bluuebunny/crossref_metadata_2025 huggingface dataset:
Ultrastructural morphometric analysis of human blood platelets exposed to minimal handling procedures
Authors: RØGER MAGNUS , HUITFELDT HENRIK S. , HOVIG TORSTEIN
Published: August 2009

A detailed morphometric study of normal human blood platelets is described. The purpose has been to evaluate the morphological characteristics of platelets exposed to minimal handling procedures in order to obtain an optimal basis for the appraisal of platelets in disease. Blood from 10 healthy volunteers was collected directly into buffered glutaraldehyde and processed for electron microscopy. This platelet fixation procedure resulted in excellent preservation of resting platelet ultrastructure with one exception: the dense bodies. Compared to platelets fixed following washing procedures, our directly fixed platelets comprised fewer pseudopods and contained more glycogen. An unexpected feature of the open canalicular system was the apparent release of blisters interpreted as microvesicles. Employing a computerized image analyzer, 300 of the platelets were examined morphologically. The morphometric data thus obtained were analyzed statistically, resulting in a set of standard values for morphological characteristics of human platelets which we have found useful in subsequent evaluations of platelet morphology in disease. Significant inter‐individual variance was, however, detected in two instances, in the section area of the alpha granules, as well as the area fraction of platelet sections occupied by channels of the open canalicular system (OCS). This should be taken into consideration when appraising platelet ultrastructure in health and disease.
17. Malignant tumours of the oesophagus
Authors: PETURSDOTTIR VIGDIS , JONASSON JON G. , HRAFNKELSSON JON
Published: August 2009

We studied all primary malignancies of the oesophagus diagnosed in Iceland between 1955 and 1984 and reclassified tumours where histological material was available using the WHO classification system. Of a total of 329 tumours diagnosed in the time period, seven were excluded for various reasons. Of the remaining 322 tumours, 178 were in males and 144 in females. The age standardized incidence was 5.3/105 for males and 3.1/105 for females. The incidence of oesophageal tumours decreased for both sexes during the time period under investigation. Of 250 reclassified tumours (142 in males and 108 in females), squamous cell carcinomas comprised 81.6%. If the undifferentiated tumours are excluded, the squamous group accounted for 89.1% of the remaining tumours. Small cell carcinomas comprised 3.2% of all cases, which was higher than expected. Most of the tumours appear to be located in the middle part of the oesophagus. The vast majority of resected tumours extended through the wall of the oesophagus. A relatively higher proportion of tumours was confined to the submucosal or muscular layers in the latter half of the period. In conclusion, the epidemiological data in our study appear to resemble what is observed in the other Nordic countries for oesophageal tumours, except for slightly higher overall incidence in Iceland, especially for women.
Detection of gastrin mRNA by in situ hybridization using radioactive‐ and digoxigenin‐labelled probes: a comparative study
Authors: XERRI LUC , MONGES GENEVIÈVE , GUIGOU VÉRONIQUE , PARC PATRICIA , HASSOUN JACQUES
Published: August 2009

The hormone gastrin is mainly produced by the G cells of the antral mucosa and plays a major role in the regulation of digestive mucosal growth. Since it permits identification of cell types containing mRNA, in situ hybridization (ISH) appears to be an interesting method for studying gastrin‐producing tissues. In this study, in situ detection of gastrin mRNA has been carried out on frozen sections of four human normal antral mucosa samples and of six colonic carcinomas removed from patients with high levels of plasma gastrin, using a gastrin oligonucleotidic DNA probe. We have compared the results provided respectively by the [35S] labelling and the digoxigenin labelling of the synthetic probe. Positive cells were found in each normal sample analysed with radioactive‐ as well as digoxigenin‐labelled antisense probes. The total number of cells expressing gastrin mRNA appeared slightly higher with the [35S]‐labelled probe, while the digoxigenin‐labelled probe gave a better definition of positive signals. In contrast, neither radioactive nor cold probes gave positive signals in the six colonic carcinoma samples, although gastrin expression had been demonstrated in these tumours using a reverse transcriptase‐PCR method. These results show that, although ISH does not seem sensitive enough to allow the detection of very low levels of gastrin expression, it would appear to be a reliable method for visualizing gastrin mRNA in human antral mucosa.
β‐lactamases in Shigella
Authors: SCHUMACHER H. , NIR M. , MANSA B. , GRASSY A.
Published: August 2009

The occurrence of resistance and production of β‐lactamases was investigated in 60 Shigella strains. Ampicillin resistance was found in 28 (47%) of the isolates, the resistance being more frequent in Sh. flexneri than in Sh. sonnei. All strains were susceptible to cefotaxime, mecillinam, and ciprofloxacin. The β‐lactamases produced by Shigella were similar to TEM‐1, OXA‐1, or the low‐level chromosomally mediated cephalosporinase produced by Escherichia coli. The β‐lactamases produced by Sh. flexneri were most often the OXA‐1‐like enzymes.
Epitope homology between bacterial heat shock protein and self‐proteins in the host cell
Authors: YAMAGUCHI HIROYUKI , YAMAMOTO TOMOKO , KONOEDA YOSHIKI , TAGUCHI HARUHIKO , OGATA SACHIO
Published: August 2009

