First Direct Detection Constraints on Planck-Scale Mass Dark Matter with Multiple-Scatter Signatures Using the DEAP-3600 Detector.

Dark matter with Planck-scale mass (≃10^{19} GeV/c^{2}) arises in well-motivated theories and could be produced by several cosmological mechanisms. A search for multiscatter signals from supermassive dark matter was performed with a blind analysis of data collected over a 813 d live time with DEAP-3600, a 3.3 t single-phase liquid argon-based detector at SNOLAB. No candidate signals were observed, leading to the first direct detection constraints on Planck-scale mass dark matter. Leading limits constrain dark matter masses between 8.3×10^{6} and 1.2×10^{19} GeV/c^{2}, and ^{40}Ar-scattering cross sections between 1.0×10^{-23} and 2.4×10^{-18} cm^{2}. These results are interpreted as constraints on composite dark matter models with two different nucleon-to-nuclear cross section scalings.

[Voice Self-Concept (FESS) in Medical Students]. / Das stimmliche Selbstkonzept (FESS) bei Studierenden der Medizin.

Introduction: The questionnaire "Fragebogen zur Erfassung des stimmlichen Selbstkonzepts (FESS)" was published in 2015 by Nusseck et al. It consists of 17 items measuring 3 scales on voice related self-concept. This paper examines the distribution of scale values in young adults by examination of medical students. Material and Methods: 96 FESS questionnaires were filled in by medical students. An additional item was added, stating whether it felt easy to answer the questionnaire. The distribution of the scales as well as percentile ranks are given in the paper. Results: In all 3 scales there were no significant differences between females and males, therefore they were analysed as one group. The distributions of all 3 scales show no relevant ceiling nor floor effects. Probands with lower scores in 2 of the three scales found it less easy to answer the questions. Discussion: The results encourage the use of the questionnaire in patients. There was no indication of relevant floor or ceiling effects and there was enough variance in each scale. If used in patients further investigation is needed on the result that patients with lower scores tend to find it more difficult to fill in the questionnaire. The percentile ranks published herein are valid for medical students at this stage. Until bigger normative data on more diverse populations are conducted we will use these data as an orientation to judge other young adults' scores, too.

Test of Electric Charge Conservation with Borexino.

Borexino is a liquid scintillation detector located deep underground at the Laboratori Nazionali del Gran Sasso (LNGS, Italy). Thanks to the unmatched radio purity of the scintillator, and to the well understood detector response at low energy, a new limit on the stability of the electron for decay into a neutrino and a single monoenergetic photon was obtained. This new bound, τ≥6.6×10^{28} yr at 90% C.L., is 2 orders of magnitude better than the previous limit.

[Universal newborn hearing screening : Definition of uniform parameters by the Association of German Hearing Screening Centers as a requirement for nationwide evaluation with valid results]. / Universelles Neugeborenen-Hörscreening : Definition einheitlicher Parameter durch den Verband Deutscher Hörscreening-Zentralen (VDHZ) als Voraussetzung für eine flächendeckende Evaluation mit validen Ergebnissen.

BACKGROUND: Since 2009, all newborns in Germany have been entitled to universal neonatal hearing screening (UNHS). UNHS with tracking of test results leads to earlier detection of hearing disorders. The Association of German Hearing Screening Centers (Verband Deutscher Hörscreening-Zentralen, VDHZ) was founded to promote nationwide tracking, validity and quality control of UNHS results. OBJECTIVES: A comparable data structure in the different screening centers, with uniform definitions of primary parameters is essential for the nationwide evaluation of UNHS results. To address the question of whether a data structure with comparable definitions already exists or still has to be created, the existing structures and primary parameter definitions in the hearing screening centers should be investigated and compared. METHODS: A survey was conducted in all hearing screening centers to assess how data on the primary UNHS parameters defined in pediatric guidelines was gathered. In the case of discrepancies, uniform definitions were created. Finally, the practicability of these definitions was evaluated. RESULTS: Due to differing definitions of primary parameters, some of the data were not comparable between the individual centers. Therefore, uniform definitions were created in a consensus process. In the centers, the screening method, the two-step first screening and the result of the first screening now correspond to these uniform definitions. Other parameters, e.g. the total number of newborns, still vary widely, rendering the comparison of screening rates almost impossible. CONCLUSION: Valid evaluation of UNHS not only requires nationwide establishment of hearing screening centers, but also unified data structures and parameter definitions.

