Image resolution of a microchannel plate x-ray image intensifier

The resolution has been evaluated for an experimental x-ray image intensifier employing three microchannel plates (MCPs) as the photon absorber and electron multipliers. The line spread function (LSF) was measured and used for determination of the modulation transfer function (MTF). The MTF was found to be independent of the incident photon energy from 20 to 150 keV. An additional measurement using a lead-bar test pattern showed that the resolution exceeded 7 line pairs (lp)/mm. The factors influencing the resolution capabilities of the intensifier are discussed. The resolution is limited primarily by the 53-micron center-to-center separation of the channels of the MCPs.     

METHYLATION STATUS OF BCL6 DISTINGUISHES DNA SAMPLES FROM BENIGN AND MALIGNANT LYMPHOPROLIFERATIVE DISORDERS

INTRODUCTION: Core biopsy of the breast has become the method of choice for tissue diagnosis of screen detected microcalcifications and some mass lesions in many breast assessment centres. Biopsy results are not available until the following day. Imprint cytology of fresh breast core samples allows same-day reporting and patient counselling.
AIM: To determine the accuracy of core imprint cytology when compared with core biopsy diagnosis when used in a breast assessment centre setting.
METHODS: Core imprints (CI) were prepared and reported on all fresh core biopsies (CB) performed at the Sir Charles Gairdner Hospital Breast Centre from May to December 2000. Fresh core samples were placed on a glass microscope slide. Core radiographs were taken for microcalcification lesions (MC). A laboratory technician gently and quickly rolled the cores on the slide with fine forceps. The cores were fixed in formalin, processed and reported next day. The imprint slide was air dried and stained with DiffQuik. CI were reported using four categories: Insufficient, Benign, Indeterminate and Malignant. Counselling and planning for management were possible on the same day in women with malignant diagnoses. Clinicians were advised not to discuss negative or indeterminate CI results with women and to defer to the final CB report.
RESULTS: Cores were performed on 381 lesions. There were 83 carcinomas (38 in MC and 45 in masses) and 56 were called malignant on CI (absolute sensitivity 67.5%; 78.9% for MC and 57.8% for masses). 3 malignancies on CB were negative on CI giving a false negative rate of 3.6%. There were no false positive diagnoses. The predictive value of a benign diagnosis was 95.3%. There were no adverse effects in the histology of CB.
CONCLUSION: CI was an accurate method of providing an immediate diagnosis of malignancy in two thirds of malignancies confirmed on CB.     

Molecular determinants of glycine receptor αβ subunit sensitivities to Zn2+-mediated inhibition

Glycine receptors exhibit a biphasic sensitivity profile in response to Zn²⁺-mediated modulation, with low Zn²⁺ concentrations potentiating (< 10 μ m ), and higher Zn²⁺ concentrations inhibiting submaximal responses to glycine. Here, a substantial 30-fold increase in sensitivity to Zn²⁺-mediated inhibition was apparent for the homomeric glycine receptor (GlyR) α1 subunit compared to either GlyR α2 or α3 subtypes. Swapping the divergent histidine (H107) residue in GlyR α1, which together with the conserved H109 forms part of an intersubunit Zn²⁺-binding site, for the equivalent asparagine residue present in GlyR α2 and α3, reversed this phenotype. Co-expression of heteromeric GlyR α1 or α2 with the ancillary β subunit yielded receptors that maintained their distinctive sensitivities to Zn²⁺ inhibition. However, GlyR α2β heteromers were consistently 2-fold more sensitive to inhibition compared to the GlyR α2 homomer. Comparative studies to elucidate the specific residue in the β subunit responsible for this differential sensitivity revealed instead threonine 133 in the α1 subunit as a new vital component for Zn²⁺-mediated inhibition. Further studies on heteromeric receptors demonstrated that a mutated β subunit could indeed affect Zn²⁺-mediated inhibition but only from one side of the intersubunit Zn²⁺-binding site, equivalent to the GlyR α1 H107 face. This strongly suggests that the α subunit is responsible for Zn²⁺-mediated inhibition and that this is effectively transduced, asymmetrically, from the side of the Zn²⁺-binding site where H109 and T133 are located.     

