bluuebunny/crossref_metadata_2025 huggingface dataset:
IgG2 Deficiency Associated with Defects in Production of Interferon‐Gamma; Comparison with Common Variable Immunodeficiency
Authors: INOUE R. , KONDO N. , KOBAYASHI Y. , FUKUTOMI O. , ORII T.
Published: July 2006

We report a novel mechanism of IgG2 deficiency. Several investigators have reported patients with IgG subclass deficiencies due to homozygous deletion of immunoglobulin heavy chain constant region genes. However, it is unclear what mechanism is responsible for IgG subclass deficiency in cases where no gene deletions have been detected and which are accompanied by recurrent infections due to aberrant immunoregulation. In the present study, we have focused our attention on production by peripheral blood mononuclear cells (PBMCs) of interferon‐gamma (IFN‐γ), which is known to induce IgG2 expression. PBMCs from four patients with IgG2 deficiency and their families were studied. Mitogen‐induced IFN‐γ production by PBMCs was decreased in all of the patients, although the proliferative responses of PBMCs and the percentages of CD3, CD4, and CD8 T cell subsets were not decreased. IgG2 production by PBMCs was restored upon addition of IFN‐γ and mitogen to the PBMCs of the patients with IgG2 deficiency though it was not restored in the patients with common variable immunodeficiency. We conclude that defects in production of IFN‐γ play an important role in IgG2 deficiency.
Clustering of B and T Epitopes Within Short Sequence Regions of the Nieotinic Acetylcholine Receptor
Authors: BELLONE M. , KARACHUNSKI P. I. , OSTLIE N. , LEI S. , CONTI‐FINE B. M.
Published: July 2006

The epitope repertoire of B cells, due to their selective ability to process their specific antigen and the potential bias imposed on the resulting peptides by the surface immunoglobulins bound to the antigen, may influence the T‐helper repertoire.

Immunization of C57B1/6 mice with Torpedo acetylcholine receptor (TAChR) causes experimental autoimmune myasthenia gravis (EAMG). Anti‐TAChR CD4+ cells recognize epitopes within three sequence regions of the TAChR a subunit (“dominant epitopes’). Immunization of mice with denatured or synthetic TAChR antigens sensitizes CD4+ cells to other TAChR sequence regions (‘cryptic epitopes’).

We investigated here whether clustering of B and T epitopes within the same short sequence segments occurs during the anti‐TAChR response, as previously described for the response to hexogenous antigens unrelated to homologous self proteins.

Twelve 19–20 residue synthetic sequences of the TAChRα, β and δ subunits, containing dominant or cryptic CD4+ epitopes for C57B1/6 mice, were tested for ability to induce anti‐peptide antibody production. C57B1/6 mice were immunized with the individual peptides. Ten peptides stimulated antibody production. Therefore >80% of these short TAChR sequences also contain B epitopes.

Therefore also in the anti‐TAChR response leading to EAMG T and B cell epitopes frequently reside within the same short sequence segment.
PKC Activity and Protein Phosphorylation in Regulation of sig Mediated B Cell Activation
Authors: LINDSBERG M.‐L. , FEUERSTEIN N. , BALAPURE A. K. , MOND J. J.
Published: July 2006

