bluuebunny/crossref_metadata_2025 huggingface dataset:
Effects of temperature and relative humidity on conidial germination and viability, colonization and sporulation ofMonilinia fructigena
Authors: Xu X.‐M. , Guerin L. , Robinson J. D.
Published: March 2003

Experiments were conducted to investigate the effects of temperature and relative humidity (RH) on thein vitrogermination and viability of conidia of the apple brown rot fungus (Monilinia fructigena), and on colonization and sporulation on detached fruits byM. fructigena. Conidia only germinated under near‐saturation humidity (≥ 97% RH) and the rate of germination initially increased with temperature to a maximum at ≈ 23–25°C and then decreased. Conidia germinated rapidly – more than 70% of viable conidia had germinated within 2 h at 20 and 25°C. The rate of colonization on detached fruits increased log‐linearly with increasing temperature. Sporulation on detached fruits was not observed at 5 or 25°C; sporulation appeared to be unaffected by either temperature (10–20°C) or RH (45–98%) once infection was established. Detached conidia remained viable for a long period of time, up to 20 days, the longest assessment time in this study, depending on storage temperature (10 or 20°C) and RH (45 or 85%). Temperature appeared to be more important than RH in affecting conidial viability. Low temperature and high RH resulted in reduced loss of conidial viability. Storage at 10°C and 85% RH for up to 20 days appeared not to affect conidial viability. These results indicate that environmental conditions during the main UK growing seasons are unlikely to be limiting factors for the development of brown rot on apple.
Biological and sequence comparisons of Potato virus Y isolates associated with potato tuber necrotic ringspot disease
Authors: Boonham N. , Walsh K. , Hims M. , Preston S. , North J. , Barker I.
Published: March 2003

The biological and molecular relationships between a large number of Potato virus Y (PVY) isolates were examined, concentrating mainly on isolates associated with potato tuber necrotic ringspot disease (PTNRD). Following detailed analysis of the coat‐protein gene, four main groups were identified which broadly corresponded to the phenotype of the different isolates. The groups comprised the ordinary strain (PVYO), the necrotic strain (PVYN), the C strain (PVYC) and a group of recombinant (between ordinary and necrotic) isolates. In the latter group, all members were associated with PTNRD. However, four nonrecombinant isolates were also identified which were associated with PTNRD or tuber necrosis. Three were from tubers showing PTNRD symptoms in the field, while the fourth originated from symptomless tubers, but could cause necrotic rings on tubers under glasshouse conditions. The results show that although coat‐protein recombination is always found associated with the PTNRD phenotype, some nonrecombinant isolates have very similar biological properties.
Detection of Citrus psorosis virus in field trees by direct tissue blot immunoassay in comparison with ELISA, symptomatology, biological indexing and cross‐protection tests
Authors: Martín S. , Alioto D. , Milne R. G. , Guerri J. , Moreno P.
Published: March 2003

Serological detection of Citrus psorosis virus (CPsV) by direct tissue blot immunoassay (DTBIA) and by double (DAS) and triple (TAS) antibody sandwich ELISA, was compared in samples from various citrus varieties growing in the glasshouse and in the field. In young shoots and leaves, CPsV was readily detected by the three procedures, whereas DTBIA detection in old leaves was less consistent. DTBIA detection and ELISA readings in nine different citrus varieties were similar, suggesting that CPsV accumulates to equivalent levels in all of them. In infected field trees from Spain or Italy, CPsV was consistently detected by TAS ELISA, even in samples of old leaves in winter, whereas DTBIA detection in the same trees was reliable only when using young shoots. Detection of CPsV by DTBIA and by DAS and TAS ELISA in previously untested field trees correlated perfectly with psorosis diagnostics based on biological indexing, specifically with the capacity of those sources to cross‐protect against challenge inoculation with psorosis B. Some trees without bark scaling were shown to be psorosis‐infected by biological indexing and to contain CPsV by serological tests; other trees showing psorosis‐like bark or leaf symptoms in the field were shown to be psorosis‐free by biological indexing and also CPsV‐free by serology. This is the first time that the presence of CPsV has been correlated with psorosis infection as diagnosed by biological indexing.
Development of a PCR‐based diagnostic test for Spongospora subterranea f.sp. nasturtii, the causal agent of crook root of watercress (Rorippa nasturtium‐aquaticum)
Authors: Down G. J. , Clarkson J. M.
Published: March 2003

The polymerase chain reaction (PCR) technique was utilized to obtain internal transcribed spacer ribosomal DNA (ITS rDNA) and small‐subunit (18S) rDNA sequences from UK isolates of Spongospora subterranea f.sp. nasturtii, a plasmodiophorid pathogen of watercress (Rorippa nasturtium‐aquaticum). ITS sequence data obtained from S. subterranea isolated from a range of UK sites were found to be identical. PCR primers were designed using these sequences and were shown to be capable of specific amplification of S. subterranea f.sp. nasturtii DNA from plant tissue and from water samples containing zoospores of the pathogen. As little as 5 ng total genomic DNA from infected plant material, or 1000 zoospores, was required for consistently successful amplification of DNA. A filtration‐based method for obtaining pathogen DNA for PCR from watercress‐bed water was developed.
Foliar aggressiveness of Northern Ireland isolates ofPhytophthora infestanson detached leaflets of three potato cultivars
Authors: Carlisle D. J. , Cooke L. R. , Watson S. , Brown A. E.
Published: March 2003