Therapeutic Effect of Doxycycline in Experimental Subclinical Canine Monocytic Ehrlichiosis: Evaluation of a 6-Week Course

The efficacy of doxycycline treatment (10 mg/kg of body weight every 24 h for 42 days) in eliminating Ehrlichia canis from four subclinically infected dogs was evaluated. One dog remained PCR positive, suggesting that 6 weeks of doxycycline treatment may not be sufficient to clear E. canis parasites from all subclinically infected dogs. Serology (indirect immunofluorescent antibody assay) was shown to be unreliable in assessing recovery from the carrier state, as anti-E. canis antibodies persisted after elimination of the parasite. Our findings suggest that an increase in the platelet count may be an important indicator for dogs that recover from subclinical ehrlichiosis.     

Amplification of Ehrlichial DNA from Dogs 34 Months after Infection with Ehrlichia canis

In order to determine whether dogs in the subclinical phase of canine monocytic ehrlichiosis (CME) are carriers of Ehrlichia canis and to determine the significance of persistent indirect immunofluorescent anti-E. canis antibody titers during this phase, PCR was performed with blood, bone marrow, and splenic aspirates collected 34 months postinoculation from six clinically healthy beagle dogs experimentally infected with E. canis. At least one of the three samples (spleen, bone marrow, and blood) from four of the six dogs was PCR positive. The spleens of all four of these dogs were PCR positive, and the bone marrow and blood of two of the four dogs were PCR positive. Indirect immunofluorescent-antibody titers increased progressively during the first 5 months postinfection, remained high for an additional period of more than 11 months, and declined thereafter, suggesting that the dogs were recovering from the disease. Five of the dogs remained seropositive 34 months postinfection. The data obtained in this study demonstrate for the first time that clinically healthy dogs in the subclinical phase of CME are carriers of the rickettsia. It was shown that dogs can harbor E. canis for years without developing the chronic clinical disease and that dogs can eliminate the parasite and recover from CME without medical treatment. Our findings suggest that the spleen is the organ most likely to harbor E. canis parasites during the subclinical phase and the last organ to accommodate the parasite before elimination. It was concluded that PCR of DNA extracted from splenic aspirates is a reliable method for determining the carrier state of CME.     

Immuno- and genetic therapy in autoimmune diseases

Animal models of autoimmune disease have been developed that mimic some aspects of the pathophysiology of human disease. These models have increased our understanding of possible mechanisms of pathogenesis at the molecular and cellular level and have been important in the testing, development and validation of new immunotherapies. The susceptibility to develop disease in the majority of these models is polygenic as is the case in humans. The exceptions to this rule are gene knock outs and transgenic models of particular genes which, in particular genetic backgrounds, have also contributed to the understanding of single gene function and their possible contribution to pathogenesis. Gene therapy approaches that target immune functions are being developed with encouraging results, despite the polygenic nature of these diseases. Basically this novel immuno-genetic therapy harnesses the knowledge of immunology with the myriad of biotechnological breakthroughs in vector design and delivery. Autoimmune disease is the result of genetic dysregulation which could be controlled by gene therapy. Here we summarize the genetic basis of these human diseases as well as some of the best characterized murine models. We discuss the strategies for their treatment using immuno- and gene therapy.     

Lack of CD 29 (beta1 integrin) and CD 54 (ICAM-1) adhesion molecules in intravascular lymphomatosis

Intravascular Lymphomatosis (IL) is a rare and usually aggressive form of non-Hodgkin's lymphoma characterized by the growth of neoplastic cells within vascular lumina that usually presents with skin or central nervous system (CNS) involvement. The mechanism(s) for the selective intravascular growth of this neoplasm remain(s) unexplained. We now report clinical and immunohistologic data on surgical material from 6 cases of IL; in 4 of 6 cases, autopsies were performed. Our IL cases shared the following features: (1) B-cell lineage; (2) lack of skin involvement at presentation; (3) aggressive behavior; and (4) lack of extravascular lymphomatous masses; in addition, 1 case had an associated gastric low-grade MALT lymphoma. We studied by immunohistochemistry formalin-fixed, paraffin-embedded sections with monoclonal antibodies to molecules known to be involved in lymphocyte and endothelial adhesion phenomena, that is, CD29 (beta1 integrin subunit), CD43 (leukosialin), CD44 (H-CAM), CD54 (ICAM-1), embryonal N-CAM (e-NCAM), and EMA (episialin). In all cases, the surfaces of IL aggregates reacted for CD44 but were consistently negative for CD29; also absent was CD54. Conversely, the integrity of the endothelial cells was underscored by their even reactivity for CD29, CD44, and CD54. Given that CD29 is currently regarded as critical for lymphocyte trafficking in general and for transvascular migration in particular, and CD54 is also involved in transvascular lymphocyte migration, we conclude that their consistent absence in IL may contribute to its intravascular and disseminated distribution pattern. The rather frequent association of IL with various conventional lymphomas is known; yet, one of our cases appears to be the first report of IL associated with a low-grade MALT lymphoma.     