The inhibitory and stimulatory elements of cellular signalling associated with activation of protein kinase C (PKC) in murine B lymphocytes were investigated by employing two PKC activators with opposing effects on cell proliferation. Being an inhibitor of anti‐Ig mediated proliferation, the phorbol ester PDBU induced a more substantial translocation of cytosolic PKC activity than the alkaloid PKC activator indolactam, which enhances anti‐Ig mediated B cell proliferation. PDBU and indolactam were equally effective kinase activators, as determined by 32P incorporation of the substrate proteins. Concentrations of indoiactam which induced an inhibition of anti‐Ig mediated B cell proliferation also induced a precipitous decline in detergent soluble cellular PKC activity, which was comparable with 1μM PDBU. The induced phosphoprotein patterns were similar, with an exception of the nuclear envelope protein lamin B, which was prominently phosphorylated by PDBU but not by stimulatory concentrations of indolactam. The enhanced phosphorylation of lamin B was associated with cellular growth arrest: inhibitory concentrations of indolactam induced the phosphorylation of lamin B equal to PDBU. whereas an increased phosphorylation of lamin B was never observed upon stimulation with anti‐Ig. Together, inhibition of anti‐Ig mediated B cell proliferation was related to down‐regulation of cytoplasmic PKC and induction of nuclear PKC‐dependent phosphorylation.
Visceral Leishmaniasis in Ververt Monkeys: Immunological Responses During Asymptomatic Infections
Authors: GICHERU M. M. , OLOBO J. O. , KARIUKI T. M. , ADHIAMBO C.
Published: July 2006

Nine vervet monkeys (Cercopithecus aethiops) were infected intradermally with 8 × 107 virulent L. donovani promastigotes. Four animals developed clinical visceral leishmaniasis and died over a period of 18 months. The remaining five animals have remained asymptomatic for a period of 3 years now. Attempts to isolate parasites from spleen and liver through biopsies were fruitless. Immunological respotises of these subclinically infected animals were examined. Enzyme‐linked itnmunosorbent assay (ELISA) and western blot analyses demonstrated Leishmania specific antibodies in these animals, but the antibody titres were low. When proliferation of peripheral blood monocytes (PBMC) to Con‐cunavalin A (Con A) of these animals was compared with control ‘disease free animals’ there were no significant differences iti response. However L. donovani antigen (fixed promastigotes) specific proliferation was demonstrated in the five subclinically infected animals. High and varying levels of interferon gamma (IFN‐γ) were secreted in PBMC cultures from the five vervet monkeys when stimulated with either Con A or L. donovani antigens. In control animals, IFN‐γ was only detected wheti PBMC were stimulated with Con A. Marked delayed‐type hypersensitivity (DTH) responses were demonstrated in the five subclinically infected animals 48 h after injection with formalin fixed promastgotes.

It was concluded that the visceral Leishmania disease spectrum due to L. donovani observed in humans could be induced in vervet monkeys and that L. donovani asymptomatic/cryptic infected animals have competent humoral and cellular responses to homologous parasites.
A Dysbalanced Immune System in Cryptogenic Lennox‐Gastaut Syndrome
Authors: VAN ENGELEN B. G. M. , WEEMAES C. M. R. , RENIER W. O. , BAKKEREN J. A. J. M. , BORM G. F. , STRENGERS P. F. W.
Published: July 2006

In children with cryptogenic Lennox‐Gastaut syndrome we found a functionally impaired humoral immune response to a primary antigen (haemocyanin), despite signs of a triggered immune system consisting of elevated IgG concentrations. This combination of immunological findings, considered to be the expression of a dysbalanced‐triggered as well as functionally impaired‐immune system, has also been described in an auto‐immune disease like systemic lupus erythaematodes in humans, and in genetically epilepsy‐prone rats. The interactions between the immune system and the nervous system in Lennox‐Gastaut syndrome will be discussed.
Reduced IL‐2 Expression Upon Antigen Stimulation is Accompanied by Deficient IL‐9 Gene Expression in T Cells of Patients with CVID
Authors: HAUBER I. , FISCHER M. B. , MARIS M. , EIBL M. M.
Published: July 2006

