bluuebunny/crossref_metadata_2025_split huggingface dataset:
A 1063G→A mutation in exon 12 of glycoprotein (GP)IIb associated with a thrombasthenic phenotype: mutation analysis of [324E]GPIIb
Authors: Tao Jianming , Arias‐Salgado Elena G. , González‐Manchón Consuelo , Iruín Gemma , Butta Nora , Ayuso Matilde S. , Parrilla Roberto 
Published: July 2010

We report the molecular, genetic and functional analysis of a case of thrombasthenic phenotype. The proband showed absence of platelet glycoprotein (GP)IIb and very low content of GPIIIa, and both his parents showed a marked reduction in the levels of platelet GPIIb‐IIIa. Single‐stranded conformational polymorphism‐polymerase chain reaction (SSCP‐PCR) analysis and direct sequencing of PCR‐amplified GPIIb exon‐12 revealed the presence of a G→A transition at position 1063 with the expected substitution of glutamate 324 with lysine (K). This mutation did not alter the level of GPIIb mRNA. Co‐expression of normal or mutant [324K] GPIIb with normal human GPIIIa in Chinese hamster ovary (CHO) cells failed to show surface exposure of [324K]GPIIb‐IIIa complexes. Pulse‐chase and immunoprecipitation analysis demonstrated that [324K]GPIIb cDNA was translated into proGPIIb, but neither mutant GPIIb heavy chain (GPIIbH) nor [324K]GPIIb‐GPIIIa complexes were detected, suggesting that this mutation is the underlying molecular basis for the thrombasthenic phenotype. Mutation analysis demonstrated that 324E of GPIIb could be replaced by other negatively charged or polar amino acids (AAs) without impairing the surface expression of GPIIb‐IIIa. However, substitution of 324E of GPIIb for a positively charged AA other than K prevented the expression of GPIIb‐IIIa complexes. These observations suggest that a domain encompassing 324E of GPIIb is essential for heterodimerization with GPIIIa and its substitution for a positively charged residue precludes normal subunit association.
Fibrinogen γ′ chain carboxy terminal peptide selectively inhibits the intrinsic coagulation pathway
Authors: Lovely Rehana S. , Boshkov Lynn K. , Marzec Ulla M. , Hanson Stephen R. , Farrell David H. 
Published: July 2010

SummaryThe minor γA/γ′ isoform of fibrinogen contains a high affinity binding site for thrombin exosite II that is lacking in the major fibrinogen isoform, γA/γA fibrinogen. The biological consequences of γ′ chain binding to thrombin were therefore investigated. Coagulation assays, thrombin activity assays, and a primate thrombosis model were used to characterize the biological effects of the γ′ 410–427 peptide. The γ′ peptide had little effect on thrombin cleavage of the small peptidyl substrate tosyl‐glycyl‐prolyl‐arginine‐4‐nitranilide acetate. However, in vitro assays demonstrated that the γ′ peptide inhibited thrombin cleavage of larger proteinaceous substrates, including fibrinogen and factor VIII. The γ′ peptide inhibited the activated partial thromboplastin time in plasma and showed greater inhibition of activated partial thromboplastin time assays than prothrombin time assays, consistent with the inhibition of factor VIII cleavage. Studies in a baboon thrombosis model showed that the γ′ 410–427 peptide inhibited fibrin‐rich thrombus formation (typical of venous thrombi) and, to a lesser extent, platelet‐rich thrombus formation (typical of arterial thrombi). These results indicate that binding of thrombin exosite II by the γ′ peptide has selective effects on the intrinsic pathway.
Identification and functional characterization of a polymorphic oestrogen response element in the human coagulation factor IX gene promoter
Authors: Adams B. , Western A. K. , Winship P. R. 
Published: July 2010

SummaryThe liver expressed procoagulant factor IX (FIX) shows inter‐individual variation in levels, some of which is heritable. Raised levels of FIX are associated with a thrombotic tendency. This study demonstrated that, in females but not males, part of this variation in FIX levels is due to polymorphic genotype at a locus in the factor IX gene (F9) promoter 698 bp upstream of the major transcription initiation site (−698C/T). The −698C allele (associated with higher FIX level) shows closer homology to a canonical ORE sequence and a higher binding affinity for oestrogen receptor α than the −698T allele. Reporter gene vectors were constructed with elements spanning residues −738 to +50 of the F9 promoter corresponding to wild type −698C and −698T alleles. A related series of vectors comprising three copies of the F9 ORE driving expression of a minimal synthetic promoter were also created. Transfection into the liver‐derived HepG2 and erythroleukaemic K562 cell lines demonstrated increased levels of expression in the presence of oestrogenic factors when compared to those found in their absence; this stimulation was more pronounced in the non‐liver derived K562 cell line and from the reporter vectors containing promoter elements corresponding to the −698C allele.
IgG subclass levels and immune reconstitution after T cell-depleted allogeneic bone marrow transplantation
Authors: KELSEY S M , LOWDELL M W , NEWLAND A C 
Published: July 2010

