bluuebunny/crossref_metadata_2025 huggingface dataset:
Errata for “Cubic polynomial maps with periodic critical orbit, Part II: Escape regions”
Authors: Bonifant Araceli , Kiwi Jan , Milnor John
Published: July 2010

In this note we fill in some essential details which were missing from our paper. In the case of an escape region

E

h

\mathcal {E}_h

with non-trivial kneading sequence, we prove that the canonical parameter

t
t

can be expressed as a holomorphic function of the local parameter

η
=

a

−
1

/

μ

\eta =a^{-1/\mu }

(where

a
a

is the periodic critical point). Furthermore, we prove that for any escape region

E

h

\mathcal {E}_h

of grid period

n
≥
2

n\ge 2

, the winding number

ν
\nu

of

E

h

\mathcal {E}_h

over the

t
t

-plane is greater or equal than the multiplicity

μ
\mu

of

E

h

\mathcal {E}_h

.
ExpressionView—an interactive viewer for modules identified in gene expression data
Authors: Lüscher Andreas , Csárdi Gábor , Morton de Lachapelle Aitana , Kutalik Zoltán , Peter Bastian , Bergmann Sven
Published: July 2010

Summary: ExpressionView is an R package that provides an interactive graphical environment to explore transcription modules identified in gene expression data. A sophisticated ordering algorithm is used to present the modules with the expression in a visually appealing layout that provides an intuitive summary of the results. From this overview, the user can select individual modules and access biologically relevant metadata associated with them.

Availability:  http://www.unil.ch/cbg/ExpressionView. Screenshots, tutorials and sample data sets can be found on the ExpressionView web site.

Contact:  sven.bergmann@unil.ch
Confocal Imaging of Pseudomonas syringae pv. phaseolicola Colony Development in Bean Reveals Reduced Multiplication of Strains Containing the Genomic Island PPHGI-1
Authors: Godfrey S. A. C. , Mansfield J. W. , Corry D. S. , Lovell H. C. , Jackson R. W. , Arnold D. L.
Published: July 2010

Pseudomonas syringae pv. phaseolicola is the seed borne causative agent of halo blight in the common bean Phaseolus vulgaris. Pseudomonas syringae pv. phaseolicola race 4 strain 1302A contains the avirulence gene hopAR1 (located on a 106-kb genomic island, PPHGI-1, and earlier named avrPphB), which matches resistance gene R3 in P. vulgaris cultivar Tendergreen (TG) and causes a rapid hypersensitive reaction (HR). Here, we have fluorescently labeled selected Pseudomonas syringae pv. phaseolicola 1302A and 1448A strains (with and without PPHGI-1) to enable confocal imaging of in-planta colony formation within the apoplast of resistant (TG) and susceptible (Canadian Wonder [CW]) P. vulgaris leaves. Temporal quantification of fluorescent Pseudomonas syringae pv. phaseolicola colony development correlated with in-planta bacterial multiplication (measured as CFU/ml) and is, therefore, an effective means of monitoring Pseudomonas syringae pv. phaseolicola endophytic colonization and survival in P. vulgaris. We present advances in the application of confocal microscopy for in-planta visualization of Pseudomonas syringae pv. phaseolicola colony development in the leaf mesophyll to show how the HR defense response greatly affects colony morphology and bacterial survival. Unexpectedly, the presence of PPHGI-1 was found to cause a reduction of colony development in susceptible P. vulgaris CW leaf tissue. We discuss the evolutionary consequences that the acquisition and retention of PPHGI-1 brings to Pseudomonas syringae pv. phaseolicola in planta.
An optical method to quantify the density of ligands for cell adhesion receptors in three-dimensional matrices
Authors: Tzeranis Dimitrios S. , Roy Amit , So Peter T. C. , Yannas Ioannis V.
Published: July 2010

The three-dimensional matrix that surrounds cells is an important insoluble regulator of cell phenotypes. Examples of such insoluble surfaces are the extracellular matrix (ECM), ECM analogues and synthetic polymeric biomaterials. Cell–matrix interactions are mediated by cell adhesion receptors that bind to chemical entities (adhesion ligands) on the surface of the matrix. There are currently no established methods to obtain quantitative estimates of the density of adhesion ligands recognized by specific cell adhesion receptors. This article presents a new optical-based methodology for measuring ligands of adhesion receptors on three-dimensional matrices. The study also provides preliminary quantitative results for the density of adhesion ligands of integrinsα1β1andα2β1on the surface of collagen-based scaffolds, similar to biomaterials that are used clinically to induce regeneration in injured skin and peripheral nerves. Preliminary estimates of the surface density of the ligands of these two major collagen-binding receptors are 5775 ± 2064 ligands µm−2for ligands ofα1β1and 17 084 ± 5353 ligands µm−2for ligands ofα2β1. The proposed methodology can be used to quantify the surface chemistry of insoluble surfaces that possess biological activity, such as native tissue ECM and biomaterials, and therefore can be used in cell biology, biomaterials science and regenerative medical studies for quantitative description of a matrix and its effects on cells.
Transcriptional-Metabolic Networks in β-Carotene-Enriched Potato Tubers: The Long and Winding Road to the Golden Phenotype
Authors: Diretto Gianfranco , Al-Babili Salim , Tavazza Raffaela , Scossa Federico , Papacchioli Velia , Migliore Melania , Beyer Peter , Giuliano Giovanni
Published: July 2010

