Neuroinflammation, Mast Cells, and Glia: Dangerous Liaisons

The perspective of neuroinflammation as an epiphenomenon following neuron damage is being replaced by the awareness of glia and their importance in neural functions and disorders. Systemic inflammation generates signals that communicate with the brain and leads to changes in metabolism and behavior, with microglia assuming a pro-inflammatory phenotype. Identification of potential peripheral-to-central cellular links is thus a critical step in designing effective therapeutics. Mast cells may fulfill such a role. These resident immune cells are found close to and within peripheral nerves and in brain parenchyma/meninges, where they exercise a key role in orchestrating the inflammatory process from initiation through chronic activation. Mast cells and glia engage in crosstalk that contributes to accelerate disease progression; such interactions become exaggerated with aging and increased cell sensitivity to stress. Emerging evidence for oligodendrocytes, independent of myelin and support of axonal integrity, points to their having strong immune functions, innate immune receptor expression, and production/response to chemokines and cytokines that modulate immune responses in the central nervous system while engaging in crosstalk with microglia and astrocytes. In this review, we summarize the findings related to our understanding of the biology and cellular signaling mechanisms of neuroinflammation, with emphasis on mast cell-glia interactions

Effects of Intracellular Calcium and Actin Cytoskeleton on TCR Mobility Measured by Fluorescence Recovery

Background: The activation of T lymphocytes by specific antigen is accompanied by the formation of a specialized signaling region termed the immunological synapse, characterized by the clustering and segregation of surface molecules and, in particular, by T cell receptor (TCR) clustering. Methodology/Principal Findings: To better understand TCR motion during cellular activation, we used confocal microscopy and photo-bleaching recovery techniques to investigate the lateral mobility of TCR on the surface of human T lymphocytes under various pharmacological treatments. Using drugs that cause an increase in intracellular calcium, we observed a decrease in TCR mobility that was dependent on a functional actin cytoskeleton. In parallel experiments measurement of filamentous actin by FACS analysis showed that raising intracellular calcium also causes increased polymerization of the actin cytoskeleton. These in vitro results were analyzed using a mathematical model that revealed effective binding parameters between TCR and the actin cytoskeleton. Conclusion/Significance: We propose, based on our results, that increase in intracellular calcium levels leads to actin polymerization and increases TCR/cytoskeleton interactions that reduce the overall mobility of the TCR. In a physiological setting, this may contribute to TCR re-positioning at the immunological synapse

Effects of combined treatment with rapamycin and cotylenin A, a novel differentiation-inducing agent, on human breast carcinoma MCF-7 cells and xenografts

INTRODUCTION: Rapamycin, an inhibitor of the serine/threonine kinase target of rapamycin, induces G(1 )arrest and/or apoptosis. Although rapamycin and its analogues are attractive candidates for cancer therapy, their sensitivities with respect to growth inhibition differ markedly among various cancer cells. Using human breast carcinoma cell line MCF-7 as an experimental model system, we examined the growth-inhibitory effects of combinations of various agents and rapamycin to find the agent that most potently enhances the growth-inhibitory effect of rapamycin. METHOD: We evaluated the growth-inhibitory effect of rapamycin plus various agents, including cotylenin A (a novel inducer of differentiation of myeloid leukaemia cells) to MCF-7 cells, using either MTT assay or trypan blue dye exclusion test. The cell cycle was analyzed using propidium iodide-stained nuclei. Expressions of several genes in MCF-7 cells with rapamycin plus cotylenin A were studied using cDNA microarray analysis and RT-PCR. The in vitro results of MCF-7 cells treated with rapamycin plus cotylenin A were further confirmed in vivo in a mouse xenograft model. RESULTS: We found that the sensitivity of rapamycin to MCF-7 cells was markedly affected by cotylenin A. This treatment induced growth arrest of the cells at the G(1 )phase, rather than apoptosis, and induced senescence-associated β-galactosidase activity. We examined the gene expression profiles associated with exposure to rapamycin and cotylenin A using cDNA microarrays. We found that expressions of cyclin G(2), transforming growth factor-β-induced 68 kDa protein, BCL2-interacting killer, and growth factor receptor-bound 7 were markedly induced in MCF-7 cells treated with rapamycin plus cotylenin A. Furthermore, combined treatment with rapamycin and cotylenin A significantly inhibited the growth of MCF-7 cells as xenografts, without apparent adverse effects. CONCLUSION: Rapamycin and cotylenin A cooperatively induced growth arrest in breast carcinoma MCF-7 cells in vitro, and treatment with rapamycin and cotylenin A combined more strongly inhibited the growth of MCF-7 cells as xenografts in vivo than treatment with rapamycin or cotylenin A alone, suggesting that this combination may have therapeutic value in treating breast cancer. We also identified several genes that were markedly modulated in MCF-7 cells treated with rapamycin plus cotylenin A

