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Lysis
Kinds of Lysis Terms modified by Lysis Selected AbstractsCell Lysis of Cyanobacteria and Its Implications for Nutrient DynamicsINTERNATIONAL REVIEW OF HYDROBIOLOGY, Issue 3 2010Linlin Ye Abstract The dynamics of nutrients, such as phosphorus, nitrogen, and carbohydrates, during cyanobacteria cell lysis was investigated under darkness incubation in the laboratory. The cell lysis rate of cyanobacteria sampled from Lake Taihu was measured using an esterase assay. Based on particulate esterase activity, the calculated cyanobacteria lysis rate was 0.094 d,1. During 30 days of darkness incubation, Chlorophyll a concentration decreased from 56 ,g L,1 to 2.0 ,g L,1. Parallel to this, total particulate carbohydrate concentration decreased rapidly. The fluctuation of dissolved organic carbon concentration was a function of the production of non-carbohydrate by cyanobacteria and the decomposition of carbohydrate by bacteria. Total dissolved carbohydrates and dissolved polysaccharides concentrations showed a similar pattern, declining at the beginning of the experiment and keeping relatively stable, thereafter. In contrast, the concentration of dissolved monosaccharides remained constant during the entire process. The concentrations of NH4+ and PO43, increased at the early stage, and then decreased afterwards. A gradual decrease in NO3, concentration after day 8 indicated that anaerobic conditions might be produced during the cell lysis process. The present results demonstrated cyanobacteria cell lysis has a big influence on the nutrient status of the surrounding water. (© 2010 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim) [source] Acute Monoplegia After Lysis of Epidural Adhesions: A Case ReportPAIN PRACTICE, Issue 5 2008DAAPM, Kok-Yuen Ho MBBS, MMed (Anaes) Abstract Lysis of epidural adhesions is an interventional procedure performed to treat refractory low back pain or radicular pain due to epidural scarring. A 39-year-old female with a diagnosis of failed back surgery syndrome underwent lysis of epidural adhesions using a Racz catheter. She developed acute monoplegia of her right lower extremity immediately after the procedure. Radiographic imaging did not reveal an epidural hematoma. Motor and sensory function in the right lower limb returned after 5 days. Large volumes of fluid injected during neuroplasty could have caused transient nerve injury from compression within loculated epidural compartments. [source] Plato and the Love of IndividualsTHE HEYTHROP JOURNAL, Issue 3 2002T. Brian Mooney A perennial problem in the philosophy of love has centred around what it is to love persons qua persons. Plato has usually been interpreted as believing that when we love we are attaching ourselves to qualities that inhere in the objects of our love and that these qualities transcend the objects. Vlastos has argued, along with Nussbaum, Price and many others that such an account tells against a true love of persons as unique and irreplaceable individuals. I argue that Plato's account of love as presented in the Lysis and Symposium is not so easily rejected. My concern is both to show that Plato can meet the objections and that his theory can still offer helpful insights into the understanding of love in our own lives. In particular I will identify two manners of loving persons; one which is context and individual specific, and another which might be termed metaphysical, thereby preserving aspects of the Platonic ascent of love. I will further argue that the two aspects are often non,controversially linked, and that such linking helps explain something of the mysterious nature of love. [source] BK Polyoma Virus Allograft Nephropathy: Ultrastructural Features from Viral Cell Entry to LysisAMERICAN JOURNAL OF TRANSPLANTATION, Issue 11 2003Cinthia B Drachenberg BK virions must enter the host cell and target their genome to the nucleus in order to complete their life cycle. The mechanisms by which the virions accomplish these tasks are not known. In this morphological study we found that BK virions localized beneath the host cell cytoplasmic membrane in 60,70-nm, smooth (non-coated) monopinocytotic vesicles similar to, or consistent with, caveolae. In the cytoplasm, the monopinocytotic vesicles carrying virions appeared to fuse with a system of smooth, vesicles and tubules that communicated with the rough endoplasmic reticulum and was continuous with the Golgi system. Membrane-bound single virions and large tubulo-reticular complexes loaded with virions accumulated in paranuclear locations. Occasional nuclei displayed virions within the perinuclear cisterna in association to the perinuclear viral accumulations. Tubular cells with mature productive infection had large nuclei, distended by daughter virions, whereas they lacked significant numbers of cytoplasmic virions. In addition to virally induced cell necrosis, there was extensive tubular cell damage (apoptosis and necrosis) in morphologically non-infected tubules. The observed ultrastructural interactions between the BK virions and host cells are remarkably similar to viral cell entry and nuclear targeting described for SV40 virus. [source] Biomass recycling from a riboflavin cultivation with B. subtilis: Lysis, extract production and testing as substrate in riboflavin cultivationBIOTECHNOLOGY & BIOENGINEERING, Issue 6 2006Karlheinz Bretz Abstract Autolysis of riboflavin-producing B. subtilis can be induced by pH, lack of carbon source, and the buffer system. Stress factors like temperature shift or oxygen dearth enhance the autolysis process. After cultivation of a riboflavin-producing strain, the pH of the whole culture broth was adjusted to 6.5,7.5. At a temperature of 40°C, autolysis started after 1 h. Adding a defined amount of commercially available endo- and exo-proteases enhanced both auto- and proteo-lysis. Optimization of endo- and exo-protease concentrations and of the time increased the degree of proteolysis. Additionally, the amount of DNA and Protein trapped in the riboflavin crystals could be significantly reduced by autolysis. After autolysis, the cultivation broth was centrifuged and the supernatant was cross-flow filtrated with a cut off of 10 kDa. Using this autolysate instead of yeast extract as a medium component for riboflavin production with B. subtilis, a riboflavin