bluuebunny/crossref_metadata_2025 huggingface dataset:
Protection from concanavalin A (Con A)-induced T cell-dependent hepatic lesions and modulation of cytokine release in mice by sodium fusidate
Authors: NICOLETTI F , BELTRAMI B , RASCHI E , DI MARCO R , MAGRO G , GRASSO S , BENDTZEN K , FIORELLI G , MERONI P L
Published: November 2003

The immunomodulatory effects of the antibiotic sodium fusidate (SF) were tested in a model of T cell-dependent hepatic injury that can be induced in normal mice by a single i.v. injection of Con A. Signs of hepatitis with elevated transaminase activities in plasma, severe infiltration of the liver by neutrophil granulocytes, lymphocytes and monocytes, and necrotic areas were observed in control mice treated intraperitoneally with PBS 24 h and 1 h before Con A challenge. T cell- and macrophage-derived cytokines (IL-2, interferon-gamma (IFN-γ), tumour necrosis factor-alpha (TNF-α), IL-1β, IL-6) were released with different kinetics in the circulation of these mice. SF, 20, 40 or 80 mg/kg, administered 24 h and 1 h before Con A challenge, protected the mice against the hepatitic effects of Con A. The protective effects of SF were dose-dependent and accompanied by profound modifications of blood levels of cytokines induced by Con A, so that, relative to control mice, SF (80 mg/kg)-treated animals showed markedly diminished plasma levels of IL-2, IFN-γ and TNF-α, along with augmented levels of IL-6. These results suggest that SF might be useful in the treatment of immunoinflammatory liver diseases in humans.
Analysis of T cell receptor Vβ usage in the autoimmune sialadenitis of non-obese diabetic (NOD) mice
Authors: YANAGI K , HANEJI N , ISHIMARU N , SAITO I , HAYASHI Y
Published: November 2003

The NOD mouse develops spontaneous autoimmune sialadenitis besides a well characterized T cell-mediated autoimmune insulitis. We used reverse transcriptase-polymerase chain reaction (RT-PCR) to analyse the repertoire of T cell receptor (TCR) Vβ chain genes expressed in the isolated infiltrating cells from affected salivary glands. Immunohistochemical analysis showed that the vast majority of inflammatory infiltrates in the salivary glands were CD4+Vβ8+ and CD4+Vβ6+ T cells, whereas CD8+T cells and B220+ B cells were fewer in number. Predominant expression of the Vβ8 and Vβ6 gene segment was detected in the infiltrating cells in the salivary glands very early, and age-related diversity of TCR Vβ gene usage was observed. Single-strand conformation polymorphism (SSCP) analysis demonstrated a strikingly symmetrical distribution of expanded clones in the PCR products of the Vβ8 and Vβ6 gene in the cells infiltrating the salivary glands. Nucleotide sequencing of amplified TCR Vβ cDNA revealed that T cell clonotypes had a high incidence of identical clones, indicating that the immune response in NOD sialadenitis is driven by common stimuli. These findings suggest that in spontaneous autoimmune sialadenitis of NOD mice, there may be a restricted usage of TCR Vβ elements in the early stage of the autoimmune lesion to recognize self-antigen in organ-specific autoimmune sialadenitis.
Analysis of VH genes rearranged by individual B cells in dermal infiltrates of patients with mycosis fungoides
Authors: FÖRSTE N , GELLRICH S , GOLEMBOWSKI S , RUTZ S , AUDRING H , STERRY W , JAHN S
Published: November 2003

In patients with cutaneous T cell lymphomas such as mycosis fungoides B cells can frequently be detected in the lymphocytic dermal infiltrate. To analyse their immunoglobulin heavy chain gene repertoire, single B cells were obtained from tissue sections of two typical patients with mycosis fungoides using hydraulic micromanipulation followed by specific amplification of the respective gene segments by single-cell polymerase chain reaction (PCR) technique. A total of 21 VHDJH genes was sequenced. From each individual B cell a single productive VHDJH rearrangement was obtained. There was no clonal relationship detected between any of these rearrangements suggesting polyclonality of the infiltrating B cells. The representation of VH families was in accordance with the germ-line complexity. A remarkably high number of VH genes (5/13 in patient 1; 3/8 in patient 2) was completely or nearly germ-line-identical. Five of seven VH4 family genes were nearly unmutated. On the other hand, most of the VH3 gene family members were somatically mutated in an antigen-driven manner. The proportion of germ-line-identical VH genes, the usage of individual VH, D, JH gene elements, and the pattern of somatic mutations found in the B cells infiltrating skin lesions of patients with mycosis fungoides resembles the peripheral blood repertoire, suggesting a bystander role of these cells.
In vitro effects of mercuric chloride (HgCl2) on human mononuclear cells
Authors: LOFTENIUS A , EKSTRAND J , MÖLLER E
Published: November 2003

Due to the release of the toxic compounds of mercury from amalgam fillings, dental amalgam has been questioned as an adequate restoration material for tooth fillings. HgCl2 has been found to be mitogenic for human blood lymphocytes in vitro. However, activation required much higher concentrations than are ever found in vivo. This study has been initiated to evaluate further the influence of HgCl2 on human immunocompetent cells in vitro. It is found that HgCl2 in a narrow concentration range has the ability to preferentially stimulate the CD4+ T cell subset to blast transformation and DNA synthesis. The reaction, when monitored during days 2–6, is maximal at day 6, and most blasts express the IL-2 receptor (IL-2R), indicating in vitro activation. The CD8+ T cell subset is not affected to the same extent. In addition, HgCl2-induced lymphocyte reactivity is dependent on accessory cells, i.e. CD14+ cells.
Relevance of local Th2-type cytokine mRNA expression in immunocompetent infiltrates in inflamed gingival tissue to periodontal diseases
Authors: Tokoro Y , Matsuki Y , Yamamoto T , Suzuki T , Hara K
Published: November 2003