Ultrasound-guided mechanical intraductal stone fragmentation and removal for sialolithiasis: a new technique.

BACKGROUND: Not all patients with sialolithiasis can be treated successfully by established minimally invasive techniques. METHODS: A forceps was used under sonographic control to fragment and retrieve salivary calculi in five cases refractory to established minimal invasive approaches. RESULTS: One patient with a sialolithiasis of the Stenon duct, two patients with a stone in the hilum region of the submandibular gland, and one patient with a sialolith in the sublingual gland were cured by this technique. For another patient, only a part of the stone in the hilum region of the submandibular gland could be removed. No relevant side effects occurred. CONCLUSIONS: To the authors' knowledge, this is the first report of a new, simple, and inexpensive minimally invasive technique that proved to be at least partially successful in the treatment of sialolithiasis in cases refractory to other therapies. The technique also seems to be suitable as a primary treatment approach.

Cell cycle response to DNA damage differs in bronchial epithelial cells and lung fibroblasts.

Epithelial cells along the conducting airways can be more or less continuously exposed to DNA-damaging agents, which should limit their proliferation by inducing cell cycle checkpoints. Yet, paradoxically, airway epithelial cells frequently show a hyperplastic response when exposed to such agents. In this in vitro study, we assessed the hypothesis that normal human bronchial epithelial cells (BECs) are more resistant to the cell cycle-arresting effects of DNA damage than are human lung fibroblasts (HLFs), a cell type often investigated in the context of cell cycle checkpoints. Using ionizing radiation as a DNA-damaging insult, we have found that BECs indeed show less pronounced G1 and G2 delays than do fibroblasts. Unlike the HLFs, which ultimately enter a condition of apparently terminal arrest in the G1 phase of the cell cycle, BECs continue proliferating following their initial, transient G1 and G2 delays. Radiation-induced p53 and p21Cip1 increases were greater in HLFs than in BECs, whereas preexposure, basal levels of p53 were higher in BECs than in HLFs. The results of this investigation indicate that BECs may be less susceptible to the cell cycle-arresting effects of DNA-damaging agents, perhaps because of their higher basal levels of p53. Extension of these findings to the in vivo condition provides a possible explanation for airway epithelial cell hyperplastic responses that occur in a background of DNA-damaging stresses. Moreover, the attenuated DNA damage-induced, cell cycle checkpoint responses in BECs potentially may favor the transmission of DNA lesions to cell progeny.

Extracellular factor(s) following exposure to alpha particles can cause sister chromatid exchanges in normal human cells.

The mechanism(s) by which alpha particles like those emitted from inhaled radon and radon progeny cause their mutagenic and carcinogenic effects remains unclear. Although direct nuclear traversals by alpha particles may be involved in mediating these outcomes, increasing evidence indicates that alpha particles can cause alterations in DNA in the absence of direct "hits" to cell nuclei. Using the occurrence of excessive sister chromatid exchanges (SCEs) as an index of DNA damage in human lung fibroblasts, we investigated the hypothesis that alpha particles may induce DNA damage via the generation of extracellular factors. We have found that a relatively low dose of alpha particles indeed results in the generation of extracellular factors, which, upon transfer to unexposed normal human cells, can cause excessive SCEs to an extent equivalent to that observed when the cells are directly irradiated with the same irradiation dose. A short-lived, SCE-inducing factor(s) was generated in alpha-irradiated culture medium containing serum in the absence of cells; it was found that the activity of this factor can be promptly inhibited by superoxide dismutase. A more persistent SCE-inducing factor(s), which can survive freeze-thawing, is heat labile and also can be inhibited by superoxide dismutase, was produced by fibroblasts after exposure to alpha particles. These results indicate that the initiating target for alpha-particle-induced genetic changes can be larger than the nuclear compartment alone and even larger than the cytoplasmic compartment. How transmissible factors like those observed here in vitro may extend to the in vivo condition in the context of alpha-particle-induced carcinogenesis in the respiratory tract and elsewhere remains to be determined.

Factors underlying the cell growth-related bystander responses to alpha particles.

