Opposing Roles for Cannabinoid Receptor Type-1 (CB1) and Transient Receptor Potential Vanilloid Type-1 Channel (TRPV1) on the Modulation of Panic-Like Responses in Rats

The midbrain dorsal periaqueductal gray (dPAG) has an important role in orchestrating anxiety-and panic-related responses. Given the cellular and behavioral evidence suggesting opposite functions for cannabinoid type 1 receptor (CB1) and transient receptor potential vanilloid type-1 channel (TRPV1), we hypothesized that they could differentially influence panic-like reactions induced by electrical stimulation of the dPAG. Drugs were injected locally and the expression of CB1 and TRPV1 in this structure was assessed by immunofluorescence and confocal microscopy. The CB1-selective agonist, ACEA (0.01, 0.05 and 0.5 pmol) increased the threshold for the induction of panic-like responses solely at the intermediary dose, an effect prevented by the CB1-selective antagonist, AM251 (75 pmol). Panicolytic-like effects of ACEA at the higher dose were unmasked by pre-treatment with the TRPV1 antagonist capsazepine (0.1 nmol). Similarly to ACEA, capsazepine (1 and 10 nmol) raised the threshold for triggering panic-like reactions, an effect mimicked by another TRPV1 antagonist, SB366791 (1 nmol). Remarkably, the effects of both capsazepine and SB366791 were prevented by AM251 (75 pmol). These pharmacological data suggest that a common endogenous agonist may have opposite functions at a given synapse. Supporting this view, we observed that several neurons in the dPAG co-expressed CB1 and TRPV1. Thus, the present work provides evidence that an endogenous substance, possibly anandamide, may exert both panicolytic and panicogenic effects via its actions at CB1 receptors and TRPV1 channels, respectively. This tripartite set-point system might be exploited for the pharmacotherapy of panic attacks and anxiety-related disorders. Neuropsychopharmacology (2012) 37, 478-486; doi:10.1038/npp.2011.207; published online 21 September 2011

MMP-7 and TIMP-1, news targets in predicting poor wound healing in apical periodontitis

Introduction: Matrix metalloproteinases (MMPs) and the tissue inhibitors of metalloproteinases (TIMPs) are strongly associated with tissue destruction because of inflammation. In this study, we investigated the expression of MMPs and TIMPs messenger RNA and protein levels in apical periodontitis lesions. Methods: Tissue samples from patients presenting clinical signs of chronic apical abscess (CAA) or asymptomatic apical periodontitis (AAP) were collected postoperatively and used for gene expression analysis of MMP-2, -3, -7, -9, -14, -16, and -25; TIMP-1; and TIMP-2 in real-time polymerase chain reaction. Immunohistochemistry was also performed to detect the expression of MMP-7 and TIMP-1 proteins. Lastly, U-937 cells were induced to terminal differentiation into macrophages, infected with purified Escherichia coli lipopolysaccharide, and assessed for the expression of MMP-7 and TIMP-1 using immunocytochemistry and confocal microscopy. Results: Significantly higher messenger RNA levels were found for all genes in AAP and CAA samples when compared with healthy control samples (P < .001). AAP cases exhibited significantly higher TIMP-1 when compared with CAA cases, whereas CAA cases showed higher MMP-2, MMP-7, and MMP-9 messenger RNA levels (P < .05). We also detected positive the expression of MMP-7 and TIMP-1 proteins in the tissue samples. The expression of both MMP-7 and TIMP-1 were increased in lipopolysaccharide-stimulated cells compared with nonstimulated cells and appear to colocalize in the Golgi apparatus. Conclusions: MMPs appear to have an influential role in CAA cases in which ongoing tissue destruction is observed. TIMPs are preferentially associated with AAP, perhaps as a subsequent defense mechanism against excessive destruction. Taken together, our findings implicate MMP and TIMP molecules in the dynamics of inflammatory periapical lesion development

Shikonin directly targets mitochondria and causes mitochondrial dysfunction in cancer cells