It has been suggested that the types of inflammatory round cell infiltrates and the divergence in the cytokine production profile by macrophages and helper T cells regulate the course of infectious or inflammatory diseases, including periodontitis and gingivitis. We examined the expression of IL-1α, IL-1β, IL-2, IL-4, IL-5, IL-6 and tumour necrosis factor-alpha (TNF-α) mRNA in the inflamed gingiva by in situ hybridization. The results of single-cell analysis were used as data sets for statistical analyses. The density of cells expressing IL-1α, IL-4 and IL-5 mRNA was higher in periodontitis than in gingivitis. IL-2 mRNA-expressing cells were almost absent in gingivitis specimens. Principal component analysis disclosed three factors explaining 84.8% of the variance: one accounting for 40.5% of the variance and mainly regulated by IL-1α, IL-1β, IL-6 and TNF-α, and two others, explaining 29.9% and 14.4% of the variance, describing the relationship between the types of cytokines derived from macrophages or Th2 type. These results suggest that the cytokines produced by inflammatory cells infiltrating in the gingival tissue are influential on the progression of gingivitis, an acute and reversible inflammatory condition, to chronic and destructive periodontitis. Thus, periodontal disease progression may be regulated by the local cytokine network, and the bias in this network towards a Th2-type cytokine dominance could be an exacerbating factor.
Increased proinflammatory cytokine gene expression in the colonic mucosa of coeliac disease patients in the early period after gluten challenge
Authors: Chowers Y , Marsh M N , De Grandpre L , Nyberg A , Theofilopoulos A N , Kagnoff M F
Published: November 2003

Activation of T cells in the intestinal mucosa in response to gluten exposure is thought to play a key role in the pathogenesis of coeliac disease. Moreover, the response of the rectal mucosa to gluten challenge has been considered a useful predictor of gluten sensitivity in coeliac disease. In the present study, we assessed early changes in the expression of proinflammatory cytokine genes and the T cell receptor (TCR) Vβ repertoire in the rectal mucosa of coeliac disease patients following experimental gluten challenge. Cytokine gene expression was assessed in rectal mucosal biopsies from coeliac disease subjects and controls before and after rectal gluten challenge using quantitative reverse transcription polymerase chain reaction analysis, and the TCR Vβ repertoire was characterized using a multiprobe RNase protection assay. Marked up-regulation of expression of the C-X-C chemokine IL-8, the proinflammatory cytokine IL-1β, and the C-C chemokine monocyte chemotactic protein-1 occurred within 2–4 h of rectal gluten challenge in coeliac disease subjects, but not in controls. Furthermore, these changes occurred in the absence of parallel changes in the expressed repertoire of TCR Vβ genes in the rectal mucosa. Thus, an increased expression of proinflammatory cytokine genes precedes the expansion of antigen-specific T cell populations in the early period following experimental exposure of the rectal mucosa of coeliac disease patients to gluten. These findings provide new insights into pathways that may be involved in the activation or reactivation of coeliac disease.
The effects of vitamin A derivatives on in vitro antibody production by peripheral blood mononuclear cells (PBMC) from normal blood donors and patients with common variable immunodeficiency (CVID)
Authors: Zhang J-G , Morgan L , Spickett G P
Published: November 2003

The underlying nature of the defect of CVID is not understood, and the treatment at present is life-long infusion of replacement immunoglobulin. Attempts have been made to use other therapeutic agents, such as IL-2 and retinoic acid (RA), with mixed results. RA is a morphogenetic signalling molecule related to vitamin A and involved in vertebrate development. We report here our in vitro evaluation of the effects of three vitamin A analogues, 9-cis retinal, 13-cis RA and all-trans RA, on antibody production of PBMC from normal donors and patients with CVID. At 10−5 m, 9-cis retinal strongly augmented IgM production of lymphocytes from normal individuals and to a much lesser extent, mild, non-granulomatous (group C) CVID patients, but IgG production was not affected. In the presence of anti-human IgM and IL-2, 9-cis retinal at 10−5 m elevated IgM and IgG production by normal PBMC, but the effect on PBMC of mild CVID was minimal. The effect of 9-cis retinal was significantly reduced at 10−7 and 10−9 m. Only minimal effects were found using 13-cis RA and all-trans RA under these conditions. No detectable antibody production was found in severe, granulomatous (group A) CVID patients under any conditions tested. Taking all data into account, 9-cis retinal is the most potent stimulator for antibody production compared with 13-cis RA and all-trans RA as tested in this in vitro study.
Antibodies to endothelial cells in Kawasaki disease lyse endothelial cells without cytokine pretreatment
Authors: Fujieda M , Oishi N , Kurashige T
Published: November 2003