Control and Manipulation of Pathogens with an Optical Trap for Live Cell Imaging of Intercellular Interactions

The application of live cell imaging allows direct visualization of the dynamic interactions between cells of the immune system. Some preliminary observations challenge long-held beliefs about immune responses to microorganisms; however, the lack of spatial and temporal control between the phagocytic cell and microbe has rendered focused observations into the initial interactions of host response to pathogens difficult. This paper outlines a method that advances live cell imaging by integrating a spinning disk confocal microscope with an optical trap, also known as an optical tweezer, in order to provide exquisite spatial and temporal control of pathogenic organisms and place them in proximity to host cells, as determined by the operator. Polymeric beads and live, pathogenic organisms (Candida albicans and Aspergillus fumigatus) were optically trapped using non-destructive forces and moved adjacent to living cells, which subsequently phagocytosed the trapped particle. High resolution, transmitted light and fluorescence-based movies established the ability to observe early events of phagocytosis in living cells. To demonstrate the broad applicability of this method to immunological studies, anti-CD3 polymeric beads were also trapped and manipulated to form synapses with T cells in vivo, and time-lapse imaging of synapse formation was also obtained. By providing a method to exert fine control of live pathogens with respect to immune cells, cellular interactions can be captured by fluorescence microscopy with minimal perturbation to cells and can yield powerful insight into early responses of innate and adaptive immunity.National Institute of Biomedical Imaging and Bioengineering (U.S.) (grant T32EB006348)Massachusetts General Hospital (Department of Medicine Internal Funds)Center for Computational and Integrative Biology (Development fund)Center for Computational and Integrative Biology (AI062773)Center for Computational and Integrative Biology (grant AI062773)Center for Computational and Integrative Biology (grant DK83756)Center for Computational and Integrative Biology (grant DK 043351)National Institute of Allergy and Infectious Diseases (U.S.)National Institutes of Health (U.S.) (grant AI057999

Short-Lived Trace Gases in the Surface Ocean and the Atmosphere

The two-way exchange of trace gases between the ocean and the atmosphere is important for both the chemistry and physics of the atmosphere and the biogeochemistry of the oceans, including the global cycling of elements. Here we review these exchanges and their importance for a range of gases whose lifetimes are generally short compared to the main greenhouse gases and which are, in most cases, more reactive than them. Gases considered include sulphur and related compounds, organohalogens, non-methane hydrocarbons, ozone, ammonia and related compounds, hydrogen and carbon monoxide. Finally, we stress the interactivity of the system, the importance of process understanding for modeling, the need for more extensive field measurements and their better seasonal coverage, the importance of inter-calibration exercises and finally the need to show the importance of air-sea exchanges for global cycling and how the field fits into the broader context of Earth System Science

In situ ozone production under free tropospheric conditions during FREETEX '98 in the Swiss Alps

The second Free Tropospheric Experiment (FREETEX '98) took place at the Jungfraujoch Observatory (3580 m above sea level), Switzerland, in March/April 1998. A number of trace gases and photolysis parameters were measured, including peroxy radicals (HO2+RO2), the photolysis rate coefficients j(NO2) and j(O1D), O3, NOx, NOy, HCHO, peroxyacetylnitrate (PAN), CO, CN, and hydrocarbons. The average midday concentration of HO2+RO2 observed during the campaign is 13±3 parts per trillion by volume (pptv). The peroxy radical concentrations measured during FREETEX '98 are discussed in terms of the respective NO measurements and box model calculations. By screening out polluted and cloudy conditions during FREETEX '98, 8 relatively clean days are selected. These 8 days represent near free tropospheric conditions. The average midday concentration of HO2+RO2 observed during these 8 days is 17±5 pptv and the modeled diurnal cycle of [HO2]+[CH3O2] simulated by a photochemical box model shows very good agreement with the mean diurnal cycle of the peroxy radicals over these 8 days. The in situ net ozone production rate is calculated for each day during FREETEX '98 using the peroxy radical, j(O1D), NO, H2O, and O3 measurements. The mean net ozone production rate during the 8 relatively clean selected days is 0.09 ppbv h-1. If more stringent criteria are applied, 3 free tropospheric days can be selected with a mean net ozone production rate of 0.05±0.01 ppbv h-1, indicating the positive role of in situ photochemistry during springtime in the free troposphere over the Alps and supporting the previous results of the first FREETEX campaign in April/May 1996. Using the experimental data, it is estimated that the ozone compensation point, that is, the point where photochemical ozone production equals destruction, occurs at 24±9 pptv of NO in good agreement with theoretical calculations