Chemotherapy is a mainstay of cancer treatment. Due to increased drug resistance and the severe side effects of currently used therapeutics, new candidate compounds are required for improvement of therapy success. Shikonin, a natural naphthoquinone, was used in traditional Chinese medicine for the treatment of different inflammatory diseases and recent studies revealed the anticancer activities of shikonin. We found that shikonin has strong cytotoxic effects on 15 cancer cell lines, including multidrug-resistant cell lines. Transcriptome-wide mRNA expression studies showed that shikonin induced genetic pathways regulating cell cycle, mitochondrial function, levels of reactive oxygen species, and cytoskeletal formation. Taking advantage of the inherent fluorescence of shikonin, we analyzed its uptake and distribution in live cells with high spatial and temporal resolution using flow cytometry and confocal microscopy. Shikonin was specifically accumulated in the mitochondria, and this accumulation was associated with a shikonin-dependent deregulation of cellular Ca(2+) and ROS levels. This deregulation led to a breakdown of the mitochondrial membrane potential, dysfunction of microtubules, cell-cycle arrest, and ultimately induction of apoptosis. Seeing as both the metabolism and the structure of mitochondria show marked differences between cancer cells and normal cells, shikonin is a promising candidate for the next generation of chemotherapy.

Search for a heavy particle decaying into an electron and a muon with the ATLAS detector in sqrt[s] = 7 TeV pp collisions at the LHC

This Letter presents the first search for a heavy particle decaying into an e ± μ(-/+) final state in sqrt[s] = 7 TeV pp collisions at the LHC. The data were recorded by the ATLAS detector during 2010 and correspond to a total integrated luminosity of 35 pb(-1). No excess above the standard model background expectation is observed. Exclusions at 95% confidence level are placed on two representative models. In an R-parity violating supersymmetric model, tau sneutrinos with a mass below 0.75 TeV are excluded, assuming all R-parity violating couplings are zero except λ(311)' = 0.11 and λ312 = 0.07. In a lepton flavor violating model, a Z'-like vector boson with masses of 0.70-1.00 TeV and corresponding cross sections times branching ratios of 0.175-0.183 pb is excluded. These results extend to higher mass R-parity violating sneutrinos and lepton flavor violating Z's than previous constraints from the Tevatron.

Tumor recovery by angiogenic switch from sprouting to intussusceptive angiogenesis after treatment with PTK787/ZK222584 or ionizing radiation

Inhibitors of angiogenesis and radiation induce compensatory changes in the tumor vasculature both during and after treatment cessation. To assess the responses to irradiation and vascular endothelial growth factor-receptor tyrosine kinase inhibition (by the vascular endothelial growth factor tyrosine kinase inhibitor PTK787/ZK222854), mammary carcinoma allografts were investigated by vascular casting; electron, light, and confocal microscopy; and immunoblotting. Irradiation and anti-angiogenic therapy had similar effects on the tumor vasculature. Both treatments reduced tumor vascularization, particularly in the tumor medulla. After cessation of therapy, the tumor vasculature expanded predominantly by intussusception with a plexus composed of enlarged sinusoidal-like vessels containing multiple transluminal tissue pillars. Tumor revascularization originated from preserved alpha-smooth muscle actin-positive vessels in the tumor cortex. Quantification revealed that recovery was characterized by an angiogenic switch from sprouting to intussusception. Up-regulated alpha-smooth muscle actin-expression during recovery reflected the recruitment of alpha-smooth muscle actin-positive cells for intussusception as part of the angio-adaptive mechanism. Tumor recovery was associated with a dramatic decrease (by 30% to 40%) in the intratumoral microvascular density, probably as a result of intussusceptive pruning and, surprisingly, with only a minimal reduction of the total microvascular (exchange) area. Therefore, the vascular supply to the tumor was not severely compromised, as demonstrated by hypoxia-inducible factor-1alpha expression. Both irradiation and anti-angiogenic therapy cause a switch from sprouting to intussusceptive angiogenesis, representing an escape mechanism and accounting for the development of resistance, as well as rapid recovery, after cessation of therapy.

BrainMaps.org - Interactive High-Resolution Digital Brain Atlases and Virtual Microscopy

BrainMaps.org is an interactive high-resolution digital brain atlas and virtual microscope that is based on over 20 million megapixels of scanned images of serial sections of both primate and non-primate brains and that is integrated with a high-speed database for querying and retrieving data about brain structure and function over the internet. Complete brain datasets for various species, including Homo sapiens, Macaca mulatta, Chlorocebus aethiops, Felis catus, Mus musculus, Rattus norvegicus, and Tyto alba, are accessible online. The methods and tools we describe are useful for both research and teaching, and can be replicated by labs seeking to increase accessibility and sharing of neuroanatomical data. These tools offer the possibility of visualizing and exploring completely digitized sections of brains at a sub-neuronal level, and can facilitate large-scale connectional tracing, histochemical and stereological analyses.

