TIP: An Umbrella Application for all SCADA-Based Applications for the CERN Technical Infrastructure

The WinCC Open Architecture (OA) SCADA package and the controls frameworks (UNICOS, JCOP) developed at CERN were successfully used to implement many critical control systems at CERN. In the recent years, the supervision and the controls of many technical infrastructure systems (electrical distribution, cooling and ventilation, etc.) were rewritten to use this standard environment. Operators at the Technical Infrastructure desk, who monitor these systems, are forced to continuously switch between the applications that allow them to monitor these infrastructure systems. The Technical Infrastructure Portal (TIP) was designed and is being developed to provide centralized access to all technical infrastructure systems and extend their functionality by linking to a powerful localization system based on GIS. Furthermore, it provides an environment for operators to develop views that aggregate data from different sources, like cooling and electricity

Macrophage-specific NOX2 contributes to the development of lung emphysema through modulation of SIRT1/MMP-9 pathways

Reactive oxygen species (ROS) participate in the pathogenesis of emphysema. Among ROS-producing enzymes, NOX NADPH oxidases are thought to be responsible for tissue injury associated with several lung pathologies. To determine whether NOX2 and/or NOX1 participate in the development of emphysema, their expression patterns were first studied by immunohistochemistry in the lungs of emphysematous patients. Subsequently, we investigated their contribution to elastase-induced emphysema using NOX2- and NOX1-deficient mice. In human lung, NOX2 was mainly detected in macrophages of control and emphysematous lungs, while NOX1 was expressed in alveolar epithelium and bronchial cells. We observed an elevated number of NOX2-positive cells in human emphysematous lungs, as well as increased NOX2 and NOX1 mRNA expression in mouse lungs following elastase exposure. Elastase-induced alveolar airspace enlargement and elastin degradation were prevented in NOX2-deficient mice, but not in NOX1-deficient mice. This protection was independent of inflammation and correlated with reduced ROS production. Concomitantly, an elevation of sirtuin 1 (SIRT1) level and a decrease of matrix metalloproteinase-9 (MMP-9) expression and activity were observed in alveolar macrophages and neutrophils. We addressed the specific role of macrophage-restricted functional NOX2 in elastase-induced lung emphysema using Ncf1 mutant mice and Ncf1 macrophage rescue mice (Ncf1 mutant mice with transgenic expression of Ncf1 only in CD68-positive mononuclear phagocytes; the MN mouse). Compared to WT mice, the lack of functional NOX2 led to decreased elastase-induced ROS production and protected against emphysema. In contrast, ROS production was restored specifically in macrophages from Ncf1 rescue mice and contributes to emphysema. Taken together, our results demonstrate that NOX2 is involved in the pathogenesis of human emphysema and macrophage-specific NOX2 participates in elastase-induced emphysema through the involvement of SIRT1/MMP-9 pathways in mice

Engineering a light-activated caspase-3 for precise ablation of neurons in vivo

The circuitry of the brain is characterized by cell heterogeneity, sprawling cellular anatomy, and astonishingly complex patterns of connectivity. Determining how complex neural circuits control behavior is a major challenge that is often approached using surgical, chemical, or transgenic approaches to ablate neurons. However, all these approaches suffer from a lack of precise spatial and temporal control. This drawback would be overcome if cellular ablation could be controlled with light. Cells are naturally and cleanly ablated through apoptosis due to the terminal activation of caspases. Here, we describe the engineering of a light-activated human caspase-3 (Caspase-LOV) by exploiting its natural spring-loaded activation mechanism through rational insertion of the light-sensitive LOV2 domain that expands upon illumination. We apply the light-activated caspase (Caspase-LOV) to study neurodegeneration in larval and adult Drosophila Using the tissue-specific expression system (UAS)-GAL4, we express Caspase-LOV specifically in three neuronal cell types: retinal, sensory, and motor neurons. Illumination of whole flies or specific tissues containing Caspase-LOV-induced cell death and allowed us to follow the time course and sequence of neurodegenerative events. For example, we find that global synchronous activation of caspase-3 drives degeneration with a different time-course and extent in sensory versus motor neurons. We believe the Caspase-LOV tool we engineered will have many other uses for neurobiologists and others for specific temporal and spatial ablation of cells in complex organisms

Bacillus calmette-guerin infection in NADPH oxidase deficiency: defective mycobacterial sequestration and granuloma formation