Vitamin A deficiency is a public health problem in a large number of countries. Biofortification of major staple crops (wheat [Triticum aestivum], rice [Oryza sativa], maize [Zea mays], and potato [Solanum tuberosum]) with β-carotene has the potential to alleviate this nutritional problem. Previously, we engineered transgenic “Golden” potato tubers overexpressing three bacterial genes for β-carotene synthesis (CrtB, CrtI, and CrtY, encoding phytoene synthase, phytoene desaturase, and lycopene β-cyclase, respectively) and accumulating the highest amount of β-carotene in the four aforementioned crops. Here, we report the systematic quantitation of carotenoid metabolites and transcripts in 24 lines carrying six different transgene combinations under the control of the 35S and Patatin (Pat) promoters. Low levels of B-I expression are sufficient for interfering with leaf carotenogenesis, but not for β-carotene accumulation in tubers and calli, which requires high expression levels of all three genes under the control of the Pat promoter. Tubers expressing the B-I transgenes show large perturbations in the transcription of endogenous carotenoid genes, with only minor changes in carotenoid content, while the opposite phenotype (low levels of transcriptional perturbation and high carotenoid levels) is observed in Golden (Y-B-I) tubers. We used hierarchical clustering and pairwise correlation analysis, together with a new method for network correlation analysis, developed for this purpose, to assess the perturbations in transcript and metabolite levels in transgenic leaves and tubers. Through a “guilt-by-profiling” approach, we identified several endogenous genes for carotenoid biosynthesis likely to play a key regulatory role in Golden tubers, which are candidates for manipulations aimed at the further optimization of tuber carotenoid content.
Use of linear scans to obtain |Fhkl|2and the integrated intensity
Authors: Shayduk Roman
Published: July 2010

An alternative method for determining the structure factor of a Bragg reflection is described. For broad reflections, the structure factor can be obtained from linear scans through the reflection in reciprocal space. The best known geometry corrections,i.e.Lorentz, polarization and area (volume) factors, are properly taken into account. In modern diffractometers, scans in reciprocal space are standard operations controlled by the diffractometer software program. The necessary corrections and diffractometer slit settings for the use of the new method are described.
New analytical approximation of diffraction size broadened peak profile for spherical crystallites with a lognormal distribution
Authors: Popa N. C. , Teodorescu C. M. , Frunza S.
Published: July 2010

A new analytical approximation of the diffraction peak size profile due to the lognormal distribution of spherical crystallites is derived. In comparison with the existing approximation constructed with the condition of conserving the volume-averaged column length of the exact profile [Popa & Balzar (2002).J. Appl. Cryst.35, 338–346], the new approximation conserves both volume- and area-averaged column lengths. Moreover, it is a better approximation for large distribution dispersions. Like the old approximation, the present one is appropriate for implementation in the whole powder pattern fitting codes, Rietveld included.
Standardization of Factor VIII I. CALIBRATION OF BRITISH STANDARDS FOR FACTOR VIII CLOTTING ACTIVITY
Authors: Barrowcliffe T. W. , Kirkwood T. B. L.
Published: July 2010

Summary.

Calibration of successive British Standards for Factor VIII clotting activity against the International Standard (concentrate) has brought to light substantial discrepancies among laboratories and between assay methods. These discrepancies were less in assays of concentrate standards than in calibration of plasma standards. Standardization of reagents in the two‐stage assays substantially improved agreement among laboratories using this method. Standardization of the phospholipid reagent and haemophilic substrate separately had little effect on variation among laboratories performing one‐stage assays. Standardization of reagents did not alter the basic discrepancy (approximately 20%)) between the one‐stage and two‐stage assay methods. Omission of the aluminium hydroxide adsorption step from the two‐stage method reduced the discrepancy between the two methods to less than 10%. Discrepancies did not occur when one plasma standard was assayed against another. Improvements in the stability of VIII: C in freeze‐dried plasma now make it feasible to establish a long‐term plasma reference standard.
Dysfibrinogenaemia Characterized by Abnormal Fibrin Monomer Polymerization and Normal Fibrinopeptide A Release
Authors: Lane D. A. , Cuddigan B. , VanRoss M. , Kakkar V. V.
Published: July 2010

Summary. Routine testing on plasma from a patient due to undergo a coronary artery bypass graft operation revealed a prolonged thrombin clotting time associated with a normal plasma fibrinogen level when this was determined by a method not dependent upon the rate of fibrin formation. Fibrinogen purified from the patient's plasma by precipitation with β‐alanine also gave a prolonged thrombin time and this confirmed the presence of a dysfibrinogenaemia. Increasing calcium chloride concentration, addition of protamine sulphate and decreasing ionic strength all produced a partial correction of the clotting defect. Addition of normal plasma to patient's plasma failed to correct the prolonged thrombin clotting time and a pH dependence of the defect was also observed. Kinetic studies of fibrinopeptide release, using a specific radioimmunoassay, demonstrated no delay in the release of patient fibrinopeptide A. The functional defect was localized as an abnormality in the polymerization of fibrin monomers by studying fibrin monomers prepared and isolated from plasma and from purified fibrinogen solution. An electrophoretic examination of the patient's fibrinogen using both agarose and polyacrylamide gels failed to demonstrate any alteration in mobility or any structural defect associated with the polypeptide chains Aα, Bβ and γ. All seven of the living siblings of the propositus and also his daughter showed no abnormality in any clotting assay. However, because the propositus did not suffer from liver disease it has been assumed that the abnormality is genetic in origin.
An Abnormal Fibrinogen with Delayed Fibrinopeptide A Release
Authors: Lane D. A. , Vanross M. , Kakkar V.V. , Bottomley J. , Dhir K. , Holt L. P. J. , MacIver J. E.
Published: July 2010

