bluuebunny/crossref_metadata_2025 huggingface dataset:
A synergistic interaction between Bacillus Cahnette‐Guérin (BCG) and NADPH oxidase stimulants on the production of reactive oxygen metabolites by human mononuclear phagocytes
Authors: Nyberg PETER W. , Nordman SUSANN A. S. , Linko LINNÉA
Published: August 2009

Bacillus Calmette‐Guerin (BCG) was added simultaneously with known NADPH oxidase stimulants to suspensions of human mononuclear leukocytes, and the subsequent production of reactive oxygen metabolites (ROMs) was studied by luminol‐dependent chemiluminescence. BCG significantly amplified the ROM responses induced by zymosan, phorbol myristate acetate (PMA), and quartz, but not by concanavalin A and asbestos fibers. The stimulatory effect occurred rapidly when BCG was added to cells already phagocytosing zymosan, and vanished rapidly when extracellular BCG was removed from adherent monocyte cultures by washing prior to the addition of zymosan. The stimulatory effect of BCG could not be reproduced with recombinant interferon‐γ, tuberculin PPD, muramyl dipeptide, nor with the apathogenic Mycobacterium tuberculosis strain RV37. BCG and zymosan or PMA that had been incubated together prior to addition to the mononuclear cell suspensions caused ROM production with faster kinetics than if the reagents were added separately without preincubation. In conclusion, the synergy between BCG and some of the NADPH oxidase stimulants seems to be due to an interaction between BCG and the NADPH oxidase stimulants rather than to an interaction between BCG and the ROM‐producing cells. Such interactions between mycobacteria and NADPH oxidase stimulants may be of importance as a factor affecting the individual susceptibility to tissue damage in tuberculosis, for example in silicotuberculosis.
Impact of the agar medium and disc type on disc diffusion susceptibility testing against teicoplanin and vancomycin
Authors: Jensen KJELD TRUBERG , SchØNheyder HENRIK , Gottschau ADAM , Thomsen VILLY FRØLUND
Published: August 2009

Susceptibility to teicoplanin and vancomycin was assessed by three disc types: two commercially available discs (NeoSensitabs and PDM disc (30 μg)) and one locally prepared 30 ug disc (SS disc) on four different medium types: Mueller‐Hinton agar (MH medium), MH medium and PDM agar II supplemented with 5% horse blood (HMB medium and PDM medium, respectively), and Danish blood agar (DBA medium). Two previously studied groups of Gram‐positive bacteria were tested: group B (N = 75) comprised miscellaneous cocci, and group C (N = 59) mostly rods. With NeoSensitabs, mean zone diameters were larger than with PDM and SS discs on all medium types, and mean zone diameters were larger on DBA medium than on MHB and PDM medium with all disc types. The impact of the medium type on the zone diameter was evaluated for 121 strains growing on MHB medium, PDM medium, and DBA medium. Bacterial groups B and C each divided into three MIC groups were analysed separately. We compared mean zone diameters for each specific group with the average zone diameter, i.e. the mean value for all zone diameters obtained. The smallest deviations from the average zone diameters were observed on PDM medium for both teicoplanin and vancomycin. Thirty‐seven percent of strains failed to grow on MH medium, but supplementation of MH medium with horse blood significantly reduced the zone diameter for group B strains both for teicoplanin and vancomycin. Poor predictability of MIC from the zone diameter was found especially for strains with MICs 1 μg/ml. The medium type hardly affected the results of regression analysis. In contrast, the medium type markedly affected the results of error‐rate bounded analysis. No errors were recorded with the SS disc on MHB medium for either teicoplanin or vancomycin, but no strains with MICs of vancomycin within the intermediate group could be correctly classified on DBA medium.
Two techniques for measuring invasion in solid tumors
Authors: Laroye GUY J. , Panzarella TONY
Published: August 2009

Two methods are described which measure invasion in solid tumors: 1)average tumor extension, i.e. the distance between the edge of the primary tumor mass and the furthest extension of tumor nests in the adjacent stroma, and 2)total surface of tumor exposed to stroma, obtained by multiplying the volume within 1 mm of the outer margin of the primary tumor by the stereologically derivedratio of tumor surface to its volume.These techniques are applied to a retrospective review of 73 consecutive cases of female breast cancer with known lymph node status and without distant metastases at the time of the original diagnosis. The study explores how these parameters discriminate 10‐year survivors from non‐survivors and compares them with other known significant parameters of tumor aggressiveness, i.e. axillary lymph node status, mitotic count, tumor diameter, tumor/desmoplasia ratio, necrosis/tumor ratio, gland space/tumor ratio, nuclear volume (= nuclear grade), and age at diagnosis. This study concludes: 1) Each parameter of invasion, adjusted in the analysis for lymph node status and mitotic count, emerges as statistically significant with low invasion in the primary tumor conferring a better prognosis than high invasion. 2) The associations low mitoses/low invasion, high mitoses/low invasion and low mitoses/high invasion tend to indicate a favorable 10‐year survival, while the combination high mitoses/high invasion results in poor survival. 3) There was a greater percentage of high invasion cases in the LN > 0 subgroup compared to the LN 0 subgroup, suggesting that high invasion in the primary tumor predisposes to metastases, but this trend did not always reach statistical significance. 4) The proposed methodology offers a practical approach for measuring invasion in solid tumors.
Determination of microbial chemical markers by gas chromatography‐mass spectrometry ‐ potential for diagnosis and studies on metabolism in situ
Authors: Larsson LENNART
Published: August 2009