Patients with chronic granulomatous disease (CGD) lack generation of reactive oxygen species (ROS) through the phagocyte NADPH oxidase NOX2. CGD is an immune deficiency that leads to frequent infections with certain pathogens; this is well documented for S. aureus and A. fumigatus, but less clear for mycobacteria. We therefore performed an extensive literature search which yielded 297 cases of CGD patients with mycobacterial infections; M. bovis BCG was most commonly described (74%). The relationship between NOX2 deficiency and BCG infection however has never been studied in a mouse model. We therefore investigated BCG infection in three different mouse models of CGD: Ncf1 mutants in two different genetic backgrounds and Cybb knock-out mice. In addition, we investigated a macrophage-specific rescue (transgenic expression of Ncf1 under the control of the CD68 promoter). Wild-type mice did not develop severe disease upon BCG injection. In contrast, all three types of CGD mice were highly susceptible to BCG, as witnessed by a severe weight loss, development of hemorrhagic pneumonia, and a high mortality (∼50%). Rescue of NOX2 activity in macrophages restored BCG resistance, similar as seen in wild-type mice. Granulomas from mycobacteria-infected wild-type mice generated ROS, while granulomas from CGD mice did not. Bacterial load in CGD mice was only moderately increased, suggesting that it was not crucial for the observed phenotype. CGD mice responded with massively enhanced cytokine release (TNF-α, IFN-γ, IL-17 and IL-12) early after BCG infection, which might account for severity of the disease. Finally, in wild-type mice, macrophages formed clusters and restricted mycobacteria to granulomas, while macrophages and mycobacteria were diffusely distributed in lung tissue from CGD mice. Our results demonstrate that lack of the NADPH oxidase leads to a markedly increased severity of BCG infection through mechanisms including increased cytokine production and impaired granuloma formation

Cytokine and chemokine responses to BCG infection.

<p>TNF-α (A), IL-17 (B), IL-12p40 (C), IFN-γ (D), CXCL1 (E) and CCL5 (F) were assessed in lung homogenates obtained 3 days and 4 weeks after BCG infection. Results are presented as the mean ± SEM (n = 4–7 mice per group). (**: p<0.05, *: p<0.01).</p

Lung parameters in response to BCG infection.

<p>(A) Lung/weight ratio of wild-type (n = 9), <i>Ncf1</i> mutant (n = 5) and <i>Ncf1</i> rescue (n = 8) mice without BCG infection and at 3 days and 4 weeks post infection. (B) Determination of alveolar space score (occupied lung tissue <i>vs.</i> free space) in lung sections at 4 weeks post infection. Data are represented as the mean of alveolar space score ± SD in 4 mice per group with at least 3 lobes analyzed per mouse. (C) Number of viable bacteria was determined at 3 days and 4 weeks following BCG infection. Data are shown as mean log of CFU per organ (±SEM; 3–5 mice per group). (D) iNOS protein expression in lung was detected by western blot 4 weeks after BCG infection. Results are expressed as mean ± SEM of relative units of iNOS/actin (n = 4, per group) after quantification by Image Quant software. (E) Nitrotyrosine quantification by ELISA was done in lungs, 4 weeks after BCG infection. Results are expressed as mean ± SEM of nM per lung (n = 4–5, per group). (***: p<0.001, **: p<0.05, *: p<0.01).</p

Analysis of published cases of mycobacterial infections in CGD patients.

<p>Our literature research identified a total of 297 published cases of mycobacterial disease in CGD patients. (A) Mycobacterial species recovered in mycobacterial disease in CGD patients. (B) Clinical presentations of <i>Mycobacterium bovis</i> BCG and <i>Mycobacterium tuberculosis</i> infections in CGD patients. The numbers indicated on top of each column represent the percentage with respect to the total number of BCG or <i>M. tuberculosis</i> cases, respectively. The terms “systemic” refers to disseminated or to lung infections.</p

Impact of CGD mutation on mortality and weight loss in response to BCG infection.

<p><i>Ncf1</i> mutant (loss of function mutation in p47^phox), <i>Ncf1</i> rescue (expression of wild-type p47^phox in mononuclear phagocytes) with C57Bl/10.Q background, <i>Ncf1</i> mutant (loss of function mutation in p47^phox) with C57Bl/6 background, <i>Cybb</i><b>-</b>deficient and their respective wild-type controls were injected intravenously with BCG (10⁷ CFU). Survival was monitored over the 4 weeks period following BCG inoculation in (A) C57Bl/10.Q wild-type (n = 15), <i>Ncf1</i> mutant (n = 15) and <i>Ncf1</i> rescue (n = 11), in (C) C57Bl/6 wild-type (n = 7), <i>Ncf1</i> mutant (n = 6) and (E) C57Bl/6 wild-type (n = 7), <i>Cybb</i><b>-</b>deficient (n = 9) mice. (B, D and F) Body weight changes as a function of time after BCG inoculation. Survival (percent of initial number of mice) is shown in brackets. Statistics shown in the figures are the comparison between respective wild-type and <i>Ncf1</i> mutant or <i>Cybb</i><b>-</b>deficient (***: p<0.001, **: p<0.05, *: p<0.01) and the comparison between <i>Ncf1</i> mutant and rescue (§§§: p<0.001 and §§: p<0.01). Note that no significant differences were observed between wild-type and <i>Ncf1</i> rescue mice.</p