The aggressiveness of 20 Northern Ireland single‐lesion isolates ofPhytophthora infestanswas compared following their inoculation onto detached leaflets of three potato cultivars chosen on the basis of their differing levels of race‐nonspecific resistance to late blight: Bintje (highly susceptible); Cara (moderately resistant); and Stirling (more resistant). Five isolates from outside Northern Ireland were included for comparative purposes: two from the Republic of Ireland; two from the USA (representing the US‐1 and US‐8 clonal lineages); and one from Mexico. To control the variation between tests, a balanced incomplete block design was used, as opposed to either a complete block design or the method of inclusion of standard isolates, where such variation would have increased the error. Highly significant variation for disease parameters, including latent period, infection frequency, area under the lesion expansion curve (AULEC) and sporulation capacity, was found between isolates. These differences were much more marked on the cultivars exhibiting higher levels of race‐nonspecific resistance. There was a significant interaction between isolate and cultivar for all parameters assessed and, overall, no one isolate was the most aggressive across all three potato cultivars. However, a group comprising seven of the 20 Northern Ireland isolates was consistently found to exhibit the highest levels of aggression towards all three cultivars for each of the disease parameters. These results demonstrate that significant variation for foliar aggressiveness exists within the Northern Ireland population ofP. infestans, and indicate the importance of selecting appropriately aggressive isolates for evaluation of host resistance to late blight within breeding programmes.
Dynamics of Polymyxa graminis and Indian peanut clump virus (IPCV) infection on various monocotyledonous crops and groundnut during the rainy season
Authors: Delfosse P. , Reddy A. S. , Thirumala Devi K. , Legrève A. , Risopoulos J. , Doucet D. , Shoba Devi P. , Maraite H. , Reddy D. V. R.
Published: March 2003

The progress of Indian peanut clump virus (Hyderabad isolate; IPCV‐H) and its vector Polymyxa graminis in various monocotyledonous crops and groundnut was studied during the 1994, 1995 and 1996 rainy seasons in a naturally infested field in India. The roles of rainfall and temperature in the dynamics of infection by both the virus and its vector were analysed by exposing young seedlings for short periods in the field. Of the host crops studied, wheat, followed by barley, showed the highest virus incidence, although P. graminis was rarely observed in roots of wheat and was not detected in those of barley. The roots of maize, pearl millet and sorghum plants infected by P. graminis showed intense colonization by sporosori. IPCV accumulated in systemically infected maize plants; the sorghum and pearl millet cultivars studied showed a transient presence of IPCV‐H. Rice was seldom infected by the virus and P. graminis was not detected in its roots. Groundnut was a systemic host for the virus, although during these experiments no P. graminis was found in its roots. Groundnut appeared to be susceptible to infection, mostly in the early stages of crop development, and the rate of IPCV‐H transmission in groundnut seeds was highest (13%) for plants infected when young. The seed‐transmission rate quickly decreased in plants showing symptoms 1 month after sowing. Time of infection had little influence on groundnut pod yield, which was always reduced by >60% in infected plants. There was some evidence that the quantity and distribution of rainfall influenced the incidences of IPCV‐H and P. graminis: high rainfall resulted in high incidences of the virus and P. graminis, and a weekly rainfall of 14 mm was sufficient for P. graminis to initiate infection. Temperatures prevailing during the rainy season ranged from 23 to 30°C and were found to be conducive to natural virus transmission. These results suggest measures to be explored for controlling peanut clump disease.
Biochemical and molecular characterization of Bacillus amyloliquefaciens, B. subtilis and B. pumilus isolates with distinct antagonistic potential against Xanthomonas campestris pv. campestris
Authors: Wulff E. G. , Mguni C. M. , Mansfeld‐Giese K. , Fels J. , Lübeck M. , Hockenhull J.
Published: March 2003

Fifty‐one Bacillus isolates were characterized by fatty acid methyl ester (FAME) analysis; universal primer polymerase chain reaction (UP‐PCR) fingerprinting; production of secondary metabolites and antagonistic activity against Xanthomonas campestris pv. campestris (causal agent of black rot in cabbage) in vitro and in vivo. Based on FAME analysis and/or PCR fingerprinting, the isolates were clustered into three different groups, named as Bacillus amyloliquefaciens, B. subtilis and B. pumilus. Seed treatment with Bacillus spp. generally reduced germination of seeds and incidence of black rot, but no relationship was found between the results of in vitro and in vivo experiments. The B. amyloliquefaciens group contained isolates that were generally the most effective at reducing attack of black rot in vivo. The metabolic profiles of these isolates suggested that they produced surfactin, iturin, bacillomycine and/or azalomycin F. Isolates belonging to the B. subtilis group were mostly able to synthesize surfactin and arthrobactin. Surfactin, amphomycin, arthrobactin and valinomycin were generally found in culture extracts of isolates belonging to the B. pumilus group. No effect on growth of the pathogen was detected when the activity of filtered culture extracts and selected metabolites produced by the three different Bacillus species was tested in vitro against X. c. pv. campestris. However, inhibition was seen when bacterial liquid cultures were used. When the ability to colonize cabbage endophytically was examined for seven selected isolates with different antagonistic potential against black rot, it was found that the ability was related to the species and not to the antagonistic activity of the isolates.
Cervical Secretions in Pregnant Women Colonized Rectally with Group B Streptococci have High Levels of Antibodies to Serotype III Polysaccharide Capsular Antigen and Protein R
Authors: Hordnes  , Tynning  , Kvam  , Bevanger  , Brown  , Jonsson  , Haneberg
Published: March 2003