Different strategies for the application of gas chromatography (GC) and gas chromatography‐mass spectrometry (GC‐MS) in medical microbiology research are discussed. GC can be used to determine specific microbial monomeric constituents and metabolites, so‐called chemical markers, in cultures of microorganisms; in particular, analysis of cellular fatty acids has proven useful for species characterization and identification. GC‐MS can be applied to study chemical markers directly in complex environmental samples, as exemplified by the analysis of airborne organic material as regards muramic acid (marker of peptidoglycan), 3‐hydroxy acids (endotoxins), and ergosterol (fungal biomass). This methodological approach represents an alternative to various biological assays for characterization of airborne microbial structures, and forms a firm basis for correlating inhalation of such structures and development of symptoms. Direct GC‐MS analysis of clinical samples provides possibilities for diagnosis (here exemplified by chiral separation of urine D‐ and L‐arabinitol in disseminated candidiasis) and insight into microbial metabolism in the infected host (exemplified by observed indications of mycobacterial build‐up of mycolic acids in vivo), with implications for drug development. Continued developments in MS technology will allow rapid advances to be made in GC‐MS research in microbiology.
Synovial sarcoma
Authors: JØRgensen LARS J. , Lyon HANS , Myhre‐Jensen OLAF , Nordentoft AXEL , Sneppen OTTO
Published: August 2009

Twenty‐five synovial sarcomas were studied with a battery of antibodies directed against keratin and epithelial membrane antigen (EMA). The keratin antibody MNF 116 showed reactivity in 24 tumors. In addition, 22 tumors showed reactivity with the antibody Keratin Wide Spectrum, 20 with the antibody Keratin 56, 64, and 19 with CAM 5.2. Seventeen tumors showed reactivity with EMA. The keratin and EMA reactivity was present in cells lining obvious cleft‐like structures in biphasic tumors. In the spindle cell areas of both biphasic and monophasic fibrous tumors, we found clusters of a few reacting cells apparently located around small clefts. In the synovioblastic tumors, clusters of plump tumor cells reactive for both the keratins and EMA were present. In conclusion, we found that proper identification of epithelial differentiation in synovial sarcomas is facilitated by an immunohistochemical application of anti‐epithelial antibodies. In most tumors, there was immunoreactivity for the same type of keratins as are normally identified in simple epithelia (the antibody CAM 5.2), but also for those found in stratified squamous epithelia (the antibody Keratin 56, 64). The results indicate that screening for epithelial features on paraffin sections in the various types of synovial sarcoma, even the poorly differentiated synovioblastic tumors, is improved if epithelial antibodies with a broad spectrum of reactivity are chosen.
The antibacterial activity of a siderophore
Authors: Hartzen SUSANNE HARTVIG , Frimodt‐MØLler NIELS , Thomsen VILLY FRØLUND
Published: August 2009

The in vitro activity of deferoxamine (DFO) both per se and in combination with the reductant ascorbic acid (AA) was determined against 10 E. coli strains, 5 P. mirabilis strains, and 10 coagulase‐negative staphylococci. In terms of interaction, the influence of DFO on the activities of cephalothin and gentamicin was furthermore investigated against the same panel of strains employing a macrobroth dilution technique and killing‐curve kinetics. The MICs of cephalothin and gentamicin were lowered for one half of the strains. Moreover, DFO prolonged the generation times of logarithmic growth phase considerably, especially when the reductant AA was present. The interactions between DFO or DFO + A A and subinhibitory concentrations of antibiotics were established by the application of growth constants, and resulted in synergy for 15 out of 25 strains with cephalothin and 9 out of 25 strains with gentamicin.
Antimicrobial susceotibilirv of bacterial isolates in South Sweden including a 13‐year follow‐up study of some respiratory tract pathogens
Authors: Forsgren ARNE , Walder MATS
Published: August 2009

The antibiotic susceptibility of consecutive isolates of the upper respiratory tract pathogens Streptococcus pyogenes, Streptococcus pneumoniae, Haemophilus influenzae, Branhamella catarrhalis, and Staphylococcus aureus, (100 strains of each species collected each year during March through April 1985, 1988 and 1992) to penicillin V, amoxycillin, cefaclor, cefuroxime, doxycycline, erythromycin, and co‐trimoxazole was investigated by MIC determination on PDM and PDM II agar. The MICs of the upper respiratory isolates from 1992 supplemented with 100 isolates each of Escherichia coli, Klebsiella spp., Enterobacter cloacae, Proteus mirabilis and Staphylococcus saprophytics collected during 1992 were determined against the above antibiotics plus cefadroxil, cefpodoxime, roxithromycin, ciprofloxacin, ofloxacin, and BAY Y 3118. Beta‐lactamase production was found in 10% of H. influenzae and 80–90% of S. aureus and B. catarrhalis in 1992. Among H. influenzae isolates, non‐beta‐lactamase‐induced resistance to all beta‐lactam antibiotics was first detected in 1988 and amounted to 3% of isolates in 1992. Decreased susceptibility of S. preumoniae to penicillin (>0.12 mg/1), co‐trimoxazole 32 mg/1, doxycycline (2 ml/1) and erythromycin (1 mg/1) was detected in 11%, 7%, and 8%, respectively, in 1992, which is significantly higher than in previous years at the same laboratory. Decreased susceptibility of S. pyogenes to doxycycline and erythromycin was detected in 11% and 9% in 1992. The two most recently developed antibiotics, cefpodoxime and BAY Y 3118, showed high antibacterial activity. The study emphasizes the need to screen for resistance mechanisms such as beta‐lactamase production and lowered penicillin affinity.
Aggressive angiomyxoma of the vulva: An unusual, deceptive and recurrence‐prone tumour with evidence of estrogen receptor expression
Authors: Havel G. , Burian P. , Kohrtz M. , Mark J.
Published: August 2009