Patients with common variable immunodeficiency (CVID) are heterogeneous in the clinical manifesta tion of the disease as well as in the underlying mechanisms leading to the immunodeficiency. In a previous study we identified a subgroup of patients with a primary immunodeficiency disease affecting IL‐2 and IFN‐γ gene expression. The T cells of these patients revealed impaired proliferative response and reduced levels of IL‐2 and IFN‐γ ‐specific mRNA after antigen stimulation in vitro, while cellular and molecular response to phorbol ester and the calcium ionophore ionomycin (PMA + IM) or anti CD3 monoclonal antibodies (MoAbs) (OKT3) were comparable to those of healthy control individuals. Here we show that stimulation of these patients T cells with tetanus toxoid (TT) resulted in dramatically reduced levels of IL‐2, IL‐9 and IFN‐γ mRNA, while IL‐3 gene expression in three patients was comparable or even increased lo the healthy controls. As expected, addition of exogeneous IL‐2 to tetanus toxoid pulsed cultures had virtually no effect on IL‐2 transcription, but corrected the defect in IL‐9 gene expression, while IFN‐γ mRNA levels were still reduced. In conclusion, these data suggest that recombinant IL‐2 alone is not able to induce the IL‐9 gene adequately in our patients, but clearly increases IL‐9 mRNA levels in combination with tetanus toxoid.
Iron Requirements of Human Lymphocytes: Relative Contributions of Intra‐ and Extra‐Cellular Iron
Authors: GOLDING S. , YOUNG S. P.
Published: July 2006

A supply of iron is of vital importance if lymphocyte proliferation is to proceed successfully and two major sources of iron are available, intracellular stores and serum transferrin. We have investigated the relative importance to the human T lymphocyte of these two sources of iron by depleting them of intracellular iron with the chelator desferrioxamine and by culturing them in medium completely depleted of transferrin iron. The chelator decreased mitogen‐stimuiated proliferation of human peripheral blood T cells, in a dose‐dependent manner, in the absence of extracellular transferrin‐iron. By culturing the cells in iron‐depleted medium, we found that normal lymphocytes proliferated, to a degree, in the absence of extracellular transferrin‐iron. We also observed that transferrin receptor mRNA expression was sustained in mitogen‐stimuiated, iron‐deprived lymphocytes, compared with untreated cells suggesting that up‐regulation of transferrin receptor may occur in these cells through stabilization of the mRNA. We propose that intra‐ and extra‐cellular iron may contribute to early and late activation processes and that a low level of intracellular iron in lymphocytes, chronically activated in the iron‐deficient environment associated with chronic inflammatory diseases such as rheumatoid arthritis, may be a factor in the abnormal cell‐mediated immunity associated with such diseases.
Apoptosis among CD45RA−/low CD3+ Progeny Accompanies Differentiation of Human Multinegative Thymocytes
Authors: PILARSKI L. M. , LADEROUTE M. P. , RUTKOWSKI D.
Published: July 2006

Apoptotic cell death is widely believed to be the fate of negatively selected cells in the thymus. In this work we have used multiparameter flow cytometry to analyse reductions in DNA content that occur among differentiating human CD3−4−8− multinegative (MN) thymocytes as they acquire CD3/TCR during in vitro culture. DNA content was measured as the intensity of the DNA‐binding dye DAPI. The position of the diploid peak was identified by comparison to chicken red blood cells and to unfractionated uncultured thymocytes which have a sharply defined diploid peak. Apoptosis, was defined as a reduction in DNA content. Apoptotic cell death occurred continuously throughout the 7 day culture period and at the latest stages of culture DNA fragmentation was apparent on gels. Although both CD3− and CD3+ progeny became apoptotic, it was more frequent among the CD3+progeny of the MN thymocytes. Apoptotic progeny included 60–70% CD3+ cells, while progeny remaining diploid were 8–36% CD3+. Fifty five per cent of CD3+ TCRδl+ progeny had less than 75% of the diploid DNA content, while for CD3+ TCRδ1− progeny only 28% were in this category. CD3+ TCRδ1+ progeny also comprised 66% of the cycling cells at days 6–7 of culture, suggesting a pattern of rapid cell division followed by apoptotic cell death for this subset. A lack of positive selection in culture may trigger apoptosis among the TCRδ1 progeny. In contrast, TCRαβ progeny arising in culture appear to be less susceptible to apoptosis, perhaps due to their lack of CD4 and CD8. The expression of CD45RA and CD45R0 isoforms were assessed on the progeny of MN thymocytes based on their DNA content. Although 30% of apoptotic progeny expressed CD45RA, it was present at relatively low density compared to that on diploid or cycling cells, 55% of which were CD45RAhi. The majority of apoptotic cells expressed neither CD45RA or CD45R0, but were CD45+, indicating the presence of an isoform not detected by our monoclonal antibodies (MoAbs). This is consistent with speculations that apoptotic cell death among thymocytes is preceded by a transition in CD45 isoform expression. These conditions may model early selective events resulting from high avidity TCR engagement that is independent of CD4 and/or CD8.
Analysis of the Proliferative T Cell Response to Human Cytomegalovirus Major Immediate‐Early Protein (IEl): Phenotype, Frequeney and Variability*
Authors: DAVIGNON J.‐L. , CLÉMENT D. , ALRIQUET J. , MICHELSON S. , DAVRINCHE C.
Published: July 2006