Summary. Investigation of the family of a patient presenting with haematuria revealed seven cases in three generations showing a prolonged thrombin clotting time. A dysfibrinogenaemia was confirmed when purified fibrinogen from an affected member of the family was shown to also exhibit a prolonged thrombin clotting time. No molecular abnormality could be demonstrated using electrophor‐etic and immunological techniques. However, using a specific radioimmunoassay to fibrinopeptide A a major defect has been localized to a delay in the rate of release of this peptide by thrombin when the abnormal fibrinogen is converted to fibrin. This case of dysfibrinogenaemia has been tentatively designated fibrinogen Manchester.
Restriction endonuclease maps of the β‐like globin gene cluster in the British and Greek forms of HPFH, and for one example of Gγβ+ HPFH
Authors: Jones R. W. , Old J. M. , Wood W. G. , Clegg J. B. , Weatherall D. J.
Published: July 2010

We report here restriction endonuclease maps of the β‐like globin gene cluster for the British form of HPFH and for a case of Gγβ+ HPFH, and also confirm and extend previous reports of the map for the Greek form of HPFH. These results show that all these conditions belong to a group lacking any substantial deletion or rearrangement of DNA sequence in this gene cluster. The absence of any gross disruption of the structure of this region of the genome, together with evidence that the HPFH genotype is either allelic with, or closely linked to, the β‐like globin gene cluster, suggests that the responsible lesions are nearer to being purely regulatory in nature than in forms of HPFH due to substantial deletions. Thus these conditions promise to provide less equivocal evidence about the regulation of β‐like globin gene expression than has so far been available.
Activation‐dependent proteolytic degradation of polymorphonuclear CD11b
Authors: Davey Penelope C. , Zuzel Mirko , Kamiguti Aura S. , Hunt John A. , Aziz Khalil A.
Published: July 2010

CD11b/CD18 is the principal integrin of polymorphonuclear (PMN) leucocytes and is involved in their adhesion, migration and phagocytosis. In quiescent cells, the receptor is stored in intracellular granules from where it is translocated to the cell surface in response to a variety of stimuli. In this study, we demonstrated that strong stimulation of PMNs not only leads to the upregulation of CD11b surface expression, but also to the subsequent time‐dependent apparent loss of this receptor, as detected by fluorescence‐activated cell sorting (FACS) using a monoclonal antibody (mAb) against an N‐terminal CD11b epitope. This epitope loss was observed following either direct stimulation of protein kinase C (PKC) with phorbol 12‐myristate 13‐acetate (PMA) or after multiple receptor stimulation using a combination of the agonist N‐formylmethionyl‐leucyl‐phenylalanine (FMLP) and the priming agents granulocyte macrophage‐colony stimulating factor (GM‐CSF) and platelet factor (PF) 4. However, upregulation following weak stimulation with FMLP alone was not followed by subsequent epitope loss of the receptor. The increases and subsequent decreases in CD11b expression induced by PMA were paralleled by an increase and a decrease in PMN adhesion to CD11b‐specific ligands, fibrinogen and intercellular adhesion molecule (ICAM)‐1. Sodium dodecyl sulphate‐polyacrylamide gel electrophoresis (SDS‐PAGE) and Western blot analysis showed that this epitope loss of PMN CD11b was the result of proteolytic degradation of the N‐terminal region of the molecule. The use of a range of proteinase inhibitors indicated that this CD11b degradation involves a cell‐associated serine proteinase. This is the first demonstration of the proteolytic alteration of CD11b in response to strong PMN stimulation. Given the central role of CD11b/CD18 in all aspects of PMN function, this alteration of the CD11b molecule and its effect on PMN adhesion are probably of considerable pathophysiological importance.
Molecular remission of Philadelphia/bcr‐abl‐positive acute myeloid leukaemia after treatment with anti‐CD33 calicheamicin conjugate (gemtuzumab ozogamicin, CMA‐676)
Authors: De Vetten M. P. , Jansen J. H. , Van Der Reijden B. A. , Berger M. S. , Zijlmans J. M. J. M. , Löwenberg B.
Published: July 2010

The anti‐CD33 monoclonal antibody linked to NAc‐gamma calicheamicin gemtuzumab ozogamicin (CMA‐676) was used to treat a patient with Philadelphia/bcr‐abl‐positive acute myeloid leukaemia. We report a morphological and cytogenetic complete remission after treatment with two doses of gemtuzumab ozogamicin as a single agent. Using real‐time polymerase chain reaction (PCR), gemtuzumab ozogamicin treatment resulted in a 3‐log tumour mass reduction in bone marrow.
An inhibitor of the toxicity of tumour necrosis factor in the serum of patients with sarcoidosis, tuberculosis and Crohn's disease
Authors: FOLEY N , LAMBERT C , MCNICOL M , JOHNSON N , ROOK G A W
Published: July 2010

The activated macrophages present in the T cell-dependent granulomata of sarcoidosis and tuberculosis are primed for enhanced release of cytokines including tumour necrosis factor (TNF or cachectin). Release of this cytokine can induce an acute-phase response, fever, and necrosis in suitably prepared sites of inflammation; if chronic, its presence may contribute to weight loss. These clinical features are characteristic of tuberculosis, but not of sarcoidosis, though alveolar macrophages from both diseases release large quantities of TNF in vitro. We therefore postulated the presence in sarcoidosis patients of an inhibitor of TNF. We have studied levels of TNF inhibitory activity by determining the quantity of TNF required to give 50% kill of 1929 cells in the presence of 20% heat-inactivated serum derived from various disease states (37 sarcoidosis, 13 tuberculosis, 13 Crohn's disease. 17 healthy donors). Normal sera used in this way do not inhibit significantly, but inhibition of TNF loxicity is caused by most sera from both sarcoidosis and tuberculosis. Used at 20%, five out of 37 sarcoidosis sera and one out of 13 tuberculosis sera caused complete inhibition of TNF, even when the latter was added at 100 times the concentration required to give 50% kill in control wells. This inhibitor may have an important physiological role.
Relation between the increase of circulating CD3+ CD57+ lymphocytes and T cell dysfunction in recipients of bone marrow transplantation
Authors: IZQUIERDO M , BALBOA M A , FERNANDEZ-RASADA J M , FIGUERA A , TORRES A , IRIONDO A , LÓPEZ-BOTET M
Published: July 2010