Mouse monoclonal antibodies (MAbs) showing distinct reactivity against the 60‐kilodalton (kDa) antigen heat shock protein of Yersinia enterocolitica, designated cross‐reacting protein antigen (CRPA), have previously been established. The reactivities of these MAbs (5C3 and 3C8) against mouse and human host cells were studied by Western blotting and flow cytometric analysis. The results indicated that epitopes on the bacterial 60‐kDa heat shock protein are present on various molecules in mouse spleen cells and human B cells. An epitope recognized by MAb 5C3 was expressed on the mouse and human host cell surface, and an epitope recognized by MAb 3C8 was also expressed on the human host cell surface.
Effects of the prostaglandin E2 analogue enprostil on the carbon tetrachloride‐induced necrosis of liver cells in mice
Authors: BANG S. , MYREN J. , LINNESTAD P. , SERCK‐HANSSEN A. , STRØMME J. H. , BERAKI K.
Published: August 2009

Female mice, eight weeks old, were injected with carbon tetrachloride (CCl4) (10 mg subcutaneously). Groups of mice (n = 10–30) were then injected with enprostil (E) 2, 20 or 50 μg/kg body weight (bw) intraperitoneally 15 min and two h after, or E 100 μg/kg bw two h after the CCl4 injection. The mice were killed after 24, 48 or 72 h. Plasma activity concentrations of alanine aminotransferase (ALAT) were determined in blood specimens from the iliac veins. The extent of liver cell necrosis in histological sections was recorded on a 100 mm Visual Analogue Scale (VAS) and measured using the electronic Mini Mop method. In the group given the highest single dose of E (100 μg/kg) a significant lowering of the CCl4‐induced liver cell necrosis was found after 24 h. No significant differences were found after 48 and 72 h. In the other groups injected with lower doses of E after CCl4, no significant differences were found compared to groups injected with CCl4 alone.
Growth of xenografted squamous cell carcinoma of the head and neck ‐ possible correlation with patient survival
Authors: ZÄTTERSTRÖM U. K. , BRAAKHUIS B. J. M. , WENNERBERG J. , VAN DONGEN G. A. M. S. , ATTEWELL R. , NAUTA J. J. P. , LAARMAN D. A. W. , ASK A.
Published: August 2009

Tumor biopsies from 100 cases of squamous cell carcinoma of the head and neck (HNSCC) were xenografted to athymic nude mice. To ascertain whether xenograft take might be a factor of clinical significance, it was compared with patient survival, the patients being divided into two groups (takeandnon‐take) according to the results of transplantation. The tumor take rate was 29%. Median survival time with respect to cancer death was 18 months in the take group, as compared with over 74 months in the non‐take group (p = 0.06). No significant differences were observed between the two groups with respect to age, tumor size, nodal status, clinical stage or histological differentiation. The findings suggest that take of HNSCC xenografts in nude mice may reflect the malignant potential of the original tumor. Moreover, the possibility cannot be excluded of a selection bias favoring the use of such malignant xenografts in therapeutic studies in nude mice.
Detection and quantitation of antibodies against Rhizopus by enzyme‐linked immunosorbent assay
Authors: SANDVEN PER , EDUARD WIJNAND
Published: August 2009

In sawmills high concentrations of spores from the mould Rhizopus microsporus may occur, causing allergic alveolitis in exposed workers. Both symptomatic and asymptomatic exposed workers may develop antibodies. An enzyme‐linked immunosorbent assay (ELISA) for the detection of antibodies to Rhizopus has been developed in order to study the relationship between antibody levels and exposure levels. The precision of the measurements of Rhizopus antibodies by ELISA carried out on the same microtiter plate was estimated to be 11%. It is therefore possible to detect changes in antibody levels of approximately 25% or more. Antibodies were studied longitudinally by ELISA in 60 wood trimmers. The observed changes in antibody levels exceeded the precision of the ELISA method substantially, indicating significant variability in antibody levels in wood trimmers. The ELISA test was compared with the double immunodiffusion test (DID). Sera from 67 wood trimmers were analyzed by both methods. Antibodies were detected by ELISA in 70% and by DID in 28% of the workers in this group, clearly demonstrating that the ELISA test is the most sensitive method for the detection of antibodies to Rhizopus.
The use of post‐embedding immunoelectron microscopy in the diagnosis of glomerular diseases
Authors: SKJØRTEN FREDRIK J. , BRORSON SVERRE‐HENNING , ROALD BORGHILD , STRØM ERIK H. , LUND BJØRG
Published: August 2009

Fifty renal biopsies were studied by immunoelectron microscopy after embedding in a partly hydrophilic polyacrylic resin (LR White). Immunofluorescence studies were carried out on frozen sections of parallel tissue samples. Polyacrylic embedding gave good preservation of the renal ultrastructure and precise localization of immunoglobulin and C3c antibodies within glomerular electron‐dense deposits. Non‐specific staining of plasma proteins within vascular lumina could easily be detected. There was good correlation between immunoelectron and immunofluorescence microscopy. Immunoelectron microscopy is a very sensitive method, which can detect small amounts of antigen. More cases were, however, positive by immunofluorescence than by immunoelectron microscopy. This discrepancy may be explained by difference in sample size, and by difference in resolution of morphological details (electron microscopy versus fluorescence microscopy).
Nationwide spread of Klebsiella oxytoca K55 in Swedish neonatal special care wards
Authors: TULLUS KJELL , AYLING‐SMITH BEVERLY , KüHN INGER , RABSCH WOLFGANG , REISSBRODT ROLF , BURMAN LARS G.
Published: August 2009

