Altered effector function of peripheral cytotoxic cells in COPD

<p>Abstract</p> <p>Background</p> <p>There is mounting evidence that perforin and granzymes are important mediators in the lung destruction seen in COPD. We investigated the characteristics of the three main perforin and granzyme containing peripheral cells, namely CD8⁺T lymphocytes, natural killer (NK; CD56⁺CD3^-) cells and NKT-like (CD56⁺CD3⁺) cells.</p> <p>Methods</p> <p>Peripheral blood mononuclear cells (PBMCs) were isolated and cell numbers and intracellular granzyme B and perforin were analysed by flow cytometry. Immunomagnetically selected CD8+ T lymphocytes, NK (CD56⁺CD3^-) and NKT-like (CD56⁺CD3⁺) cells were used in an LDH release assay to determine cytotoxicity and cytotoxic mechanisms were investigated by blocking perforin and granzyme B with relevant antibodies.</p> <p>Results</p> <p>The proportion of peripheral blood NKT-like (CD56⁺CD3⁺) cells in smokers with COPD (COPD subjects) was significantly lower (0.6%) than in healthy smokers (smokers) (2.8%, p < 0.001) and non-smoking healthy participants (HNS) (3.3%, p < 0.001). NK (CD56⁺CD3^-) cells from COPD subjects were significantly less cytotoxic than in smokers (16.8% vs 51.9% specific lysis, p < 0.001) as were NKT-like (CD56⁺CD3⁺) cells (16.7% vs 52.4% specific lysis, p < 0.001). Both cell types had lower proportions expressing both perforin and granzyme B. Blocking the action of perforin and granzyme B reduced the cytotoxic activity of NK (CD56⁺CD3^-) and NKT-like (CD56⁺CD3⁺) cells from smokers and HNS.</p> <p>Conclusion</p> <p>In this study, we show that the relative numbers of peripheral blood NK (CD56⁺CD3^-) and NKT-like (CD56⁺CD3⁺) cells in COPD subjects are reduced and that their cytotoxic effector function is defective.</p

Sequential Metabolism of 7-Dehydrocholesterol to Steroidal 5,7-Dienes in Adrenal Glands and Its Biological Implication in the Skin

Since P450scc transforms 7-dehydrocholesterol (7DHC) to 7-dehydropregnenolone (7DHP) in vitro, we investigated sequential 7DHC metabolism by adrenal glands ex vivo. There was a rapid, time- and dose-dependent metabolism of 7DHC by adrenals from rats, pigs, rabbits and dogs with production of more polar 5,7-dienes as detected by RP-HPLC. Based on retention time (RT), UV spectra and mass spectrometry, we identified the major products common to all tested species as 7DHP, 22-hydroxy-7DHC and 20,22-dihydroxy-7DHC. The involvement of P450scc in adrenal metabolic transformation was confirmed by the inhibition of this process by DL-aminoglutethimide. The metabolism of 7DHC with subsequent production of 7DHP was stimulated by forscolin indicating involvement of cAMP dependent pathways. Additional minor products of 7DHC metabolism that were more polar than 7DHP were identified as 17-hydroxy-7DHP (in pig adrenals but not those of rats) and as pregna-4,7-diene-3,20-dione (7-dehydroprogesterone). Both products represented the major identifiable products of 7DHP metabolism in adrenal glands. Studies with purified enzymes show that StAR protein likely transports 7DHC to the inner mitochondrial membrane, that 7DHC can compete effectively with cholesterol for the substrate binding site on P450scc and that the catalytic efficiency of 3βHSD for 7DHP (Vm/Km) is 40% of that for pregnenolone. Skin mitochondria are capable of transforming 7DHC to 7DHP and the 7DHP is metabolized further by skin extracts. Finally, 7DHP, its photoderivative 20-oxopregnacalciferol, and pregnenolone exhibited biological activity in skin cells including inhibition of proliferation of epidermal keratinocytes and melanocytes, and melanoma cells. These findings define a novel steroidogenic pathway: 7DHC→22(OH)7DHC→20,22(OH)27DHC→7DHP, with potential further metabolism of 7DHP mediated by 3βHSD or CYP17, depending on mammalian species. The 5–7 dienal intermediates of the pathway can be a source of biologically active vitamin D3 derivatives after delivery to or production in the skin, an organ intermittently exposed to solar radiation

Vitamin D-VDR signaling inhibits Wnt/beta-catenin-mediated melanoma progression and promotes anti-tumor immunity

1α,25-dihydroxyvitamin D3 signals via the Vitamin D Receptor (VDR). Higher serum vitamin D is associated with thinner primary melanoma and better outcome, although a causal mechanism has not been established. As melanoma patients commonly avoid sun exposure, and consequent vitamin D deficiency might worsen outcomes, we interrogated 703 primary melanoma transcriptomes to understand the role of vitamin D-VDR signalling and replicated the findings in TCGA metastases. VDR expression was independently protective for melanoma death in both primary and metastatic disease. High tumor VDR expression was associated with upregulation of pathways mediating anti-tumor immunity and correspondingly with higher imputed immune cell scores and histologically detected tumor infiltrating lymphocytes (TILs). High VDR expressing tumors had downregulation of proliferative pathways, notably Wnt/beta-catenin signaling. Deleterious low VDR levels resulted from promoter methylation and gene deletion in metastases. Vitamin D deficiency (< 25 nmol/l ~ 10 ng/ml) shortened survival in primary melanoma in a VDR-dependent manner. In vitro functional validation studies showed that elevated vitamin D-VDR signaling inhibited Wnt/beta-catenin signaling genes. Murine melanoma cells overexpressing VDR produced fewer pulmonary metastases than controls in tail vein metastasis assays. In summary, vitamin D-VDR signaling contributes to controlling pro-proliferative/immunosuppresive Wnt/beta-catenin signaling in melanoma and this is associated with less metastatic disease and stronger host immune responses. This is evidence of the causal relationship between vitamin D-VDR signaling and melanoma survival which should be explored as a therapeutic target in primary resistance to checkpoint blockade