Positive staining for estrogen receptors is reported in a new case of aggressive angiomyxoma of the vulva. Clinical, light‐ and electron microscopic features as well as immunohistochemical reactions in this case are compared with the 28 earlier published cases, and the main differential diagnosis, angiomyofibroblastoma of the vulva, is discussed.
Rules for peptide binding to MHC class II molecules
Authors: Sinigaglia FRANCESCO , Hammer JUERGEN
Published: August 2009

CD4+ T lymphocytes recognize peptide fragments of antigens that lie in the antigen binding pocket of class II major histocompatibility complex (MHC) molecules expressed on antigen‐presenting cells. Specificity of T cells is determined by structural features of both the MHC molecule and antigenic peptide. MHC class II amino acid sequences are highly polymorphic within a population, and correlate with individual differences in response to infectious agents, vaccines, tumour antigens, and autoantigens. In the last few years several important breakthroughs and technological advances have made it possible to clarify the role of polymorphism and the molecular events in peptide interaction with major histocompatibility complex (MHC) class II proteins. The X‐ray structural analysis of a MHC class II molecule together with the use of peptide libraries has permitted determination of the structural features of the complex between peptide antigen fragments and MHC class II proteins. The purpose of this article is to review our own studies and those of others on the requirements for peptide‐class II molecule interaction and discuss possible implications for active immune intervention.
Leishmania major surface protease Gp63 interferes with the function of human monocytes and neutrophils in vitro
Authors: SØRensen ANNE L. , Hey ADAM S. , Kharazmi ARSALAN
Published: August 2009

In the present study the effect of Leishmania major surface protease Gp63 on the chemotaxis and oxidative burst response of human peripheral blood monocytes and neutrophils was investigated. It was shown that prior incubation of cells with Gp63 inhibited chemotaxis of neutrophils but not monocytes towards the chemotactic peptide f‐met‐leu‐phe. On the other hand, chemotaxis of both neutrophils and monocytes towards zymosan‐activated serum containing C5a was inhibited by Gp63. Monocyte and neutrophil chemiluminescence response to opsonized zymosan was reduced by preincubation of the cells with Gp63 in a concentration‐dependent manner. Notably, monocytes were inhibited to a much greater degree than neutrophils by a given concentration of Gp63, and they were also inhibited at much lower concentrations of the protease. The inhibitory effect of Gp63 on chemotaxis and chemiluminescence was completely abolished by heat inactivation of the protease at 70 °C for 15 min. Neither neutrophil nor monocyte chemiluminescence was inhibited by Gp63 when cells were stimulated with PMA. Our data suggest that the major surface protease Gp63 might play an important role in the initial stages of Leishmania/macrophage interactions and the intracellular survival of the parasite.
Identification of coagulase‐negative staphylococci and typing of Staphylococcus epidermidis by a 4 h micromethod
Authors: JarlØV JENS OTTO , Nissen BENT , Rosdahl VIBEKE THAMDRUP , Espersen FRANK
Published: August 2009

A new 4 h micromethod (Minibact‐S) for identification of coagulase‐negative staphylococci (CNS) important in human medicine, Staphylococcus epidermidis, Staphylococcus hominis. Staphylococcus haemolyticus and Staphylococcus saprophytics, has been investigated. The reproducibility for species identification was 100% and the reproducibility for the single reactions was 95%. When compared to a modified Kloos & Schleifer conventional identification method, discrepant identification was obtained with the Minibact‐S in only 8 of 382 clinical and reference strains. The system has been used for typing and biotyping of 201 S. epidermidis strains, and gave a subdivision in 69 different types; 4 related types accounted for 91 (45%) of the isolates, whereas 44 types were represented by only 1 isolate. The discriminatory index was 0.82. In a comparison with other typing methods performed on 81 isolates from 15 patients Minibact‐S gave about the same number of types as antibiograms. but in 20% of the patients extra types were obtained with Minibact‐S alone compared to antibiogram alone and vice versa. The Minibact‐S is of some value in the routine clinical microbiology laboratory as a first screening method for identification of coagulase‐negative staphylococci and typing of S. epidermidis in combination with the antibiogram.
Immunoelectron microscopy of the receptor for urokinase plasminogen activator and cathepsin D in the human breast cancer cell line MDA‐MB‐231
Authors: Bastholm LONE , Elling FOLMER , BrÜNner NILS , Nielsen MORTEN H.
Published: August 2009

Receptors for urokinase‐type plasminogen activator (uPAR) are present on the surface of many cell types and appear to be the key determinant controlling extracellular proteolysis catalyzed by the urokinase‐type plasminogen activator (uPA). Receptor‐bound uPA may be inhibited by the specific inhibitors PAI‐1 and PAI‐2, and the complex thus formed may subsequently be internalized and degraded in lysosomes. Biochemical evidence has recently indicated that also uPAR is internalized with the uPA/uPAI complex. We report here the subcellular localization of uPAR and cathepsin D in the MDA‐MB‐231 human breast cancer cell line studied by immuno‐electron microscopy of ultrathin cryosections using single or double immunostaining techniques. Cell surface uPAR was preferentially localized at cell‐cell junctions; cytoplasmic uPAR was inside large vesicles of different morphology and in flat Golgi saccules. A number of vesicles also contained cathepsin D. The uPAR was exclusively membrane‐bound at the cell surface and in cytoplasmic vesicles without cathepsin D. In lysosomal vesicles with both cathepsin D and u‐PAR, uPAR was probably degraded as it was observed in the luminal contents.
Prevalence of antibodies to Borrelia burgdorferi in Danish deer
Authors: Webster PIA , Frandsen FLEMMING
Published: August 2009