Group B streptococci (GBS) colonizing the female genital tract will often infect newborn infants during delivery. In 200 pregnant women studied, 14% were colonized with GBS in the cervix, 12% in the rectum, and 9% in both cervix and rectum. We have previously reported that antibody levels to GBS serotypes Ia, II, and III in sera and cervical secretions were increased in women colonized in the rectum and/or cervix, when analyzed by a whole‐cell ELISA. Here, we report the levels of antibodies to GBS serotype III capsular polysaccharide antigen (CPS III) and to protein antigen R4, which are present in most GBS III strains. Compared to culture‐negative women, the group of women colonized rectally had markedly elevated levels of immunoglobulin (Ig)A and IgG antibodies in cervical secretions to both CPS III and protein R4 (P < 0.01 and P < 0.001, respectively). In sera, the corresponding differences between culture‐negative and culture‐positive women were less pronounced, or not present. In contrast to antibody levels to whole‐cell GBS, antibody levels to CPS III and protein R4 in cervical secretions were not significantly increased in women colonized only in the cervix, except that IgA antibodies to protein R4 were slightly elevated (P < 0.05). These findings suggest that capsular type‐specific polysaccharides and protein R4 in a mucosal vaccine might induce protective antibodies against GBS colonization of the uterine cervix.
Birth of the Major Histocompatibility Complex
Authors: Klein  , Sato
Published: March 2003

Klein J, Sato A. Birth of the Major Histocompatibility Complex. Scand J Immunol 1998;47:199–209
Phenotypic and Functional Analysis of Activated B Cells of Autoimmune NZB × NZW F1 Mice
Authors: Ye  , Suen  , Chen  , Chiang
Published: March 2003

Polyclonal B‐cell activation is the central theme in the production of autoantibodies and possible activation of autoreactive T cells in both human and murine lupus. The abnormal expansion of CD5+ B cells in murine lupus has been suggested, in particular, to be one of the most characteristic findings in these mice. Activated B cells can be separated from the B cells of resting stage by the difference in cell density. The aim of this study was to investigate the characteristics of different densities of the spleen cells separated by gradient density. Furthermore, the ability of anti‐DNA antibody secretion in each percoll gradient fraction of B cells was also analysed. The results showed: a higher percentage of CD5+ B cells, which corresponded to the activated B‐cell population, in percoll gradient 1 and 2 fractions; that splenic B cells of NZB/W F1 mice had proliferative response to interleukin (IL)‐4 or IL‐5 but not to IL‐10 or interferon‐γ (IFN‐γ); and that B cells isolated by percoll gradient produced anti‐DNA antibody after stimulation with lipopolysaccharide (LPS) plus IL‐5 and IFN‐γ, but not IL‐4 and IL‐10. These data suggest that B cells at different stages of activation express differential characteristics and functions.
Acquisition of Interleukin‐5 Secretion by Human Naive T‐Helper Cells is Regulated by Distinct Signals from both the T‐Cell Receptor/CD3 Complex and CD2
Authors: Semnani R. Tolouei , Svoboda  , Khoshnood  , Van Seventer
Published: March 2003

We developed a human naive T‐helper (Th) cell differentiation model with anti‐CD3 monoclonal antibody (MoAb), using a B‐cell line as source of costimulation. In this system, we examined the contribution of the T‐cell receptor (TCR)/CD3‐derived signals and that of lymphocyte function‐associated antigen‐1 (LFA‐1) and CD2 in regulating Th‐cell subset differentiation. We found that lowering the level of anti‐CD3 MoAb decreased tumour necrosis factor‐α (TNF‐α) production, while increasing secretion of the Th2 cytokines, interleukin‐5 (IL‐5) and interleukin‐13 (IL‐13). Secretion of interferon‐γ (IFN‐γ) was not influenced by the strength of the anti‐CD3 signal. Under conditions where Th0 cells are generated, co‐culture with anti‐CD2 F(ab′)2 MoAb led to the generation of Th cells that secreted 30–35% less IL‐5, while not affecting secretion of IFN‐γ or granulocyte–macrophage colony‐stimulating factor (GM‐CSF). By contrast, anti‐CD18 MoAb F(ab′)2 inhibited IFN‐γ and GM‐CSF levels only in the primary stimulation, but did not affect cytokine levels after restimulation. Neither anti‐CD2 nor anti‐CD18 F(ab′)2 MoAbs could alter cytokine secretion profiles of peripheral blood‐derived memory/effector Th cells. Our results indicate that acquisition of IL‐5 secretion capability by Th cells is regulated mainly by signals transduced by the TCR/CD3 complex and by the presence of interleukin‐4 (IL‐4), while the CD2/LFA‐3 pathway plays an additional, but minor, role. These regulatory CD2‐derived signals, however, are distinct from those generated by the TCR/CD3 complex.
Hepatoprotective Effect of Propagermanium on Corynebacterium parvum and Lipopolysaccharide‐Induced Liver Injury in Mice
Authors: Yokochi  , Ishiwata  , Hashimoto  , Ninomiya  , Suzuki
Published: March 2003

Propagermanium is an organic germanium compound with immunopotentiating activity. We examined the hepatoprotective effect of propagermanium and its mechanism in an experimental animal model of acute liver injury induced with Corynebacterium parvum (C. parvum) and lipopolysaccharide (LPS) injection. Oral pretreatment with propagermanium decreased alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activity in a dose‐dependent manner. Significant attenuation of ALT and AST activity was obtained at a dose of 3 mg/kg. Administration of propagermanium also inhibited the infiltration of mononuclear cells into the liver of mice induced by C. parvum/LPS. Immunohistochemical examination revealed infiltration of the liver by CD4‐, CD8‐, CD11b‐ and Gr‐1‐positive cells. Propagermanium prevented CD4‐ and CD11b‐positive cells from infiltrating the liver. In this animal model, blood cytokine levels increased rapidly after LPS injection, causing severe hepatitis. Notably, tumour necrosis factor‐α (TNF‐α) and interferon‐γ (IFN‐γ) are important mediators of the progress of liver injury. We demonstrated that propagermanium reduced IFN‐γ production by 53% at a dose of 3 mg/kg and also significantly inhibited the production of interleukin‐12 (IL‐12). These results indicate that propagermanium inhibits cell infiltration in the liver and cytokine production, and improves massive liver injury in C. parvum/LPS mice.
In Vitro Leishmania major Promastigote‐Induced Macrophage Migration is Modulated by Sensory and Autonomic Neuropeptides
Authors: Ahmed  , Wahbi  , Nordlind  , Kharazmi  , Sundqvist  , Mutt  , Lidén
Published: March 2003