Increases in cell proliferation are widely viewed as being of importance in carcinogenesis. We report that exposure of normal human lung fibroblasts to a low dose of alpha particles like those emitted by radon/radon progeny stimulates their proliferation in vitro, and this response also occurs when unirradiated cells are treated with supernatants from alpha-irradiated cells. We attribute the promitogenic response to superoxide dismutase- and catalase-inhibitable a particle-induced increases in the concentrations of transforming growth factor beta1 (TGF-beta1) in cell supernatants. TGF-beta1 at concentrations commensurate with those in the supernatants capably induces increases in intracellular reactive oxygen species (ROS) in unirradiated cells. Furthermore, the addition of supernatants from alpha-irradiated cells to unirradiated cells decreases cellular levels of TP53 and CDKN1A and increases CDC2 and proliferating cell nuclear antigen in the latter. Like the increased intracellular ROS bystander effect, this "decreased TP53/CDKN1A response" can be mimicked in otherwise untreated cells by the addition of low concentrations of TGF-beta1. Our results indicate that alpha particle-associated increases in cell growth correlate with intracellular increases in ROS along with decreases in TP53 and CDKN1A, and that these cellular responses are mechanistically coupled. As well, the proliferating cell nuclear antigen and CDC2 increases that occur along with the decreased TP53/CDKN1A bystander effect also would expectedly favor enhanced cell growth. Such processes may account for cell hyperplastic responses in the conducting airways of the lower respiratory track that occur after inhalation exposure to radon/ radon progeny, as well as, perhaps, other ROS-associated environmental stresses.

Alpha particles initiate biological production of superoxide anions and hydrogen peroxide in human cells.

The mechanism(s) by which high-linear energy transfer a particles, like those emitted by inhaled radon and radon daughters, cause lung cancer has not been elucidated. Conceivably, DNA damage that is induced by a particles may be mediated by the metabolic generation of reactive oxygen species (ROS), in addition to direct a particle-DNA interactions and hydroxyl radical-DNA interactions. Using normal human lung fibroblasts, we investigated the hypothesis that densely ionizing alpha particles may induce the intracellular generation of superoxide (O2.-) and hydrogen peroxide (H2O2). Ethidium bromide and 2',7'-dichlorofluorescein, fluorescent products of the membrane-permeable dyes hydroethidine and 2',7'-dichlorofluorescin diacetate, respectively, were used to monitor the intracellular production of O2.- and H2O2, respectively, by flow cytometry. Compared to sham-irradiated cells, fibroblasts that were exposed to alpha particles (0.4-19 cGy) had significant increases in intracellular O2.- production, along with concomitant increases in H2O2 production. Further analyses suggest that the plasma membrane-bound NADPH-oxidase is primarily responsible for this increased intracellular generation of ROS and that the ROS response does not require direct nuclear or cellular "hits" by the a particles. In this latter regard, we additionally report that unirradiated cells also show the ROS response when they are incubated with serum-containing culture medium that has been exposed to a particles or when they are incubated with supernatants from a-irradiated cells. Our overall results support the possibility that a particles, at least in part, may mediate their DNA-damaging effects indirectly via a ROS-related mechanism.

Control of radiation-induced G1 arrest by cell-substratum interactions.

Ionizing radiation has been reported to cause an irreversible, senescence-like G1 arrest in human fibroblasts, which is accompanied by elevated p21CIP1 amounts. In further support of a senescence-like arrest, we show that expression of p53 and cyclin D1 is elevated in gamma-irradiated, arrested fibroblasts. However, we also demonstrate that the arrest is reversible if the irradiated cells are trypsinized and replated, which may implicate cellular-extracellular matrix interactions in cell cycle control after irradiation.

Requirements for p53 and the ATM gene product in the regulation of G1/S and S phase checkpoints.