yield of 77% was obtained in comparison with the standard cultivation on yeast extract. © 2006 Wiley Periodicals, Inc. [source] Diagnosing PNH with FLAER and multiparameter flow cytometryCYTOMETRY, Issue 3 2007D. Robert Sutherland Abstract Background: PNH is an acquired hematopoietic stem cell disorder leading to a partial or absolute deficiency of all glycophosphatidyl-inositol (GPI)-linked proteins. The classical approach to diagnosis of PNH by cytometry involves the loss of at least two GPI-linked antigens on RBCs and neutrophils. While flow assays are more sensitive and specific than complement-mediated lysis or the Hams test, they suffer from several drawbacks. Bacterial aerolysin binds to the GPI moiety of cell surface GPI-linked molecules and causes lysis of normal but not GPI-deficient PNH cells. FLAER is an Alexa488-labeled inactive variant of aerolysin that does not cause lysis of cells. Our goals were to develop a FLAER-based assay to diagnose and monitor patients with PNH and to improve detection of minor populations of PNH clones in other hematologic disorders. Methods: In a single tube assay, we combined FLAER with CD45, CD33, and CD14 allowing the simultaneous analysis of FLAER and the GPI-linked CD14 structure on neutrophil and monocyte lineages. Results: Comparison to standard CD55 and CD59 analysis showed excellent agreement. Because of the higher signal to noise ratio, the method shows increased sensitivity in our hands over single (CD55 or CD59) parameter analysis. Using this assay, we were able to detect as few as 1% PNH monocytes and neutrophils in aplastic anemia, that were otherwise undetectable using CD55 and CD59 on RBC's. We also observed abnormal FLAER staining of blast populations in acute leukemia. In these cases, the neutrophils stained normally with FLAER, while the gated CD33bright cells failed to express normal levels of CD14 and additionally showed aberrant CD45 staining and bound lower levels of FLAER. Conclusion: FLAER combined with multiparameter flow cytometry offers an improved assay for diagnosis and monitoring of PNH clones and may have utility in detection of unsuspected myeloproliferative disorders. © 2007 Clinical Cytometry Society [source] Flow cytometric measurement of circulating endothelial cells: The effect of age and peripheral arterial disease on baseline levels of mature and progenitor populationsCYTOMETRY, Issue 2 2006Rebecca Gusic Shaffer Abstract Background: Age and cardiovascular disease status appear to alter numbers and function of circulating endothelial progenitor cells (EPCs). Despite no universal phenotypic definition, numerous studies have implicated progenitors with apparent endothelial potential in local responses to vascular injury and with cardiovascular disease in general. To further define the role of this lineage in peripheral artery disease (PAD), we developed a multiparameter flow cytometry assay to analyze multiple phenotypic definitions of progenitor cells (PCs), EPCs, and mature endothelial cells (ECs) and evaluate effects of age and PAD on baseline levels of each subset. Methods: Blood was collected from young healthy subjects (N = 9, mean age 33 ± 8 years), older healthy subjects (N = 13, mean age 66 ± 8 years), and older subjects with PAD (N = 15, mean age 69 ± 8 years). After ammonium chloride lysis, cells were stained and analyzed on a Becton-Dickinson LSR II with a 5-color antibody panel: FITC-anti-CD31, PE-anti-CD146, PE-anti-CD133, PerCP-Cy5.5-anti-CD3,-CD19,-CD33 (lineage panel), PE-Cy7-anti-CD34, and APC-anti-VEGF-R2. Viability was assessed by propidium iodide exclusion, and only viable, low to medium side scatter lineage-negative singlets were analyzed. In some studies, cells were sorted for morphological studies. Subsets were defined as indicated later. Results: Our results, using a comprehensive flow cytometric panel, indicate that CD133+, CD34+, and CD133+/CD34+ PCs are elevated in younger healthy individuals compared to older individuals, both healthy and with PAD. However, the number of EPCs and mature ECs did not significantly differ among the three groups. Assessment of endothelial colony forming units and dual acLDL-lectin staining supported the flow cytometric findings. Conclusions: We describe a comprehensive flow cytometric method to detect circulating mature and progenitor endothelial populations confirmed by conventional morphological and functional assays. Our findings suggest that aging may influence circulating levels of PCs, but not EPCs or ECs; PAD had no effect on baseline levels of any populations investigated. This study provides the basis for evaluating the potential effects of acute stress and therapeutic intervention on circulating progenitor and endothelial populations as a biomarker for cardiovascular status. © 2005 International Society for Analytical Cytology [source] TOGp regulates microtubule assembly and density during mitosis and contributes to chromosome directional instabilityCYTOSKELETON, Issue 8 2009Lynne Cassimeris Abstract TOGp, a member of the XMAP215 MAP family, is required for bipolar mitotic spindle assembly. To understand how TOGp contributes to spindle assembly, we examined microtubule dynamics after depleting TOGp by siRNA. Fluorescence recovery after photobleaching of GFP-tubulin demonstrated that spindle microtubule turnover is slowed two-fold in the absence of TOGp. Consistent with photobleaching results, microtubule regrowth after washout of the microtubule depolymerizing drug nocodazole was slower at the centrosomes and in the vicinity of mitotic chromatin in cells depleted of TOGp. The slower microtubule turnover is likely due to either nucleation or the transitions of dynamic instability because TOGp depletion did not effect the rate of plus end growth, measured by tracking EB1-GFP at microtubule ends. In contrast, microtubule regrowth after nocodazole washout was unaffected by prior depletion of TACC3, a centrosomal protein that interacts with TOGp. Kinetochore fibers in both untreated and TOGp-depleted cells were stable to incubation at 4°C or lysis in buffer containing calcium indicating that stable kinetochore-microtubule attachments are formed in the absence of TOGp. Depletion of TOGp, but not TACC3, reduced kinetochore oscillations during prometaphase/metaphase. Defects in oscillations are not due simply to multipolarity or loss of centrosome focus in