The Analysis of Receptor-binding Cancer Antigen Expressed on SiSo Cells (RCAS1) immunoreactivity within the microenvironment of the ovarian cancer lesion relative to the applied therapeutic strategy

RCAS1 is involved in generating the suppressive profile of the tumor microenvironment that helps cancer cells evade immune surveillance. The status of the cells surrounding the cancer nest may affect both the progression of the cancer and the development of metastases. In cases of ovarian cancer, a large number of patients do not respond to the applied therapy. The patient’s response to the applied therapy is directly linked to the status of the tumor microenvironment and the intensity of its suppressive profile. We analyzed the immunoreactivity of RCAS1 on the cells present in the ovarian cancer microenvironment in patients with the disease; these cells included macrophages and carcinoma-associated fibroblasts. Later we analyzed the immunoreactivity levels within these cells, taking into consideration the clinical stage of the cancer and the therapeutic strategy applied, such as the number of chemotherapy regiments, primary cytoreductive surgery, or the presence of advanced ascites. In the patients who did not respond to the therapy we observed significantly higher immunoreactivity levels of RCAS1 within the cancer nest than in those patients who did respond; moreover, in the non-responsive patients we found RCAS1 within both macrophages and carcinoma-associated fibroblasts. RCAS1 staining may provide information about the intensity of the immuno-suppressive microenvironment profile found in cases of ovarian cancer and its intensity may directly relate to the clinical outcome of the disease

Optimization of the return streams for the Hengelo Brine Field

Civil Engineering and GeosciencesGeoscience & Engineerin

Measure Guideline: Transitioning to a Tankless Water Heater

This Measure Guideline provides information to help residential builders and retrofitters with the design, specification, selection, implementation, installation, and maintenance issues of transitioning from tank-type water heaters to tankless water heaters. The report compares the differences between tankless and tank-type water heaters, highlighting the energy savings that can be realized by adopting tankless water heaters over tank-type water heaters. Selection criteria and risks discussed include unit sizing and location, water distribution system, plumbing line length and diameter, water quality, electrical backup, and code issues. Cost and performance data are provided for various types of tankless and tank-type water heaters, both natural gas fired and electric. Also considered are interactions between the tankless water heater and other functional elements of a house, such as cold water supply and low-flow devices. Operating costs and energy use of water distribution systems for single- and two-story houses are provided, along with discussion of the various types of distribution systems that can be used with tankless water heaters. Finally, details to prepare for proper installation of a tankless water heater are described

Measure Guideline: Transitioning to a Tankless Water Heater

This Measure Guideline provides information to help residential builders and retrofitters with the design, specification, selection, implementation, installation, and maintenance issues of transitioning from tank-type water heaters to tankless water heaters. The report compares the differences between tankless and tank-type water heaters, highlighting the energy savings that can be realized by adopting tankless water heaters over tank-type water heaters. Selection criteria and risks discussed include unit sizing and location, water distribution system, plumbing line length and diameter, water quality, electrical backup, and code issues. Cost and performance data are provided for various types of tankless and tank-type water heaters, both natural gas fired and electric. Also considered are interactions between the tankless water heater and other functional elements of a house, such as cold water supply and low-flow devices. Operating costs and energy use of water distribution systems for single- and two-story houses are provided, along with discussion of the various types of distribution systems that can be used with tankless water heaters. Finally, details to prepare for proper installation of a tankless water heater are described

Measure Guideline: Transitioning from Three-Coat Stucco to One-Coat Stucco with EPS

This Measure Guideline has been developed to help builders transition from using a traditional three-coat stucco wall-cladding system to a one-coat stucco wall-cladding system with expanded polystyrene (EPS) insulated sheathing. The three-coat system uses a base layer, a fill layer, and a finish layer. The one-coat system maintains the look of a traditional stucco system but uses only a base layer and a finish coat over EPS insulation that achieves higher levels of energy efficiency. Potential risks associated with the installation of a one-coat stucco system are addressed in terms of design, installation, and warranty concerns such as cracking and delamination, along with mitigation strategies to reduce these risks

Measure Guideline: Transitioning from Three-Coat Stucco to One-Coat Stucco with EPS

This Measure Guideline has been developed to help builders transition from using a traditional three-coat stucco wall-cladding system to a one-coat stucco wall-cladding system with expanded polystyrene (EPS) insulated sheathing. The three-coat system uses a base layer, a fill layer, and a finish layer. The one-coat system maintains the look of a traditional stucco system but uses only a base layer and a finish coat over EPS insulation that achieves higher levels of energy efficiency. Potential risks associated with the installation of a one-coat stucco system are addressed in terms of design, installation, and warranty concerns such as cracking and delamination, along with mitigation strategies to reduce these risks