We investigated the requirements for protein p53 and the ATM gene product in radiation-induced inhibition of DNA synthesis and regulation of the cyclin E/ and cyclin A/cyclin dependent kinases (Cdks). Wild type (WT) mouse lung fibroblasts (MLFs), p53(-/-) knock-out MLFs, normal human skin fibroblasts (HSF-55), and human AT skin fibroblasts (GM02052) were used in the investigations. The absence of p53 had no significant effect on the inhibition or recovery of DNA synthesis throughout the S phase, as determined from BrdU labeling and flow cytometry, or the rapid inhibition of cyclin A/Cdks. Gamma radiation (8 Gy) inhibited DNA synthesis and progression into G2 during the first 3 h after irradiation, and the recovery of these processes occurred at similar rates in both WT and p53(-/-) MLFs. The cyclin A/Cdks were inhibited 55-70% at 1 h after irradiation in both cell types, but p21WAF1/Cip1 levels or p21 interaction with Cdk2 did not increase in the irradiated p53(-/-) MLFs. Although p53(-/-) MLFs do not exhibit prolonged arrest at a G1 checkpoint, radiation did induce a rapid 20% reduction and small super-recovery of cyclin E/Cdk2 within 1-2 h after irradiation. Similar inhibition and recovery of cyclin E/Cdk2 previously had been associated with regulation of transient G1 delay and the inhibition of initiation at an apparent G1/S checkpoint in Chinese hamster cells. In contrast, loss of the ATM gene product abrogated transient cyclin E/Cdk2 inhibition, most inhibition of DNA synthesis and all, but a 10-15% inhibition, of the cyclin A/Cdks. The results indicate that neither p53 nor p21 is required for transient inhibition of cyclin E/Cdk2 associated with the G1/S checkpoint or for inhibition of DNA synthesis at 'checkpoints' within the S phase. Conversely, the ATM gene product appears to be essential for regulation of the G1/S checkpoint and for inhibition of DNA replication associated with the inhibition of cyclin A/Cdk2. Differential aspects of DNA synthesis inhibition among cell types are presented and discussed in the context of S phase checkpoints.

Pulmonary interstitial macrophages: isolation and flow cytometric comparisons with alveolar macrophages and blood monocytes.

Pulmonary interstitial macrophages (IM) were isolated from rat lungs by an Fc gamma receptor-based affinity technique coupled with multiparameter flow cytometry. Single cell suspensions obtained by collagenase digestion of extensively perfused and lavaged lungs were applied to carpets of opsonized sheep red blood cells (SRBC-IgG) bound to plastic tissue culture flasks. At 0-4 degrees C, optimal binding of lung cells occurred within 60 min at plating densities of 1-2 X 10(6) lung cells/cm2 when the SRBC substrate was opsonized with 10 micrograms/ml anti-SRBC IgG. Nonadherent cells were removed by gently rinsing the plates and adherent cells were recovered by lysing the SRBC-IgG substrata. By light microscopy, the mixture of adherent cells was comprised of mononuclear cells (approximately 54%), many of which appeared to be macrophages, lymphocytes (approximately 20%), polymorphonuclear leukocytes (approximately 15%), plasma cells (approximately 8%), eosinophils (approximately 2%), and mast cells (approximately 0.5%). The cells which adhered to the SRBC-IgG monolayers were further resolved into subpopulations by multiparameter flow cytometry and sorted according to their electro-optical characteristics. One subpopulation appeared morphologically to be macrophages, and greater than 90% of these cells readily phagocytized SRBC-IgG in vitro. Peroxidase staining of this population was minimal, indicating that these cells were not blood monocytes (BM). Using a method by which alveolar macrophages (AM) were prelabeled with SRBC-IgG in situ, we demonstrated that alveolar macrophages constituted only approximately 5% of the total adherent cell population. We concluded from these observations that the macrophage population harvested in this manner were neither BM nor AM, but, rather, were harvested from the lung's interstitial compartment. Flow cytometric analyses indicated that the IM exhibited electro-optical characteristics intermediate between those of BM and AM, which is consistent with the concept of the lung's interstitium as a maturation compartment for the BM prior to migration into the alveolar compartment. However, the IM more closely resembled the BM than the AM, indicating that if the IM is in fact a precursor to the AM population, substantial maturation or differentiation must occur subsequent to its migration into the alveolar compartment. This isolation technique will be useful for harvesting highly purified IM for in vitro investigations.

Flow cytometric characterization of alveolar macrophages.