To estimate the prevalence of antibodies to Borrelia burgdorferi in the most common species of Danish deer, blood samples were collected from roe deer (Capreolus capreolus), fallow deer (Dama dama), and red deer (Cervus elaphus). A total of 156 blood samples were collected primarily from hunts in three areas of Denmark from November 1990 to December 1991. Presence of deer IgG antibodies to the Borrelia burgdorferi strain DK ECM 1 were shown by an indirect immunofluorescence assay at serum titre 1:64. Antibodies to spirochaetes were detected in 52% of roe deer, 38% of fallow deer, and 27% of red deer. There were significant differences between the number of seropositive animals in the three areas of Denmark examined, but no differences could be shown for age, sex or season. The high antibody prevalence indicates that deer are exposed to tickborne Borrelia burgdorferi throughout Denmark.
The herpes simplex virus type 1 particle: structure and molecular functions
Authors: Haarr LARS , Skulstad SIRI
Published: August 2009

This review is a summary of our present knowledge with respect to the structure of the virion of herpes simplex virus type 1. The virion consists of a capsid into which the DNA is packaged, a tegument and an external envelope. The protein compositions of the structures outside the genome are described as well as the functions of individual proteins. Seven capsid proteins are identified, and two of them are mainly present in precursors of mature DNA‐containing capsids. The protein components of the 150 hexamers and 12 pentamers in the icosahedral capsid are known. These capsomers all have a central channel and are connected by Y‐shaped triplexes. In contrast to the capsid, the tegument has a less defined structure in which 11 proteins have been identified so far. Most of them are phosphorylated. Eleven virus‐encoded glycoproteins are present in the envelope, and there may be a few more membrane proteins not yet identified. Functions of these glycoproteins include attachment to and penetration of the cellular membrane. The structural proteins, their functions, coding genes and localizations are listed in table form.
Long‐term cyclosporin A nephrotoxicity in the rat
Authors: Starklint HENRIK , Hansen HANNE V. , Kemp MICHAEL , Leyssac PAUL PETER , Kemp EJVIND , Dieperink HANS
Published: August 2009

Cyclosporin A (CyA) nephrotoxicity was examined in Spraque‐Dawley rats given CyA (12.5 (n = 45) or 25 (n = 45) mg/kg/day perorally for 16 weeks. Control rats (n = 45) received CyA vehicle. All rats were given either isradipine (ISRA) 1 or 5 mg/kg/day orally, or isradipine vehicle. Fifteen rats died from interstitial pneumonia caused by Staphylococcus xylosus. A predefined morphological CyA nephrotoxicity scoring system, based on semiquantitative scores for basophilic tubules and for interstitial fibrosis, performed on hematoxylin‐eosin‐stained tissue, yielded mean scores for basophilic tubules of 0.2 (range 0–1) in controls, 1.4 (range 0–3) in rats given CyA 12.5 mg/kg/day (p< 0.001), and 1.7 (range 0–3) in CyA 25 mg/kg/day rats (p< 0.001 as compared to controls). Rats given CyA were grouped according to their score for interstitial fibrosis: 0.2 (range 0–1) in CyA 12.5 mg/kg/ day and 1.7 (range 0–3) in CyA 25 mg/kg/day rats (p<0.001). When scores for basophilic tubules and interstitial fibrosis were pooled, none of the control rats had a score above 1, while 47% of the low‐dose and 95% of the high‐dose rats scored above 1. Thus, this CyA nephrotoxicity scoring system provided an easy, efficacious, and reproducible identification of rats with morphological CyA nephrotoxicity, and may be of clinical interest in the assessment of CyA nephrotoxicity. Kidney tissue from rats not treated with isradipine was further investigated with periodic acid‐Schiff (PAS) with and without diastase treatment, and with Sirius Red. The latter confirmed the increase in connective tissue following tubular atrophy in CyA‐treated rats. PAS reaction disclosed diastase‐resistant positiv‐ity in the glomerular arterioles (score in controls: mean 0.4, range 0–1, in CyA 12.5 mg/kg/day mean 2.2, range 1–3, p<0.001 as compared to controls; in CyA 25 mg/kg/day mean 1.1, range 0–2, p< 0.005 as compared to controls, p<0.05 as compared to CyA 12.5 mg/kg/day). Furthermore, the straight part of the distal tubules of rats given the highest CyA dose contained considerable amounts of glycogen. The significance of this finding is unknown. Renal functional studies confirmed previous results since CyA decreased inulin clearance (Cin) from 1.2±0.5 to 0.8 + 0.3 ml/min/g kidney weight (kW) (p<0.05), and lithium clearance (CLi) was reduced from 263±113 to 119 ± 61 μl/min/gKW (p< 0.001). Isradipine had no significant effect.
Immunopathological patterns in the rectal mucosa of patients with shigellosis: expression of HLA‐DR antigens and T‐lymphocyte subsets
Authors: Raqib RUBHANA , Reinholt FINN P. , Bardhan PRADEEP KUMAR , KÄRnell ANDERS , Lindberg ALF A.
Published: August 2009