Association study of a SNAP‐25 microsatellite and attention deficit hyperactivity disorder

Several lines of evidence implicate synaptosomal‐associated protein of 25 kDa (SNAP‐25) in the etiology of attention deficit hyperactivity disorder (ADHD). Most notably, the coloboma mouse mutant, considered to be a good animal model of hyperactivity, has a deletion spanning this gene. Introducing a SNAP‐25 transgene into these animals alleviates hyperlocomotion. We have identified a novel microsatellite repeat in SNAP‐25 located between the 5′UTR and the first coding exon, and tested for association with ADHD. Case‐control analyses suggest there may be a role of this polymorphism in ADHD, with one allele over‐represented in controls and another over‐represented in probands. Within‐family tests of linkage and association confirmed these findings. Further work is needed to ascertain the role of SNAP‐25 in ADHD and assess the functional significance of this polymorphism. © 2002 Wiley‐Liss, Inc.     

The intermediate filament complement of the spectrum of nerve sheath neoplasms

The intermediate filament complement of the spectrum of nerve sheath neoplasms including 12 typical benign schwannomas, 1 ancient schwannoma, 2 cellular schwannomas, 6 neurofibromas and 4 malignant schwannomas was investigated by immunofluorescence microscopy, two dimensional electrophoresis, and immunoblot analysis. Studies were performed on freshly frozen tumor tissue samples; a broad spectrum of antibodies against all classes of intermediate filaments was utilized. Samples were also studied by electron microscopy, and immunohistochemically for S-100 protein and desmoplakins. By immunofluorescence microscopy, all nerve sheath neoplasms revealed intense positivity for vimentin throughout the cytoplasm while 2 benign schwannomas displayed co-expression of vimentin and glial filament proteins. Two-dimensional gel electrophoresis and immunoblot analysis confirmed the presence of vimentin and showed that it was the predominant protein in all tumors. Electrophoretic analysis of the 2 benign schwannomas that immunostained for glial filament proteins confirmed the presence of this protein which was shown to comigrate with a known human control sample. Neither immunofluorescence microscopy nor biochemical analyses revealed cytokeratin polypeptides, neurofilament proteins, desmin, or desmoplakin in any of the tumors. We conclude that while vimentin is the predominant intermediate filament expressed by the entire spectrum of nerve sheath neoplasms, at least occasional benign schwannomas are capable of co-expressing glial filament proteins. It remains to be determined whether the subgroup of nerve sheath neoplasms that co-expresses vimentin and glial filament proteins is otherwise distinguishable from their more frequent counterparts that express vimentin exclusively.     

Malgun (clear) cell change in Helicobacter pylori gastritis reflects epithelial genomic damage and repair

Cancers may develop in the background of genomic instability with accumulated mutations. Helicobacter pylori gastritis is characterized by acute foveolitis of the proliferative zone, which is found in any stage of the gastritis as long as the infection persists. Because acute foveolitis targets specifically the proliferative zone of pits, the proliferating epithelial cells are under severe and persistent mutagenic pressure. In H. pylori gastritis, a characteristic morphological change of epithelial cells, the malgun (clear) cell change is frequently present in association with acute foveolitis. Malgun cells have enlarged euchromatic nuclei and abundant cytoplasm. The expression of proliferating cell nuclear antigen and cytokeratin 8 are typically up-regulated in them indicating that they are mitotically and metabolically active. Here, we report evidence for DNA damage and repair in malgun cells. Significant double-strand DNA breaks were shown by the consistent terminal dUTP nick-end labeling in the nuclei of malgun cells. Proteins related to DNA damage and repair, such as Ku, poly(ADP-ribosyl) polymerase, OGG1, and MSH2 were selectively up-regulated in malgun cells. Inducible nitric oxide synthase was also up-regulated. There were occasional bcl2- and p53-expressing cells suggesting that further steps of carcinogenesis took place at the single cell level. Our results suggest that the malgun cell change represents a characteristic morphological sign of cellular genomic damage and repair, and may be implicated in an early stage of carcinogenesis. It is suggested that acute foveolitis of the proliferative zone is a major pathogenetic step of gastric carcinogenesis in H. pylori gastritis.