Kawasaki disease (KD) is characterized by diffuse vasculitis and marked immune activation. To confirm the presence of antiendothelial cell antibodies (AECA) and cytotoxicity of AECA, we investigated the presence of AECA using ELISA and cytotoxicity of AECA in KD. Sera from patients with acute KD were assessed for binding to human umbilical vein endothelial cells (HUVEC) using a cell-based ELISA, and for cytotoxicity against HUVEC as indicated by the conversion of a tetrazolium salt (MTT) into a coloured product. IgM AECA were detected in 8/19 KD sera, IgG AECA were detected in 5/19 KD sera. Significant differences in both AECA titres existed between patients and febrile and afebrile controls. Six out of 19 patients showed complement-dependent cytotoxicity against HUVEC. Cytotoxicity was significantly enhanced by pretreating HUVEC with tumour necrosis factor (TNF), and reduced by incubation with gammaglobulin. Serum titres of IgM AECA in the KD patients were positively correlated with cytotoxicity. Findings suggest that IgM AECA mediates complement-dependent cytotoxicity against endothelial cells in patients with KD, and gammaglobulin may reduce complement-dependent cytotoxicity of AECA against endothelial cells.
Experimental acute glomerulonephritis induced in the rabbit with a specific streptococcal antigen
Authors: Yoshizawa N , Oshima S , Takeuchi A , Kondo S , Oda T , Shimizu J , Nishiyama J , Ishida A , Nakabayashi I , Tazawa K , Sakurai Y
Published: November 2003

FITC-labelled IgG obtained from patients convalescing from acute poststreptococcal glomerulonephritis (APSGN) stains glomeruli of patients with early APSGN. We previously reported a streptococcal antigen (preabsorbing antigen (PA-Ag)) that preabsorbed the stain out of sera from the convalescent patients and thus prevented glomerular staining. To confirm the nephritogenicity of PA-Ag, we administered up to 40 mg of this antigen to rabbits for 8 days and observed them for up to 9 weeks. Immunohistological analysis showed diffuse and global glomerular staining for C3 without notable staining for γ-globulin. Light microscopic examinations revealed slight to moderate proliferative glomerulonephritis with exudative change. Control rabbits, which received similar doses of bovine serum albumin, did not show significant staining for C3. A transient and significant decrease in CH50 was observed from weeks 3 to 7 (9.7 ± 0.3 U/ml at week 3; normal range 12.9 ± 0.6 U/ml). This experimental model showed a resemblance to immunological and immunohistological features of APSGN in humans. Although the precise mechanisms are yet to be determined, complement activation by PA-Ag seems to hold a key position in this model and in the human disease.
Koilocytosis; an indication for conservative management
Authors: DUDDING N. , SUTTON J. , LANE S.
Published: November 2003

The management of women with mild dyskaryosis continues to be the subject of debate. The management of cases with such smears could be improved if additional morphological features were found to be indicative of low/high grade cervical intraepithelial neoplasia (CIN). Our own experience in the routine audit of the diagnostic accuracy of cervical smears had suggested that cases of dyskaryosis with koilocytosis were associated with a low risk of CINIII. Two hundred and forty‐four cervical smears reported as showing evidence of wart virus infection were reviewed and their subsequent histories noted. Of these smears, 173 demonstrated clear evidence of koilocytosis. Irrespective of the initial grade of dyskaryosis, only 28 cases (16.2%) had underlying CIN or had progressed to CIN over a minimum follow‐up period of 21 months. Only two cases (1.2%) showed evidence of CINIII. Koilocytosis is associated with a low risk of CIN and an especially low risk of CINIII. The management of women with koilocytic smears should be modified accordingly.
Gender differences in resting metabolic rate and noradrenaline kinetics in older individuals
Authors: POEHLMAN E. T. , TOTH M. J. , ADES P. A. , CALLES‐ESCANDON J.
Published: November 2003

The physiological factors mediating gender differences in resting metabolic rate (RMR) in older individuals are presently unclear. We examined the contribution of sympathetic nervous system activity to gender differences in resting metabolic rate in older men and women and its relation to body fat distribution. We performed measurements of noradrenaline (NA) kinetics from infusions of [3H]‐NA, RMR, body fat distribution, body composition, peak Vo2 and dietary intake in 29 older men (69 ± 6 years) and 26 older women (65 ± 5 years). Older men weighed more (P < 0.01) and had a greater fat‐free mass (P < 0.01) and a larger waist circumference (P < 0.01) than older women. Older men had a higher RMR (P < 0.05) than older women, which persisted after controlling for differences in fat‐free mass and fat mass. Older men also showed a greater NA appearance rate (P < 0.01) at rest than older women. The higher NA appearance rate in older men was partly related to their greater waist circumference (r = 0.50, P < 0.01). We explored the sympathetic contribution to gender differences in RMR by statistically controlling for differences in body composition and NA appearance rate. After this procedure, we found no gender differences in adjusted RMR between older men (4.3 ± 0.5 kJ min–1) and older women (4.3 ± 0.4 kJ min–1). Our results suggest that: (a) older men have a higher RMR than older women independent of differences in body composition; (b) the higher RMR in older men may be partly due to higher levels of sympathetic nervous system activity; (c) the higher sympathetic nervous system activity in older men is partly related to their greater waist circumference, a proxy measure of central body fatness.
Relationship between fasting insulin and cardiovascular risk factors is already present in young men: the Verona Young Men Atherosclerosis Risk Factors Study
Authors: BONORA E. , TARGHER G. , ZENERE M. B. , SAGGIANI F. , CACCIATORI V. , TOSI F. , TRAVIA D. , ZENTI M. G. , BRANZI P. , SANTI L. , MUGGEO M.
Published: November 2003