Some patients undergoing bone marrow transplantation (BMT) persistently present increased proportions of circulating CD57+ T cells. We analysed the cell surface phenotype in peripheral blood mononuclear cells (PBMC) from 69 allogeneic and 11 autologous BMT recipients. In parallel samples from 49 patients, the proliferative response to T cell mitogens was assessed, either in the presence or absence of exogenous interleukin-2 (IL-2). PBMC samples from long-term allogeneic BMT patients with increased proportions of CD57+ cells displayed significantly (P < 0.001) lower proliferative responses, compared with samples from patients with normal proportions of CD57+ cells and from healthy subjects. Elimination of the CD57+ population by C′-dependent lysis did not normalize the proliferative response. After positive selection by cell sorting. CD57+ cells responded poorly, but in the presence of IL-2 the proliferation appeared to be similar to that displayed by the CD57- subset and still suboptimal compared with normal controls. These data suggest that the hyporesponsiveness to mitogenic stimuli in the presence of exogenous IL-2 of PBMC from allogeneic BMT recipients cannot be simply attributed either to the putative suppressor activity of CD57+ cells, or to a poor proliferative capacity of this subset. Supporting this notion we report that PBMC from long-term autologous BMT recipients containing high proportions of CD57+ T cells respond normally to T cell mitogens.
Quantitative measurement of cytomegalovirus-specific IgG and IgM antibodies in relation to cytomegalovirus antigenaemia and disease activity in kidney recipients with an active cytomegalovirus infection
Authors: VAN DER GIESSEN M , VAN DEN BERG A P , VAN DER BIJ W , POSTMA S , VAN SON W J , THE T H
Published: July 2010

In a longitudinal investigation 103 kidney recipients were studied with respect to the development of cytomegalovirus (CMV) specific antibodies of the IgG and IgM class, in relation to the detection of CMV antigenaemia (immediate early antigen. IEA), in weekly obtained blood samples during the first 3 months after transplantation. In 15 out of 49 (31%) seronegative patients a primary infection occurred, which was characterized by a quick rise in IgM antibody followed by a slower production of IgG antibody, high maximum numbers of IEA+ cells, and a CMV syndrome in 11 patients, ln 35 out of 54 (65%) seropositive patients a secondary infection occurred. After a post-operative fall in the IgG antibody, which was also found in patients without an active infection and which was accompanied by a similar drop in scrum albumin and IgG, a second dip in IgG antibody was found 6 days before the first IEA+ leucocyte appeared in the blood. This was followed by a significant increase, indicative of an active immune response in consequence of the infection. 18 days later. In 31 of these 35 patients an IgM response was found. This could be ascribed to the presence of rheumatoid factor activity in 20 of them. Eight patients who showed a transient rise in IgG antibody between the two dips could be distinguished from the remaining ones by a lower maximum number of IEA+ cells and less severe disease symptoms. The described results suggest that (i) an adequate humoral immune system may prevent symptomatic CMV disease in secondary infections; and (ii) CMV-specific antibodies may be removed from the circulation by antigens present in infected tissues before CMV antigenaemia becomes detectable.
Elevated binding activity of CD8+ cells with phytohaemagglutinin by asbestos fibrein vitro
Authors: KINUGAWA K , HYODOH F , ANDOH A , UEKI A , TANAKA H , MOCHIZUKI Y
Published: July 2010

Asbestos fibres are known to depress the mitogenic stimuli of phytohaemagglutinin (PHA) to lymphocytes. We examined effects of asbestos (chrysotile) fibre on the proliferation of PHA-stimulated lymphocytes and the PHA binding activity of lymphocytes in vitro. The incorporation of 3H-thymidine and the expression of interleukin-2 receptor were depressed when the cells were exposed to 50 μg/ml of chrysotile fibre. The PHA binding activity of lymphocytes was significantly enhanced after chrysotile fibre exposure, as compared with non-exposed group. These results indicate that the depression of PHA stimuli with chrysotile fibre was not due to blocking of PHA binding to lymphocytes. The enhancement of PHA binding activity by chrysotile fibre was observed on CD8+ but not on CD4+ cells. It is possible, therefore, that treatment of peripheral blood mononuclear cells with chrysotile fibre induces increased reactivity of CDS+ cells with PHA, and that intensely activated CD8+ cells suppress proliferation of lymphocytes.
Acute shift in immune response to microbial activators in very-low-birth-weight infants
Authors: VEBER M B , CUNNINGHAM-RUNDLES S , SCHULMAN M , MANDEL F , AULD P A M
Published: July 2010

Investigation of lymphocyte activation in vitro to microbial pathogens was undertaken in very-low-birth-weight infants during the first 2 weeks of life. Twenty-three infants with birth weights less than 1500 g were studied on day 1. Normal adults (n= 23) and cord blood from seven full-term infants were used as controls. Longitudinal studies were also carried out on seven of the 23 infants 2 weeks following delivery. Results indicated that lymphocyte responses of very-low-birth-weight infants on day 1 of life were significantly greater than those of both adult controls and full-term infants, particularly to Haemophilus influenzae. Staphylococcus epidermidis and Staphylococcal protein A. In contrast, response to the T cell mitogen phytohaemagglutinin (PHA) was significantly less in very-low-birth-weight infants than in adult controls and full-term infants. The seven very-low-birth-weight infants studied showed a down-regulation of immune response in the 2 weeks following birth, such that responses on day 14 were significantly less than those on day 1 for the same activators. This shift in immune response appears to have important implications for the immune development and host defence in the post-natal period.
Characterization of antibodies reacting with HIV gag proteins occasionally found in the serum of non-infected subjects
Authors: BÜRGISSER P , FREI P C
Published: July 2010