Phenotypes of lung free cells (FC) harvested from Fischer-344 rats by episodic lavage were characterized by flow cytometry. Parameters evaluated included electronic volume (EV), axial light loss (ALL), 90 degrees light scatter (LS), blue autofluorescence (BA), and green-yellow autofluorescence (G-YA). Three phenotypic populations, FC-A, FC-B, and FC-C were identified by their differing LS characteristics. FC-C represented 90% of the cells and were exclusively alveolar macrophages. Two subpopulations in FC-C, FC-CI and FC-CII, were further distinguished by their unique ALL features. The morphologic appearances of these subpopulations by light microscopy clearly differed in sorted preparations. Based on their patterns of autofluorescence, these FC-CI and FC-CII phenotypes were found to be composed of eight subpopulations. In FC populations harvested during further lavage episodes of the same lungs, the relative contributions of FC-CI to the FC-C subpopulation decreased as FC-CII correspondingly increased. This study demonstrates 1) that subpopulations of lavaged AM can be categorized according to their optical phenotypes by flow cytometry and 2) that the relative frequency of retrieval of some phenotypes depends on how exhaustively the lungs are lavaged. With regard to the latter, bronchoalveolar lavage does not randomly sample the underlying AM population in the alveolar compartment.

Size, adherence, and phagocytic characteristics of alveolar macrophages harvested 'early' and 'later' during bronchoalveolar lavage.

Alveolar macrophages (AM) harvested by sequential lung lavage procedures are physically, biochemically, and functionally heterogeneous. The present studies were undertaken in order to determine if such AM heterogeneity is a function of their sequential removal from the lung during episodic bronchoalveolar lavage. Our results demonstrate that AM harvested during 'early' lung wash cycles and during 'later' sequential lung wash cycles are essentially identical in terms of their: (1) abilities to exclude a vital dye; (2) volume distributions; (3) plastic substrate adherence characteristics; and (4) Fc gamma receptor-mediated phagocytic activities.

Quantitative evaluation of opsonin-independent phagocytosis by alveolar macrophages in monolayer using polystyrene microspheres.

Macrophages can bind and engulf a variety of particles in the absence of specific opsonins. Polystyrene-type microspheres are often employed to quantitate opsonin-independent phagocytic activities of macrophages in vitro. Reliable measurement of this cell function, however, requires the ability of the investigator to distinguish between particles that are merely attached to the cell surface and those that are actually internalized. We have developed a simple, rapid, and reproducible method for quantitating phagocytosis using polystyrene microspheres and adherent alveolar macrophages. Basically, particles associated with macrophages after a given incubation time are microscopically quantitated on a cell-by-cell basis before and after toluene dissolution of external particles. Particle/macrophage values obtained after toluene treatment exclusively index phagocytosis.

Discrimination of damaged/dead cells by propidium iodide uptake in immunofluorescently labeled populations analyzed by phase-sensitive flow cytometry.

We report a flow cytometric fluorescence lifetime-based method to discriminate damaged/dead from viable cells in immunofluorescently labeled populations using propidium iodide as a dye-exclusion viability probe. Fluorescence signals from propidium iodide and the anti-thymus cell-surface immunofluorescence marker fluorochromes, phycoerythrin and phycoerythrin/Texas Red (tandem conjugate), which have overlapping emission spectra with propidium iodide, are resolved based on differences in their fluorescence emission lifetimes using phase-sensitive detection. Mouse thymus cell samples were first labeled separately with anti-Thy 1.2 antibody directly conjugated to phycoerythrin and to phycoerythrin/Texas Red and propidium iodide. Labeled cells were then analyzed to determine the lifetimes of the immunofluorescence markers and propidium iodide. Based on these results, rat and mouse thymocytes labeled with anti-Thy 1.1 conjugated to phycoerythrin and anti-Thy 1.2 conjugated to phycoerythrin/Texas Red, respectively, were suspended in phosphate buffered saline containing propidium iodide, and were analyzed as they passed through a flow chamber and crossed a high-frequency, intensity-modulated (sinusoidal) laser excitation beam. The resulting immunofluorescence and propidium iodide signals were resolved based on differences in fluorescence lifetimes expressed as phase shifts using phase-sensitive detection and displayed as frequency distribution histograms and bivariate contour diagrams. This technology provides a new method to resolve immunofluorescence and propidium iodide signals from overlapping fluorescence emission spectra and a flow cytometric lifetime-based technique to quantify damaged/dead cells in immunofluorescence studies.