The associations between fasting plasma insulin concentration and risk factors for cardiovascular diseases were examined in 979 18‐year‐old men participating in the Verona Young Men Atherosclerosis Risk Factors Study, a cross‐sectional population‐based study. Body mass index (BMI), waist‐to‐hip ratio (WHR), plasma triglycerides and uric acid concentrations, and blood pressure values significantly increased, and the high‐density lipoprotein (HDL)–total cholesterol ratio decreased, across quartiles of fasting insulin. Total and low‐density lipoprotein cholesterol concentrations did not change significantly with the increase in fasting insulin levels. After adjustment for BMI, WHR, smoking, alcohol intake and physical activity, only plasma triglycerides significantly increased across insulin quartiles (F =7.1; P <0.001). However, systolic blood pressure and uric acid were close to statistical significance (P =0.06–0.07). Multiple linear regression analysis confirmed that plasma insulin was independently correlated with plasma triglycerides and, to a lesser extent, with blood pressure and uric acid concentration. This analysis pointed out that BMI was a stronger independent predictor of all cardiovascular disease risk factors than fasting insulin. When subjects were categorized according to the number of metabolic and haemodynamic disorders occurring within the same individual, subjects with multiple disorders (i.e. three or four) had higher plasma insulin levels than those with none or few disorders, even after adjusting for BMI, WHR and behavioural variables (F =4.0; P <0.01). These results indicate that hyperinsulinaemia is already associated with a cluster of cardiovascular disease risk factors in young adulthood, the strongest independent association being with plasma triglycerides.
Lecithin–cholesterol acyltransferase activity in normocholesterolaemic and hypercholesterolaemic roosters: modulation by lipid apheresis
Authors: KOSTNER K. M. , SMITH J. L. , DWIVEDY A. K. , SHAFEY T. M. , FANG N. X. , MAHON M. G. , IANNUZZI C. I. , COLQUHOUN D. M. , CHAM B. E.
Published: November 2003

Lipid apheresis, a recently described procedure for the elimination of lipid but not apolipoproteins from plasma, was applied to normocholesterolaemic and hypercholesterolaemic roosters. Lipid apheresis resulted in an immediate reduction in plasma unesterified cholesterol concentration, which was sustained for 150 min. The reduction in unesterified cholesterol concentration was higher in the normocholesterolaemic animals than in the hypercholesterolaemic animals. Lipid apheresis induced changes in the ratio of plasma unesterified to total cholesterol in normocholesterolaemic animals but not in hypercholesterolaemic animals. In hypercholesterolaemic animals, lecithin–cholesterol acyltransferase (LCAT) activity was not affected by lipid apheresis, whereas in normocholesterolaemic animals LCAT activity was acutely reduced for 150 min after lipid apheresis. Saturated LCAT kinetics occurred in the hypercholesterolaemic animals but not in the normocholesterolaemic animals. LCAT obeyed Michaelis–Menten kinetics. After lipid apheresis, there was a pool of unesterified cholesterol that was available as substrate for LCAT to a greater extent in hypercholesterolaemic animals than in normocholesterolaemic animals. These observations may have important implications for lipid apheresis as a treatment for atherosclerosis.
Gender, light and water effects in carbon isotope discrimination, and growth rates in the dioecious tree Ilex aquifolium
Authors: Retuerto R. , Lema B. Fernandez , Roiloa S. Rodriguez , Obeso J. R.
Published: November 2003

1. Detailed understanding of the specific physiology of sexes in dioecious species is required to explain patterns in gender dimorphism. Under controlled‐environment conditions we tested the hypothesis that sexes of the dioecious tree holly Ilex aquifolium L. (Aquifoliaceae) differed in growth and long‐term potential water‐use efficiency, as measured by carbon isotope discrimination (Δ13C), and that these differences were dependent on the environmental context.

2. Patterns of response in Δ13C to the various combinations of light and water were gender‐specific. Under more xeric conditions, females maintained significantly higher Δ13C than males.

3. Female plants exhibited significantly greater relative diameter growth rates than male plants.

4. As expected, Δ13C significantly increased with decreasing irradiance, and decreased with increasing limitation in water supply. Light and water effects were not independent, with a more pronounced drought effect in decreasing leaf Δ13C under unshaded than under shaded conditions.

5. Our results suggest that between‐sex differences in physiology are context‐dependent. Future studies attempting to assess gender dimorphism should take more account of gender‐specific interactions with the environment. Gender‐specific efficiency in water use could play a decisive role in explaining gender differences in growth and ecological interactions.
The small GTP‐binding protein Rho1 is a multifunctional protein that regulates actin localization, cell polarity, and septum formation in the fission yeast Schizosaccharomyces pombe
Authors: Nakano Kentaro , Arai Ritsuko , Mabuchi Issei
Published: November 2003

Background:

The small GTP‐binding protein Rho has been shown to regulate the formation of the actin cytoskeleton in animal cells. We have previously isolated two rho genes, rho1+ and rho2+ , from the fission yeast Schizosaccharomyces pombe in order to investigate the function of Rho using genetic techniques. In this paper, we report the cellular function of Rho1.

Results:

We found that Rho1 is essential for cell viability and cell polarity using gene disruption and by exogenous expression of botulinum C3 ADP‐ribosyltransferase. In cells expressing either a constitutively active Rho1 or a dominant‐negative Rho1, actin patches were delocalized. Both the cell wall and secondary septum were thick and stratified in cells expressing the constitutively active Rho1, while the cell wall of cells expressing the dominant‐negative Rho1 seemed to be loosely organized. Furthermore, inactivation of Rho1 is apparently required for the separation of daughter cells. Cell fractionation studies suggested that Rho1 is predominantly membrane‐bound. Moreover, we observed that Rho1 is localized to the cell periphery and to the septum.