The use of serological tests for the diagnosis of HIV infection has revealed that some non-infected persons have antibodies that react with HIV-1 gag proteins. Here, the sera of three non-infected subjects reacting with p 17 and 11 non-infected subjects reacting with p24 were investigated, using an enzyme immunoassay (EIA) with six recombinant gag antigens and Western blot analysis of proteolytic peptides of two of these gag antigens. The results indicate that whereas all p 17-reactive sera could react with an unique epitope, individual p24-reactive sera recognize different epitopes. Investigations by EIA also demonstrated the role of sequences located far from the epitopes in making these epitopes accessible to the antibodies or in providing them with an antigenic conformation. In addition to the 14 subjects mentioned above, another subject was shown to have antibodies reacting with the p9 (NC) gag protein. Several proteins are known as having homology with HIV-1 gag proteins. Their possible role in eliciting cross-reactive antibodies is discussed.
Genetic control of Coxsackievirus B3-induced heart-specific autoantibodies associated with chronic myocarditis
Authors: TRAYSTMAN M D , BEISEL K W
Published: July 2010

Cardiac-specific autoantibodies to sarcolemmal and cardiac myosin antigens observed during the chronic phase of Coxsackievirus B3-induced myocarditis appear to be under autosomal recessive control. This observation is based on examination of F1 hybrids bred from A/J mice which develop chronic myocarditis and C57BL/6J mice which resolve the virus-induced lesions. Previous mouse studies demonstrated that the prevalence of heart-specific autoantibodies varied with the H-2 complex. However, in 25 H-2 congenic mouse strains the strain background was the predominant determinant of autoantibody presence. Recently, we extended our genetic evaluation of the chromosomal locations governing autoantibody responses by examining 25 AXB and BXA recombinant inbred strains. Two populations of heart-specific autoantibodies were demonstrated against sarcolemmal and cardiac myosin antigens. Analyses of the AXB/BXA strain distribution patterns for these two traits revealed that the anti-sarcolemmal response was controlled by a gene(s) linked to Np-2 and Terα loci on chromosome 14. Linkage could not be assigned for the anti-cardiac myosin response.
Alpha-1 anti-trypsin and CD30 expression occur in parallel in activated T cells
Authors: BASHIR M S , JONES D B , WRIGHT D H
Published: July 2010

We have performed an immunohistochemical study of immunoblastic T cell lymphoma and enteropathy-associated T cell lymphoma for alpha-1 anti-trypsin and CD30. Cytoplasmic staining for α1 anti-trypsin is present in the malignant cells in both types of T cell neoplasm which also express CD30, a marker of lymphoid activation. Peripheral blood T lymphocytes on stimulation with mitogen also show granular cytoplasmic expression of α1 anti-trypsin. Time course studies show that this parallels the expression of CD30. α1 anti-trypsin expression appears therefore to be associated with activation in T cells. Further studies of sub-fractionated T lymphocytes in vitro suggest that the expression of α1 anti-trypsin on activation is not restricted to an individual lymphocyte subset.
Antigen-dependentin vitroculture of protective T cells from BCG-primed mice
Authors: D'SOUZA S , IVANYI J
Published: July 2010

Induction of protective immunity against pathogenic mycobacteria depends on vaccination with live organisms such as Bacille Calmette-Guerin (BCG). However, it is not known how many and which antigens are involved in the protective host response. In this study, we developed a system of antigen-dependent in vitro culture which is suitable for the analysis of protective subunits, presented in a soluble form. Spleen cells from Mycobacterium bovis BCG-immune mice, enriched for T cells and depleted of adherent cells on a column of G-10 Sephadex, were cultured for periods varying between 3 and 14 days before transfer and challenge with M. tuberculosis in irradiated hosts. Following 10 days in culture, immune T cells sustained their capacity to transfer protection to tuberculous infection when incubated in the presence of either live BCG or a soluble extract from M. tuberculosis, but lost this ability when cultured in the absence of antigen, or in the presence of the polyclonal mitogen concanavalin A. One immunodominanl antigen, represented by the recombinant 38-kD antigen, failed to sustain the adoptive protection, despite pronounced stimulation of lymphoproliferation in culture. Antigenic in vitro stimulation of protective T cells was accompanied by enhanced responsiveness to exogenous IL-2. The experimental system described may be generally suitable to test in vitro the protective potentials of soluble molecular subunits of mycobacteria.
Ro/SSA inhibits the autologous mixed lymphocyte reaction
Authors: KARSH J , HARLEY J B , GOLDSTEIN R , LAZAROVITS A I
Published: July 2010

To test the hypothesis that the Ro/SSA autoantigen can be recognized as antigenic by the human immune system, lymphocytes obtained from normal volunteers were used in in vitro assays evaluating the ability of Ro/SSA (mol. wt 60 kD) to induce B and/or T cell responses. Bovine Ro/SSA strongly inhibited the autologous mixed lymphocyte reaction in a dose-dependent manner without similar effects on concurrently performed allogeneic mixed lymphocyte reactions or T cell proliferation induced by phytohaemagglutinin. Using three colour FACS analysis, Ro/SSA was found to decrease the percentage of CD4+CD45+RA+ T cells in the proliferative, S+(G2+ M), phase of the cell cycle. Associated with the decrease in the percentage of suppressor-inducer cells, was the finding that Ro/ SSA was able to augment RF production in pokeweed mitogen stimulated cultures of peripheral blood lymphocytes.
Role of tumour necrosis factor and reactive oxygen intermediates in lipopolysaccharide-induced pulmonary oedema and lethality
Authors: GATTI S , FAGGIONI R , ECHTENACHER B , GHEZZI P
Published: July 2010

