and diabetes. Lipid uptake is the first step to adipose tissue lipid storage. The aim of this study was to analyze the gene expression of factors involved in lipid uptake and processing in subcutaneous (SAT) and visceral (VAT) adipose tissue according to body mass index (BMI) and the degree of insulin resistance (IR). METHODS AND PRINCIPAL FINDINGS: VLDL receptor (VLDLR), lipoprotein lipase (LPL), acylation stimulating protein (ASP), LDL receptor-related protein 1 (LRP1) and fatty acid binding protein 4 (FABP4) gene expression was measured in VAT and SAT from 28 morbidly obese patients with Type 2 Diabetes Mellitus (T2DM) or high IR, 10 morbidly obese patients with low IR, 10 obese patients with low IR and 12 lean healthy controls. LPL, FABP4, LRP1 and ASP expression in VAT was higher in lean controls. In SAT, LPL and FABP4 expression were also higher in lean controls. BMI, plasma insulin levels and HOMA-IR correlated negatively with LPL expression in both VAT and SAT as well as with FABP4 expression in VAT. FABP4 gene expression in SAT correlated inversely with BMI and HOMA-IR. However, multiple regression analysis showed that BMI was the main variable contributing to LPL and FABP4 gene expression in both VAT and SAT. CONCLUSIONS: Morbidly obese patients have a lower gene expression of factors related with lipid uptake and processing in comparison with healthy lean persons

Susceptibility to ozone-induced airway inflammation is associated with decreased levels of surfactant protein D

BACKGROUND: Ozone (O(3)), a common air pollutant, induces exacerbation of asthma and chronic obstructive pulmonary disease. Pulmonary surfactant protein (SP)-D modulates immune and inflammatory responses in the lung. We have shown previously that SP-D plays a protective role in a mouse model of allergic airway inflammation. Here we studied the role and regulation of SP-D in O(3)-induced inflammatory changes in the lung. METHODS: To evaluate the effects of O(3 )exposure in mouse strains with genetically different expression levels of SP-D we exposed Balb/c, C57BL/6 and SP-D knockout mice to O(3 )or air. BAL cellular and cytokine content and SP-D levels were evaluated and compared between the different strains. The kinetics of SP-D production and inflammatory parameters were studied at 0, 2, 6, 12, 24, 48, and 72 hrs after O(3 )exposure. The effect of IL-6, an O(3)-inducible cytokine, on the expression of SP-D was investigated in vitro using a primary alveolar type II cell culture. RESULTS: Ozone-exposed Balb/c mice demonstrated significantly enhanced acute inflammatory changes including recruitment of inflammatory cells and release of KC and IL-12p70 when compared with age- and sex-matched C57BL/6 mice. On the other hand, C57BL/6 mice had significantly higher levels of SP-D and released more IL-10 and IL-6. Increase in SP-D production coincided with the resolution of inflammatory changes. Mice deficient in SP-D had significantly higher numbers of inflammatory cells when compared to controls supporting the notion that SP-D has an anti-inflammatory function in our model of O(3 )exposure. IL-6, which was highly up-regulated in O(3 )exposed mice, was capable of inducing the expression of SP-D in vitro in a dose dependent manner. CONCLUSION: Our data suggest that IL-6 contributes to the up-regulation of SP-D after acute O(3 )exposure and elevation of SP-D in the lung is associated with the resolution of inflammation. Absence or low levels of SP-D predispose to enhanced inflammatory changes following acute oxidative stress

Drosophila Lipophorin Receptors Mediate the Uptake of Neutral Lipids in Oocytes and Imaginal Disc Cells by an Endocytosis-Independent Mechanism