Conclusions:

Rho1 is involved in actin patch localization, the control of cell polarity, the regulation of septation, and cell wall synthesis.
Primary spinal paragangliomas: a clinicopathological and immunohistochemical study of 30 cases
Authors: MORAN C.A. , RUSH W. , MENA H.
Published: November 2003

Aims: Extra‐adrenal paragliomas are neoplasms which have been the subject of much debate regarding parameters to establish their biological behaviour. This study describes the clinicopathological and immunohistochemical features of 30 cases of spinal paragliomas. Methods and results: There were 15 male and 15 female patients. The median age at diagnosis was 46 years (range 20–74 years). Fourteen patients presented with back pain, two with numbness of the lower extremities, one with difficulty in walking and one with spinal cord compression. Nineteen tumours were located in the lumbar region, six in the cauda equina, two in the filum terminale, two in the thoracic region and one in the cervical region. All patients underwent gross total excision. The size of the tumours ranged from 10 to 50 mm. Histologically, 18 neoplasms showed alveolar (Zellballen) pattern, seven a spindle component, two eosinophilic granular cells suggestive of oncocytic metaplasia, two melanin pigment and one ganglion cells. Positive immunohistochemical results include: neuron‐specific enolase 23/23 (100%), synaptophysin 21/23 (91%), S100 protein 22/23 (95%, sustentacular cells), leu‐enkephalin 11/23 (47%), somatostatin 8/23 (34%), focal glial fibrillary acidic protein 7/23 (30%), focal keratin 5/23 (21%), neurofilament proteins 3/23 (13%) and adrenocorticotrophic hormine (ACTH) 1/23 (4%). Follow‐up information obtained in 20 patients show 17 patients alive over a period of 6–216 months. One patient had bone metastases. Two patients died of unrelated causes, including one of congestive heart failure and one of myocardial infarction. Conclusions: In our experience, spinal paragangliomas behave as slow‐growing tumours susceptible to potential cure by total excision. We agree with the current World Health Organization (WHO) classification as grade I tumours. Less than 1% may be locally aggressive. Spinal paragangliomas immunoreact not only for conventional neuroendocrine markers but also for peptides including somatostatin and ACTH and focally for the epithelial marker keratin.
Improved prognosis of solid‐type poorly differentiated colorectal adenocarcinoma: a clinicopathological and immunohistochemical study
Authors: SUGAO Y. , YAO T. , KUBO C. , TSUNEYOSHI M.
Published: November 2003

Aims: We rarely encounter solid‐type poorly differentiated colorectal carcinoma, and their histogenesis and biological behaviour are not fully disclosed. Methods and results: A review of 60 poorly differentiated carcinomas of the colorectum was undertaken, 36 (59%) of which were located in the right side of the colorectum. Although, on the basis of the World Health Organization (WHO) classification solid carcinomas are included among undifferentiated carcinomas, the poorly differentiated carcinomas were divided into four types; 27 solid carcinomas (Sol.), 17 poorly differentiated adenocarcinomas (PDA), six signet‐ring cell carcinomas (Sig.) and 10 mucinous carcinomas (Muc.). Solid carcinomas revealed a solid alveolar growth of fairly uniformly sized tumour cells with occasional mitotic figures. This type of tumour had a relatively lower percentage of lymphatic permeation and lymph node metastasis compared with the other three types. The 5‐year survival rates were 31% for all poorly differentiated carcinomas, 47% for the Sol. type, 32% for the PDA type, and 0% for both the Sig. and the Muc. types, with a rate of 72% for well‐differentiated adenocarcinomas selected as controls. Immunohistochemically, bcl‐2 protein expression was demonstrated in 38% of the Sol. type, but in only 12 % of the other three non‐solid types, this difference being significant (P < 0.05). Conclusions: These findings suggest that solid carcinomas of the colorectum should be regarded as a distinct type of poorly differentiated carcinoma, leading to a good prognosis.
Immunohistochemical staining of hepatocellular carcinoma with monoclonal antibody against inhibin
Authors: McCLUGGAGE W.G. , MAXWELL P. , PATTERSON A. , SLOAN J.M.
Published: November 2003

Inhibin is a peptide hormone produced by ovarian granulosa cells. During a recent study investigating the immunohistochemical staining of ovarian granulosa cell tumours and other neoplasms with an anti‐inhibin monoclonal antibody, we identified strong cytoplasmic staining of hepatocytes. In the present study we investigated the immunostaining of hepatocellular carcinoma and other neoplasms involving the liver with anti‐inhibin to determine whether the antibody may be of value in the differential diagnosis of hepatic neoplasms. Immunostaining for α‐fetoprotein was also performed. With anti‐inhibin there was positive, generally strong, cytoplasmic staining of 17 of 19 cases of hepatocellular carcinoma, including the pleomorphic and glandular variants. There was positive staining of six of 20 cases of adenocarcinoma. In these, positive staining was generally focal, of weak intensity and involved the luminal surface of neoplastic glands. There was no staining of five cases of neuroendocrine tumour. There was positive staining for α‐fetoprotein in 13 of 19 cases of hepatocellular carcinoma and in two of 20 cases of adenocarcinoma but no staining of neuroendocrine tumours. Immunostaining with anti‐inhibin antibody may be of value in the differentiation of hepatocellular carcinoma from other neoplasms involving the liver. The antibody is a more sensitive, but less specific, immunohistochemical marker for hepatocellular carcinoma than is α‐fetoprotein.
Metastatic carcinoma of the prostate presenting as parotid tumour
Authors: SIMPSON R.H.W. , SKÁLOVÁ A.
Published: November 2003