The purpose of this study was to characterize the role of tumour necrosis factor (TNF) and neutrophils (PMN) in the pathogenesis of pulmonary oedema induced by endotoxin (lipopolysac-charide (LPS)). Intraperitoneal administration to BALB/c mice of 0–6–1 mg of LPS caused pulmonary oedema and lethality. This was associated with production of TNF in serum and bronchoalveolar lavage fluid and with accumulation of PMN in the lung. In this experimental model, we could block TNF production by different means: pretreatment 30 min before LPS with 4 mg/kg of i.p. chlorpromazine (CPZ), 3 mg/kg of i.p. dexamethasone (DEX), 1 g/kg p.o. of N-acctylcysteine (NAC, an antioxidant precursor of glutathione), or an anti-TNF MoAb. CPZ, DEX and anti-TNF completely prevented LPS lethality but not pulmonary oedema or pulmonary PMN infiltration, indicating that: (i) lung oedema is not the main cause of death after LPS; and (ii) lung oedema induced by LPS is not mediated by TNF. Pretreatment with NAC not only inhibited TNF production but also protected against LPS-induced pulmonary oedema, indicating that reactive oxygen intermediates are implicated. NAC also blocked TNF production in blood and in bronchoalveolar lavage. We also tested the effect of PMN depletion induced with cyclophosphamide (CP) or 5-fluorouracil (5-FU). While no pulmonary PMN infiltrate was observed in PMN-depleted mice, neutropenia did not prevent LPS lethality or oedema, indicating PMN do not play an important role in the toxic effects of LPS in this experimental model.
The putative oncoprotein DEK, part of a chimera protein associated with acute myeloid leukaemia, is an autoantigen in juvenile rheumatoid arthritis
Authors: SIERAKOWSKA H , WILLIAMS K R , SZER I S , SZER W
Published: July 2010

The 45-kD autoantigen associated with juvenile rheumatoid arthritis (JRA) has been isolated from HeLa cell nuclei and purified about 2500-fold to near homogeneity in a five-step chromatographic procedure. Purification of the antigen was monitored by immunoblot assays using a nearly monospecific anti-45-kD serum from a child with JRA. Tryptic peptide mapping and partial amino acid sequencing of the purified 45-kD antigen demonstrated its identity with the DEK protein. DEK is a 43-kD protein of unknown function expressed by the putative oncogene dek located on chromosome 6. As a result of a (6;9) translocation associated with a rare subtype of acute myeloid leukaemia a chimeric protein containing most of DEK amino acids at the N-terminus is found in leukaemic cells (von Linden et al., Mol Cell Biol. 1992; 12: 1687–97). The 43-kD DEK was detected by immunoblotting with serum from a patient with JRA in a variety of rat tissues, and was most abundant in the spleen and in bone marrow.
The γδ T cell repertoire in Graves' disease and multinodular goitre
Authors: McINTOSH R S , TANDON N , PICKERILL A P , DAVIES R , BARNETT D , WEETMAN A P
Published: July 2010

γδ T cells are a subset of T cells with unknown function, and restriction of the γδ T cell receptor (TCR) repertoire has been described in rheumatoid arthritis and multiple sclerosis. Elevated numbers of γδ cells have been reported in the peripheral blood and thyroids of patients with Graves' disease. We have carried out flow cytometric analysis on peripheral blood mononuclear cells (PBMC) and intrathyroidal lymphocytes (ITL) from 12 patients with Graves' disease and nine patients with multinodular goitre (MNG). a thyroid disease of unknown etiology. There was no significant difference between the proportion of γδ T cells in the PBMC of Graves' and MNG patients, nor between the PBMC and ITL populations in either patient group. We have also carried out polymerase chain reaction amplification on RNA prepared from matched PBMC. ITL and the activated (CD25+) subset of ITL using six TCR γδ-family specific primers. Although there were differences in the amounts of each γδ transcript amplified from PBMC and ITL, there was no difference between the two patient groups. No consistent differences were therefore found between the γδ T cell populations in Graves' and MNG patients, arguing against the direct involvement of this T cell subset in the pathogenesis of Graves' disease.
Bone marrow transplantation therapy using resistant donors for retrovirus-induced leukaemia in mice
Authors: IWAI H , DAY N K , HAMADA N , INABA M M , IKEHARA S , GOOD R A
Published: July 2010

Infection with Rauscher leukaemia virus (RLV) causes erythroleukaemia, anaemia, viraemia, and splenomegaly, features which develop very quickly in mice of strains susceptible to RLV but more slowly or not al all in strains resistant to RLV. We compared the effects of hone marrow transplantation (BMT) of donor cells obtained from a mouse strain resistant lo the virus with the effects of BMT of donor cells obtained from strains susceptible to the virus, in recipient virus-susceptible mice which had previously been inoculated with RLV. Our results indicated that bone marrow transplantation from a marrow-resistant donor, but not from a susceptible donor, delayed the expression of leukaemia and delayed death, although such treatment did not appear to inhibit viral infection in susceptible recipients.
Influence of theIty/Lsh/Bcggene on the development of suppressor cell precursors in the early phase of the infection of mice withMycobacterium lepraemurium
Authors: GOSSELIN D , TURCOTTE R , LEMIEUX S
Published: July 2010