Lipids are constantly shuttled through the body to redistribute energy and metabolites between sites of absorption, storage, and catabolism in a complex homeostatic equilibrium. In Drosophila, lipids are transported through the hemolymph in the form of lipoprotein particles, known as lipophorins. The mechanisms by which cells interact with circulating lipophorins and acquire their lipidic cargo are poorly understood. We have found that lipophorin receptor 1 and 2 (lpr1 and lpr2), two partially redundant genes belonging to the Low Density Lipoprotein Receptor (LDLR) family, are essential for the efficient uptake and accumulation of neutral lipids by oocytes and cells of the imaginal discs. Females lacking the lpr2 gene lay eggs with low lipid content and have reduced fertility, revealing a central role for lpr2 in mediating Drosophila vitellogenesis. lpr1 and lpr2 are transcribed into multiple isoforms. Interestingly, only a subset of these isoforms containing a particular LDLR type A module mediate neutral lipid uptake. Expression of these isoforms induces the extracellular stabilization of lipophorins. Furthermore, our data indicate that endocytosis of the lipophorin receptors is not required to mediate the uptake of neutral lipids. These findings suggest a model where lipophorin receptors promote the extracellular lipolysis of lipophorins. This model is reminiscent of the lipolytic processing of triglyceride-rich lipoproteins that occurs at the mammalian capillary endothelium, suggesting an ancient role for LDLR–like proteins in this process

Peptide microarrays for the profiling of cytotoxic T-lymphocyte activity using minimum numbers of cells

The identification of epitopes that elicit cytotoxic T-lymphocyte activity is a prerequisite for the development of cancer-specific immunotherapies. However, especially the parallel characterization of several epitopes is limited by the availability of T cells. Microarrays have enabled an unprecedented miniaturization and parallelization in biological assays. Here, we developed peptide microarrays for the detection of CTL activity. MHC class I-binding peptide epitopes were pipetted onto polymer-coated glass slides. Target cells, loaded with the cell-impermeant dye calcein, were incubated on these arrays, followed by incubation with antigen-expanded CTLs. Cytotoxic activity was detected by release of calcein and detachment of target cells. With only 200,000 cells per microarray, CTLs could be detected at a frequency of 0.5% corresponding to 1,000 antigen-specific T cells. Target cells and CTLs only settled on peptide spots enabling a clear separation of individual epitopes. Even though no physical boundaries were present between the individual spots, peptide loading only occurred locally and cytolytic activity was confined to the spots carrying the specific epitope. The peptide microarrays provide a robust platform that implements the whole process from antigen presentation to the detection of CTL activity in a miniaturized format. The method surpasses all established methods in the minimum numbers of cells required. With antigen uptake occurring on the microarray, further applications are foreseen in the testing of antigen precursors that require uptake and processing prior to presentation

Genome-wide promoter analysis of histone modifications in human monocyte-derived antigen presenting cells

<p>Abstract</p> <p>Background</p> <p>Monocyte-derived macrophages and dendritic cells (DCs) are important in inflammatory processes and are often used for immunotherapeutic approaches. Blood monocytes can be differentiated into macrophages and DCs, which is accompanied with transcriptional changes in many genes, including chemokines and cell surface markers.</p> <p>Results</p> <p>To study the chromatin modifications associated with this differentiation, we performed a genome wide analysis of histone H3 trimethylation on lysine 4 (H3K4me3) and 27 (H3K27me3) as well as acetylation of H3 lysines (AcH3) in promoter regions. We report that both H3K4me3 and AcH3 marks significantly correlate with transcriptionally active genes whereas H3K27me3 mark is associated with inactive gene promoters. During differentiation, the H3K4me3 levels decreased on monocyte-specific CD14, CCR2 and CX3CR1 but increased on DC-specific TM7SF4/DC-STAMP, TREM2 and CD209/DC-SIGN genes. Genes associated with phagocytosis and antigen presentation were marked by H3K4me3 modifications. We also report that H3K4me3 levels on clustered chemokine and surface marker genes often correlate with transcriptional activity.</p> <p>Conclusion</p> <p>Our results provide a basis for further functional correlations between gene expression and histone modifications in monocyte-derived macrophages and DCs.</p

Enhancement of Tumour-Specific Immune Responses In Vivo by ‘MHC Loading-Enhancer’ (MLE)