Pulmonary and thoracic macrophage subpopulations and clearance of particles from the lung.

Pulmonary macrophages consist of several subpopulations that can be defined by their anatomical locations as well as by other criteria. In addition to the well-known alveolar macrophages that reside on the alveolar surface, pulmonary macrophages also occur in the conducting airways, in various pulmonary interstitial regions, and, in some mammalian species, in the lung's intravascular compartment. Other thoracic macrophages of relevance to pulmonary defense and some lung disease processes are the pleural macrophages resident in the pleural space and macrophages present in regional lymph nodes that receive lymphatic drainage from the lung. Of the above subpopulations of pulmonary and thoracic macrophages, the alveolar macrophages have received the most experimental attention in the context of the pulmonary clearance and retention of deposited particles. Accordingly, less information is currently available regarding the roles other pulmonary and thoracic populations of macrophages may play in the removal of particles from the lower respiratory tract and associated tissue compartments. This report provides an overview of the various subpopulations of pulmonary and thoracic macrophages, as defined by their anatomical locations. The known and postulated roles of macrophages in the pulmonary clearance and retention of particles are reviewed, with particular emphasis on macrophage-associated processes involved in the pulmonary clearance of relatively insoluble particles.

A new mechanism for DNA alterations induced by alpha particles such as those emitted by radon and radon progeny.

The mechanism(s) by which alpha (alpha) particles like those emitted from inhaled radon and radon progeny cause their carcinogenic effects in the lung remains unclear. Although direct nuclear traversals by alpha-particles may be involved in mediating these outcomes, increasing evidence indicates that a particles can cause alterations in DNA in the absence of direct hits to cell nuclei. Using the occurrence of excessive sister chromatid exchanges (SCE) as an index of DNA damage in human lung fibroblasts, we investigated the hypothesis that alpha-particles may induce DNA damage through the generation of extracellular factors. We have found that a relatively low dose of alpha-particles can result in the generation of extracellular factors, which, upon transfer to unexposed normal human cells, can cause excessive SCE to an extent equivalent to that observed when the cells are directly irradiated with the same irradiation dose. A short-lived, SCE-inducing factor(s) is generated in alpha-irradiated culture medium containing serum in the absence of cells. A more persistent SCE-inducing factor(s), which can survive freeze-thaw and is heat labile is produced by fibroblasts after exposure to the alpha-particles. These results indicate that the initiating target for alpha-particle-induced genetic changes can be larger than a cell's nucleus or even a whole cell. How transmissible factors like those observed here in vitro may extend to the in vivo condition in the context of a-particle-induced carcinogenesis in the respiratory tract remains to be determined.

Correlation between particle size, in vivo particle persistence, and lung injury.

Dosimetry parameters such as deposition, clearance, retention, and translocation and dissolution of inhaled particles in and to different lung compartments may be important for the persistence of particles in the lung and may correlate with adverse pulmonary effects. We investigated such correlations using a model involving TiO2 particles of two particle sizes (20 nm diameter, ultrafine; 250 nm diameter, fine) of the same crystalline structure (anatase). A 12-week inhalation experiment in rats resulted in a similar mass deposition of the two particle types in the lower respiratory tract. The ultrafine particles elicited a persistently high inflammatory reaction in the lungs of the animals compared to the larger-sized particles. In the postexposure period (up to 1 year) retention in the alveolar space per se was not different between fine and ultrafine TiO2. However, the following differences between the particle types were noted: a significantly different total pulmonary retention, both quantitatively (significantly prolonged retention of the ultrafine TiO2) and qualitatively (increased translocation to the pulmonary interstitium and persistence there of the ultrafine TiO2); greater epithelial effects (Type II cell proliferation; occlusion of pores of Kohn) and the beginning of interstitial fibrotic foci with ultrafine TiO2; significant sustained impairment of alveolar macrophage function after ultrafine TiO2 exposure as measured by the clearance of test particles. A correlation between particle surface area and effects was observed. A comparison of the adverse reactions with dosimetric parameters of TiO2 in different lung compartments in the postexposure period showed a correlation of the persistence of effects in both the alveolar and interstitial space with the persistence of particles in the respective compartment.