the TOGp-depleted cells, since kinetochore oscillations appear normal in cells treated with the proteosome inhibitor MG132, which also results in multipolar spindles and centrosome fragmentation. We hypothesize that TOGp is required for chromosome motility as a downstream consequence of reduced microtubule dynamics and/or density. Cell Motil. Cytoskeleton 2009. © 2009 Wiley-Liss, Inc. [source] Mesotherapy and Phosphatidylcholine Injections: Historical Clarification and ReviewDERMATOLOGIC SURGERY, Issue 4 2006ADAM M. ROTUNDA MD BACKGROUND Mesotherapy was originally conceived in Europe as a method of utilizing cutaneous injections containing a mixture of compounds for the treatment of local medical and cosmetic conditions. Although mesotherapy was traditionally employed for pain relief, its cosmetic applications, particularly fat and cellulite removal, have recently received attention in the United States. Another treatment for localized fat reduction, which was popularized in Brazil and uses injections of phosphatidylcholine, has been erroneously considered synonymous with mesotherapy. Despite their attraction as purported "fat-dissolving" injections, the safety and efficacy of these novel cosmetic treatments remain ambiguous to most patients and physicians. OBJECTIVE To distinguish mesotherapy from phosphatidylcholine injections by reviewing their history and the relevant experimental or clinical findings. METHODS A comprehensive search of Medline indexed literature and conference proceedings. RESULTS All the published studies evaluating the clinical efficacy of traditional mesotherapy currently originate from Europe. These reports focus primarily on musculoskeletal pain and vascular disease, rather than cosmetic applications. Although experimental data suggest that a number of traditional mesotherapy ingredients may theoretically reduce fat, these effects have not been supported in peer-reviewed studies. An increasing number of reports demonstrate that subcutaneous injections of a formula containing phosphatidylcholine combined with its emulsifier, deoxycholate, are effective in removing small collections of adipose tissue. Cell lysis, resulting from the detergent action of deoxycholate, may account for this clinical effect. CONCLUSIONS Mesotherapy is distinct from a method of treating adipose tissue with subcutaneous injections of deoxycholate alone or in combination with phosphatidylcholine. Additional clinical and experimental studies are necessary to more definitively establish the safety and efficacy of these treatments. [source] Ex vivo TCR-induced leukocyte gene expression of inflammatory mediators is increased in type 1 diabetic patients but not in overweight childrenDIABETES/METABOLISM: RESEARCH AND REVIEWS, Issue 1 2010Jaime S. Rosa Abstract Background Abnormal systemic concentrations of proinflammatory cytokines/chemokines have been implicated in the development of long-term cardiovascular complications in type 1 diabetes (T1DM) and obesity. Whether leukocyte white blood cell (WBC) gene expression of these proinflammatory mediators contributes to their increased systemic levels, however, remains unclear, especially in the pediatric patient populations. This study examines mRNA changes of 9 cytokines and chemokines in WBCs following ex vivo immunostimulation from 9 T1DM (13.4 ± 0.5 year, 4F/5 M), 23 overweight (OW, 12.3 ± 0.5 year, 10F/13M, BMI% 97.1 ± 0.5 and > 90.0), and 21 healthy (CL, 13.8 ± 0.7 year, 9F/12 M, BMI% 59.6 ± 4.6 and < 85.0) children. Methods All subjects had been maintained in euglycemic conditions for at least 90 min before blood draws. Whole blood was then sampled and incubated with anti-T-cell receptor (TCR) antibody or heat-aggregated IgG (HAG) to stimulate T-cell and Fc receptors (FcR), respectively. After lysis of leukocytes, mRNA levels of six tumor necrosis factor superfamily cytokines (TNFSF2, 5, 6, 7, 9, 14) and three chemokines (CCL8, 20, and CXCL10) were measured using RT-PCR. Results Following TCR stimulation, T1DM displayed significantly greater mRNA responses than CL for TNFSF5, 7, 9, and CCL8, and CXCL10; TNFSF9, CCL8, and CXCL10 were also significantly higher in T1DM than OW; no difference was observed between OW and CL. FcR stimulation induced similar responses across groups. Conclusions Leukocytes of T1DM children displayed exaggerated gene expression in response to ex vivo TCR induction of five key proinflammatory cytokines/chemokines. This elevated leukocyte gene expression may be one of the pathophysiological contributors to the development of vascular complications in T1DM. Copyright © 2009 John Wiley & Sons, Ltd. [source] Viral infections as potential triggers of type 1 diabetesDIABETES/METABOLISM: RESEARCH AND REVIEWS, Issue 3 2007Nienke van der Werf Abstract During the last decades, the incidence of type 1 diabetes (T1D) has increased significantly, reaching percentages of 3% annually worldwide. This increase suggests that besides genetical factors environmental perturbations (including viral infections) are also involved in the pathogenesis of T1D. T1D has been associated with viral infections including enteroviruses, rubella, mumps, rotavirus, parvovirus and cytomegalovirus (CMV). Although correlations between clinical presentation with T1D and the occurrence of a viral infection that precedes the development of overt disease have been recognized, causalities between viruses and the diabetogenic process are still elusive and difficult to prove in humans. The use of experimental animal models is therefore indispensable, and indeed more insight in the mechanism by which viruses can modulate diabetogenesis has been provided by studies in rodent models for T1D such as the biobreeding (BB) rat, nonobese diabetic (NOD) mouse or specific transgenic mouse strains. Data from experimental animals as well as in vitro studies indicate that various viruses are clearly able to modulate the development of T1D via different mechanisms, including direct ,-cell lysis, bystander activation of autoreactive T cells, loss of regulatory T cells and molecular mimicry. Data obtained in rodents and in vitro systems have improved our insight in the possible role of viral infections in the pathogenesis of human T1D. Future studies will hopefully reveal which human viruses are causally involved in the induction of T1D and this knowledge may provide directions on