In vitro inducible suppressor cell precursors were detected in the spleen of BALB/c but not in DBA/2 mice infected intraperitoneally with 108Mycobacterium lepraemurium bacilli, thus suggesting that their development is genetically controlled. Two pairs of mouse strains congenic at the Ity/Lsh/Bcg locus (BALB/c-C.D2 and B10.A-B10.A.Beg) were used to investigate whether this phenomenon is influenced by this gene known to control the relative susceptibility of mice to Myco. lepraemurium infection. This seems likely, as the detection of culture-induced suppressor activity was delayed for 5–6 weeks in C.D2 and B10.A.Bcgr mice infected intravenously with 104Myco. lepraemurium bacilli. However, despite the retardation in the detection of suppressor cell precursors, the level of in vitro induced suppressor activity at onset in spleen cell suspensions of mice carrying the resistant allele was higher than in cell cultures derived from susceptible mice. As the resistant allele has a different effect when found on BALB/c or DBA/2 background, other genetic factors are apparently involved in the development of suppressor cell precursors. We finally observed that, in spleen cell cultures from intravenously infected Ity/Lsh/Bcg congenic mice on the BALB/c background, adherent and non-adherent cells were required in the inductive phase of suppressor cell development, whereas in vitro induced suppressor activity was found exclusively in the adherent cell fraction. Given these properties, we thus conclude that suppressor cell precursors detected in the spleen of these intravenously infected mice are similar to those previously observed in C3H mice infected intraperitoneally with a thousand times more bacilli.
The immunomodulatory activity of human amniotic fluid can be correlated with transforming growth factor-beta 1 (TGF-β1) and β2 activity
Authors: LANG A K , SEARLE R F
Published: July 2010

The role of alphafetoprotein (AFP) in the immunomodulatory activity of amniotic fluids (AE) from normally progressing human pregnancy (weeks 14–16) was investigated. A panel or42 AF (25% v/v) reduced significantly phytohaemagglutinin (PHA)-induced peripheral blood mononuclear cell (PBMC) proliferation in serum-free cultures with a mean per cent inhibition of 68·4 ± 5·5%. In contrast, AEP preparations, with one exception (U.AFP), failed to display inhibitory activity. Pretreatment of AF with anti-TGF-β1 and β2 antibodies used alone resulted in the mean per cent loss of inhibition of 33·1 ± 3·9% and 52·3 ± 7·5%, respectively. A summative loss of AF-mediated inhibition was detected when anti-TGF-β1 and β2 antibodies were used in combination, but immunomodulation was rarely abolished 100% by this treatment. Anti-TGE-β2 antibody treatment, unlike anti-TGF-β1 antibody treatment, reversed the inhibitory activity of U.AFP. The amount of TGF-β1 and β2 contained in human AF was studied by growth inhibition of Mv1 Lu cells. The mean levels of TGF-β1 and β2 in AF were 11 ± 0·9 U/ml and 2·3 ± 0·4 U/mI, respectively, which corresponds with a mean per cent inhibition of 49 ±4·7%. U.AFP also significantly inhibited Mv1 Lu cell growth. To investigate the mechanism of AF-mediated inhibition, the effect of AF and AEP on IL-2 production by concanavalin A (Con A)-stimulated PBMC blasts was determined by the CTLL-2 cell bioassay. IL-2 production was reduced 55·5% in AF-treated blasts and 61% in U.AEP-treated blasts compared with controls. Our findings indicate that the immunomodulatory activity of human AF can be correlated with TGE-β1 and β2 and not with AFP, the inhibitory activity of U.AFP preparation reflecting copurifying TGF-β2 activity.
Analysis of idiotope structure of ovarian cancer antibodies: recognition of the same epitope by two monoclonal antibodies differing mainly in their heavy chain variable sequences
Authors: SLOBBE R , POELS L , TEN DAM G , BOERMAN O , NIELAND L , LEUNISSENJ J , RAMAEKERS F , VAN EYS G
Published: July 2010

Two MoAbs, independently raised against ovarian carcinoma cells and referred to as OV-TL3 and OV-TL16, display an identical reaction pattern with a membrane-associated protein in both normal and malignant ovarian cells. Also, a similar binding affinity constant and a similar number of binding sites per cell indicate that both MoAbs bind to the same antigen. Competition assays reveal that OV-TLI6 is able to compete with OV-TL3 for binding to OVCAR-3 cells. Epitope mapping using a filamentous phage hexapeptide epitope library showed that both MoAbs are able to select identical phages, suggesting that their epitopes are identical or at least overlapping. However, purified polyclonal and monoclonal anti-idiotypic antibodies directed against OV-TL3 failed lo recognize the OV-TL16 idiotype, indicating that the structure of the antigen-binding regions of both antibodies is distinct. This was corroborated by molecular cloning and sequencing of the variable heavy (VH) and light (VL) chain immunogiobulin regions of both MoAbs. The VH regions of both antibodies were found to be distinct, whereas the VL regions are almost identical. Computer modelling of the idiotypes suggests that the complementarity determining regions (CDR), with the exception of VHCDR3, have (almost) identical spatial configurations. Our data indicate that, although structurally different in their VH regions, OV-TL3 and OV-TL16 are able to bind to identical epitopic regions on the antigen, because differences in primary structure do not exclude the formation of sufficient and similar spatial structures for the interaction with an epitope.
Soluble CD4 antigen reactivity in intravenous immunoglobulin preparations: is it specific?
Authors: Perosa F , Rizzi R , Pulpito V , Dammacco F
Published: July 2010