In a nationwide survey of invasive bacterial infections in Swedish neonates, 36% of Klebsiella spp. were Klebsiella oxytoca serotype K55. This unexpectedly high proportion of K55 infections was due to clusters of infection in neonatal special care wards, and at first seemed attributable to nosocomial spread of a K. oxytoca strain of high virulence. Factors predisposing infants to infection were, however, found irrespective of whether the infecting strain was of serotype K55 or not. Additionally, the prevalence rates of a potential virulence factor, siderophore production, were similar among the two groups of strains. During the same period of time a K. oxytoca K55 with similar biochemical phenotype and drug resistance pattern was found to be spread among the neonates in 12 of 22 neonatal wards in Sweden. The increased proportion of invasive neonatal K. oxytoca K55 infections thus seemed to reflect a high rate of colonization rather than an increased virulence of the K55 strain.
Mucosal immunoreactivity in experimental pneumococcal otitis media
Authors: SVINHUFVUD M. , HELLSTRÖM S. , HERMANSSON A. , PRELLNER K.
Published: August 2009

A rat model was used to investigate immunological events in the middle ear mucosa during pneumococcal acute otitis media (AOM). Twelve healthy male Sprague‐Dawley rats were challenged in the right middle ear with Streptococcus pneumoniae type 3, the presence of AOM being confirmed by otomicroscopy. Randomly selected rats, four at a time, were sacrificed on days 7, 14 and 56 after bacterial challenge. Immunohistochemical staining for IgG, secretory IgA (SIgA), and IgM was performed on tissue specimens from four separate locations in the middle ear and tubal mucosa. Immunoglobulins, especially IgG, were found around blood vessels in the middle ear mucosa. Immunoreactive lymphoid cells of all three Ig classes investigated, undetectable before bacterial challenge, appeared within 7–14 days after middle ear challenge, and the location of these cells in the middle ear mucosa suggests the presence of IgG, SIgA, and IgM, respectively. On the other hand, reactivity with anti‐SIgA (but not with anti‐IgG or anti‐IgM) in Eustachian epithelial cells, and also in the subepithelial glands of tubal mucosa was present both before and after challenge. The results suggest that the middle ear mucosa is an immunoreactive site only after it has been activated with pathogens. In contrast, the tubal mucosa exhibits immunological activity also prior to the antigenic stimulation of present interest.
Enzyme‐linked immunosorbent assay for determination of antibodies to Vibrio cholere toxin‐coregulated pili
Authors: OSEK JACEK , JONSON GUNHILD , SVENNERHOLM ANN‐MARI , HOLMGREN JAN
Published: August 2009

An ELISA for determination of antibodies to V. cholerae TCP was developed. Since purified TCP preparations contained detectable amounts of LPS (as shown by ELISA and immunoelectron microscopy with anti‐LPS polyclonal serum), a capture ELISA was used. In this test the plate was coated with anti‐TCP monoclonal antibody followed by incubation with TCP fimbriae. By this procedure no LPS bound to the solid phase as shown by the loss of reactivity with anti‐LPS serum. The capture ELISA allowed sensitive and specific determination of TCP antibodies in sera of rabbits immunized with classical but not El Tor V. cholerae strains. There was good agreement between results in the TCP ELISA and reactivity with the TcpA band in immunoblot analyses when antisera raised against classical and El Tor vibrios were studied.
Neuronal uptake of plasma proteins in cryogenic brain lesions
Authors: LØBERG E. M. , BRORSON S.‐H. , SKJØRTEN F. , TORVIK A.
Published: August 2009

A previous light microscopic study on cryogenic brain lesions in rats demonstrated uptake of plasma proteins into damaged neurons within a few minutes after the lesion. The protein concentration was much higher inside the nerve cell bodies than in the surrounding neuropil. This is puzzling since the neuropil to a large extent consists of damaged neuronal processes. The present investigation describes the intracellular localization of albumin in this model using a post‐embedding immunoelectron microscopic technique. The distribution of albumin in the lesions was studied after 1, 6 and 12 h survival periods. The intraneuronal albumin was mainly bound to the particulate elements of the cytoplasm and nuclei, while the watery parts of the cells showed no immunoreactivity. The intracellular organelles contained very little albumin, indicating that their membranes may be more resistant to freezing than those of the cells. Most of the neuronal and glial processes in the neuropil were swollen and contained almost no albumin. This explains the contrast between the strong immunoreactivity of the neurons and the vague reactivity of the neuropil in light microscopy.
Experimental studies of survival of anaerobic bacteria at 4°C and 22°C in two different transport systems
Authors: TVEDE MICHAEL , HØIBY NIELS
Published: August 2009

The survival of anaerobic bacteria on swabs in two different transport systems at 4°C and 22°C was evaluated. The transport systems were a charcoal‐impregnated cotton swab in modified Stuart transport medium (MST), and a viscose swab in modified SIFF transport medium (BTM) (Bionor™). The following eight clinical strains of anaerobic bacteria were tested for quantitative recovery at 24 h, 48 h, 72 h, and 96 h; Fusobacterium necrophorum, Bacteroides melaninogenicus, Bacteroides intermedius, Peptostreptococcus anaerobius, Peptococcus magnus, Clostridium perfringens, Clostridium tetani, and Actinomyces israelii. Additionally, a mixture of Staphylococcus aureus and Escherichia coli was tested together with Bacteroides fragilis, Fusobacterium necrophorum and Clostridium perfringens. Both transport systems preserved the bacteria, but the BTM medium gave a better quantitative recovery of the bacteria than the MST in 29/64 (45%) of the experiments, whereas the opposite was the case in 15/64 (23%) of the experiments (p < 0.05). There was no significant strain‐related difference between the recovery of 10 different B. fragilis strains. There was no major difference in the recovery of the anaerobes in the two systems at 4°C compared to 22°C, except for Fusobacterium necrophorum, which survived best at 4°C in the Bionor™ transport system.
Rate‐limiting components and reaction steps in complement‐mediated haemolysis
Authors: NIELSEN HANS ERIK , LARSEN SEVERIN OLESEN , VIKINGSDOTTIR THORA
Published: August 2009