Two patients are described in each of whom metastatic prostatic carcinoma presented as a tumour of the parotid gland. In one, primary prostate cancer had been diagnosed three years prior to the appearance of what was believed clinically to be an unrelated salivary lesion. In the other a neck swelling was the first indication of disease, although subsequent investigation revealed widespread metastases. Biopsy of each tumour showed an adenocarcinoma with immunohistochemical expression of prostate markers. These cases, and the six previous reports, illustrate the need to consider metastatic carcinoma from the prostate in the assessment of any malignant tumour of the salivary glands in men of advancing years.
The in vivo antibody response against exogenous antigens is not influenced by the mouse Bcg (Nramp1) gene
Authors: ŠÍROVÁ M. , HOVORKA O. , ŘÍHA I. , ŘÍHOVÁ B. , BAUDYŠ M. , KIM S.‐W. , SKAMENE E.
Published: November 2003

The mouse Nramp1 (Bcg) gene on chromosome 1 exerts pleiotropic effects on macrophage function. The gene is known to affect presentation of mycobacteria, and other antigens in vitro, so that macrophages carrying the resistant Bcg allele better support the proliferation of antigen‐specific T cells compared with macrophages of the sensitive phenotype. To determine whether the Bcg allele could affect in vivo the antibody response to antigens not related to mycobacterial infections, we tested the primary and secondary responses to sheep red blood cells (SRBC) and glycosylated bovine insulin (G‐insulin) in two pairs of Bcg congenic strains: BALB/c (Bcgs) versus BALB/c.CD2 (Bcgr), and B10.A (Bcgs) versus B10Ar (Bcgr), and in C57BL/10ScSn (B10; Bcgs) and A/J (Bcgr) mice. Furthermore, the antigen‐specific proliferative responses of T cells primed in vivo by protein antigens were also tested in Bcg congenic mice. We found no significant difference in in vivo antibody response either to SRBC or G‐insulin between the Bcgrand Bcgs strains. The magnitude of in vitro antigen‐specific proliferation of lymph node cells sensitized in vivo by hen egg lysozyme (HEL) or chicken ovalbumin (OVA) was also similar in Bcgs and Bcgr congenic mice. However, we have documented a higher antigen‐presenting capacity of Bcgr macrophages in in vitro antigen‐specific proliferation to OVA. Since the macrophages are the only cells in which the Nramp1 gene is expressed, we suggest that the activity of other types of antigen‐presenting cells masks the effect of the Bcgr allele on antigen‐presentation in vivo.
Phenotypic and functional characterization of a new human macrophage cell line K1m demonstrating immunophagocytic activity and signalling through HLA class II
Authors: DIALYNAS D. P. , LEE M.‐J. , SHAO L.‐E. , TAN P. C. , YU J.
Published: November 2003

A human macrophage line, designated K1m, has been established from peripheral blood. K1m expresses a number of lineage‐specific markers as well as a broad array of intercellular adhesion molecules. In particular, K1m expresses high levels of human leucocyte antigen (HLA) class I and class II. In response to ligation of HLA class II (HLA‐DR), but not in response to ligation of HLA class I, K1m forms tighter homotypic aggregates and develops a striking ‘stellate’ culture phenotype. K1m also expresses Fc receptors for immunoglobulin G (IgG) (CD64, CD32, and CD16) and can be shown to phagocytose polystyrene latex beads, as well as neuroblastoma cells in the presence of tumour‐specific monoclonal antibody (mAb). The K1m cell line should therefore prove useful for studying both signalling through macrophage HLA class II and immunophagocytosis.
Rejection of cardiac allografts by T cells expressing a restricted repertoire of T‐cell receptor Vβ genes
Authors: SHIRWAN H. , BARWARI L. , CRAMER D. V.
Published: November 2003

We have recently shown that T cells infiltrating cardiac allografts early in graft rejection use a limited T‐cell receptor (TCR) Vβ repertoire. In this study we tested whether this limited repertoire of Vβ genes is important for graft rejection. A cell line, AL2‐L3, was established from LEW lymphocytes infiltrating ACI heart allografts 2 days after transplantation. This cell line is composed of CD4+ T cells that primarily recognize the class II RT1.B major histocompatibility complex (MHC) molecule expressed by the donor graft. This cell line precipitated acute rejection of donor hearts with a median survival time (MST) of 10·5 days following adoptive transfer to sublethally irradiated LEW recipients. This rate of graft rejection was significantly (P<0·0007) accelerated when compared with a MST of 60 days for allografts in irradiated control recipients. The AL2‐L3‐mediated acceleration of graft rejection was donor specific as WF third‐party heart allografts were rejected with a delayed tempo (MST=28·5 days). The Vβ repertoire of this cell line was primarily restricted to the expression of Vβ4, 15 and 19 genes. The nucleotide sequence analysis of the β‐chain cDNAs from this cell line demonstrated that the restricted use of the V gene repertoire was not shared with the N, D and J regions. A wide variety of CDR3 loops and Jβ genes were used in association with selected Vβ genes. These data provide evidence for the role a restricted repertoire of Vβ genes plays in cardiac allograft rejection in this model. The restricted usage of the Vβ repertoire in an early T‐cell response to allografts may provide the opportunity to therapeutically disrupt the rejection reaction by targeting selected T‐cell populations for elimination at the time of organ transplantation.
Differential G‐protein expression during B‐ and T‐cell development
Authors: GRANT K. R. , HARNETT W. , MILLIGAN G. , HARNETT M. M.
Published: November 2003