Size-Dependent Uptake of Particles by Pulmonary Antigen-Presenting Cell Populations and Trafficking to Regional Lymph Nodes

The respiratory tract is an attractive target organ for novel diagnostic and therapeutic applications with nano-sized carriers, but their immune effects and interactions with key resident antigen-presenting cells (APCs) such as dendritic cells (DCs) and alveolar macrophages (AMs) in different anatomical compartments remain poorly understood. Polystyrene particles ranging from 20 nm to 1,000 nm were instilled intranasally in BALB/c mice, and their interactions with APC populations in airways, lung parenchyma, and lung-draining lymph nodes (LDLNs) were examined after 2 and 24 hours by flow cytometry and confocal microscopy. In the main conducting airways and lung parenchyma, DC subpopulations preferentially captured 20-nm particles, compared with 1,000-nm particles that were transported to the LDLNs by migratory CD11blow DCs and that were observed in close proximity to CD3+ T cells. Generally, the uptake of particles increased the expression of CD40 and CD86 in all DC populations, independent of particle size, whereas 20-nm particles induced enhanced antigen presentation to CD4+ T cells in LDLNs in vivo. Despite measurable uptake by DCs, the majority of particles were taken up by AMs, irrespective of size. Confocal microscopy and FACS analysis showed few particles in the main conducting airways, but a homogeneous distribution of all particle sizes was evident in the lung parenchyma, mostly confined to AMs. Particulate size as a key parameter determining uptake and trafficking therefore determines the fate of inhaled particulates, and this may have important consequences in the development of novel carriers for pulmonary diagnostic or therapeutic applications.

Th17 cells and tissue remodeling in atopic and contact dermatitis

BACKGROUND: Eczematous skin lesions of atopic dermatitis (AD) as well as allergic and irritant contact dermatitis (ACD, ICD) are characterized by the same typical clinical signs, although due to different causes. In both AD and ACD, the presence of T helper 17 cells which play an important role in host defense, has been reported. Furthermore, IL-17 is involved in tissue repair and remodeling. This study aimed to investigate IL-17 expression in acute eczematous skin lesions and correlate it with markers of remodeling in AD, ACD, and ICD. METHODS: Skin specimens were taken from positive patch test reactions to aeroallergens, contact allergens, and irritants at days 2, 3, and 4. Inflammatory cells as well as the expression of cytokines and extracellular matrix proteins were evaluated by immunofluorescence staining and confocal microscopy. RESULTS: Allergic contact dermatitis and ICD were characterized by IFN-γ expression, whereas in AD lesions, IL-13 expression and high numbers of eosinophils were the prominent phenotype. Expression of IL-17, but also IL-21 and IL-22, was observed in all eczema subtypes. The number of IL-22+ T cells correlated with the number of eosinophils. Markers of remodeling such as MMP-9, procollagen-3, and tenascin C were observed in all acute eczematous lesions, while a correlation of IL-17+ T cell numbers with tenascin C-expressing cells and MMP-9+ eosinophils was apparent. CONCLUSION: The expression of IL-17 and related cytokines, such as IL-22, was demonstrated in acute eczematous lesions independent of their pathogenesis. Our results suggest a potential role for IL-17 in remodeling of the skin.

Electron and photon energy calibration with the ATLAS detector using LHC Run 1 data

This paper presents the electron and photon energy calibration achieved with the ATLAS detector using about 25 fb−1 of LHC proton–proton collision data taken at centre-of-mass energies of √s = 7 and 8 TeV. The reconstruction of electron and photon energies is optimised using multivariate algorithms. The response of the calorimeter layers is equalised in data and simulation, and the longitudinal profile of the electromagnetic showers is exploited to estimate the passive material in front of the calorimeter and reoptimise the detector simulation. After all corrections, the Z resonance is used to set the absolute energy scale. For electrons from Z decays, the achieved calibration is typically accurate to 0.05% in most of the detector acceptance, rising to 0.2% in regions with large amounts of passive material. The remaining inaccuracy is less than 0.2–1% for electrons with a transverse energy of 10 GeV, and is on average 0.3% for photons. The detector resolution is determined with a relative inaccuracy of less than 10% for electrons and photons up to 60 GeV transverse energy, rising to 40% for transverse energies above 500 GeV.