The aims of this study were to identify the rate‐limiting components and reaction steps in the integrated activation sequence of the alternative (AP) and classical (CP) pathways of the complement (C) system. In an initial correlation analysis we found that the haemolysis rate in AP was correlated with the concentrations of C5 and IgM. In CP, the haemolysis rate was correlated with the concentrations of C2‐C6, factors I and B, and IgM. In order to identify the rate‐limiting components, we added single, purified C components and IgM to pooled, normal human serum and measured the resultant change in the haemolysis rate. We found that a large number of different components, rather than a single one, were rate‐limiting in AP and CP. In reconstitution experiments we found that in CP the rate‐limiting reaction steps are the activation of C4 and C2. In AP we cannot identify the rate‐limiting step precisely, but can only state that it is at the C3 activation step or earlier.
Comparison of detection speed and yield in agitated and non‐agitated aerobic blood culture bottles
Authors: ARP MAGNUS , LARSEN SEVERIN OLESEN
Published: August 2009

The influence of agitation on detection speed and yield was evaluated in 7,033 paired, identical aerobic blood culture bottles (Media Department, Statens Seruminstitut, Copenhagen, Denmark). One bottle was agitated and the other was incubated stationary. Of a total of 943 organisms isolated in aerobic blood culture bottles, 74% were clinically significant. No difference in the total yield of significant organisms was observed between agitated and non‐agitated bottles. In the evaluation of detection speed, only cultures where organisms were isolated from both bottles, but at different times, were included, to ensure that no other factors influenced the result. Staphylococci, Pseudomonas spp., and Candida spp. were detected significantly faster in agitated bottles, on average 0.5‐1 day earlier (p<0.05), and in the majority of the cases within the first incubation day. These species are frequently found and are important causes of severe generalized infections, especially in immunocompromised patients, where early detection is of great importance. The detection principle in agitated bottles in our system, darkening of blood, proved to be an easy, reliable, and fast method to detect positive aerobic blood cultures, which could probably lead to increased automation.
Fimbriae of Branhamella catarrhalis as possible mediators of adherence to pharyngeal epithelial cells
Authors: AHMED KAMRUDDIN
Published: August 2009

This study attempted to elucidate the role of fimbriae in the adherence of B. catarrhalis to human oropharyngeal epithelial cells. Antifimbrial immune serum was prepared by immunization of rabbit with whole fimbriated bacteria, and adsorption of the serum with a nonfimbriated B. catarrhalis strain. After pretreatment with the antifimbrial antiserum, the adherence of fimbriated B. catarrhalis to human epithelial cells was significantly decreased (p<0.05). The adherence was also significantly (p< 0.001) decreased by trypsin treatment. Electron microscopy revealed destruction of fimbriae after trypsin treatment. These observations suggest that fimbriae are involved in the adherence of B. catarrhalis to epithelial cells.
Superficial perineal leiomyosarcoma in an adolescent female and a review of the literature including vulvar leiomyosarcomas
Authors: GROVE ANNI , NØHR SUSANNE BACKMAN
Published: August 2009

Superficial perineal leiomyosarcomas are rare, with only three previously reported examples. We encountered a superficial (deep subcutaneous) perineal leiomyosarcoma in a 17‐year‐old female. At follow‐up two years after a wide excision, there were no signs of recurrence. The tumour was well differentiated and showed immunoreactivity for alpha‐smooth muscle actin and desmin. A review of the literature on superficial leiomyosarcomas indicates that superficial perineal leiomyosarcomas may be more aggressive than superficial leiomyosarcomas in general. As the presented tumour occurred in a female, it was compared with vulvar leiomyosarcomas.
Primary non‐Hodgkin's lymphoma of the breast (PLB): a clinicopathological study of seven cases
Authors: HANSEN TOM GIEDSING , OTTESEN GYDA LOLK , PEDERSEN NIELS TINGGAARD , ANDERSEN JOHAN A.
Published: August 2009

Seven cases of PLB in females were reviewed. Six cases involved the breast alone (stage 1E), whereas one case also involved the ipsilateral lymph nodes (stage 2E). None had B symptoms. The age range was 37–70 years (mean 52 years). The clinical course was indistinguishable from that of breast carcinomas, though the tumors were relatively large and the duration of symptoms was relatively short. Histologically, all cases were non‐Hodgkin's B‐cell lymphomas, comprising five cases of diffuse centroblastic type, one case of diffuse centroblastic‐centrocytoid type and one case of follicular centroblastic‐centrocytic type, according to the updated Kiel classification, corresponding to six cases of the diffuse large cell type and one case of the follicular mixed, small cleaved and large cell type, according to the modified International Working Formulation (IWF). None of the cases was a lymphoma of mucosa‐associated lymphoid tissue (MALT). The mammary glandular tissue outside the tumor showed ductectasia in four cases. Changes resembling fibrous disease of the breast were seen in four cases, too. The inflammatory infiltration in all these cases was mainly composed of T cells. At the time of diagnosis, all patients were receiving cytoreductive treatment. The mean followup time was 34 months (range 5–79 months). At present, six of the patients are alive and well.
Immunohistology of skin and rectum biopsies in bone marrow transplant recipients
Authors: LESKINEN RITVA , TASKINEN EERO , VOLIN LIISA , RUUTU TAPANI , HÄYRY PEKKA
Published: August 2009