Cellular immune responses are important in the recovery from human cytomegalovirus (HCMV) infection. However, little is known about the CD4+T cell response and the target antigens (Ag) recognized. In this paper, we have analysed the proliferative T cell response of healthy HCMV seropositive (HCMV+) blood donors to recombinant immediate‐early proteins expressed in trans‐fected astrocytoma cells and to total HCMV Ags expressed in infected astrocytoma cells. We found that CD4+T cells were the major cell population that proliferated in the presence of IE or total HCMV Ags. Among healthy HCMV seropositive blood donors with anti‐HCMV specific proliferative response, 33–44% also responded to IE Ags. Moreover, in high responders, the precursor frequencies of cells which proliferated in the presence of total HCMV, IE, or IEl Ags were high (1/103 to 1/255, 1/2785 to 1/7744 and 1/5190 to 1/13531, respectively). In some donors, the anti‐IE response was variable over time, whereas the anti‐total HCMV Ags response remained constant, which suggests regulation of the anti‐IE response in immunocompetent subjects. Our results suggest that the CD4+anti‐IEl response represents a significant part of the anti‐HCMV proliferative response, both at the population level, and within individual immune systems.
Monospecific Polyclonal Anti‐Anti‐Idiotypic Antibodies to the Carboxyterminal Undecapeptide of the SV40 Large Tumour Antigen
Authors: AGOSTINI H. T. , GERSTENECKER B. , HAESSLER C. , BRAUN D. G. , BRANDNER G. , HESS R. D.
Published: July 2006

The murine monoclonal antibody PAb1605 defines an epitope, peptide Lys(698)‐Thr(708) (KT), on the carboxyterminus of the tumour(T)antigen of SV40‐transformed cells. In vivo and in vitro experiments had shown that this sequence represents an epitope for both humoral and cellular immune responses. When injected into rabbits PAb1605 induces anti‐idiotypic antibodies (Ab‐2). Ab‐2β (internal image type) was purified by adsorption chromatography and characterized by the ability of KT to compete with the binding of ab‐2 with ab‐1. Murine anti‐anti‐idiotypic antibodies (ab‐3) were obtained by immunization of mice with ab‐2β. Both ab‐1 and ab‐3 JgG showed affinities to immunoprecipitated SV40 T antigen by immunoblot analysis and to nuclear SV40 T antigen by the immunofluorescence assay. The binding of ab‐3 to SV40 T antigen was completely inhibited by competition with KT. We conclude that the polyclonal ab‐3 is of the ab‐3 subtype and specific for only one epitope which is represented by KT and defined by ab‐1. The results demonstrate that the specificity for a defined peptide epitope of an antibody was conserved even after two consecutive steps of anti‐idiotypic‐antibody formation in two host species. Since this postulate of network theory could be verified for a sequence of a tumour‐associated antigen which represents a B‐ and T cell epitope, this model is of great interest for further tumour immunological studies.
Native Human Serum Amyloid P Component is a Single Pentamer
Authors: SØRENSEN I. J. , ANDERSEN O. , NIELSEN E. HOLM , SVEHAG S. E.
Published: July 2006

Serum amyloid P component (SAP) and C‐reactive protein (CRP) are members of the pentraxin protein family. SAP is the precursor protein to amyloid P component present in all forms of amyloidosis. The prevailing notion is that SAP in circulation has the form of a double pentameric molecule (decamer) whereas CRP is a single pentameric molecule.