Binding of the HIV-1 gp120 -V3 to cellular glycosphingolipids is necessary for CXCR4 recruitment and infection

Infection by human immunodeficiency virus type 1 (HIV-1) is a multi-step process, and detailed analyses of the various events critical for productive infection are necessary to clearly understanding the infection process and identifying novel targets for therapeutic interventions. Evidence from this study reveals binding of the viral envelope protein to host cell glycosphingolipids (GSLs) as a novel event necessary for the orderly progression of the host cell-entry and productive infection by HIV-1. Data obtained from co-immunoprecipitation analyses and confocal microscopy showed that the ability of viral envelope to interact with the co-receptor CXCR4 and productive infection of HIV-1 were inhibited in cells rendered GSL-deficient, while both these activities were restored after reconstitution of the cells with specific GSLs like GM3. Furthermore, evidence was obtained using peptide-inhibitors of HIV-1 infection to show that binding of a specific region within the V3-loop of the envelope protein gp120 to the host cell GSLs is the trigger necessary for the CD4-bound gp120 to recruit the CXCR4 co-receptor. Infection-inhibitory activity of the V3 peptides was compromised in GSL-deficient cells, but could be restored by reconstitution of GSLs. Based on these findings, a revised model for HIV-1 infection is proposed that accounts for the established interactions between the viral envelope and host cell receptors while enumerating the importance of the new findings that fill the gap in the current knowledge of the sequential events for the HIV-1 entry. According to this model, post-CD4 binding of the HIV-1 envelope surface protein gp120 to host cell GSLs, mediated by the gp120-V3 region, enables formation of the gp120-CD4-GSL-CXCR4 immune-complex and productive infection. The identification of cellular GSLs as an additional class of co-factors necessary for HIV-1 infection is important for enhancing the basic knowledge of the HIV-1 entry that can be exploited for developing novel antiviral therapeutic strategies. ^

(Table 1) Microplankton biomass determined from ATP and by microscopy

Simultaneous determinations of the microplankton biomass by direct microscopy and by measuring concentration of adenosine triphosphate (ATP) in samples from different depths of the Eastern Pacific revealed considerable differences between values obtained. With the exception of 3 of 62 determinations, the biomass determined from ATP exceeded that measured by microscopy; the latter averaged about 10% of the former for the region as a whole. This sharp difference is largely due to underestimation of ultramicroscopic cells by direct microscopy and to substantial variations in the factor used to convert ATP concentrations to cell biomass.

Structural protein 4.1 is located in mammalian centrosomes

Structural protein 4.1 was first characterized as an important 80-kDa protein in the mature red cell membrane skeleton. It is now known to be a member of a family of protein isoforms detected at diverse intracellular sites in many nucleated mammalian cells. We recently reported that protein 4.1 isoforms are present at interphase in nuclear matrix and are rearranged during the cell cycle. Here we report that protein 4.1 epitopes are present in centrosomes of human and murine cells and are detected by using affinity-purified antibodies specific for 80-kDa red cell 4.1 and for 4.1 peptides. Immunofluorescence, by both conventional and confocal microscopy, showed that protein 4.1 epitopes localized in the pericentriolar region. Protein 4.1 epitopes remained in centrosomes after extraction of cells with detergent, salt, and DNase. Higher resolution electron microscopy of detergent-extracted cell whole mounts showed centrosomal protein 4.1 epitopes distributed along centriolar cylinders and on pericentriolar fibers, at least some of which constitute the filamentous network surrounding each centriole. Double-label electron microscopy showed that protein 4.1 epitopes were predominately localized in regions also occupied by epitopes for centrosome-specific autoimmune serum 5051 but were not found on microtubules. Our results suggest that protein 4.1 is an integral component of centrosome structure, in which it may play an important role in centrosome function during cell division and organization of cellular architecture.