Recruitment, migration and adherence of macrophages and their interaction with inoculated promastigotes are key steps in the initiation of the inflammatory process in cutaneous leishmaniasis. Parasite‐ and nervous system‐derived factors might be involved in this process. In the present study the chemotactic activities of live, killed and sonicated Leishmania major promastigotes and of the promastigote culture supernatant as well as the L. major surface protease gp63 towards a murine macrophage cell line, Raw 264.7, were investigated, using the Boyden technique. The sensory neuropeptides SOM, CGRP and SP, and the autonomic neuropeptides VIP and NPY, were also investigated for possible modulatory effects on this chemotaxis, using the living promastigotes. Living promastigotes were the most efficient attractants for macrophages compared with other forms of the parasites. Prior incubation of the macrophages with the parasites completely abolished the chemotactic activity. This might indicate that the living promastigote chemotaxis is a receptor‐mediated process. On the other hand, paraformaldehyde‐killed promastigotes not only failed to induce macrophage chemotaxis but also inhibited it in comparison with the control. The surface protease gp63 tended to inhibit the macrophage chemotactic activity and the sonicate tended to stimulate it compared with controls. The culture supernatant had no effect, indicating that the chemoattractive factors putatively synthesized by the living promastigotes are not released to the surrounding medium. Somatostatin inhibited L. major promastigote‐induced macrophage migration at a high concentration, 10−6 M, while substance P inhibited it at both low concentrations, 10−10 and 10−9 M, and a high one, 10−6 M, the last‐mentioned having the greatest inhibitory effect. A stimulatory effect of calcitonin gene‐related peptide was found at high concentrations, 10−5 and 10−6 M. Vasoactive intestinal peptide stimulated macrophage chemotactic activity at both a high, 10−5 M, and at a low, 10−9 M, concentration, the same concentration at which neuropeptide Y exerted its maximum inhibitory effect.
Assessment of the Role of Leucocyte Function‐Associated Antigen‐1 in Homotypic Adhesion of Activated B Lymphocytes
Authors: Greicius  , Tamosiunas  , Severinson
Published: March 2003

In this study we investigated how T‐cell‐dependent stimuli, via interleukin‐4 (IL‐4) or CD40 ligation, influence homotypic B‐cell adhesion when compared with induction by the T‐cell‐independent stimulus lipopolysaccharide (LPS). Using primary murine B cells, we found that T‐cell‐dependent stimulation led to increased aggregation as compared to that induced by LPS. The adhesion was to a large extent dependent on the adhesion molecule, lymphocyte function‐associated antigen‐1 (LFA‐1). We found that activation of B cells with the mitogenic stimuli induced an increased avidity of LFA‐1 for its ligand, intercellular adhesion molecule‐1 (ICAM‐1). The increase was stable and different from that induced by phorbol esters. Although adhesion was reduced using B cells from LFA‐1 –/– mice, aggregation occurred in response to T‐cell‐dependent stimuli. Our data suggest that adhesion of B lymphocytes is regulated in different modes. One is induced by antigen and leads to a transient conformational change of the LFA‐1 molecule. Another is induced by mitogenic stimuli and leads to stable avidity increase of LFA‐1, possibly via activation of cytoskeletal anchorage. A third is LFA‐1 independent, of low avidity and is induced by T‐cell‐dependent stimuli.
Complement Receptor Type 1 Gene Regulation: Retinoic Acid and Cytosine Arabinoside Increase CR1 Expression
Authors: FUNKHOUSER  , VIK
Published: March 2003

Complement receptor type 1 (CR1) is expressed principally on erythrocytes, monocytes, neutrophils and B cells, where it acts as a negative regulator of the complement cascade and as a clearance mechanism for immune complexes. As CR1 expression occurs in a number of lineages, its regulation may parallel other steps in haematopoiesis. Several leucocyte‐cell lines (including K‐562, THP‐1, U‐937 and HL‐60) and B‐cell lines, as well as peripheral blood cells (PBCs), were tested for the ability of various compounds to up‐regulate their CR1 expression. While most of the compounds tested had minimal effects on CR1 message level, retinoic acid induced increases in mRNA levels in nearly all cell lines studied. Furthermore, in K‐562 cells, the cytosine analogue Ara‐C, an inducer of erythroid differentiation, caused the highest increases in CR1 mRNA levels as compared to untreated cells. Ara‐C also induced significant increases in CR1 message in PBCs. These data suggest that induction of specific CR1 expression could be an integral part of blood cell differentiation and that the identified cell induction systems may be useful as models to study the regulation of CR1 gene expression.
Structured Reactions of Serum IgM Repertoires to Immunization are Dependent on Major Histocompatibility Complex Genes
Authors: FESEL  , COUTINHO
Published: March 2003