We have investigated by gel permeation chromatography the Mr of SAP in freshly collected human serum and of SAP purified by carbohydrate affinity chromatography and anion exchange chromatography. SAP was monitored by quantitative immunoelectrophoresis and ELISA. and SAP peak fractions were analysed by use of SDS‐PAGE, Western blotting, and electron microscopy. The results indicate that native SAP circulates as a single pentamer, a part of which forms complexes with C4b‐binding protein. The properties of SAP changed during purification as indicated by rocket immuno‐electrophoresis and electron microscopy. Thus, electron micrographs of purified SAP showed a predominance of decamers. However, the decamer form of SAP reversed to single pentamers when purified SAP was incorporated into SAP‐depleted serum.
Characteristics of Immune Responses to Native and Protein Conjugated Pneumococcal Polysaccharide Type 14
Authors: VAN DEN DOBBELSTEEN G. P. J. M. , KROES H. , VAN REES E. P.
Published: July 2006

The immune response to pneumococcal polysaccharide type 14 (PPS‐14), induced by native PPS‐14 was compared with the response induced by PPS‐14 conjugated with CRM197 (PPS‐14‐CRM197). In our animal model, immunization with PPS‐14‐CRM197 gave a significant enhancement of anti‐PPS‐14 serum titres for IgM and IgG. but not for IgA. Also an increase in total number of anti‐PPS‐14 antibody‐secreting ceils was found. Using immunohistochemical techniques, a different distribution pattern of specific antibody‐containing cells in spleen section after immunization with PPS‐I4‐CRM197 was observed. Furthermore, a higher number of IFN‐γ producing cells was found after immunization with PPS‐14‐CRM197, as compared with immunization with PPS‐14. This enhanced IFN‐γ production may be the cause for enhanced IgG response observed after immunization with PPS‐14‐CRM197.

The specific immune response was less affected by splenectomy in animals immunized with PPS‐14‐CRM197 than with PPS‐14. However, an age‐related response to the native as well as the conjugated form of the PPS‐14 was observed, since no effect of conjugation with CRM197 was seen in the onset of the immune response to PPS‐14 in young animals.

In conclusion, our results affirm the hypothesis that conjugation of polysaccharides changes the characteristics of the antigen towards a thymus‐dependent antigen.
In Vivo Evidence for a Non‐T Cell Origin of Interleukin‐5
Authors: CASTRO A. G. , SILVA R. A. , MINÓPRIO P. , APPELBERG R.
Published: July 2006

Eosinophil myelopoiesis is to a great extent regulated by interleukin (IL)‐5. Analysis of IL‐5 mRNA in spleen cell preparations by reverse transcription‐polymerase chain reaction (RT‐PCR) revealed the presence of message for this cytokine in uninfected severe combined immunodeficiency (SCID) mice. This message was increased following Mycobacterium avium infection. Normal BALB/c mice had higher levels of expression of IL‐5 but the expression of this cytokine was reduced during M. avium infection. Anti‐IL‐5 monoclonal antibody administration in vivo to SCID mice reduced the number of peritoneal and splenic eosinophils. Gamma interferon (IFN‐γ) had an inhibitory effect of eosinophi‐lopoiesis during infection of SCID mice by M. avium since neutralization of this cytokine increased the number of eosinophils detected in the peritoneal cavity of infected animals. Our results suggest that IL‐5 may be produced by cells other than T cells that are both able to respond to infection and are under the control of IFN‐γ.
Specific and Non‐Specific T‐Cell Activation in High and Low Antibody‐Producing Mice (Selection IV‐A)
Authors: CABRERA W. H. , SIQUEIRA M. , TAKAHASHI N. S. H. , RIBEIRO O. G. , ARAUJO L. M. , MOUTON D. , IBANEZ O. M.
Published: July 2006