Expression of HLA‐DR antigens and infiltration of T‐lymphocyte subsets (CD4, CD8), cell activation marker (CD25), B cells (CD20), macrophages (CD68 and Ber‐Mac 3) and natural killer cells (CD56) in the rectal mucosa of patients with bacillary dysentery and in healthy controls were studied in an effort to interpret the immunopathological changes taking place in the rectal mucosa during the acute phase of shigellosis. The epithelium of the rectal mucosa from 21 of 32 patients was HLA‐DR+. Conventional histology showed acute inflammation in 16 of these patients, chronic inflammation in 3, and in 2 histology was normal. In 7 of 20 controls the epithelium was HLA‐DR+; 4 of these 7 were found to suffer from chronic inflammation, whilst in 3 the mucosa was normal. The number of HLA‐DR+ intraepithelial lymphocytes in biopsies from patients with Shigella infection was significantly higher (p = 0.005) than in controls. The infiltration of CD8+ cells in the surface epithelium and in the lamina propria, and of CD4+ cells in the lamina propria alone, was significantly higher (p< 0.001) in patients than in controls. The results demonstrate that infiltration of T cells with suppressor/cytotoxic or helper/inducer phenotype in the epithelium and in the lamina propria in Shigella‐infected patients may be related to the induction of HLA‐DR expression in non‐lymphoid cells during acute Shigella infection.
Microwave processing for immunohistochemical demonstration of proliferating cell nuclear antigen (PCNA) in formalin‐fixed and paraffin‐embedded tissue
Authors: Haerslev TORBEN , Jacobsen GRETE KRAG
Published: August 2009

The effect of microwave irradiation on the immunoreactivity of proliferating cell nuclear antigen (PCNA) in five primary breast carcinomas, a tonsil with hyperplasia, and a seminoma of the testis was evaluated in formalin‐fixed and paraffin‐embedded sections using the monoclonal antibody PC‐10. The tissues were fixed in 4% buffered formaldehyde for 6, 24, 28, 72 h, and 1 week. Before incubation with the primary antibody, sections were microwaved for × 2 5 min in distilled water, in citrate buffer, or they were processed routinely without microwave irradiation. In sections microwaved in distilled water the immunoreactivity was found to be independent of the length of fixation. PCNA immunoreactivity in sections microwaved twice in citrate buffer was not satisfactory. The PCNA immunoreactivity in sections which were routinely processed without microwave irradiation was markedly decreased after 48 h of fixation, and was nearly absent in sections fixed for up to 1 week. In sections processed without microwave irradiation the optimal dilution of the primary antibody was 1:10, whereas in sections microwaved twice in distilled water the optimal dilution was 1:600. The morphology of the tissue was well preserved after microwave processing. We therefore recommend microwave processing in distilled water for × 2 5 min for demonstration of PCNA in formalin‐fixed and paraffin‐embedded sections using the monoclonal antibody PC‐10, since this technique yields optimal staining reactions independent of the length of fixation.
Evaluation of methods for the detection of nasal carriage of Staphylococcus aureus
Authors: Eriksen N. H. RIEWERTS , Espersen F. , Rosdahl V. THAMDRUP , Jensen K.
Published: August 2009

In the present study we investigate the optimal methodology for determination of the nasal carriage rate of Staphylococcus aureus. Tests were performed on 91 healthy laboratory staff. The reproducibility of different sampling, transportation, storage and culture methods was examined. We compared sterile dry cotton wool swabs with sterile dry cotton wool swabs impregnated with charcoal and 5% blood agar plates with mannitol salt agar plates after different incubation periods. Finally, we investigated the detection rate for S. aureus following direct plating compared to storage in Stuart's transport medium for 7 days. There were no differences in isolation rates from the right or left nostril using either cotton or charcoal swabs. Charcoal swabs gave an increased isolation rate as compared to cotton swabs, and incubation in broth enrichment medium containing 6.5% NaCl also increased the isolation rate. Storage in Stuart's transport medium for 7 days gave an increase in isolation rate as compared to direct plating on blood agar. With mannitol salt agar plates the increase in isolation rate when incubation was performed for from 2 to 4, 2 to 7, and 4 to 7 days was 5.9%, 16.7%, and 11.5%, respectively. For the detection of S. aureus nasal carriers we find the use of charcoal swabs and Stuart's transport medium combined with cultivation on mannitol salt agar for 7 days to be the optimal method.
Folate receptor of human mammary adenocarcinoma
Authors: Holm JAN , Hansen STEEN I. , HØIer‐Madsen MIMI , SØNdergaard KNUD , Bzorek MICHAEL
Published: August 2009

Binding of 3H‐folate to human mammary tumor homogenate was of a high‐affinity type (K= 1010 M−1) and displayed apparent positive cooperativity. Radioligand dissociation was slow at pH 7.4, but rapid at pH 3.5. As compared to methotrexate, 5‐formyltetrahydrofolate acted as a strong inhibitor of radioligand binding. Gel chromatography of radioligand‐labeled homogenate of tumor tissue revealed three peaks: a small 110 kDa peak and two major peaks of folate‐binding activity (Mr∼ 25 kDa and Mr∼ 100 kDa). Mammary tumor tissue showed immunostaining with rabbit antibodies against human milk folate binder. A parallel elevation in the concentrations of folate‐binding protein and triglyceride in tumor tissue as compared to normal tissue adjacent to the tumor was compatible with the localization of folate‐binding protein in the triglyceride‐rich fraction of mammary gland homogenate.
Antisecretory factor enhances in vivo internalization of cholera toxin and of horseradish peroxidase into rat enterocytes
Authors: Lange STEFAN , Jennische EVA , LÖNnroth IVAR
Published: August 2009