how to deal with viral infections in diabetes-susceptible individuals in order to delay or even prevent the diabetogenic process. Copyright © 2007 John Wiley & Sons, Ltd. [source] Dynamics of P2X7 receptor pore dilation: Pharmacological and functional consequencesDRUG DEVELOPMENT RESEARCH, Issue 2-3 2001I.P. Chessell Abstract The biophysical and functional properties of the human P2X7 receptor, expressed recombinantly in HEK-293 cells or natively in THP-1 pro-monocytic cells, were investigated in the context of pore dilation and externalisation of mature interleukin 1, (IL1,). In HEK-293 cells, the agonist 2,- and 3,-O-(4-benzoylbenzoyl)-ATP (BzATP) caused concentration-dependent inward currents (EC50 59 ,M) and with prolonged application this agonist caused a gradual increase in inward current culminating in a plateau. This increase in current was associated with pore dilation, determined by intracellular accumulation of YO-PRO-1. BzATP displayed increased potency at the pore-dilated form of the P2X7 receptor (EC50 17 ,M), and positive correlations between apparent receptor density and speed of pore dilation were observed. A monoclonal antibody selectively blocked current mediated by the naïve receptor, while currents through pore-dilated receptors were not significantly affected, which together suggest a conformational change at the level of the receptor during the dilation event. The release of mature IL1, from THP-1 cells was independent of P2X7 -mediated cell lysis, as determined by study of lactate dehydrogenase release. Moreover, using conditions designed to minimise pore dilation (using buffers containing 2 mM Ca2+ and 1 mM Mg2+), BzATP caused significant release of IL1,, but without concomitant YO-PRO-1 accumulation, indicating pore dilation is not required for IL1, release. In addition, short (4-min) incubation of THP-1 cells with BzATP (terminated by enzymatic degradation of BzATP using apyrase) resulted in significant quantities of IL1, release some 60 min later, suggesting commitment of cells to release IL1, can be triggered with only brief receptor ligation. These findings suggest that receptor expression and ligation time are critical factors for selecting multiple functional states of P2X7. Drug Dev. Res. 53:60,65, 2001. © 2001 Wiley-Liss, Inc. [source] Incremental Value of Live/Real Time Three-Dimensional Transthoracic Echocardiography in the Assessment of Right Ventricular MassesECHOCARDIOGRAPHY, Issue 5 2009Venkataramana K. Reddy M.D. This case series demonstrates the incremental value of three-dimensional transthoracic echocardiography (3D TTE) over two-dimensional transthoracic echocardiography (2D TTE) in the assessment of 11 patients with right ventricular (RV) masses or mass-like lesions (three cases of RV thrombus, one myxoma, one fibroma, one lipoma, one chordoma, and one sarcoma and three cases of RV noncompaction, which are considered to be mass-like in nature). 3D TTE was of incremental value in the assessment of these masses in that 3D TTE has the capacity to section the mass and view it from multiple angles, giving the examiner a more comprehensive assessment of the mass. This was particularly helpful in the cases of thrombi, as the presence of echolucencies indicated clot lysis. In addition, certainty in the number of thrombi present was an advantage of 3D TTE. Also, sectioning of cardiac tumors allowed more confidence in narrowing the differential diagnosis of the etiology of the mass. In addition, 3D TTE allowed us to identify precise location of the attachments of the masses as well as to determine whether there were mobile components to the mass. Another noteworthy advantage of 3D TTE was that the volumes of the masses could be calculated. Additionally, the findings by 3D TTE correlated well with pathologic examination of RV tumors, and some of the masses measured larger by 3D TTE than by 2D TTE, which was also validated in one case by surgery. As in the case of RV fibroma, another advantage was that 3D TTE actually identified more masses than 2D TTE. RV noncompaction was also well studied, and the assessment with 3D TTE helped to give a more definitive diagnosis in these patients. [source] On-line cell lysis and DNA extraction on a microfluidic biochip fabricated by microelectromechanical system technologyELECTROPHORESIS, Issue 9 2008Xing Chen Dr. Abstract Integrating cell lysis and DNA purification process into a micrototal analytical system (,TAS) is one critical step for the analysis of nucleic acids. On-chip cell lysis based on a chemical method is realized by sufficient blend of blood sample and the lyzing reagent. In this paper two mixing models, T-type mixing model and sandwich-type mixing model, are proposed and simulation of those models is conducted. Result of simulation shows that the sandwich-type mixing model with coiled channel performs best and this model is further used to construct the microfluidic biochip for on-line cell lysis and DNA extraction. The result of simulation is further verified by experiments. It asserts that more than 80% mixing of blood sample and lyzing reagent which guarantees that completed cell lysis can be achieved near the inlet location when the cell/buffer velocity ratio is less than 1:5. After cell lysis, DNA extraction by means of a solid-phase method is implemented by using porous silicon matrix which is integrated in the biochip. During continuous flow process in the microchip, rapid cell lysis and PCR-amplifiable genomic DNA purification can be achieved within 20,min. The potential of this microfluidic biochip is illustrated by pretreating a whole blood sample, which shows the possibility of integration of sample preparation, PCR, and separation on a single device to work as portable point-of-care medical diagnostic system. [source] Thiol redox status evaluation in red blood cells by capillary electrophoresis-laser induced fluorescence detectionELECTROPHORESIS, Issue 10 2005Angelo Zinellu Abstract Thiols and in particular glutathione (GSH) play a central role in human metabolism, including the detoxification of xenobiotics, cell homeostasis, radioprotection, and antioxidant defence. Here, a new method is provided for the measurement of reduced and total forms of thiols in red blood cells. In order to minimize oxidation of reduced thiols, a water erythrocyte lysis (15 min at 4°C) was performed followed by a