The T‐cell regulatory function has been evaluated in lines of mice genetically selected according to the High (H) and Low (L) antibody responsiveness to heterologous erythrocytes. The inhibition of antibody production following CD4+ subset depletion was stronger in L than in H mice. The dose of CD4‐specific monoclonal antibody (MoAb) needed for a 50% inhibition of the anti‐sheep erythrocyle antibody response was much lower in L mice, even when their responsiveness was improved by repeated antigen injections or by co‐injection of lipopolysaccharide as an adjuvant. Lymph‐node cells from L mice responded normally or even better than those from H mice to in vitro non‐specific triggering via lectins, exogenous superantigens or CD3‐specific MoAb. In contrast, the antigen‐specific responses of the lymph‐node cells from protein‐primed mice were consistently lower in the Low responder mice. Altogether the results suggest that a defective stimulation of T helper lymphocytes by the antigen contributes to the weak antibody response of L mice.
Induction of Anti‐Tumour Immunity in Syngeneic Mice by Leukaemic Cell Line
Authors: MONGINI C. , WALDNER C. I. , ALVAREZ E. , ROIG M. I. , LOCKHART M. SÁNCHEZ , GRAVISACO M. J. , HAJOS S. E.
Published: July 2006

Induction of anti‐tumour immunity in syngeneic mice by LBC cell line derived from a non‐immunogenic T cell leukaemia was studied. The immunization of BALB/c mice with LBC irradiated cells induced in them anti‐tumour spleen cells, cytotoxic T lymphocytes and anti‐LBC antibodies. The anti‐LBC antibodies reacted with components of 14, 16 and 27kDa present on LB tumour cells, LBC cell line and normal thymocytes, but not with normal lymph node cells. Furthermore, immunization of the autologous hosts with LBC cells partially protected them against subsequent challenge with the original LB leukaemic cells. These findings demonstrate that culture conditions induced modifications in the antigenic properties of the leukaemic cells, allowing LBC cells to stimulate an immune response directed against components expressed at early stages during T cell maturation. These results also suggest that the Immune response is responsible for the prolongation of the survival time of the mice inoculated with the parental leukaemic cells.
Spleen Cells from Antithymocyte Serum Pretreated Mice Do Not Induce GVHD but Exert Increased Repopulating Activity in Irradiated Semiallogeneic Recipients
Authors: VIZLER C. , JÁNOSSY T. , VÉGH P.
Published: July 2006

Graft‐versus‐host disease (GVHD) due to allogeneic bone marrow transplantation can be prevented by depleting the T cells from the marrow graft in vitro. However, the elimination of the donor T cells results in a higher frequency of graft failure, secondary infections and, in case of leukaemia, relapse. We found, that, in contrast to normal spleen cells, spleen cells from A or B10 donor mice pretreated with xenogeneic antithymocyte serum (ATS) in vivo did not induce GVHD in non‐irradiated (B10 x A)F1 hybrids. Spleen cells of ATS‐pretreated A donors did not cause GVHD in allogeneic CBA mice made neonataily tolerant to the A donor strain either. Furthermore, spleen cells from ATS‐treated donors did not cause GVHD in irradiated F1 hybrid recipients, moreover, they decreased the lethal effect of irradiation. The in vivo ATS pretreatment improved the repopulating capacity of spleen cells in irradiated syngeneic recipients, too. The effect of the ATS treatment does not rely solely upon the elimination of T cells, since flow cytofluorometric analysis revealed only a partial depletion of both the CD4+ and CD8+ T cells of the ATS‐pretreated animals. These observations may also have clinical relevance.
Anti‐IL‐10 Treatment Does Not Block Either the Induction or the Maintenance of Orally Induced Tolerance to Ova
Authors: AROEIRA L. S. , CARDILL F. , DE ALBUQUERQUE D. A. , VAZ N. M. , MENGEL J.
Published: July 2006