Soluble CD4 antigen (sCD4) was measured in seven commercially available intravenous immunoglobulin preparations (IVIg) by means of a double determinant immunoassay (DDIA), whereby two MoAbs recognizing two distinct and spatially distant epitopes on CD4 were used to capture and detect the antigen, respectively, Preincubation of six out of seven IVIg, which were found to be apparently positive for sCD4, with mouse- and bovine-derived serum or purified immunoglobulins completely neutralized DDIA reactivity for sCD4. The inhibition was specific since it was not or only partially observed when IVIg were mixed with whole serum or purified IgG from rabbit. Extensive absorption of six IVIg on insolubilized mouse IgG (mIgG) resulted in a complete loss of reactivity. Eluted human anti-mouse antibodies (HAMA) from any of the IVIg displayed a dose-dependent binding in a DDIA, though its extent varied from one preparation to another. Western blot analysis showed that HAMA from all IVIg contained no component with a molecular weight identical with or close to that of recombinant CD4. Purified mIgG markedly influenced the sCD4 reactivity of two IVIg (Sandoglobulin and Globuman I.V.) when sCD4 was measured with a purchased ‘CD4-specific Test Kit’, thus suggesting that HAMA can exceed the absorbing capacity of the sample diluent. Taken as a whole, these data indicate that sCD4-based DDIA signal is mostly, if not completely, generated by the presence of human immunoglobulin with anti-mouse immunoglobulin reactivity, thus casting doubts on the actual occurrence of sCD4 in IVIg.
Response ofscidmice to establishment ofLeishmania majorinfection
Authors: GUY R A , BELOSEVIC M
Published: July 2010

The initiation of Leishmania major infection in susceptible BALB/c mice is regulated by interferon-gamma (IFN-γ). To examine further the mechanisms of IFN-γ-dependent regulation of the establishment of L. major, we studied the characteristics of the infection in severe combined immunodeficient (scid) mice. In the first 2 weeks of infection, we observed a delay in the development of the lesions in the footpads and lower numbers of parasites in scid compared with BALB/c mice. By week 5 after infection, the size of the leishmanial lesion was similar in both strains of mice, but the number of parasites in scid mice was 100-fold higher than in BALB/c. Treatment with anti-IFN-γ during the establishment of L. major did not alter the course of infection in scid mice, while it exacerbated lesion development in BALB/c mice. Macrophages from scid mice were unable to kill L. major when stimulated with IFN–γin vitro, and produced lower levels of nitric oxide compared with macrophages from susceptible BALB/c or the resistant C57B1/6 mice. We examined whether delayed lesion development in scid mice was due to their inability to mount appropriate inflammatory responses. While significantly fewer nucleated cells were present in the footpads of scid mice compared with BALB/c. 2 and 3 weeks after infection, no difference in inflammatory response between scid and BALB/c mice was observed in response to L. major antigen in the footpads. In contrast, there was a dramatic increase in the number of cells in the popliteal lymph nodes of BALB/c mice. Decreased inflammatory responses of scid mice in the footpad (at the site of infection) may contribute to slower development of leishmanial lesions during the first 2 weeks of infection.
Circulating intercellular adhesion molecule-1 (ICAM-1) as an early and sensitive marker for virus-induced T cell activation
Authors: Christensen J p , Johansen J , Marker O , Thomsen A R
Published: July 2010

The effect of systemic virus infection on the level of circulating ICAM-1 (cICAM-1) in serum, and the role of virus-activated T cells in this context, were studied using the murine lymphocytic choriomeningitis virus infection as primary model system. A marked virus-induced elevation in cICAM-1 in serum was revealed, the presence of which coincided with the phase of virus-induced T cell activation. However, high levels of cICAM-1 in serum were observed well before maximal T cell activation could be demonstrated. No increase in cICAM-1 was observed in the serum of infected T cell-deficient nude mice, clearly demonstrating that T cells were mandatory. Analysis of MHC class I and MHC class II-deficient mice revealed that either CD4+ or CDS+ T cells alone are sufficient, despite a markedly reduced inflammatory exudate in the former animals. These results indicate that virus-activated T cells induce shedding of ICAM-1 into the circulation, and this parameter may be used as an early and sensitive marker for immune activation.
Molecular characteristics of anti-self antibody fragments against neutrophil cytoplasmic antigens from human V gene phage display libraries
Authors: Finnern R , Bye J M , Dolman K M , Zhao M-H , Short A , Marks J D , Wood M C LOCK , Ouwehand W H
Published: July 2010

Recently it has been demonstrated that human antibody fragments with binding activities against self antigens can be isolated from repertoires of rearranged V genes from non-immunized humans. We have applied phage display technology to study the B cell repertoire for antibody activity against neutrophil cytoplasmic antigens. These antibodies may play an important role in Wegener's granulomatosis (WG) and related forms of vasculitides. Autoantibodies in patients with WG are directed against proteinase 3. The immunodominant antigen in other forms of vasculitis is myeloperoxidase, but the B cell response can also be directed against other neutrophil enzymes, e.g. lysozyme, human neutrophil elastase, lactoferrin and cathepsin G. We show here that anti-self reactivity against neutrophil cytoplasmic antigens can be detected in the rearranged V gene repertoire of healthy individuals and that the reactivity can be directed against structural related epitopes which are present on different neutrophil cytoplasmic antigens. The scFv with binding activities were sequenced and the V gene usage, the level of somatic mutations and the immunoserological characteristics of the antibody fragments are discussed. Further evidence is presented that antibody fragments consisting only of a heavy chain variable domain can recognize neutrophil cytoplasmic antigens in a specific manner. These single-domain antibody fragments were used in experiments designed to establish the relative role of the light chain variable domains in antigen binding.