The molecular mechanisms underlying B‐ and T‐cell development are, as yet, poorly understood. However, as G proteins regulate a diverse range of biological responses including growth, proliferation and differentiation, we have investigated differential expression of G proteins during B‐ and T‐cell development with the aim of identifying key signals involved in lymphocyte maturation. Differential expression of β1/2 and α‐subunits of the Gs‐, i‐ and q‐families was found throughout lymphoid development. Most strikingly, Gαi1 and Gα11 were very weakly, or not expressed in pre‐, immature and mature B cells, thymocytes or mature T cells, but strongly induced in mature B‐lymphoblastoid cell lines, some of which have been used as models of germinal centre B cells, suggesting that expression of these G proteins may correlate with the later stages of B‐cell development. In contrast, Gα16 expression was highest in T cells and pre‐B cells and progressively declined with B‐cell maturation. These findings suggest that G proteins, and the signals they regulate, such as ion channels and/or adenylate cyclase (Gαs/i ) and phospholipase C (Gβγ and Gα11/16) are differentially regulated in lymphoid cells in a maturation‐ and lineage‐dependent manner.
Langerhans’ cells produce type IV collagenase (MMP‐9) following epicutaneous stimulation with haptens
Authors: KOBAYASHI Y.
Published: November 2003

For initiation of the contact hypersensitivity response, epidermal Langerhans’ cells (LC) migrate from the epidermis to draining nodes via afferent lymphatics by passing through the basement membrane. In this study, we examined production of matrix metalloproteinases (MMPs) in LC‐enriched epidermal cells to clarify the type of enzymes involved in LC transmigration through the basement membrane. Using gelatine enzymography and immunoblotting analysis, 95 000 MW type IV collagenase (MMP‐9) was found to be produced by LC‐enriched epidermal cells. Analysis of the kinetics of MMP‐9 expression showed that its production was induced within 6 hr after application of 2,4,6‐trinitrochlorobenzene (TNCB), substantially increased between 12 hr and 24 hr, and then decreased to the normal level by 7 to 10 days. Other haptens, such as 2,4‐dinitrochlorobenzene and 2,4‐dinitrofluorobenzene, also induced MMP‐9 expression. Fluoroescence‐activated cell sorter analysis revealed that LC were one of the major cell types to express MMP‐9 in response to TNCB. In addition, highly enriched LC from sensitized skin were shown to express strong gelatinolytic activity. These results indicate that LC by themselves, as well as other epidermal cells, are capable of producing MMP‐9, and suggest that MMP‐9 may contribute to proteolysis associated with transmigration of LC in the induction phase of contact dermatitis.
Microorganisms in closed periapical lesions
Authors: Abou‐Rass M. , Bogen G.
Published: November 2003

The purpose of this study was to investigate the microorganisms of strictly selected closed periapical lesions associated with both refractory endodontic therapy and pulpal calcification. Definitive criteria were established that assured complete clinical isolation of the periapical lesion from the oral and periodontal environment. A total of 13 criteria‐referenced lesions were selected from 70 patients with endodontic surgical indications. A well controlled culturing method was used in all cases and samples were taken by one clinician at three separate sites during each surgery. Samples taken at the surgical window and within the body of the lesion served as controls, whilst a third sample was taken at the apex. In all 13 cases, samples taken from the apex yielded microorganisms comprising 63.6% obligate anaerobes and 36.4% facultative anaerobes. Prevalence of the isolated species was 31.8% for Actinomyces sp., 22.7%, Propionibacterium sp., 18.2%Streptococcus sp., 13.6%Staphlyococcus sp., 4.6%Porphyromonas gingivalis, 4.6%Peptostreptococcus micros and 4.6% Gram‐negative enterics. The results of this investigation indicate that closed periapical lesions associated with calcified teeth or those resistant to root canal treatment harbour bacteria. The inability to eradicate all root canal microorganisms during root canal treatment, along with anatomical factors, may allow further bacterial colonization of the root apex and surrounding periapical tissues, and consequently prevent healing.
Root positioning and leakage to dye in extracted teeth using reduced pressure
Authors: Katz A. , Rosenwasser R. , Tamse A.
Published: November 2003

A study was conducted in vitro to assess the root apex position on leakage to dye under reduced pressure (560 mm Hg). A total of 132 extracted single‐rooted teeth were randomly allocated to four groups. The teeth were prepared using a step‐back technique and sealed with the cold lateral condensation technique, using gutta‐percha without sealer. No significant differences (P < 0.813) were found between horizontally positioned experimental groups under reduced pressure and those in passive immersion. In the group in which the apices were in an upright position, the mean leakage was significantly different (P < 0.024) from the mean dye penetration in the group of teeth positioned horizontally under reduced pressure. Those teeth with apices upright allowed the most linear leakage (0.51 mm), whereas the groups in which the roots were horizontally positioned (with and without reduced pressure) had the least dye penetration (0.21 mm and 0.19 mm, respectively). These results show that tooth positioning has a significant effect on linear dye penetration under reduced pressure and emphasize the need to standardize factors that may influence penetration when assessing leakage study methodology.
Infra‐red microspectroscopy of hydrated biological systems: design and construction of a new cell with atmospheric control for the study of plant cell walls
Authors: Chen L. , Wilson R. H. , McCann M. C.
Published: November 2003

A new hydration cell has been constructed that allows wet biological samples, or samples of controlled moisture content, to be analysed in situ using infra‐red microspectroscopy. The cell has been used to show that there are minor spectral changes associated with the hydration of pectin and tomato pericarp cell walls and slightly more significant changes in onion and carrot epidermal walls. The cell was also used to show that molecular orientations of polymers, previously observed in dry cell walls, were also to be seen in hydrated walls. For cell walls of onion and carrot epidermis, it was shown that the orientations of cell wall polymers are not affected by hydration. Furthermore, the polymer orientations in cell walls of fully elongated onion epidermal cells are different from those of elongating carrot epidermis. By using the hydration cell, it is now possible to investigate both fresh samples and wet systems routinely. The applications of this to the study of biological materials with infra‐red microspectroscopy are discussed.
Co‐localization analysis of complex formation among membrane proteins by computerized fluorescence microscopy: application to immunofluorescence co‐patching studies
Authors: Lachmanovich E. , Shvartsman D. E. , Malka Y. , Botvin C. , Henis Y. I. , Weiss A. M.
Published: November 2003