protein precipitation step with acetonitrile. The supernatant was rapidly derivatized with 5-iodoacetoamidefluorescein that trapped thiol groups, thus minimizing auto-oxidation. Derivatized samples were separated in a 57 cm × 75 ,m ID capillary by using 5 mmol/L sodium phosphate, 4 mmol/L boric acid as electrolyte solution with 75 mmol/L N -methyl- D -glucamine at pH 11.0. Under these conditions, cysteinylglycine (CysGly), cysteine (Cys), glutathione, and ,-glutamylcysteine (GluCys) were baseline-resolved in , 4 min. Precision tests showed a good repeatability of our method both for migration times (coefficient of variation CV < 0.8%) and areas (CV < 3.3%). Furthermore, a good reproducibility of intrassay and interassay tests was obtained (CV < 5% and CV < 8%, respectively). The method was employed to investigate the effect of acidic precipitation on intracellular thiol concentration. Our data suggest that sample acidification causes a modification of the measured redox thiol status due to the development of a pro-oxidant environment; moreover, the thiol redox status of red blood cells was evaluated in 22 healthy volunteers. [source] Plants, gall midges, and fungi: a three-component systemENTOMOLOGIA EXPERIMENTALIS ET APPLICATA, Issue 1 2008Odette Rohfritsch Abstract Larvae of gall midges (Diptera: Cecidomyiidae) induce the activation of plant cells, partial cell lysis, and differentiation of nutritive tissue. Specialized nutritive tissue is essential for larval development and plays a key role in gall organization. Midges of the tribes Lasiopterini and Asphondyliini, however, do not induce nutritive tissues as part of the formation of their galls. Instead, these ,ambrosia galls' contain fungal mycelia that line the interior surface of the chambers. The fungi not only provide Lasiopterini with nutrition, they also penetrate the stems, induce the lysis of the middle lamella of host cells, and open a channel to the vascular bundles. Larvae of Lasioptera arundinis (Schiner) (Lasiopterini) follow the fungus and feed on its mycelium along with adjoining stem cells of Phragmites australis (Cav.) Trin. (Poaceae). Eggs together with fungal conidia are deposited by the imago on the host. Asphondyliini use a needle-like ovipositor to introduce fungal conidia and eggs into the organs they attack. Larvae of Schizomyia galiorum Kieffer (Asphondyliini) are unable to initiate the gall or to develop in the flowers of Galium mollugo L. (Rubiaceae) without their fungal associate. In this article, I provide an overview of oviposition behaviour in the Asphondyliini, as well as descriptions of the ovipositor and the female post-abdominal segments. Gall formation by Lasiopterini and Asphondyliini and the role of associated fungi are discussed, as is the role of the fungus as an inquiline or an organizer of gall tissues and a nutritive device. [source] Ultrastructural and electron energy-loss spectroscopic analysis of an extracellular filamentous matrix of an environmental bacterial isolateENVIRONMENTAL MICROBIOLOGY, Issue 9 2007Uta Böckelmann Summary Strain F8, a bacterial isolate from ,river snow', was found to produce extracellular fibres in the form of a filamentous network. These extracellular filaments, which were previously shown to be composed of DNA, have been studied for the first time by ultrastructural and electron energy-loss spectroscopy in the present work. ,Whole mount' preparations of strain F8 indicate these polymers are ultrastructurally homogeneous and form a network of elemental filaments, which have a width of 1.8,2.0 nm. When incubated at pH 3.5 with colloidal cationic ThO2 tracers they become intensely stained (electron dense), affording direct evidence that the fibres are negatively charged and thus acidic chemically. Elemental analysis of the extracellular filaments by Energy-filtered Transmission Electron Microscopy revealed phosphorus to be the main element present and, because pretreatment of F8 cells with DNase prevented thorium labelling, the fibres must be composed of extracellular DNA (eDNA). Neither ultrathin sections nor ,whole mount negative stain' caused DNA release by general cell lysis. Additionally, cells infected with phages were never observed in ultrathin sections and phage particles were never detected in whole mount samples, which rules out the possibility of phages being directly involved in eDNA release. [source] Towards clarification of the biological role of microcystins, a family of cyanobacterial toxinsENVIRONMENTAL MICROBIOLOGY, Issue 4 2007Daniella Schatz Summary Microcystins constitute a serious threat to the quality of drinking water worldwide. These protein phosphatase inhibitors are formed by various cyanobacterial species, including Microcystis sp. Microcystins are produced by a complex microcystin synthetase, composed of peptide synthetases and polyketide synthases, encoded by the mcyA-J gene cluster. Recent phylogenetic analysis suggested that the microcystin synthetase predated the metazoan lineage, thus dismissing the possibility that microcystins emerged as a means of defence against grazing, and their original biological role is not clear. We show that lysis of Microcystis cells, either mechanically or because of various stress conditions, induced massive accumulation of McyB and enhanced the production of microcystins in the remaining Microcystis cells. A rise in McyB content was also observed following exposure to microcystin or the protease inhibitors micropeptin and microginin, also produced by Microcystis. The extent of the stimulation by cell extract was strongly affected by the age of the treated Microcystis culture. Older cultures, or those recently diluted from stock cultures, hardly responded to the components in the cell extract. We propose that lysis of a fraction of the Microcystis population is sensed by the rest of the cells because of the release of non-ribosomal peptides. The remaining cells respond by raising their ability to produce microcystins thereby enhancing their fitness in their ecological niche, because of their toxicity. [source] Grazer and virus-induced mortality of bacterioplankton accelerates development of Flectobacillus populations in a freshwater communityENVIRONMENTAL MICROBIOLOGY, Issue 3 2007Karel, imek Summary We present a detailed analysis of the effects of distinct bacterial mortality factors, viral lysis and