The in vivo effect of antisecretory factor (ASF, derived from pig plasma) on the ability of cholera toxin (CT) and of horseradish peroxidase (HRP) to bind to and penetrate into epithelial cells of the rat small intestine was evaluated in the absence of anesthetics. The potencey of intravenously administrated ASF was demonstrated by some 70% inhibition of CT‐induced secretion in ligated small intestinal loops. Using immunohistochemical methods for visualization, we found ASF to enhance internalization of both CT and HRP after 30 to 60 min of challenge, without interfering with the initial binding to the enterocyte brush border region. The internalization process started in the upper 2/3 of the villus region. After 5 h, no CT or HRP could be seen bound to the enterocytes. The results suggest that ASF might enhance small intestinal absorption.
Flow cytometric measurement of the production of reactive oxygen intermediates in activated rat mast cells
Authors: Tsinkalovsky OLEG R. , Laerum OLE DIDRIK
Published: August 2009

The production of reactive oxygen intermediates (ROI) in compound 48/80‐ or calcium ionophore A 23187‐ activated pleural or peritoneal mast cells was monitored using flow cytometry and the fluorescence indicator dihydrorhodamine 123 (a derivate of rhodamine 123). Mast cell degranulation and ROI production were estimated by flow cytometric fluorescence and light‐scatter analysis. In addition, activation was monitored by spectrofluorimetric measurement of histamine secretion from the cells. We used flow cytometric measurement of narrow and wide angle scatter and fluorescence intensity to distinguish between degranulated and resting mast cells during activation and to monitor the two populations separately. Both stimuli induced a dose‐dependent elevation of ROI production in mast cells which was accompanied by cell degranulation and histamine secretion. These alterations were completely abolished by preincubation of the mast cells with diethyldithiocarbamate (DTC). Using low concentrations of DTC partial decrease of degranulation and histamine secretion was observed while ROI production was blocked. D‐mannitol increased ROI production in stimulated mast cells without a marked effect on degranulation or histamine secretion. The data suggest that mast cell degranulation (histamine secretion) and ROI production are independent processes.
Immunity to experimental Salmonella typhimurium infections in rats
Authors: THYGESEN PETER , JØRGENSEN TINA , CHRISTENSEN H. BJØRN , BRANDT LISE , RYGAARD JØRGEN , HOUGEN HANS PETTER , JENSEN ELSEBETH T.
Published: August 2009

The protective effect of primed CD4+ T lymphocytes against a lethal dose of 108 viable Salmonella typhimurium was studied in Lewis rats. Primed CD4+ T lymphocytes were obtained by inoculating Lewis rats with a non‐lethal dose of 106 viable S. typhimurium. Four weeks after the infection, spleen CD4+ T lymphocytes were separated using magnetic microspheres coated with an antibody against the CD4 molecule (W3/25). Subsequent sorting into activated and non‐activated subpopulations using the p55 α‐chain of the interleukin‐2 receptor (CD25) as an activation marker was performed by a fluorescence‐activated cell sorter. Untreated Lewis rats were injected with 104 different primed CD4+ T‐cell populations 24 h prior to the lethal dose of 108 viable S. typhimurium. Blood samples were drawn from the orbital plexus 1, 2, 3, and 4 weeks after the infection, and analysed for specific IgM and IgG antibodies. Cell sorting revealed that 2/3 of the primed CD4+ T lymphocytes expressed high levels of CD25. Cell transfer revealed that both CD25high and CD25low expression populations could induce immunity against a lethal dose of S. typhimurium, whilst antibody analysis revealed that antibody levels were not correlated with protection against S. typhimurium infections, although it showed that a higher and more persistent level of specific IgG antibodies was produced in animals receiving the CD4+CD25high fraction. It is concluded that 104 primed CD4+ T lymphocytes can induce immunity in animals challenged with a lethal dose of S. typhimurium and that antibodies do not seem to be correlated with the immunity induced. The CD4+CD25high fraction was, however, associated with a higher and more persistent level of specific IgG antibodies.
Epstein‐Barr virus and Hodgkin's disease: A comparative immunological, in situ hybridization, and polymerase chain reaction study
Authors: LAURITZEN ANNE F. , HØRDING ULLA , NIELSEN HENRIK W.
Published: August 2009

During recent years numerous studies have demonstrated the presence of Epstein‐Barr virus (EBV) in tissues affected by Hodgkin's disease (HD). The percentage of cases with evidence of EBV infection has varied among the different studies, a positive result being highly dependent on the sensitivity of the method employed. In this study three different methods of detecting EBV in 48 cases of ‘classical’ HD (33 cases of nodular sclerosis and 15 cases of mixed cellularity) were compared: Immunohistochemistry (IH) for detection of latent membrane protein‐1 (LMP‐1), in situ hybridization (ISH) for detection of Epstein‐Barr virus early RNAs (EBER 1 and 2), and polymerase chain reaction (PCR) for detection of a reiterated 110 base‐pair EBV genomic sequence of the BamHI region. In 14 cases (29%) Hodgkin's (H) and Reed‐Sternberg (RS) cells were positive for LMP‐1 using IH, and in 21 cases (44%) positive signals were seen in H‐RS cells with EBER 1 and 2 probes using ISH. A few EBER‐positive non‐malignant lymphocytes were seen in 17 cases. Thirty‐two cases (71%) were EBV‐positive by PCR. It is concluded that the PCR technique is the most sensitive method for detecting EBV in HD. However, this method cannot provide information about the cellular localization of EBV. ISH with EBER 1 and 2 probes is superior to immunohistochemical detection of LMP‐1 with regard to sensitivity. The advantage that the latter two methods have over the PCR techniques is that it is possible to analyse whether the EBV infection occurs in the H‐RS cells or in the admixed non‐malignant cell population. Furthermore, this study supports the observation that EBV is associated with a considerable number of HD cases.
Serum antibodies from MS patients do not recognize HTLV‐I, HIV‐1, HIV‐2 and SIV
Authors: BROKSTAD KARL ALBERT , KALLAND KARL HENNING , HAAHEIM LARS , PAGE MARK , NYLAND HARALD
Published: August 2009