Fluorescence imaging of two independently labelled proteins is commonly used to determine their co‐localization in cells. Antibody‐mediated crosslinking can mediate the patching of such proteins at the cell surface, and their co‐localization can serve to determine complex formation among them. However, manual analysis of such studies is both tedious and subjective. Here we present a digital co‐localization analysis that is independent of the fluorescence intensity, is highly consistent and reproducible between observers, and dramatically reduces the analysis time. The approach presented is based on a segmentation procedure that creates binary objects, and then determines whether objects belonging to two different groups (e.g. green‐ and red‐labelled) are co‐localized. Two methods are used to determine co‐localization. The ‘overlap’ analysis defines two objects as co‐localized if the centre of mass of one falls within the area of the other. The ‘nearest‐neighbour distance’ analysis considers two objects as co‐localized if their centres are within a threshold distance determined by the imaging modality. To test the significance of the results, the analysis of the actual images is tested against randomized images generated by a method that creates images with uncorrelated distributions of objects from the two groups. The applicability of the algorithms presented to study protein interactions in live cells is demonstrated by co‐patching studies on influenza haemagglutinin mutants that do or do not associate into mutual oligomers at the cell surface via binding to AP‐2 adaptor complexes. The approach presented is potentially applicable to studies of co‐localization by other methods (e.g. electron microscopy), and the nearest‐neighbour distance method can also be adapted to study phenomena of correlated placement.
Monitoring cell movements and volume changes with pulse‐mode scanning ion conductance microscopy
Authors: Happel P. , Hoffmann G. , Mann S. A. , Dietzel I. D.
Published: November 2003

Here we describe the use of pulse‐mode scanning ion conductance microscopy (SICM) to observe volume changes and cell membrane movements during the locomotion of cultured cells in the range of minutes to several hours. The microscope is based on the pulse‐mode SICM previously developed for stable imaging of single cells in culture. Our instrument uses current pulses to control the distance between cell surface and electrode tip as well as a back‐step mode to prevent contact of tip and membrane during lateral movements of the probe. We performed repeated scans of cell surfaces using feedback‐controlled piezoactors to position the electrode. Using patch‐clamp‐type electrode tips the height of cells could reproducibly be measured with a standard deviation of 50 nm. To quantify and separate changes in cell position and volume occurring between consecutive scans, a program was written to subtract images and calculate volume changes. Examples of repeated scans show that membrane movements in the range of 30 min to a few hours can be quantitatively monitored with a lateral resolution of 500 nm using difference images and that faster movements in the range of minutes can be recorded at defined cell sections using the line scan mode. Difference images indicate that volume changes can affect cell surfaces inhomogeneously, emphazising the role of the cytoskeleton in the stabilization of cell shape.
Antibody responses to Rhoptry‐Associated Protein‐1 (RAP‐1) of Plasmodium falciparum parasites in humans from areas of different malaria endemicity
Authors: JAKOBSEN P.H. , KURTZHALS J.A.L. , RILEY E.M. , HVIID L. , THEANDER T.G. , MORRIS‐JONES S. , JENSEN J.B. , BAYOUMI R.A.L. , RIDLEY R.G. , GREENWOOD B.M.
Published: November 2003

Plasma IgM and IgG antibody reactivities against the recombinant Plasmodium falciparum protein, Rhoptry Associated Protein‐1 (rRAP‐1) were measured by ELISA in individuals from Sudan, Indonesia, Kenya and The Gambia living in areas of different malaria endemicity. IgG and IgM reactivities to rRAP‐1 increased with malaria endemicity. IgG reactivities were associated with spleen rates in Indonesia with high malaria endemicity while IgM reactivities were associated with spleen rates in Kenya with low malaria endemicity. IgG and IgM reactivities to rRAP‐1 increased during acute episodes of P. falciparum malaria in Sudanese adults and IgG reactivities remained high one month after treatment in all adults tested. Antibody reactivities to rRAP‐1 in Gambian children in the dry season were higher in children with parasitaemia than in children without detectable parasitaemia. Antibody reactivities were not associated with protection against clinical episodes in the following rainy season but higher antibody reactivities were detectable at the end of the rainy season. There was no difference in antibody reactivity to rRAP‐1 between Gambian children with mild or severe malaria.
Antigenic variation during malaria infection—the contribution from the murine parasite Plasmodium chabaudi
Authors: PHILLIPS R.S. , BRANNAN L.R. , BALMER P. , NEUVILLE P.
Published: November 2003

P. chabaudi AS strain in laboratory mice provides an accessible and useful model for investigating antigenic variation in malaria parasites. Evidence that P. chabaudi AS undergoes antigenic variation is summarized. A live indirect fluorescent test (IFAT) detects a variable antigen on the surface of schizont‐infected erythrocytes. Five different variable antigen types (VATS) (detected using the live IFAT) are described from a cloned mosquito transmitted parent population. Even during the rising primary parasitaemia VATS switch at high and variable rates (1.2–1.6%). Work towards cloning genes coding for the variable antigen is briefly summarized. Acquired immunity to blood‐stage P. chabaudi AS is initially mediated through Th1 CD4+ T cells and after the primary parasitaemia there is a switch to Th2 CD4+ T cells. Acquired immune effector mechanisms are discussed in the context of antigenic variation by the parasite.