We have studied histological and immunohistological specimens of 39 skin biopsies from 21, and 30 rectal biopsies from 17 bone marrow transplant recipients. The biopsies were taken before transplantation, during acute and chronic graft‐versus‐host disease (GVHD), and at times with no GVHD. In biopsies taken during cutaneous aGVHD grade I to III, epithelial changes were seen in 16/23 biopsies. The cutaneous infiltrates during aGVHD consisted of CD2‐, CD4‐, CD8‐ and FMC‐33‐positive cells both in the epithelium and in the dermis. CD57‐positive NK cells were also detected in most biopsies. During chronic GVHD the cutaneous cellular infiltrates were similar to those seen in moderate aGVHD, i.e. both CD4‐ and CD8‐positive lymphoid cells were present. When the biopsy was taken after the beginning of corticosteroid treatment for aGVHD, or at times when the patient did not have GVHD symptoms, the cellular infiltrates were considerably smaller in the dermis. During clinical intestinal aGVHD mucosal epithelial changes were relatively uncommon; instead, increased numbers of both CD4‐ and CD8‐positive lymphocytes in the lamina propria (LP) were seen in 11/13 samples. During chronic GVHD the number of CD4‐positive cells exceeded that of CD8‐positive cells in the LP, and the large lymphoid infiltrates also reached the muscularis mucosae. In rectal biopsies the differences were not so prominent because most of the pretransplant biopsies showed CD2‐, CD4‐, CD8‐ and CD57‐positive lymphocytes both in the lamina propria and epithelium.
Cell kinetics of the rat corneal epithelium
Authors: HAASKJOLD ERLING , REFSUM HELGA , REFSUM SIGVALD B. , BJERKNES ROLF
Published: August 2009

In the rat corneal epithelium the mitotic rate (MR) is almost equal throughout the epithelium in the morning (Haaskjold et al.1988). The labelling index (LI) shows a marked reduction in the central cornea, which could suggest a lack of uptake of tritiated thymidine via the salvage pathway (Haaskjold et al.1989). In the present study we have used [3H]deoxycytidine, and [3H]thymidine after prior treatment with a methotrexate regimen to elucidate this discrepancy. Deoxycytidine is incorporated into DNA independent of thymidine kinase, while methotrexate, which depletes the cells of reduced folates, makes the cells completely dependent on the salvage pathway. With both techniques the same pattern of labelling was observed, confirming that in the morning the ratio between the MR and the LI differs throughout the cornea. Based on previous observations, an analysis of the MR/LI ratio during 24 h was performed, showing that these parameters were strongly correlated. This suggests that there may be different circadian variations in the cell proliferation parameters throughout the corneal epithelium. The methotrexate regimen may be a useful tool to investigate the salvage pathway.
Binding of human IgA to HCl‐extracted c protein from group B streptococci (GBS)
Authors: KVAM AUGUSTA IRENE , IVERSEN OLE‐JAN , BEVANGER LARS
Published: August 2009

The cβ protein of group B streptococci obtained by HCl extraction appears as a ladder‐like pattern in SDS‐PAGE when detected by a rabbit anti‐cβ serum, and a similar picture is seen when the crude extract is incubated with human IgA and an anti‐human IgA conjugate. Affinity‐purified cβ antigen and IgA receptors from GBS gave identical pictures in Western blots using rabbit anti‐cβ serum. Both the cβ antigen and the IgA receptor are exposed on the surface of GBS as demonstrated by immunofluorescence.
Interferon‐gamma and resistance to bacterial infections
Authors: CZARNIECKI CHRISTINE W. , SONNENFELD GERALD
Published: August 2009

Since its initial description as an antiviral, it has become clear that Interferon‐γ (IFN‐γ) has potent immunoregulatory and cell growth regulatory activities. As a result of these additional activities, it is now apparent that IFN‐γ plays a major role in regulation of bacterial infections. IFN‐γ can be both induced by bacteria and bacterial products; endogenous IFN‐γ production has been shown to play a protective role in the natural host response to several bacterial infections; and administration of exogenous IFN‐γ is effective in the prevention and treatment of bacterial infections in numerous animal model systems. Although it is now clear that IFN‐γ plays a role in regulation of bacterial infections, the mechanisms of its anti‐bacterial effects in vivo remain to be established due to the pleiotropic nature of IFN‐γ activity.
Demonstration of toxin A and B by polymerase chain reaction and McCoy cell assay in clinical isolates of Clostridium difficile from Denmark
Authors: KNUDSEN J. D. , TVEDE M.
Published: August 2009

A polymerase chain reaction (PCR) for demonstration of gene fragments of Clostridium difficile was established. One hundred and sixty‐eight clinical isolates of C. difficile from three population groups were tested for production of cytotoxins by McCoy cell line assay (MCA) and for fragments of toxin A and B genes by PCR. The fragments for PCR amplification were at the 5′ end of the toxin genes, which was found to be specific for C. difficile. Full agreement between the PCR and MCA results was found with respect to toxicity. Fifty‐eight of the 168 strains were cytotoxin positive. Isolates from 41 normal healthy children did not differ regarding cytotoxicity compared to isolates from hospitalized children. In adult hospitalized patients a much higher frequency of toxin‐producing isolates was found. In two strains from two children, not of the same family, only the toxin A gene fragment was demonstrated, indicating that some strains of C. difficile only harbour the gene for enterotoxin. When two isolates from different periods of time were tested from 36 of the healthy children, a variation in cytotoxicity was found: in seven children strains changed from non‐toxic to toxic and in four vice versa. This may be explained by a fluctuating colonization of toxic and non‐toxic C. difficile strains, or it may indicate the need for examination of more than one strain from a positive faecal sample to demonstrate cytotoxicity.
Stereological estimation of nuclear volume in benign and atypical meningiomas
Authors: MADSEN CLAUS , SCHRØDER HENRIK D.
Published: August 2009