SUMMARY
Defects of humoral immunity are well documented after bone marrow transplantation (BMT). Immunoglobulin recovery can be impaired and selective deficiencies of IgG subclasses have been reported. The nature of these deficiencies may reflect patterns of infection in the post-BMT period. We studied immunoglobulin and IgG subclass recovery in 20 long term (&gt; 100 days) survivors of T depleted allogeneic BMT. Although there was no fall in mean levels of IgG, IgM or IgA for the patient group, 14 patients (70%) developed a deficiency of one or more immunoglobulin isotype at some stage post-BMT. Eight patients (40%) had deficiency of IgG, IgA and IgM and six had selective deficiencies. When IgG subclasses were measured it was seen that mean levels of IgG2 and IgG4 fell post-BMT with trough levels occurring at around 120 days post-transplant. Sixty per cent of patients developed IgG2 subclass deficiency and of these patients 78% had an associated IgG4 deficiency. Deficiencies of IgG1 and IgG3 were less common and less prolonged than those of lgG2 and lgG4; in addition, mean levels of IgG1 and IgG3 showed a rise early post-BMT. In conclusion, a majority of our patients developed immunoparesis following BMT, usually at around 120 days after transplantation. IgG2 subclass deficiency, often in association with IgG4 deficiency, is common and may occur despite normal total IgG levels. Deficiencies of immunoglobulin and IgG subclasses may persist for longer than 1 year post-BMT. Differing profiles of immunoglobulin and IgG subclass recovery may help dictate patterns of infection in long-term survivors of BMT.
Modulation of leucocyte locomotion by interleukin-1
Authors: ZIELINSKI C C , PESAU B , KALINOWSKI W , MÜLLER C 
Published: July 2010

SUMMARYThe influence of interleukin-1 (IL-1) upon leucocyte locomotion in vitro was studied, using either casein or zymosan-activated serum (ZAS) as chemotaxigens. A pre-incubation of polymorpho-nuclear cells (PMNC) with ultrapure (purity 99%) human IL-1 (1 U/ml) for 2 and 5 h followed by a washing step resulted in a significant decrease in leucocyte locomotion against casein (P &lt; 0.0005 and P&lt;0.01, respectively), but not against ZAS (P&gt;0.1). Moreover, the direct addition of this IL-1 preparation to leucocyte locomotion assays without pre-incubation produced a similar and significant inhibition of leucocyte locomotion directed against casein (P &lt; 005). This inhibitory effect could not be augmented further by higher concentrations of ultrapure IL-1. In order to exclude the effect of possibly contaminating cytokines in the ultrapure IL-1 preparation used, additional assays with recombinant human IL-1 (rIL-1) alpha and rIL-1 beta (1 U/ml) were performed to investigate their influence upon leucocyte locomotion. It was found that both rIL-1α and rIL-1β inhibited leucocyte locomotion directed against casein significantly (P &lt; 0.005). Similar to the previous experiments, leucocyte locomotion could not be further inhibited by higher concentrations (10 U/ml) of rIL-1. Thus, both ultrapure and rIL-1 were found to have the ability to inhibit leucocyte locomotion in vitro.
Oestrogen is a potent disease accelerator in SLE-prone MRL lpr/lpr mice
Authors: CARLSTEN H , TARKOWSKI A , HOLMDAHL R , NILSSON L-Å 
Published: July 2010

SUMMARYThe influence of oestrogen on the lupus disease in MRL/1 mice has been investigated. Adult, castrated male and female MRL/1 mice were administered with s.c. injections of 3–2 μg of 17β-oestradiol twice a week. The results clearly demonstrate that a relatively small dose of oestrogen is a potent accelerator of the lupus disease in this mouse strain. Thus, administration of oestrogen accelerates glomerulonephritis, lymphoproliferation and mortality. Our results also indicate that oestrogen exerts a dual effect on the immune system of MRL/1 mice by depression of antigen-specific and mitogen-induced T cell responses as well as enhancement of polyclonal Bcell activation and autoantibody formation. In addition, even short-term administration of oestrogen in the preclinical phase of the disease resulted in long-lasting effects as evaluated by reduced longevity and aggravation of renal disease.
Depletion of CD8+ T cells suppresses growth of Trypanosoma brucei brucei and interferon-gamma) production in infected rats
Authors: BAKHIET M , OLSSON T , VAN DER MEIDE P , KRISTENSSON K 
Published: July 2010