heterotrophic nanoflagellates (HNF) bacterivory, associated with the development of filamentous Flectobacillus populations. Reservoir bacterioplankton communities were subjected to additions of both HNF and viruses together, or HNF alone, and then incubated in situ in dialyses bags. For distinct bacterial groups, mortality or growth stimulation was analysed by examining bacterial prey ingested in HNF food vacuoles with fluorescence in situ hybridization (FISH) and via FISH with microautoradiography (MAR-FISH). We also developed a semi-quantitative MAR-FISH-based estimation of relative activities of Flectobacillus populations (targeted by the R-FL615 probe). Bacterial groups vulnerable to HNF predation (mainly clusters of Betaproteobacteria), or discriminated against (Actinobacteria), were detected. Bacterial lineages most vulnerable to virus-lysis (mainly the Betaproteobacteria not targeted by the R-BT065 probe, of the Polynucleobacter cluster) were identified by comparing treatments with HNF alone to HNF and viruses together. Filaments affiliated with the Flectobacillus cluster appeared in both treatments, but were about twice as abundant, long and active as in incubations with viruses and HNF as compared with HNF alone. Viruses appeared to selectively suppress several bacterial groups, perhaps enhancing substrate availability thus stimulating growth and activity of filamentous Flectobacillus. [source] DNA extraction procedure: a critical issue for bacterial diversity assessment in marine sedimentsENVIRONMENTAL MICROBIOLOGY, Issue 2 2006Gian Marco Luna Summary In order to evaluate whether different DNA extraction procedures can affect estimates of benthic bacterial diversity, based on 16S rRNA gene terminal restriction fragment length polymorphism (T-RFLP) fingerprinting technique, we compared two in situ lysis procedures (a SDS-based protocol and a commercial kit for DNA recovery) and one cell-extraction protocol on a variety of marine sediments. Despite the two in situ lysis procedures resulted in significantly different DNA yields (highest with the SDS in situ lysis), estimates of bacterial diversity provided a not significantly different ribotype richness, as well as similar values of the Shannon-Wiener (H,) and Margalef (d) indices of biodiversity and of evenness (Pielou index, J). Conversely, the cell-extraction procedure for DNA extraction resulted always in a significantly lower ribotype richness and diversity. The analysis of similarities (anosim) among the T-RFLP electropherograms allowed concluding that ribotypes composition did not change significantly using different protocols. However, the analysis of ,-diversity (turnover diversity) revealed that a large number of ribotypes was observed exclusively with one of the three protocols utilized. When unshared ribotypes from in situ lysis and cell extraction were pooled together, total ribotype richness resulted much higher (up to 80%). Our results indicate that estimates of ribotype diversity based on a single protocol of DNA extraction can significantly underestimate the total number of bacterial ribotypes present in the benthic domain. We recommend that future studies will not only integrate different DNA extraction procedures, but also will explore the possibility of integrating two or more different genetic markers in order to increase our ability to detect the actual bacterial diversity in environmental samples. [source] Phenotypical variation in a toxic strain of the phytoplankter, Cylindrospermopsis raciborskii (nostocales, cyanophyceae) during batch cultureENVIRONMENTAL TOXICOLOGY, Issue 6 2001Peter R. Hawkins Abstract A nonaxenic strain of Cylindrospermopsis raciborskii Woloszynska (AWT 205) was grown in batch culture, with and without nitrate as the primary N source. Rapid log-phase growth with nitrate was 1.0 doubling/day versus 0.3 doubling/day without nitrate. Cylindrospermopsin (CYN) production was measured by HPLC. The rate of intracellular CYN production matched cell division rate for both the diazotrophies at cell densities less than 107 cell/ml. At cell density >107 cell/ml, additional resource limitation in batch culture slowed log-phase growth to 0.04 division/day and cell division and CYN production decoupled. Intracellular CYN concentration increased at a rate of 0.08 doubling/day, twice the cell division rate. Extracellular CYN as a proportion of the total CYN increased from 20% during the rapid growth phase, to 50% during the slow growth phase. The total CYN yield from cultures grown out to stationary phase (55 days) exceeded 2 mg CYN/l. C. raciborskii cells in log-phase growth, exposed to 1 ppm copper (as copper sulphate), lysed within 24 hours. After copper treatment, all CYN was in the filterable fraction. These findings imply that in naturally occurring blooms of C. raciborskii, the movement of intracellular CYN into solution will be the greatest during stationary phase, when intracellular concentrations are highest and cell lysis is more frequent. The application of algicides that promote cell lysis will exacerbate this effect. © 2001 John Wiley & Sons, Inc. Environ Toxicol 16: 460,467, 2001 [source] Tiludronate infusion in the treatment of bone spavin: A double blind placebo-controlled trialEQUINE VETERINARY JOURNAL, Issue 5 2010M. R. GOUGH Summary Reasons for performing study: Tiludronate regulates bone remodelling through a decrease of the resorptive process and should therefore ameliorate the remodelling processes active in osteoarthritis of the distal tarsal joints (,bone spavin') and alleviate pain associated with abnormal bone lysis. Objective: To confirm the efficacy of tiludronate, administered as a single infusion at a dose of 1 mg/kg bwt, in the treatment of bone spavin in the horse. Methods: A double blind placebo controlled trial on 108 clinical cases of bone spavin was undertaken. The lameness score of the lamest limb was assessed following distal tarsal analgesia of the contralateral limb and followed-up using the same procedure throughout the study. Bone spavin in the lamest limb was confirmed by distal tarsal analgesia and radiography. Horses were treated at Day 0 and reassessed 60 days later after controlled exercise. A second nonblinded treatment was given to unresponsive horses and all horses were re-examined at Day 120. Exercise levels were recorded at each examination. Results: Eighty-seven