In normal animals, responses to immunization include alterations in the serum IgM antibody repertoire, as scored on autologous tissue antigens with no respect for the immunizing antigen. These immunogen‐nonspecific antibody reactions were found previously to display specific structures dependent on strain and immunization protocols. Using major histocompatability complex (MHC)‐congenic Lewis rats, we show that such IgM repertoire reactions are under the control of MHC‐linked genes, including a class I locus. This strongly suggests the involvement of T cells restricted by both class I and class II MHC, in regulating serum IgM repertoires. Immunogen‐nonspecific repertoire reactions to immunization may, therefore, represent degenerate, but prototypical, reactions or regulatory mechanisms embodying the natural repertoires of T‐ and B cells connected to autoantigens. Natural (auto)immunity could so serve to regulate the effector class of adaptive immune responses, particularly in order to avoid pathogenic autoreactivity following specific immunization with self‐cross‐reacting antigens. Appropriate analysis of nonspecific repertoire reactions could therefore contribute to the understanding of general structures of immune regulation and natural tolerance.
Fourier Transform Infrared Spectroscopic Investigation of Temperature‐ and Pressure‐Induced Disaggregation of Amyloid A
Authors: DUBOIS  , ISMAIL  , CHAN  , ALI‐KHAN
Published: March 2003

The conformation‐sensitive amide I band in the Fourier transform infrared (FTIR) spectra of amyloid A suspensions in D2O was examined as a function of temperature (25–95 °C) and applied hydrostatic pressure (1–12 kbar) to assess the stability of the peptide. The principal changes observed upon heating were a significant loss of intermolecular β‐sheet structure, and an increase in the broad band centred at 1644 cm−1 assigned to unordered structure and α‐helices of the dissociated species. Application of hydrostatic pressure at ambient temperature resulted in a limited degree of aggregate dissociation. These structural changes were partially reversible with cooling or release of the applied pressure. Dissolving the aggregated peptide in alkaline solution (pH 12) also resulted in disaggregation. Dissociation of organ‐deposited amyloid substance bears clinical relevance. The present data indicate that residual amounts of undissociated amyloid in the milieu at physiological and acidic pH may act as nucleating foci rendering dissociated amyloid to reaggregate into organized amyloid.
Splenic Marginal Metallophilic Macrophages and Marginal Zone Macrophages are the Major Interferon‐α/β Producers in Mice upon Intravenous Challenge with Herpes Simplex Virus
Authors: ELORANTA  , ALM
Published: March 2003

The interferon (IFN)‐α/β‐producing cells (IPCs) are localized predominantly in the spleen, in particular in the marginal zones (MZ), in C57BL/6 mice injected intravenously (i.v.) with UV‐inactivated herpes simplex virus (HSV). We defined the phenotype of these murine IPCs using simultaneous immunohistochemical labelling of intracellular IFN‐α/β and various surface antigens. We found that the IPCs in the MZ are not dendritic cells because they did not express major histocompatibility complex (MHC) class II and CD11c molecules. Furthermore, they did not express antigenic markers typical for T cells, B cells or red pulp macrophages. In contrast, the majority of IPCs were stained by the anti‐sialoadhesin monoclonal antibody (MoAb) SER‐4, which is specific for marginal metallophilic macrophages. In addition, a minor part of the IPCs with a more outward localization in the MZ were stained by a MoAb specific for MZ macrophages. We conclude that the massive IFN‐α/β production in the MZ of the spleen upon in vivo stimulation by HSV is mainly exerted by marginal metallophilic macrophages and to a lesser extent by MZ macrophages.
The Kinetics and Magnitude of the Synergistic Activation of the Serum Amyloid A Promoter by IL‐1β and IL‐6 is Determined by the Order of Cytokine Addition
Authors: UHLAR  , WHITEHEAD
Published: March 2003

Human serum amyloid A protein (A‐SAA) is a major hepatic acute‐phase protein, the concentration of which increases by up to 1000‐fold during inflammation. This induction is primarily due to synergistic transcriptional up‐regulation by pro‐inflammatory cytokines, principally interleukin (IL)‐1 and IL‐6. Using HepG2 hepatoma cells transfected with pGL2‐SAA2pt, a cytokine‐responsive human SAA2 promoter/luciferase reporter gene construct, we show that stimulation with IL‐1β prior to IL‐6 is essential for maximal synergistic transcriptional induction of the SAA2 gene. The reciprocal treatment, i.e. stimulation of the promoter with IL‐6 before IL‐1β, results in significantly less synergistic activation of the SAA2 promoter. These findings strongly suggest that in vitro studies of acute‐phase‐protein induction using combinations of cytokines should be designed to reflect the chronology of their participation in the cytokine cascade.
Possible Role of Interleukin‐10 in Autoantibody Production and in the Fate of Human Cord Blood CD5+ B Lymphocytes
Authors: VILLASEÑOR‐BUSTAMANTE  , ALVARADO‐DE LA BARRERA  , RICHAUD‐PATIN  , MARTÍNEZ‐AYALA  , LLORENTE
Published: March 2003

Mononuclear cell subsets in 25 human umbilical cord blood samples and 10 healthy adults were studied. We found a decreased percentage of CD3+ cells, CD8+ cells and γδ T cells in cord blood compared with blood from healthy adults. The CD16+56+ and CD19+CD5+ phenotypes were overexpressed in cord blood. We then measured spontaneous gene expression and the production of interleukin‐10 in mononuclear cells from cord blood and adult subjects. Although we found no difference between cord blood cells and those from healthy adults, a tendency towards spontaneous interleukin‐10 production was observed in cord blood. Phorbol 12‐myristate 13‐acetate stimulation revealed that, among lymphocytes, cord blood B cells are the main cellular source of interleukin‐10. Finally, we found no evidence of augmented spontaneous apoptosis but an increased bcl‐2 gene expression in non‐T cells from cord blood. Interleukin‐10 might protect CD19+CD5+ B cells from apoptosis by inducing bcl‐2 and promoting autoantibody production in this B‐cell subpopulation.
Light Chain Variable (VL) Sequences of Rheumatoid Factors (RF) in Patients with Primary Sjögren's Syndrome (pSS): Moderate Contribution of Somatic Hypermutation
Authors: Elagib  , Boslash;rretzen  , Thompson  , Natvig
Published: March 2003