A retroviral aetiology has been proposed for multiple sclerosis (MS). Although there is as yet no definitive evidence of viral involvement, there have been preliminary reports of antiretroviral antibody detection in sera from MS patients. Such sera have, for example, been found to react with HTLV‐I. We here describe investigations involving various immunological techniques which attempt to confirm the virus‐specific nature of these antibodies against a range of human and macaque retroviruses. Sera from 25 MS patients, 25 patients with non‐associated neurological diseases and 16 patients with non‐neurological conditions were tested by immunoblotting methods using lysates of HIV‐1‐, HIV‐2‐, HTLV‐I‐ and SIV‐infected cells as antigens. None of the sera reacted against any of these retroviral antigens but each serum demonstrated a distinctive and reproducible reaction pattern against cellular components of the cells in which the viruses were propagated. Further examination of the sera was carried out by ELISA using synthetic oligopeptides covering the HIV‐1 Gag p24 protein as antigens. None of the sera reacted with the peptides. Our results suggest that in some MS patients the repeated seropositivity to HTLV‐I may be due to the reaction with host cell proteins.
Detection of gastrin mRNA in paraffin‐embedded samples of normal antral mucosae using polymerase chain reaction technique
Authors: BIAGINI PHILIPPE , MONGES GENEVIÈVE , CANTALOUBE JEAN‐FRANÇOIS , PARRIAUX DANIELLE , HASSOUN JACQUES , CHICHEPORTICHE COLETTE
Published: August 2009

Gastrin, a peptide hormone produced by the G cells of the gastric antrum, plays a major role in the regulation of digestive mucosal growth. Although some light has been shed on the peptidic aspects of this hormone's mode of action and the co‐regulatory activity in which it is involved along with the other gastrointestinal hormones, little is known at present about the modes of expression of its mRNA at the tissue level. A few attempts have been made so far to detect the transcript, mostly using molecular hybridization techniques. Here it was proposed to detect gastrin mRNA using a RT‐PCR technique on a series of paraffin‐embedded samples of normal human antrum processed with various fixatives commonly used in histology. The transcript was detectable in all the 7‐μm sections of the samples treated with either formalin or Carnoy's solution, whereas Bouin's solution, which is also used as a fixative in histology, was found to have inhibitory effects on the method described here.
Epithelial antigens in carcinomas of the ovaries. Relation to histological classification
Authors: JØRGENSEN ANNEMETTE , NIELSEN KARSTEN
Published: August 2009

In an attempt to assess and improve the histological classification of ovarian tumors the value of immunohistochemical techniques has been examined in 50 ovarian tumors. A panel of six immuno‐histochemical markers (two cytokeratins, EP4, EMA, CEA, and vimentin) seems to have no additional value in differential diagnosis and typing of ovarian tumors.
Mucosal mast cell proliferation following normal and heterotopic infections of the nematode Nippostrongylus brasiliensis in rats
Authors: ARIZONO NAOKI , YAMADA MINORU , TEGOSHI TATSUYA , OKADA MANABU , UCHIKAWA RYUICHI , MATSUDA SHINJI
Published: August 2009

Infections of intestinal nematodes induce the T cell‐dependent proliferation of intestinal mucosal mast cells (MMC). To examine whether nematode‐induced MMC proliferation is affected by the site of infestation, adult‐stage nematode Nippostrongylus brasiliensis (NB) was transplanted into the normal infection site, the duodenum, or into heterotopic sites, the peritoneal cavity (i.p.) or subcutaneous tissue (s.c.), of rats. Two weeks after duodenal inoculation, MMC numbers in the small intestine had increased 6.5‐fold. In contrast, i.p. and S.C. inoculation induced only slight increases of intestinal MMC. After i.p. inoculation, worm granulomas developed in the connective tissues adhering to stomach and duodenum, and large numbers of mast cells appeared around the granulomas. The majority of the latter mast cells showed histochemical features similar to MMC: they were formalin sensitive, berberine sulfate‐, alcian blue+isafranine‐, and rat mast cell protease (RMCP) I I +. After S.C. inoculation, worm granulomas developed at the inoculation site, but the number of mast cells around the granulomas was not significantly increased. These results suggest that intense proliferation of MMC or MMC‐like cells is induced only by the infections on mucosa or in mucosa‐associated tissues.
c‐erbB‐2 oncoprotein is not an independent prognostic parameter in primary breast carcinoma. An immunohistochemical study
Authors: HAERSLEV TORBEN , JACOBSEN GRETE KRAG
Published: August 2009

The prognostic significance of c‐erbB‐2 oncoprotein expression was studied using the monoclonal antibody, anti‐c‐erbB‐2 (CB‐11, BioGenex) and the avidin‐biotin‐complex (ABC) technique. Four hundred and ninety patients with primary breast carcinoma diagnosed at Gentofte Hospital in the period 1980–1985 were included. Information about treatment, relapse‐free period and overall survival was obtained from the Danish Breast Cancer Co‐operative Group (DBCG). The mean follow‐up period was more than 10 years. Fifteen per cent of the tumours showed positive immunoreactivity for c‐erbB‐2. Oncoprotein expression was correlated with presence of lymph node metastases, type of tumour, high number of mitoses, severe nuclear pleomorphism, high histological grade (poor differentiation), and absence of steroid hormone receptors. By univariate analysis, expression of c‐erbB‐2 oncoprotein, was correlated with poorer overall survival and shorter disease‐free period in the entire cohort and in patients with lymph metastases, but not in the group of patients without lymph node metastases. By multivariate analysis, c‐erbB‐2 failed to be an independent prognostic marker for either disease‐free period or overall survival, whereas classical histopathological parameters such as presence of lymph node metastases, high number of mitoses, high histological grade (poor differentiation) and absence of progesterone receptors turned out to be of independent prognostic significance.
Immunohistochemical detection of progesterone receptors in paraffin sections
Authors: VON BOGUSLAWSKY KRISTINA
Published: August 2009