Herein, the role of IL‐10 in the induction and maintenance of oral tolerance was evaluated. The results show that: (1) mice treated with MoAb anti‐IL‐10 are permissive to the induction of oral tolerance to OVA; (2) anti‐IL‐10 treatment did not reverse the in vitro blocking of T cell proliferative response found in orally‐tolerized mice; and (3) orally‐induced tolerance could not be broken by anti‐IL‐10 treatment. Taken together, these results suggest that IL‐10 is not a fundamental cytokine for the establishment and maintenance of oral tolerance.
The T Cell Receptor and AIDS Pathogenesis
Authors: HOFFMANN G. W.
Published: July 2006

An idiotypic network model of AIDS pathogenesis is described in which the T cell receptor plays a role both in infection and as a target of autoimmunity. This is an extension of a previously published autoimmunity model, and provides explanations for several otherwise puzzling aspects of AIDS pathogenesis. In the model HIV–specific T cells are preferentially infected, and HIV, acting as an antigen, stimulates the expansion of the infectable pool of T cells. The HIV variants that are most strongly selected are those that are recognized by the most helper T cells. HIV and suppressor T cells are subject to the same selective environment, and consequently undergo a process of convergent selection to resemble each other more and more with time. Eventually immunity against HIV cross–reacts with suppressor T cell idiotypes, disrupting the normal regulation of helper T cells. Autoimmunity ensues. The model leads to novel vaccine and therapy approaches involving the targeting and elimination of HIV–specific T cells.
Expression of the Human Interleukin–2 Receptor Gamma Chain in Insect Cells Using a Baculovirus Expression Vector
Authors: RAIVIO E. , OETKEN C. , OKER‐BLOM C. , ENGBERG C. , ÅKERMAN K. , LINDQVIST C.
Published: July 2006

The gene encoding the γ‐chain of the human lnterleukin–2 receptor was expressed in lepidopteran insect cells using the baculovirus expression vector system. The corresponding gene was inserted under the polyhedrin promoter of the Autographa californica nuclear polyhedrosis virus and expressed in the Spodoptera frugiperda insect cell line Sf9 during viral infection. The recombinant receptor protein was identified by immunoblotting in cell lysates, prepared from insect cells infected with the recombinant virus. At 40 h post infection the corresponding protein was detected as two major bands with apparent molecular weights of 50–60 kDa using a rabbit anti–human IL–2R γ‐receptor specific antiserum. Metabolic labelling with [35S]–methionine and SDS–PAGE analysis of the recombinant baculovirus infected insect cells verified the immunoblotting data. The expressed IL–2Rγ‐protein could also be determined on the surface of infected insect cells by flow cytometer analysis.
Interleukin‐4 and Interferon‐Gamma Production by Leishmania Stimulated Peripheral Blood Mononuclear Cells from Nonexposed Individuals
Authors: KURTZHALS J. A. L. , KEMP M. , POULSEN L. K. , HANSEN M. B. , KHARAZMI A. , THEANDER T. G.
Published: July 2006