We have characterized and sequenced the variable (V) region genes of the light (L) chains of 10 immunoglobulin (IgM) rheumatoid factor (RF) monoclonal antibodies (MoAb) derived by the hybridoma/Epstein‐Barr virus (EBV) technique from the peripheral blood of patients with primary Sjögren's syndrome (pSS). Six out of 10 RFs used lambda (λ) L chains, while four RFs used kappa (κ) L chains. Five out of the six λ RFs were encoded by Vλ3 gene segments, the sixth one was encoded by a Vλ1 gene segment. This preferential utilization of the Vλ3 family genes suggests selective expansion of the B cell in pSS. Three of the κ RFs used Vκ3 gene segments, while the fourth used a Vκ2 gene segment. Half of the RFs were found as unmutated copies of their closest germline (GL) gene. Interestingly these RFs were previously shown to use heavy (H) chains in GL gene configuration. Three RFs have very few mutations (2–3) and only two RFs have substantial numbers of mutations (6 and 11). This also correlated with the number of mutations in the respective H chains. In contrast to RFs in normal and RA these results further suggest the somatic mutation to be of moderate importance in the generation of RF from the peripheral blood of pSS patients.
α‐Difluoromethylornithine Blocks Thymocyte Apoptosis Via a Reduction in Tyrosine Phosphorylation
Authors: Jan  , Wing  , Lin  , Lin
Published: March 2003

The effect of α‐difluoromethylornithine on cell apoptosis was investigated. Freshly isolated mouse thymocytes were cultured in the medium alone or with dexamethasone, and apoptotic cell death was monitored after 6 h. A correlation was seen between cell apoptosis and a reduction in the polyamine levels of thymocytes. Addition of exogenous polyamines decreased the levels of apoptosis induced spontaneously in the culture medium or by dexamethasone. However, addition of α‐difluoromethylornithine, an irreversible inhibitor of ornithine decarboxylase, to the cultures did not enhance apoptosis but rather caused inhibition of thymocyte apoptosis. Analysis of the mechanism of α‐difluoromethylornithine‐mediated inhibition of apoptosis indicated that α‐difluoromethylornithine treatment blocked protein tyrosine phosphorylation, which was elevated drastically during the first hour of thymocyte cultivation. Treatment with the phosphotyrosine phosphatase inhibitor phenylarsine oxide reversed this inhibitory effect of α‐difluoromethylornithine on apoptotic cell death. Our results provide an alternative mechansim for α‐difluoromethylornithine showing the inhibition of apoptosis via reduction of protein tyrosine phosphorylation.
Intranasal Administration of Recombinant Mouse Interleukin‐12 Increases Inflammation and Demyelination in Chronic Experimental Autoimmune Neuritis in Lewis Rats
Authors: Pelidou  , Zou  , Deretzi  , Nennesmo  , Wei  , Mix  , Van Der Meide  , Zhu
Published: March 2003

To examine whether interleukin (IL)‐12 modulates ongoing chronic experimental autoimmune neuritis (EAN), we evaluated the effects of recombinant mouse IL‐12 (rmIL‐12) in Lewis rats with chronic EAN, induced by immunization with P0 peptide (180–199) plus complete Freund's adjuvant. Rats were treated intranasally with either 0.1 or 1 μg/rat/day rmIL‐12 for 6 days from the onset of clinical chronic EAN, on days 5–10 postimmunization (p.i.). Only high‐dose rmIL‐12 exacerbated chronic EAN. This clinical effect was associated with higher numbers of inflammatory cells and more severe demyelination in sciatic nerve sections on days 15 and 80 p.i. compared with low‐dose rmIL‐12‐treated rats and phosphate‐buffered saline (PBS)‐treated control rats. High‐dose rmIL‐12 increased significantly the lymph node mononuclear cell proliferation in response to P0 peptide 180–199 and IFN‐γ production in the sciatic nerves. These data indicate that intranasally administered IL‐12 acts as a proinflammatory cytokine in chronic EAN. Effective inhibition of IL‐12 in vivo could be considered for therapeutic use in chronic inflammatory demyelinating polyradiculoneuropathy.
Distinct Innate and Acquired Immune Responses to Leishmania in Putative Susceptible and Resistant Human Populations Endemically Exposed to L. (Viannia) panamensis Infection
Authors: Bosque  , Saravia  , Valderrama  , Milon
Published: March 2003

Mechanisms of constitutive and acquired susceptibility/resistance to Leishmania Viannia panamensis (L. (V ) p.) were investigated in endemically exposed human populations presenting either recurrent disease (putative susceptible) or subclinical infection (clinically resistant). Cutaneous delayed type hypersensitivity response to leishmanin was significantly lower among individuals experiencing recurrent leishmaniasis than among those whose skin test converted without developing the disease. Monocyte derived macrophages from individuals with recurrent disease were more permissive in vitro to the entry of parasites than macrophages from subclinically infected individuals. In vitro proliferation of CD4 and CD8 T lymphocytes in response to intracellular amastigotes was significantly lower among individuals with a history of recurrent disease compared with subclinically infected individuals. Linear regression analyses revealed a strong direct relationship between the production of interferon (IFN)‐γ, tumour necrosis factor (TNF)‐α and interleukin (IL)‐10 by peripheral blood mononuclear cells (PBMC) from resistant (subclinically infected) individuals and no correlation in the production of these cytokines by PBMC from individuals who experienced recurrent disease. The results provide evidence of differences in the innate and acquired responses to Leishmania according to the outcome of the natural infection. These findings support the feasibility of identifying the immunological bases of innate and acquired resistance through studies in naturally exposed human populations.
Identification of Seven Genes in the Major Histocompatibility Complex Class I Region of the Zebrafish
Authors: Sültmann  , Murray  , Klein
Published: March 2003