SUMMARY
Sprague-Dawley rats infected with Trypanosoma brucei brucei showed a strong and rapid induction of splenocyte IFN-γ (within 12 h post-infection) as measured by a single cell assay for IFN-γ secretion. Depiction of CD8+ cells in infected rats abrogated the IFN-γ production, suppressed parasite growth and increased survival of the animals. Induction of MHC class I antigens in the paraventricular and supra-optic hypothalamic nuclei caused by the trypanosome infection was also inhibited by the CD8+ cell depletion. It is suggested that the CD8+ cells are involved directly or indirectly in growth regulation of the parasite and that IFN-γ induced by the parasite may be one of the factors that trigger MHC expression and immunosuppression.
CD4+ T cells play a major role for IgM and IgG anti-DNA production in mice infected withPlasmodium yoelii
Authors: FOSSATI L , MERINO J , IZUI S 
Published: July 2010

SUMMARYThe infection by a non-lethal strain of Plasmodium yoelii induces the formation of autoantibodies such as anti-DNA and anti-Sm antibodies in mice. The extent of the relative increase in serum levels of IgM and IgG anti-DNA and anti-Sm antibodies and their kinetics were found to be similar to those of anti-hapten antibodies and of total IgM and IgG levels. This strongly suggested that anti-DNA and anti-Sm autoantibody responses observed in malaria-infected mice are a result of polyclonal activation of B cells. The analysis of the IgG subclasses reacting with DNA antigen showed significant levels of the T cell-dependent isotypes, IgG1 and IgG2. The role of T cells in the activation of autoreactive B cells was confirmed by using athymic nude mice. Indeed, BALB/c-nu/nu and C57BL/6-nu/nu mice failed to produce IgG anti-DNA antibodies after infection with P. yoelii. Moreover, the reconstitution of BALB/c nude mice with lymph node cells from congenic euthymic BALB-Igb mice showed the activation of autoreactive B cells in nude mice by T cells from euthymic mice. Studies in mice depleted of CD4+ T cells strongly suggested that malaria-induced anti-DNA antibodies were almost entirely dependent on the presence of CD4+ T cells, as this depletion significantly decreased IgM anti-DNA antibodies and completely abolished the IgG anti-DNA production, including the IgG3 subclass in infected mice. In contrast, depletion of the CD8+ T cell subset had no effect on the production of autoantibody in malaria-infected mice. Our results indicate that CD4+ T cells play a major role for both IgM and IgG anti-DNA production during the course of malaria infection.
Mononuclear cells in glomeruli and cytokines in urine reflect the severity of experimental proliferative immune complex glomerulonephritis
Authors: NOBLE B , REN K , TAVERNE J , DIPIRRO J , VAN LIEW J , DUKSTRA C , JANOSSY G , POULTER L W 
Published: July 2010

SUMMARYImmunohistochemical methods were used to investigate the role of macrophages in the progression of proliferative immune complex glomerulonephritis. The mononuclear cell component of glomerular inflammation was analysed in three different stages of chronic serum sickness, each of which was clearly distinguished by criteria of kidney function. Urinary excretion of the macrophage secretory products interleukin-1 and tumour necrosis factor was also evaluated in relation to the functional severity of kidney disease. T lymphocytes and macrophages began to accumulate in glomeruli at the onset of proteinuria, but not before. Urinary excretion of interleukin-1 also began with proteinuria. Proteinuria increased in direct correlation with increases in the number of glomerular macrophages. Development of the most severe stage of glomerulonephritis, characterized by cachexia, declining kidney function, and necrotizing glomerular pathology, was accompanied by the disappearance of T cells from glomeruli and the expression of highly abnormal phenotypes by most macrophages. In addition, there was a switch from urinary excretion of interleukin-1 to excretion of tumour necrosis factor. The progression of proliferative immune complex glomerulonephritis was associated with qualitative as well as quantitative changes in glomerular macrophage populations. Differentiation and/or activation of those glomerular macrophages may have resulted from local T cell-mediated immunoregulation. Measurements of urinary cytokine excretion provided a reliable means of monitoring disease progression. The local action of tumour necrosis factor probably contributed to declining kidney function in the most severe stage of disease.
Isolation of a Trypanosoma cruzi antigen by affinity chromatography with a monoclonal antibody. Preliminary evaluation of its possible applications in serological tests
Authors: CARBONETTO C H , MALCHIODI E L , CHIARAMONTE M , DE ISOLA E DURANTE , FOSSATI C A , MARGNI R A 
Published: July 2010