horses completed the trial as per the protocol. The tiludronate horses were significantly less lame than the placebo horses (P = 0.0318). Horses treated at Day 60 with tiludronate showed further improvement in lameness at Day 120 (P = 0.0096 and P = 0.0034 for horses treated with tiludronate and placebo at Day 0, respectively). The only significant difference in radiographic findings between tiludronate and placebo was for presence of periarticular osteophytes (P = 0.006). Conclusions: Tiludronate treatment is proven to be effective in bone spavin in horses in association with a controlled exercise programme. Clinical relevance: Tiludronate in combination with controlled exercise offers an alternate medical treatment for bone spavin. [source] Disruption of nasopharyngeal epithelium by pneumococci is density-linkedEUROPEAN JOURNAL OF CLINICAL INVESTIGATION, Issue 4 2003K. Lagrou Abstract Background The aim of this project was to study the influence of pneumococci on nasopharyngeal epithelial integrity as a function of time and pneumococcal density. Materials and methods Cell layers of an in vitro model of human nasopharyngeal epithelium were inoculated with different pneumococcal strains. The transepithelial electrical resistance (TEER), a measure of the integrity of the cell layers, and the pneumococcal concentration in the apical fluid on the epithelial cells were measured at different times after inoculation. Results Pneumococci caused a decrease in the TEER when a density of 1 × 107 CFU mL,1 was reached. The growth rate of pneumococci in our in vitro model differed between the strains tested and, for the same strain, between in vitro culture on the epithelial cells and broth culture. Differences in timing of the onset of decrease in the TEER between strains were the result of differences in growth rate on the epithelial cells. Antibiotic-induced lysis of pneumococci caused an immediate decrease in the TEER of the cell layers. Conclusion Pneumococci cause a decrease in the TEER at a density of 1 × 107 CFU mL,1. Our hypothesis is that this decrease in the TEER is the result of quorum-induced lysis of the pneumococci. [source] Natural killer cells become tolerogenic after interaction with apoptotic cellsEUROPEAN JOURNAL OF IMMUNOLOGY, Issue 6 2010Wai Po Chong Abstract NK cells are effectors in innate immunity and also participate in immunoregulation through the release of TGF-,1 and lysis of activated/autoreactive T cells. Apoptotic cells (AC) have been shown to induce tolerogenic properties in innate immune cells, including macrophages and dendritic cells, but not NK cells. In this study, we demonstrated that after interaction with AC, NK cells released TGF-,1, which in turn suppressed the production of IFN-, by NK cells upon IL-12 and IgG activation. We further identified phosphatidylserine as a potential target on AC for the NK cells, as phosphatidylserine could stimulate NK cells to release TGF-,1, which in turn suppressed CD4+ T-cell proliferation and activation. Moreover, AC-treated NK cells displayed cytotoxicity against autologous-activated CD4+ T cells by upregulating NKp46. This lysis occurred in part through the NKp46-vimentin pathway, as activated CD4+ T cells expressed vimentin on the cell surface and blocking of vimentin or NKp46, but not other NK-cell receptors, significantly suppressed the NK-cell cytotoxicity. We report here a novel interaction between NK cells and AC, resulting in the tolerogenic properties of NK cells required for immune contraction. [source] DNAX accessory molecule-1 (CD226) promotes human hepatocellular carcinoma cell lysis by V,9V,2 T cellsEUROPEAN JOURNAL OF IMMUNOLOGY, Issue 5 2009Olivier Toutirais Abstract Human V,9V,2 T lymphocytes can be activated by nonpeptidic antigens such as the mevalonate pathway-derived isopentenyl pyrophosphate or synthetic phosphoantigen such as bromohydrin pyrophosphate. They display a strong cytotoxic activity against several tumor types, including hepatocellular carcinoma (HCC). Little is known about the mechanisms underlying V,9V,2 T-cell recognition of tumor cells, but there is strong evidence that activating NK receptors play a role in ,, T-cell cytotoxicity. In this study, we showed that the two NK receptors DNAX accessory molecule-1 (DNAM-1) and CD96 were expressed by V,9V,2 T cells. The ligands Nectin-like-5 specific of both DNAM-1 and CD96, and also Nectin-2, an additional ligand of DNAM-1, were present on all HCC cell lines analyzed. Furthermore, we demonstrated by mAb-mediated masking experiments that cytotoxicity against HCC cells as well as IFN-, production in ,, T cells were dependent on DNAM-1. Our experiments indicated that Nectin-like-5 but not Nectin-2 was involved in DNAM-1-dependent ,, T-cell functions. We did not reveal a role for CD96 in the killing of HCC cells. Finally, we showed by combined mAb-mediated blockade that DNAM-1 and NKG2D could cooperate in the cell lysis of HCC. [source] Both CD133+ and CD133, medulloblastoma cell lines express ligands for triggering NK receptors and are susceptible to NK-mediated cytotoxicityEUROPEAN JOURNAL OF IMMUNOLOGY, Issue 11 2007Roberta Castriconi Abstract Adoptive cellular immunotherapy has been proposed as an additional treatment of medulloblastoma, an intracranial tumor characterized by a particularly poor prognosis. However, little is known on the ability of the immune system to effectively attack this tumor. In this study, we show that activated human NK cells efficiently kill medulloblastoma cell lines in vitro. NK-mediated killing involved different activating receptors (including NKp46, NKp30, DNAM-1 and NKG2D) and correlated with the presence of their specific ligands on tumor cells. In contrast, the absence of major adhesion interactions, such as LFA-1/ICAM did not impair the NK-mediated cytotoxicity. Medulloblastoma expressed a number of tumor-associated molecules including CD146 and CD133, considered a marker for cancer stem cells. Remarkably, both CD133-positive and CD133-negative cell lines were susceptible to lysis. Tumor cells also expressed molecules that are currently used as diagnostic tools for neuroblastoma cell identification. In particular, B7 homolog 3 (B7-H3) was expressed by all the medulloblastoma cell lines analyzed, while the presence of GD2 and NB84 was restricted to given cell lines and/or marked a defined tumor cell