Physical linkage of genes whose products are involved in similar physiological pathways may have functional significance. The identification of conserved gene linkage in distantly related organisms can therefore strengthen the hypothesis of selection acting towards keeping genes on a chromosome. We used the cDNA selection technique and the polymerase chain reaction (PCR) with generic primers for the identification of new genes on the genomic clones bearing the major histocompatibility complex (Mhc) class I genes of the zebrafish (Danio rerio). We found six new genes (BING1, DAXX, TAPBP, KNSL2, TAP2B and KE6) whose orthologues are known to be linked to the Mhc class II region in humans and mice. In addition, a new zebrafish Mhc class I gene, termed Dare‐UFA, was detected. By contrast, a search for the human leucocyte antigen (HLA)‐linked BING3, KE3 and SACM2L genes revealed that these loci are not located on the class I clones of the zebrafish. The zebrafish class I region contains repetitive elements with similarity to the DANA, SATA and LINE repeats, as well as Tc1 transposable elements. Our findings indicate a high degree of linkage conservation between the zebrafish class I and the mammalian class II regions.
T‐Lymphocyte Subpopulations Do Not Express Identical Combinations of Interleukin‐2 Receptor Chains in the Early Phase of Their Activation and Proliferation
Authors: Perola  , Ripatti  , Pelkonen
Published: March 2003

Owing to the contrasting observations in the field of interleukin(IL)‐2 receptor research, the expression of IL‐2 receptor chains was analysed on resting and anti‐CD3 antibody (OKT3) activated CD4 and CD8 T cells by flow cytometry. Prior to the stimulation, 49% of CD4+ cells expressed IL‐2Rα (CD25), whereas the expression of IL‐2Rβ (CD122) was very low (8%). The reverse was true for CD8 cells: 48% of them were positive for CD122, but only a fraction (10%) expressed CD25. Practically all lymphocytes expressed IL‐2Rγ (CD132). Interestingly, the unbalanced expression of IL‐2Rα and ‐β continued throughout the stimulation period of 2 days. In addition, the expression of CD45 isoforms in combination with the IL‐2R chains and CD71 was followed during the activation of CD4+ T cells. Although CD45RA+/RO− CD4 cells were effectively activated, they retained their naive phenotype up to 2 days of stimulation. On the other hand, CD45RA+low/RO+low (Ddull) CD4+ cells shifted to the memory phenotype rapidly after being activated. However, by day 6 of stimulation the shift of both naive and Ddull cells to memory ones was obvious. The role of the IL‐2 receptor in the activation of CD4 subpopulations is discussed.
Polyreactivity of Disease‐Associated Anti‐RBC IgG Autoantibodies of Patients with Warm Autoimmune Haemolytic Anaemia and Natural Anti‐RBC IgG Autoantibodies of Healthy Individuals
Authors: Stahl  , Lacroix‐Desmazes  , Kaveri  , Kazatchkine
Published: March 2003

Anti‐red blood cell (RBC) immunoglobulin G (IgG) autoantibodies are present in patients with warm autoimmune haemolytic anaemia (WAIHA), and, as natural autoantibodies, in healthy individuals. This study investigated whether the feature of polyreactivity discriminates disease‐associated from natural anti‐RBC IgG autoantibodies. The patterns of reactivity of purified anti‐RBC IgG eluted from the RBC of WAIHA patients and from the RBC of healthy individuals were analysed using quantitative immunoblotting on a panel of whole human tissue or bacterial cell extracts as antigen sources. In parallel, the reactivity patterns of IgG purified from plasma were analysed. Anti‐RBC IgG of WAIHA patients and of healthy individuals recognized a wide range of self‐ and nonself‐antigens. The reactivity patterns of anti‐RBC IgG were homogeneous among patients and controls, did not differ between patients and controls, and were similar to those of IgG purified from plasma in the case of both patients and healthy individuals. The data demonstrate that the anti‐RBC IgG autoantibodies of WAIHA patients share extensive similarity with those of healthy individuals. Polyreactivity is a common feature of both disease‐associated and natural anti‐RBC IgG autoantibodies.
Immune Activation in the Intestinal Mucosa Before the Onset of Colitis in Gαi2‐Deficient Mice
Authors: Öhman  , Franzén  , Rudolph  , Harriman  , Hultgren Hörnquist
Published: March 2003