SUMMARY
By affinity chromatography with a monoclonal antibody (163B6), obtained in our laboratory, we have isolated a T. cruzi antigen which could be useful for differential diagnosis of Chagas' disease from leishmaniasis. This antigen, a 52-kD protein, reacted with all sera from Chagas1 disease patients tested but not with sera from patients with leishmania, in ELISA. The 52-kD antigen is widely distributed in the Trypanosoma genus since the 163B6 monoclonal antibody reacts with T. rangeli and 8 strains and a clone of T. cruzi epimastigotes.
The specificity of anti-neutrophil cytoplasm autoantibodies in systemic vasculitides
Authors: LAI K N , JAYNE D R W , BROWNLEE A , LOCKWOOD C M 
Published: July 2010

SUMMARY
Autoantibodies to neutrophil cytoplasmic antigens are found in patients with small or medium-size vessel systemic vasculitis such as Wegcner's granulomatosis and microscopic polyartcritis. We studied the specificity of anti-neutrophil cytoplasm autoanlibodies (ANCA) by examining the binding of sera from patients with different forms of systemic vasculitis, to neulrophil extract size-fractionated by gel filtration for high performance liquid chromatography. Sequential fractions were collected for solid-phase radioimmunoassay. Patients with Wegcner's granulomatosis showed two peaks of activity over fractions of 12 and 2 kD. Patients with microscopic polyarteritis had two different peaks of aclivity over fractions of 100 and 25 kD. Patients with Takayasu's arteritis and dermal lymphocytic vasculitis were studied as controls (all were ANCA negative by indirect immunofluorescence studies and solid-phase radioimmunoassay). However, the patient with lymphoeytic vasculilis showed a distinct peak of activity over 22 kD and patients with Takayasu's arteritis had a single peak of activity over 200 kD. These results suggest the binding pattern of sera to fractionated neutrophil extract could be useful in differentiating between different diseases within the spectrum of systemic vasculilis. Cryptic antigen recognition in certain vasculitides may only be revealed by this procedure. We conclude that the association of ANCA and systemic vasculitis may not be limited to diseases of small or medium-size vessels, but could also include large vessel disease such as Takayasu's arteritis.
CD3-T cell receptor modulation is selectively induced in CD8 but not CD4 lymphocytes cultured in agar
Authors: OUDRHIRI N , FARCET J P , GOURDIN M F , M'BEMBA E , GAULARD P H , KATZ A , DIVINE M , GALAZKA A , REYES F 
Published: July 2010

SUMMARYThe CD3-T cell receptor (TcR) complex is central to the immune response. Upon binding by specific ligands, internalized CD3-TcR molecules increase, and either T cell response or unresponsiveness may ensue depending on the triggering conditions. Using semi-solid agar culture, we have shown previously that quiescent CD4 but not CDS lymphocytes generate clonal colonies under phyto-haemagglutinin stimulation. Here we have demonstrated that the agar induces selective CD3-TcR modulation in the CD8 and not in the CD4 subset. CDS lymphocytes preactivated in liquid culture and recultured in agar with exogenous recombinant interleukin-2 generate colonies with a modulated CD3-TcR surface expression. The peptides composing the CD3-TcR complex are synthesized in CD8 colonies as well as in CD4; however, the CD3 gamma chain is phosphorylated at a higher level in CD8 colonies. A component of the agar polymer, absent in agarose, appears to be the ligand that induces differential CD3-TcR modulation in the CD8 subset. In contrast to agar culture, CD8 colonies can be derived from quiescent CD8 lymphocytes in agarose. These CD8 colonies express unmodulated CD-TcR. CD3-TcR modulation with anti-CD3 monoclonal antibody prior to culturing in agarose inhibits the colony formation. We conclude that given triggering conditions can result in both CD3-TcR modulation and inhibition of the proliferative response selectively in the CD8 lymphocyte subset and not in the CD4.
Immunochemical characterization of human antibodies to lymphobiastoid interferon
Authors: THURMOND L M , REESE M J 
Published: July 2010