A stereological estimation of nuclear volume in benign and atypical meningiomas was made. The aim was to investigate whether this method could discriminate between these two meningeal neoplasms. The difference was significant and it was moreover seen that there was no overlap between the two groups. The results demonstrate that atypical meningiomas can be distinguished from benign meningiomas by an objective stereological estimation of nuclear volume.
Susceptibility pattern of Scandinavian Francisella tularensis isolates with regard to oral and parenteral antimicrobial agents
Authors: SCHEEL OLAF , HOEL TERJE , SANDVIK TORSTEIN , BERDAL BJØRN P.
Published: August 2009

Some recently introduced antimicrobial agents have only been incompletely evaluated for use in Francisella tularensis infections. The present study evaluated the susceptibility pattern of Scandinavian human, rodent, and hare F. tularensis isolates with respect to a selection of traditional as well as recently introduced antimicrobial agents. All strains were resistant to the following β‐lactams: penicillin, cephalexin, cefuroxime, ceftazidime, aztreonam, imipenem, and meropenem with minimal inhibitory concentrations > 32 mg/1. Against macrolides, a mixed susceptibility/resistance pattern appeared. All strains were susceptible to gentamicin, chloramphenicol, doxycycline, and four quinolones. Since the quinolones showed the lowest MIC values, and in addition give a good intracellular penetration, we conclude that future drugs to consider against tularemia should definitely include this group of antibiotics. The outpatient mode of antibiotic treatment is especially relevant as the Scandinavian variant of F. tularensis infection is nonlethal, usually pustuloglandular, and not septicemic. Therefore, oral drugs must be sought, and the quinolone group also satisfies this requirement.
Chlamydial cervicitis: Role of culture, enzyme immunoassay and Giemsa cytology in diagnosis
Authors: MITTAL ARUNA , KAPUR SUJALA , GUPTA SUNANDA
Published: August 2009

Three different laboratory tests were carried out to find the occurrence of chlamydial infections in a selected group of 150 female patients presenting clinically with cervicitis. The tests included isolation using a cell culture system, enzyme immunoassay (EIA) for antigen detection and Giemsa cytology of endocervical smears. Contrary to earlier reports on cytological diagnosis of chlamydial cervicitis, endocervical smears stained by Giemsa stain and EIA for antigen detection were found to be of comparable sensitivity and specificity (73.1% and 86.3% for smear, and 76.1% and 90.9% for EIA, respectively) when cell culture was used as the gold standard.
Antibody profiles to the group B streptococcal beta antigen in maternal and infant paired sera
Authors: FLORES AUREA E. , NELSON JO ANN , WU XIAO YAN , FERRIERI PATRICIA
Published: August 2009

Antibody profiles to the purified β antigen of the c protein of group B streptococci (GBS) were studied by ELISA and Western immunoblot (WB). The sera from 139 parturient women colonized with GBS, 35 non‐colonized parturients and their newborn infants were studied by ELISA; WB was done on 76 maternal and 26 infant sera. Enzyme‐labeled anti‐IgA (α), ‐IgG (γ), ‐IgM (μ), or ‐IgG (H&L) were used as secondary antibodies. A high prevalence of antibody to the β antigen was observed by both ELISA and WB among parturient women and their newborns. IgG (H&L) ELISA titers ≥200 were found in 84% and ≥800 in 31% of the maternal sera. A significantly higher percentage of women colonized than those non‐colonized with GBS had IgG (γ) titers ≥800. A significantly higher percentage of women colonized with c protein‐positive than c‐negative strains of GBS had IgG (H&L) titers ≥3200. Twelve of 27 women with IgM antibody to the β antigen also had IgG (γ) titers ≥800 and were, in addition, colonized with GBS. Multiple molecular forms of the antigen from 25 to 140 kDa were blotted by the maternal and infant sera. Concordance in the IgG but not in IgA or IgM antibody profiles of maternal and infant paired sera was observed in the overall blotting patterns and ELISA titers. The same titer as the mother was found in 55% of the infant sera and within one dilution in 97%. This suggests active transfer of IgG antibody to the β antigen across the placenta from mother to baby.
Intracellular levels and extracellular release of lysosomal enzymes from peripheral blood monocytes in pulmonary tuberculosis patients
Authors: JASWAL S. , DHAND R. , SETHI A. K. , KOHLI K. K. , GANGULY N. K.
Published: August 2009

The intracellular activity and extracellular release (basal and latex‐stimulated) of B‐glucuronidase (BG) and N‐acetylglucosaminidase (NAG), measured fluorimetrically, were observed to be significantly (P<0.05) higher in blood monocytes (BM) of untreated pulmonary tuberculosis (TB) patients compared to those of age‐ and sex‐matched controls and Mantoux‐positive subjects without any evidence of active disease. After completion of antituberculous therapy, BG and NAG activities declined appreciably (P<0.05) and their levels became comparable to those in control subjects. The present results suggest the potentiation of the oxygen‐independent defense mechanism of BM in pulmonary TB.
Survival and ultrastructural changes of Helicobacter pylori after phagocytosis by human polymorphonuclear leukocytes and monocytes
Authors: ANDERSEN LEIF PERCIVAL , BLOM JENS , NIELSEN HENRIK
Published: August 2009