subset. [source] Modulation of 2B4 (CD244) activity and regulated SAP expression in human NK cellsEUROPEAN JOURNAL OF IMMUNOLOGY, Issue 1 2007Johanna Endt Abstract The adapter protein SAP is important for the signal transduction of the family of SLAM-related receptors (SRR), which have important immune-modulating functions. The importance of SAP and SRR for a functional immune reaction becomes obvious in patients suffering from X-linked lymphoproliferative disease, which is characterized by non-functional SAP. Here we investigate the regulation of SAP expression in human NK cells. We demonstrate that SAP mRNA expression and protein levels are low in freshly isolated resting NK cells. IL-2 stimulation leads to an up-regulation of SAP expression, which can be enhanced by IL-12, the stimulation of TLR3 by polyinosinic-polycytidylic acid (poly(I:C))and to a lesser extent by IFN-,. EAT-2, a SAP-related adapter protein, is already detectable in resting NK cells and does not change its expression after IL-2 stimulation. The regulation of SAP has functional consequences for the stimulation of NK cell cytotoxicity by 2B4. In resting NK cells, 2B4 stimulation can only enhance NK cell lysis when co-triggered with other activating NK cell receptors. In IL-2-activated NK cells with high SAP expression the triggering of 2B4 alone is sufficient to induce NK cell cytotoxicity, demonstrating a correlation between the regulated SAP expression and the function of 2B4. [source] Cross-linking tumor cells with effector cells via CD55 with a bispecific mAb induces ,-glucan-dependent CR3-dependent cellular cytotoxicityEUROPEAN JOURNAL OF IMMUNOLOGY, Issue 4 2006Abstract Complement (C) regulatory proteins decrease the effectiveness of immunotherapeutic anti-cancer antibodies. Bispecific mAb (bi-mAb) that target a tumor antigen and simultaneously inhibit a C regulator increase the effectiveness of such a therapy. Here we investigated the mechanism by which bi-mAb increase tumor cell lysis. Apart from C-dependent cytotoxicity, C activation can lead to complement receptor 3 (CR3)-dependent cellular cytotoxicity (CR3-DCC) by CR3-positive effector cells in the presence of ,-glucan. Here we show that an anti-Ep-CAM*anti-CD55 bi-mAb induced more than threefold higher CR3-DCC (71%) of human colorectal cancer cells compared with anti-Ep-CAM alone (20%). This CR3-DCC was dependent on the binding of the anti-CD55 arm of tumor-bound anti-Ep-CAM*anti-CD55 bi-mAb to effector cell CD55, CR3 priming by ,-glucan and the presence of iC3b on the target cell. Comparable lysis could be obtained in the absence of iC3b, when CR3 and CD55 were cross-linked on the effector cells, suggesting cooperation between CD55 and CR3 in signal transduction. Tumor cells with low antigen expression were effectively lysed via this mechanism in contrast to direct C-dependent cytotoxicity. These data imply that the effectiveness of mAb immunotherapy can be improved using anti-tumor antigen*anti-CD55 bi-mAb and ,-glucan, thereby initiating CR3-DCC as an additional effector mechanism that is efficient for eradication of tumor cells with lower antigen expression. [source] Antibody-mediated bacterial clearance from the lower respiratory tract of mice requires complement component C3EUROPEAN JOURNAL OF IMMUNOLOGY, Issue 1 2004Elizabeth Abstract To assess the contribution of complement to respiratory immunity in the context of a natural bacterial infection, we used mice genetically deficient in complement components and the murine pathogen Bordetella bronchiseptica. Complement component C3 was not required for the control of bacterial infection or for the generation of infection-induced protective immunity. However, C3-deficient (C3,/,) mice were severely defective, compared to wild type, in vaccine-induced protective immunity. Adoptively transferred immune serum from convalescent wild-type or C3,/, animals rapidly cleared B.,bronchiseptica from the lungs of wild-type mice but did not affect its growth in C3,/, mice, indicating that the defect is not in the generation of protective immunity, but in its function. Immune serum was effective in C5-deficient mice but had little effect in the lungs of mice lacking either Fc, receptors (Fc,R) or CR3, suggesting bacterial clearance is not via direct complement-mediated lysis. Together, these data indicate that complement is required for antibody-mediated clearance of Bordetella and suggest the mechanism involves C3 opsonization of bacteria for phagocytosis that is both CR3- and Fc,R-dependent. [source] Efficient and selective tumor cell lysis and induction of apoptosis in melanoma cells by a conditional replication-competent CD95L adenovirusEXPERIMENTAL DERMATOLOGY, Issue 8 2010Lothar F. Fecker Please cite this paper as: Efficient and selective tumor cell lysis and induction of apoptosis in melanoma cells by a conditional replication-competent CD95L adenovirus. Experimental Dermatology 2010; 19: e56,e66. Abstract:, The high mortality of melanoma demands the development of new strategies, and gene therapy may be considered provided improvements in efficacy and selectivity. Overexpression of the death ligand CD95L/FasL has been shown in previous studies as highly effective for apoptosis induction in melanoma cells. For efficient and selective targeting of melanoma, a conditional replication-competent adenoviral vector was constructed (Ad5-FFE-02), which drives CD95L expression by a tetracycline-inducible promoter. For restricting its replication to melanoma cells, the adenoviral E1A gene is controlled by a tyrosinase-derived promoter. Furthermore, adenoviral E1B was deleted and a mutated E1A was used to preferentially support replication in tumor cells. Proving its high selectivity and efficiency, strong expression of E1A and doxycycline-dependent induction of CD95L were characteristic for tyrosinase-positive melanoma cells after Ad5-FFE-02 transduction, whereas absent in non-melanoma cell lines. Importantly, Ad5-FFE-02-mediated cell lysis was restricted to melanoma cells, and induction of apoptosis was found only in tyrosinase and CD95 expressing cells. Finally, the combination of adenoviral replication and CD95L-mediated apoptosis resulted in an enhanced repression of melanoma cell growth. This new adenoviral vector may provide a basis for an efficient targeting of melanoma. [source] | ||||||||||||||||||||||||||||||||||||||||