BACKGROUND:Class II MHC molecules (MHC II) are cell surface receptors displaying short protein fragments for the surveillance by CD4+ T cells. Antigens therefore have to be loaded onto this receptor in order to induce productive immune responses. On the cell surface, most MHC II molecules are either occupied by ligands or their binding cleft has been blocked by the acquisition of a non-receptive state. Direct loading with antigens, as required during peptide vaccinations, is therefore hindered. PRINCIPAL FINDINGS:Here we show, that the in vivo response of CD4+ T cells can be improved, when the antigens are administered together with 'MHC-loading enhancer' (MLE). MLE are small catalytic compounds able to open up the MHC binding site by triggering ligand-release and stabilizing the receptive state. Their enhancing effect on the immune response was demonstrated here with an antigen from the influenza virus and tumour associated antigens (TAA) derived from the NY-ESO-1 protein. The application of these antigens in combination with adamantane ethanol (AdEtOH), an MLE compound active on human HLA-DR molecules, significantly increased the frequency of antigen-specific CD4+ T cells in mice transgenic for the human MHC II molecule. Notably, the effect was evident only with the MLE-susceptible HLA-DR molecule and not with murine MHC II molecules non-susceptible for the catalytic effect of the MLE. CONCLUSION:MLE can specifically increase the potency of a vaccine by facilitating the efficient transfer of the antigen onto the MHC molecule. They may therefore open a new way to improve vaccination efficacy and tumour-immunotherapy

HER2-based recombinant immunogen to target DCs through FcγRs for cancer immunotherapy

Dendritic cell (DC)-based immunotherapy is an attractive approach to induce long lasting antitumor effector cells aiming to control cancer progression. DC targeting is a critical step in the design of DC vaccines in order to optimize delivery and processing of the antigen, and several receptors have been characterized for this purpose. In this study, we employed the FcγRs to target DCs both in vitro and in vivo. We designed a recombinant molecule (HER2-Fc) composed of the immunogenic sequence of the human tumor-associated antigen HER2 (aa 364–391) and the Fc domain of a human IgG1. In a mouse model, HER2-Fc cDNA vaccination activated significant T cell-mediated immune responses towards HER2 peptide epitopes as detected by IFN-γ ELIspot and induced longer tumor latency as compared to Ctrl-Fc-vaccinated control mice. Human in vitro studies indicated that the recombinant HER2-Fc immunogen efficiently targeted human DCs through the FcγRs resulting in protein cross-processing and in the activation of autologous HER2-specific CD8+ T cells from breast cancer patients

Effectiveness of IT-based diabetes management interventions: a review of the literature

Background : Information technology (IT) is increasingly being used in general practice to manage health care including type 2 diabetes. However, there is conflicting evidence about whether IT improves diabetes outcomes. This review of the literature about IT-based diabetes management interventions explores whether methodological issues such as sample characteristics, outcome measures, and mechanisms causing change in the outcome measures could explain some of the inconsistent findings evident in IT-based diabetes management studies.Methods : Databases were searched using terms related to IT and diabetes management. Articles eligible for review evaluated an IT-based diabetes management intervention in general practice and were published between 1999 and 2009 inclusive in English. Studies that did not include outcome measures were excluded.Results : Four hundred and twenty-five articles were identified, sixteen met the inclusion criteria: eleven GP focussed and five patient focused interventions were evaluated. Nine were RCTs, five non-randomised control trials, and two single-sample before and after designs. Important sample characteristics such as diabetes type, familiarity with IT, and baseline diabetes knowledge were not addressed in any of the studies reviewed. All studies used HbA1c as a primary outcome measure, and nine reported a significant improvement in mean HbA1c over the study period; only two studies reported the HbA1c assay method. Five studies measured diabetes medications and two measured psychological outcomes. Patient lifestyle variables were not included in any of the studies reviewed. IT was the intervention method considered to effect changes in the outcome measures. Only two studies mentioned alternative possible causal mechanisms.Conclusion : Several limitations could affect the outcomes of IT-based diabetes management interventions to an unknown degree. These limitations make it difficult to attribute changes solely to such interventions.<br /