The benefits of microwave oven pretreatment of formalin‐fixed routinely processed paraffin sections were assessed using citric acid solution to demonstrate progesterone receptors (PR). Parallel frozen and paraffin sections from 25 cases of breast cancer were stained with the PR‐ICA kit (Abbott). Compared with untreated paraffin sections a great enhancement of PR immunostaining occurred after microwave irradiation. In 21 cases the percentage of positively stained tumor cell nuclei in microwave oven‐pretreated paraffin sections was practically identical to that in the frozen ones. After microwave irradiation the PR score in paraffin sections was in three cases higher than in the frozen material. In comparison with the frozen sections, only one tumor of 25 showed a lower percentage of stained malignant cells after the irradiation. It is concluded that microwave oven pretreatment is a simple, reliable and reproducible method to detect PR in routinely fixed paraffin‐embedded tissues.
Identification of a soluble Fcy‐binding molecule (annexin II) in huan serum using a competitive ELISA
Authors: ULVESTAD ELLING , KRISTOFFERSEN EINAR K. , JENSEN TONE SKEIE , MATRE ROALD
Published: August 2009

We have previously produced a monoclonal antibody (mAb), B1D6, reactive with a 37 kD placental IgG Fc‐binding molecule (FcR), recently identified as annexin II. Annexin II is an intracellular molecule found in several cell types, including endothelium and monocytes. Since soluble Fc‐binding molecules are of importance in the regulation of the immune response, we have now used B1D6 in a competitive ELISA to study levels of soluble annexin II in human sera. Soluble annexin II was detected in all sera studied. The highest levels were observed in patients with infectious mononucleosis. Gel filtration of sera revealed annexin II in fractions corresponding to a molecular weight of 40–60 kD. In Western blot analysis a molecule of approximately 37 kD was found. The pi of soluble annexin II was about 7.5–8 as demonstrated by chromatofocusing. Annexin II belongs to a family of phospho‐lipid‐binding molecules involved in anti‐inflammatory responses, and elevated levels of annexin II in serum may be important for the suppression of an immune response.
Development of antibiotic resistance in Pseudomonas aeruginosa during two decades of antipseudomonal treatment at the Danish CF Center
Authors: CIOFU O. , GIWERCMAN B. , HØIBY N. , PEDERSEN S. S.
Published: August 2009

At the Danish CF Center patients with chronic Pseudomonas aeruginosa lung infection were treated 3–4 times a year (from 1976) with a 2‐week intravenous antipseudomonal course which included preferentially an aminoglycoside and a β‐lactam antibiotic. We investigated the development of antibiotic resistance in P. aeruginosa strains isolated from Danish CF patients over a period of 18 years by testing the in vitro efficacy of carbenicillin, piperacillin, ceftazidime, tobramycin and ciprofloxacin against P. aeruginosa strains collected in 1973 (51 strains), 1980 (80 strains), 1985 (58 strains), and 1991 (100 strains). All the strains were screened and assayed semiquantitively for β‐lactamase activity by use of nitrocefin. We found a significant (p<0.005) increase in the MIC values of the P. aeruginosa strains against piperacillin and ceftazidime. However, no significant correlation was found between the MIC and the number of antipseudomonal courses of antibiotics. The proportion of resistant in vivo selected P. aeruginosa strains, presumed to be stably derepressed producers of chromosomal β lactamase, also increased significantly during the period studied. Our results confirm that the β lactamase production is an important mechanism of antibiotic resistance in P. aeruginosa.
The influence of endotoxin upon middle ear fibroblasts cultured in normal middle ear gas and atmospheric air
Authors: OVESEN THERESE , GAIHEDE MICHAEL , ELBRØND OLE
Published: August 2009

Otitis media represents a continuum of inflammatory stages frequently in association with bacteria and/or endotoxin. Furthermore, the disease is often treated with insertion of ventilation tubes, which causes hyperoxia relative to the physiological state in the tympanic cavity. The present study was undertaken to quantitate the interaction between endotoxin and relative hyperoxia in cultures of rabbit middle ear fibroblasts incubated in normal middle ear gas and atmospheric air, respectively. Growth was monitored by determination of DNA, cell protein and cell division. The synthetic activity was estimated by collagen production. The antioxidant defense was determined by measuring the intra‐and extracellular concentrations of superoxide dismutase (SOD). The results demonstrated that hyperoxia significantly impaired the growth of middle ear fibroblasts, which was compensated for by addition of endotoxin stimulating the growth. The collagen synthesis increased significantly in atmospheric air with a synergistic effect of endotoxin. Hyperoxia induced intracellular SOD formation, while endotoxin tended to reduce the synthesis. Finally, exposure to atmospheric air caused significantly larger amounts of reducing agents extracellularly in cultures without endotoxin compared to endotoxin incubated cultures. It is suggested that endotoxin possesses both synergistic and antagonistic potential as regards the effects of relative hyperoxia, and that the interaction between endotoxin and hyperoxia may be an important factor in otitis media.