Few studies have been carried out on the phagocytosis and killing of Helicobacter pylori by both polymorphonuclear leukocytes (PMNs) and monocytes. In this study, H. pylori was incubated for up to 60 min either alone or with phagocytes in the presence or absence of human serum. Both non‐immune serum and immune serum were used. Reduction in the number of H. pylori, which corresponds to the killing of H. pylori, was analysed by a colony count and ultrastructural changes were studied by electron microscopy. No reduction in the number of H. pylori was found when the bacteria were incubated alone or with phagocytes in the absence of serum. It is remarkable that unopsonized H. pylori was phagocytosed. When immune serum was added to the suspensions of bacteria and phagocytes, the killing rate of H. pylori was found to depend on the ratio of H. pylori to phagocytes. Thus an excess of monocytes reduced the number of H. pylori, whereas an excess of PMNs resulted in complete killing of H. pylori. On incubation with PMNs and serum, ultrastructural changes were observed in the majority of the bacteria whether they were phagocytosed or not. Controls without serum did not show any changes in the morphology of H. pylori, indicating that components in the serum play an important role in the phagocytosis and killing of H. pylori. In contrast, several of the phagocytosed bacteria were found to be unaffected after incubation with monocytes and serum. Such preparations often contained large aggregates of platelets surrounding unaffected H. pylori. In the gastric mucosa, H. pylori is often found in excess as compared to the phagocytes. If these results can be compared to the situation in vivo, the phagocytes seem to be ineffective in the killing of H. pylori, and other immune mechanisms may therefore be of importance for the elimination of H. pylori from the gastric epithelium. The possible intracellular survival of H. pylori should be taken into account when treatment regimes for H. pylori infections are chosen.
Thymic epithelial abnormalities in patients with congenital heart disease and Down's syndrome
Authors: DYBDAHL H. , HENRIQUES U. V.
Published: August 2009

Among 85 children (< 5 years) operated on for congenital heart disease, we found 15 with structural changes of the thymus consisting of epithelial ducts and cysts. Nine of the children had Down's syndrome. In only two of these did we find epithelial abnormalities. In conclusion, these changes were not more frequent in children with Down's syndrome than in other patients with congenital heart disease.
Hypoxia‐associated proteins in human cells cultivated in vitro: lack of association with hypoxia‐induced cell cycle regulation
Authors: SHI YOUJIU , ÅMELLEM ØYSTEIN , PETTERSEN ERIK O.
Published: August 2009

The synthesis of proteins expressed in human NHIK 3025 cells following exposure to extremely hypoxic conditions (<4 ppm O2) has been studied. Populations of cells, either in exponential growth or synchronized by the method of detaching mitotic cells, were exposed to extremely hypoxic conditions for up to 20 h. The rate of total protein synthesis was measured at various time points after reoxygenation, and it appeared to be relatively constant and similar to the control level. The protein expression in cells was studied by pulse labelling for 1 h with [35S]‐methionine, and subsequently visualized by SDS‐PAGE and autoradiography. Six proteins appeared to have a changed expression after exposure to extreme hypoxia as compared to control cells; four of them (45, 80, 100 and 150 kD) showed increased, while two (46 and 90 kD) showed decreased expression. The response of these proteins to extreme hypoxia seems to be relatively slow, i.e. with half‐times of several hours. Since extreme hypoxia influences cell cycle progression by instantaneous blockage at the G1/S border as well as halting DNA synthesis in S cells, these proteins can hardly cause these effects. Neither is the altered expression of these proteins due to the accumulation of G1 cells caused by hypoxia.
Mortality rate and bacteremia, endotoxin, and endothelin‐1 levels in antibiotic therapy for E. coli septic peritonitis
Authors: TSUKADA KATSUHIKO , KATOH HIROYUKI , SHIOJIMA MASAYUKI , TAKENOSHITA SEI‐ICHI , NAGAMACHI YUKIO
Published: August 2009

In order to study the mortality rate and bacteremia, plasma endotoxin, and plasma endothelin‐1 levels in antibiotic therapy for E. coli peritonitis, blood samples were obtained from rats given intraperitoneal injections of latamoxef or placebo. Intraperitoneal injections of latamoxef improved the prognosis of peritonitis rats. Two h after treatment, bacteremia levels were noticeably higher in rats treated with placebo than in rats treated with latamoxef, but the latamoxef‐treated group manifested a significant elevation of plasma endotoxin and endothelin‐1 levels compared to the placebo‐treated group. The results of this study demonstrate that treating E. coli septic peritonitis with selected antibiotics induces increased plasma endotoxin levels, which are associated with elevation of plasma endothelin‐1 levels.
Antibodies from chronically infected cystic fibrosis patients react with lipopolysaccharides extracted by new micromethods from all serotypes of Pseudomonas aeruginosa
Authors: FOMSGAARD ANDERS , SHAND GEOFFREY H. , FREUDENBERG MARINA A. , GALANOS CHRIS , CONRAD ROBERT S. , KRONBORG GITTE , HØIBY NIELS
Published: August 2009