G‐protein subunit Gαi2‐deficient mice spontaneously develop an inflammatory bowel disease that clinically and histopathologically resembles ulcerative colitis in humans. The aim of this study was to determine whether immunological changes precede the development of colitis in Gαi2‐deficient mice. Therefore, Gαi2‐deficient mice with no clinical or histopathological signs of colitis were compared with Gαi2‐deficient mice with established colitis and wild‐type animals, concerning immunological parameters. Healthy Gαi2‐deficient mice displayed an increased frequency of CD4+ T cells and a decreased frequency of CD19+ B lymphocytes in the intestinal mucosa compared with control mice. The CD4+ population was characterized by a memory phenotype, i.e. increased expression of CD44 and decreased expression of CD45RB and CD62L, as well as increased expression of the mucosal homing receptors integrins α4β7 and αEβ7. Production of pro‐inflammatory cytokines, interleukin (IL)‐1β and interferon (IFN)‐γ, were increased in Gαi2‐deficient mice before clinical signs of disease were evident. In addition, total immunoglobulin (Ig)G and IgA levels in large intestinal secretions were increased significantly compared with wild‐type mice, and antibodies specific for the normal intestinal flora in large intestinal secretions were present in Gαi2‐deficient mice several weeks before the onset of colitis. In contrast, antibodies against tropomyosin, a putative autoantigen in human ulcerative colitis, were not found in Gαi2‐deficient mice before the onset of colitis, although they were present in animals with established disease. In conclusion, activation of the intestinal immune system precedes histopathological and clinical signs of inflammation in Gαi2‐deficient mice, suggesting that immune abnormalities play an important role in the induction of colitis.
Analysis of the Tissue Distribution of the Rat C5a Receptor and Inhibition of C5a‐Mediated Effects Through the Use of Two MoAbs
Authors: Rothermel E. , Götze O. , Zahn S. , Schlaf G.
Published: March 2003

The C5‐anaphylatoxin C5a is a protein of 74 (human) or 77 (rat) amino‐acid residues, respectively, the generation of which may be induced by either the classical and/or the alternative pathways. C5a binds specifically to its receptor (C5aR/CD88) which belongs to the superfamily of G‐protein‐coupled receptors with seven transmembrane segments. In this study we describe the tissue distribution of the rat C5aR (rC5aR) and the blocking of its ligand by the application of two monoclonal antibodies (MoAbs). The first antibody (MoAb R63) which is directed against the amino‐terminal domain Ex1 of the rat C5aR was generated in mice immunized with RBL–2H3 cells which had been stably transfected with the rat C5a receptor gene. Checking the rC5aR expression in various tissues bronchial epithelial cells stained positive only in tissue samples from animals with a mycoplasm infection indicating that the receptor may be induced in this cell type as a consequence of an inflammatory process. Using immunohistochemistry there was no evidence for nonmyeloid expression in the large or small intestine, heart, lung, kidney or liver of the normal rat. The MoAb R63 was found to be a reliable tool for the investigation of the expression of the receptor by FACS analyses or immunohistochemistry. Despite numerous attempts neutralizing antibodies could not be generated against the receptor. Therefore a C5a‐ligand neutralizing MoAb was generated against the synthesized carboxyterminal 20mer peptide. This antibody (6–9F) recognized the carboxy terminus of C5a/C5a‐FLUOS and prevented its binding at a three‐fold molar excess as evidenced by FACS‐analyses. It also blocked the C5a‐mediated signal transduction as demonstrated by the inhibition of intracellular Ca2+‐release (at a 16‐fold molar excess) and the release of N‐Acetyl‐β‐
d‐glucosaminidase (at a 25‐fold molar excess).
Neural Tissue Xenografting
Authors: Larsson  , Widner
Published: March 2003

Neural transplantation may become an important treatment alternative for focal brain disorders. To date, the most successful grafts have been obtained in patients with Parkinson's disease. Completely normalized dopamine production and reduction of Parkinsonian symptoms have been demonstrated 10 years after grafting. However, the allogeneic donor tissue has to be obtained from induced abortions, and there are logistical difficulties, risks of infection, and ethical constraints limiting a wider clinical use. Xenografting is an alternative that could bridge these limitations if immunological rejection could be prevented. Pig embryonic neural tissue has been grafted to patients with Parkinson's disease, but no functional benefits have clinically been proven so far. The immune reactions to neural xenografts were incompletely characterized at the time of these early clinical trials, and it is likely that the treatments used were insufficient and that the grafts were rejected. In this article we will review new experiments addressing the immune responses against porcine neural tissue grafted to the adult brain, including the role of antibodies, complement, natural killer (NK) cells, lymphocytes, as well as the effects of immunosuppressive drugs and donor tissue modifications.
Alternatively Activated Macrophages Differentially Express Fibronectin and Its Splice Variants and the Extracellular Matrix Protein βIG‐H3
Authors: Gratchev A. , Guillot P. , Hakiy N. , Politz O. , Orfanos C. E. , Schledzewski K. , Goerdt S.
Published: March 2003

Alternative activation of macrophages, induced by Th2 cytokines and glucocorticoids, is essential for the proper functioning of anti‐inflammatory immune reactions. To this end, alternatively activated macrophages (aaMΦ) express a not yet fully unravelled set of genes including cytokines such as alternative macrophage activation‐associated CC‐chemokine (AMAC)‐1 and pattern recognition molecules such as the scavenger receptor CD163. In order to further characterize the molecular repertoire of aaMΦ, differential gene expression was analyzed by combining subtractive suppression cloning and differential hybridization. We show here that aaMΦ induced by interleukin (IL)‐4 overexpress the prototype extracellular matrix (ECM) protein fibronectin on the mRNA and protein level. This overall increase is accompanied by a shift in fibronectin splice variants from an embryonic to a mature pattern. In addition, the expression of another ECM protein, βIG‐H3, is also upregulated by IL‐4 in aaMΦ. In contrast to IL‐4 and in line with its inhibitory effect on wound healing, dexamethasone exerts a strongly suppressive effect on fibronectin and βIG‐H3 expression. In conclusion, overexpression of ECM proteins induced by IL‐4 in macrophages suggests that aaMΦ may be involved in ECM deposition and tissue remodelling during the healing phase of acute inflammatory reactions and in chronic inflammatory diseases.