Oxidized nitrogen and ozone production efficiencies in the springtime free troposphere over the Alps

The Free Tropospheric Experiment (FREETEX'98) was conducted at the Jungfraujoch Observatory in the Swiss Alps (3580 m above sea level) during the well-documented spring maximum in ozone. In spring the Jungfraujoch frequently lies in the free troposphere but can also be influenced by air from the planetary boundary layer. Measurements of NOx, NOy, peroxyacetylnitrate (PAN), HCHO, O3, CO, nonmethane hydrocarbons, peroxy radicals, j(O1D), j(NO2), and a variety of other tropospheric constituents crucial to ozone photochemical cycles were made over a 1-month period. Two independent measurements of NOx, NOy, and PAN showed good agreement. Average free tropospheric daytime NO levels were about 50 pptv, sufficient to sustain photochemical ozone formation. Although high mixing ratios were encountered, PAN decomposition did not contribute to NOx production during FREETEX'98. Ozone production efficiencies (EN) derived from observed ?O3/(NOz) ratios in free tropospheric air were 20-30 molecules of O3 produced per NOx molecule oxidized and agreed well with a photochemical model. A much lower ozone production efficiency of 4 was determined in a photochemically aged air mass arriving from southern Europe, in line with other measurements and calculations in regimes containing high levels of oxidized nitrogen. Model simulations indicated that by sequestering NOx and HO2, low-temperature formation of peroxynitric acid (PNA) decreased ozone production by 20% and instantaneous ozone production efficiencies by 40%, whereas PAN formation had little effect. The model reproduced well the observed sharp transformation from ozone production to ozone destruction (defined as ?O3/?(NOz) = 0) at 20-25 pptv NO. The observed and calculated strong dependence of EN on NOx concentration in the low-NOx regime highlights the difficulty in assigning a single O3 production efficiency value to remote regions, where most of the CO and CH4 in the atmosphere are oxidized

Influence of clouds on the spectral actinic flux density in the lower troposphere (INSPECTRO): Overview of the field campaigns

Ultraviolet radiation is the key factor driving tropospheric photochemistry. It is strongly modulated by clouds and aerosols. A quantitative understanding of the radiation field and its effect on photochemistry is thus only possible with a detailed knowledge of the interaction between clouds and radiation. The overall objective of the project INSPECTRO was the characterization of the three-dimensional actinic radiation field under cloudy conditions. This was achieved during two measurement campaigns in Norfolk (East Anglia, UK) and Lower Bavaria (Germany) combining space-based, aircraft and ground-based measurements as well as simulations with the one-dimensional radiation transfer model UVSPEC and the three-dimensional radiation transfer model MYSTIC. During both campaigns the spectral actinic flux density was measured at several locations at ground level and in the air by up to four different aircraft. This allows the comparison of measured and simulated actinic radiation profiles. In addition satellite data were used to complete the information of the three dimensional input data set for the simulation. A three-dimensional simulation of actinic flux density data under cloudy sky conditions requires a realistic simulation of the cloud field to be used as an input for the 3-D radiation transfer model calculations. Two different approaches were applied, to derive high- and low-resolution data sets, with a grid resolution of about 100 m and 1 km, respectively. The results of the measured and simulated radiation profiles as well as the results of the ground based measurements are presented in terms of photolysis rate profiles for ozone and nitrogen dioxide. During both campaigns all spectroradiometer systems agreed within ±10% if mandatory corrections e.g. stray light correction were applied. Stability changes of the systems were below 5% over the 4 week campaign periods and negligible over a few days. The J(O1D) data of the single monochromator systems can be evaluated for zenith angles less than 70°, which was satisfied by nearly all airborne measurements during both campaigns. The comparison of the airborne measurements with corresponding simulations is presented for the total, downward and upward flux during selected clear sky periods of both campaigns. The compliance between the measured (from three aircraft) and simulated downward and total flux profiles lies in the range of ±15%