Micromethods were developed to extract lipopolysaccharide (LPS, endotoxin) from single bacterial colonies of the 20 recognized Pseudomonas aeruginosa type strains. The appearance of these LPSs in polyacrylamide gel electrophoresis (PAGE) and their reactivity with serum of cystic fibrosis (CF) patients chronically infected with P. aeruginosa was studied. Silver staining of LPS after PAGE showed that 13 of the P. aeruginosa LPSs had high numbers of O‐repeating units arranged in 1–4 clusters of banding. Low‐molecular‐weight LPS fractions were more prominent in six of the serotype strains, of which O:7 and O:14 appeared semi‐rough. Corresponding immunoblots using the CF sera showed LPS patterns very similar to the silver‐stained appearance, indicating an immune reaction to all P. aeruginosa LPS including that from the newly discovered O:18, O:19 and O:20. This was unexpected since only a few serotype strains (mostly O:3, O:6 and O:9) had been isolated from the patients. Absorption experiments using purified and chemically defined P. aeruginosa rough LPS and smooth LPS suggested these immune reactions were due to antibodies cross‐reactive to core/lipid A as well as to lower molecular weight O‐polysaccharides or “A‐bands”. However, in some cases O:3, O:6, and O:9 LPSs were also found to contain additional distinct O‐epitopes. Immune recognition of various polyagglutinable P. aeruginosa LPSs seemed also to be caused by cross‐reactive antibodies. The described microextraction methods, followed by PAGE and silver staining or immunoblotting, are easy and convenient techniques with which to study antibodies against LPS epitopes and to screen for LPS phenotypic appearance using only a few bacterial colonies from larger numbers of Gram‐negative bacterial strains.
Acridine orange‐mediated photodamage to cultured cells
Authors: ZDOLSEK JOHANN M.
Published: August 2009

Photosensitization mediated by the lysosomotropic, weakly basic dye acridine orange (AO) was studied on cultured J‐774 cells. The phototoxicity was found to be potentiated by elevated oxygen tension and reduced at low oxygen tension. Moreover, cell cultures pre‐exposed to the singlet oxygen scavenger sodium azide showed pronounced protection against the loss of viability induced by AO and blue light. AO‐mediated photosensitization was neither increased by pre‐exposure of cell cultures to ferric chloride or the catalase‐inhibitor aminotriazole nor decreased by exposure to deferoxamine. These observations suggest that type II (singlet oxygen‐mediated) reactions predominate over type I reactions (radical‐mediated). A rapid and pronounced decrease in lysosomal cathepsin L activity (up to 60%) was observed after an initial 10 min irradiation, indicating the lysosomal compartment to be an early target. This irradiation time did not, however, result in any substantial loss of viability. Levels of cytosolic lactate dehydrogenase were unaffected even after 30 min irradiation, indicating that neither cytosol nor plasma membrane is a primary target of the AO‐mediated photodamage. Glutathione depletion by pre‐exposure to buthionine‐S, R‐sulfoximine (BSO) much enhanced the sensitivity of J‐774 cells to AO‐mediated photosensitization, indicating a protective role for thiolcontaining compounds against AO‐mediated photodamage.
Stereologic studies on collagen in bovine articular cartilage
Authors: HEDLUND HÅKAN , MENGARELLI‐WIDHOLM SILWA , REINHOLT FINN P. , SVENSSON OLLE
Published: August 2009

In an ultrastructural stereologic study on bovine articular cartilage we found that collagen volume density increased with increasing distance from the joint surface and from the chondrocyte. These results not only corroborate previous biomechanical data of a vertical stiffness gradient, but they also suggest that the mechanical forces are unevenly distributed horizontally. On the other hand, although mean collagen fibril diameter showed large differences between the interterritorial compartments of the three zones, there was a population of slender fibrils in all zones and compartments. Since the coarser fibrils provide the high tensile strength (Nimni 1988), the role of the slender fibrils may be to enhance the deformability of the tissue. Moreover, in spite of substantial differences in mean fibril diameter, collagen surface densities were in the same order of magnitude in the territorial and interterritorial compartments, and only slightly lower in the pericellular compartments. The surface parameter may be important for specific molecular interactions. The collagen fibrils have different polarity, i.e. the direction of the fibrils appears to be parallel and antiparallel, about 50% running in each direction. This, together with the very high length/diameter ratio (Clark 1985), may indicate that each fibril is assembled by the concerted action of many cells. The characteristic properties of articular cartilage depend on interactions between its macromolecular components, and the present quantitative data form a basis for discussions on the specificity and regulation of such interactions.
Comparison of growth and susceptibility testing of pyrazinamide in different Bactec media using strains of the M. tuberculosis complex
Authors: FUURSTED KURT
Published: August 2009

The aim of this study was to evaluate and compare growth and susceptibility testing of pyrazinamide (pza) in different Bactec media using 85 strains of the M. tuberculosis complex. Fifty‐four were clinical isolates of M. tuberculosis, of which 34 strains were selected as they failed to grow in primary pza susceptibility testing. Fourteen were strains of M. bovis, 10 were different strains of M. bovis BCG, five were strains of M. africanum, and two were strains of M. microti. The following Bactec media were evaluated: (1) Bactec PZA test medium, (2) acidified Bactec 12B medium with pH reduced to 5.6, (3) acidified 12B medium supplemented with fresh egg yolk, (4) acidified 12B medium supplemented with a commercial egg yolk enrichment. Demonstration of pyrazinamidase activity was included for comparison. All strains of M. tuberculosis, M. microti and M. africanum were inhibited by pza at 100 μg/ml, and all strains of M. bovis and M. bovis BCG were found to be resistant to pza at 100 μg/ml. Bactec PZA test medium supported growth for all 85 strains tested. The two egg yolk‐supplemented media supported growth in all strains except one strain of M. microti, and acidified but otherwise unsupplemented Bactec 12B medium supported growth in 64 strains (75%). A pza sensitivity test result could be obtained within one week in > 79% of strains. An overall correlation of 89% between susceptibility to pza and demonstration of pyrazinamidase activity was demonstrated.