SUMMARYAntibodies produced in recurrent respiratory papillomatosis (RRP) patients treated with Iymphoblastoid interferon (1yIKN) may neutralize antiviral activity or may only bind to 1yIFN. These antibodies were characterized for immunoglobulin class. IgG subclass. and light chain type by an indirect immunoassay. Serum dilutions were incubated on 1yIFN-coated plates and the presence of antibody detected using peroxidase-conjugated goat antibodies to each human immunoglobulin class and light chain isotype, or using MoAbs to each human IgG subclass. Neutralizing activity was measured as the inhibition of 1yIFN antiviral activity for Vervet monkey cells challenged with Semliki Forest Virus. Among antibody-positive patients, 12% produced IgM coincident with IgG, and 25% produced IgA coincident with IgG. Thus, antibody responses in patients treated with 1yIFN are not exclusively of IgG class. The predominant 1yIFN-specific subclasses were IgG1 and IgG3, which occurred in 70% and 83% of patients, respectively. An IgG4 response was detected in two patients who also had antibody of other isotypes; no IgG2 antibody was detected in any patient. Antibodies were not IgG subclass-restricted, a trend which was more pronounced in patients having neutralizing antibody than non-neutralizing antibody. Light chain molecules of 1yIFN-specific antibody were of both k and λ isotypes. with k chains occurring most frequently. Among patients having non-neutralizing antibodies, monotypic light chains occurred in 65% of the patients, whereas no patient with neutralizing antibody had monotypic light chain antibody. Sera from 599 normal human volunteers were assayed for antibody, and seven were found to be immunoreactive to 1yIFN. Only-one serum of the seven was positive for neutralizing activity.
Cytotoxic CD4+ T cells specific forFrancisella tularensis
Authors: SURCEL H M , TAPANINAHO S , HERVA E 
Published: July 2010

SUMMARYThe specific cell-mediated cytotoxicity of tularaemia-immune human T lymphocytes were studied in vitro. Peripheral blood mononuclear cells (PBMC) of six tularaemia-vaccinated healthy subjects were stimulated with F. tularensis LVS whole cell antigen for 6 days and used as effector cells in a conventional 4-h 5lCr release cytotoxicity assay. The target cells were phagocyting autologous monocytes, which were pulsed with F. tularensis or PPD antigen. The specific lysis of the F. tularensis pulsed cells (42.6%± 11.7) was significantly higher (P &lt; 0.05) than that of the PPD-pulsed ones (22.2%± 8.3) or unpulsed control cells (15.9%± 5.2). The cytotoxicity was associated with CD4+F. tularensis-specific T cell clones (TLC), which killed 36.3%± 12.3 of the F. tularensis-pulsed targets but only 6.9%± 6.5 of the unpulsed control targets. Their lysing was inhibited by monoclonal anti-HLA-DR and anti-HLA-DQ antibodies, but not by CD 15 (monocyte/macrophage) antibody. The functional role of CD4+ lymphocytes in tularaemia immunity is discussed.
Cellular origins of human polymeric and monomeric IgA: enumeration of single cells secreting polymeric IgA1 and IgA2 in peripheral blood, bone marrow, spleen, gingiva and synovial tissue
Authors: TARKOWSKI A , MOLDOVEANU Z , KOOPMAN W J , RADL J , HAAIJMAN J J , MESTECKY J 
Published: July 2010

SUMMARYUsing modified ELISA and spot-ELISA, which permit the parallel determination of heavy chain subclass and the presence of covalently linked J chain, we analysed IgA found in cell culture supernatants or secreted by individual cells from peripheral blood, spleen, bone marrow, gingiva and synovial tissue, with respect to its polymeric or monomeric IgA form (plgA, mlgA) and IgA I or IgA2 subclass. The ELISA for determination of J chain in tissue culture supcrnalants was specific and highly sensitive (detection limit in pg). The results demonstrated that IgA1-producing cells predominated in the tissues examined, and that J chain could be detected in association with the majority of IgA1 and lgA2 secreted by individual cells. With respect to the frequency of cells secreting polymeric, J chain-containing IgA, only 20-30% of cells from the bone marrow were engaged in the synthesis of pIgA. In other tissues the frequency of cells secreting pIgA1 and plgA2 was considerably higher. Peripheral blood mononuclear cells secreting pIgA2 were easily inducible during stimulation with T cell-dependent pokeweed mitogen, whereas Epstein-Barr virus-transformed cells secreted preferentially mIgA1. When the frequencies of plgA-, pIgA 1 - or pIgA 2-secret ing cells (determined by spot-ELISA technique) from different tissues were correlated with the proportion of pIgA EG mIgA (and IgA subclasses) secreted in tissue culture supernatants. data obtained suggest that many individual IgA-producing cells could be engaged in simultaneous secretion of mIgA and pIgA.