Interferon‐gamma (IFN‐γ) and interleukin‐4 (IL‐4) production by Leishmania reactive peripheral blood mononuclear cells (PBMC) from non‐exposed individuals was investigated. IFN‐γ was measured in culture supernatants after antigen stimulation. For the measurement of IL‐4, antigen stimulated cells were pulsed with PMA and ionomycin before IL‐4 release was measured. L. donovani and L. major antigens induced IL‐4 production (105–1748pg/ml) in 13 and seven cultures, and IFN‐γ production (1.7‐ > 66IU/ml) in 14 and 11 of 20 cultures, respectively. IL‐4 production rose steeply after 6 days of antigen stimulation suggesting a response due to antigen recognition. Both IL‐4 and IFN‐γ production was abrogated by depletion of CD2+ or CD4+ but not CD8+ cells. CD2+ or CD4+ but not CD8+ enriched cultures produced cytokines as unseparated PBMC. Thus, in non‐exposed individuals circulating Leishmania reactive CD4+ T cells could be demonstrated. The cells from different individuals showed different patterns of IFN‐γ and/or IL‐4 production upon antigenic stimulation. In experimental leishmaniasis the early balance between IFN‐γ and IL‐4 is important for the clinical outcome. Our findings call for studies of the importance of cytokine production by cross‐reactive T cells for the outcome of L. donovani infections in humans and show that the method for IL‐4 detection is useful for this purpose.
The Influence of Tumour Necrosis Factor‐γ, Interleukin‐1β and Interferon‐γ on the Expression and Function of the Complement Regulatory Protein CD59 on the Human Colonic Adenocarcinoma Cell Line HT29
Authors: BJØRGE L. , JENSEN T. SKEIE , ULVESTAD E. , VEDELER C. A. , MATRE R.
Published: July 2006

CD59 is a 18–25kDa glycoprotein which, by inhibiting the formation of the membrane attack complex, protects homologous cells from complement mediated damage. We have described recently the expression and complement regulatory function of CD59 on colonic adenocarcinoma cells both in vivo and in vitro. In this study we have examined the influence of cytokines on the expression and complement regulatory function of CD59 on the colonic adenocarcinoma cell line HT29. CD59 expression on the HT29 cells was up–regulated after stimulation by mononuclear cells activated by mixed lymphocyte reaction and by culture supernatants from activated mononuclear cells. Similarly, a dose–dependent increase in CD59 expression was observed after stimulation with both tumour necrosis factor‐γ and interleukin‐1β. A dose–dependent increase in the level of CD59 expression was also seen using low concentrations of interferon‐γ (IFN‐γ), while CD59 expression on cells cultured with high IFN‐γ concentrations was comparable to non–stimulated cells. Cytokine treated cells were more resistant to lysis by homologous complement than non–stimulated cells, and the increase in CD59 expression was shown to be partially responsible for this. The present data strengthen the role of CD59 as a possible participant in tumour escape.
Complete Coding Sequence of Rainbow Trout Mhc II β Chain
Authors: GLAMANN J.
Published: July 2006

Several cDNA clones comprising the entire coding sequence of the rainbow trout (Oncorh ynchus mykiss) major histocompatibility comlex (Mhc) class II B gene have been isolated from different sources. A single B gene appears to be transcribed in the rainbow trout and it encodes a 247 amino acid long polypeptide, which is of similar size to mammalian, avian, and amphibian and other teleost δ chains. The amino acid sequence identity to mammalian, amphibian, and avian class II δ chains is only about 30%. Despite the low similarity, a striking pattern of conservation is observed, both in the putative peptide‐binding domain and in the Ig‐like domain. Most of the conserved residues are located in the Ig like domain and in the transmembrane segment. The majority of polymorphic residues reside in the δ1 domain, with the greatest variability found in the amino‐terminal half of the domain. The sequence data are compatible with a rather limited polymorphism of a single, expressed Mhc class II B gene.
Acute Graft‐versus‐Host Reaction in SCID Mice Leads to an Abnormal Expansion of CD8+ Vβ14+ and a Broad Inactivation of Donor T Cells Followed by a Host‐Restricted Tolerance and a Normalization of the TCR Vβ Repertoire in the Chronic Phase
Authors: SCHNEIDER M. K. J. , GRÖNVIK K.‐O.
Published: July 2006

The persistence and selection of allogeneic CBA/J T lymphocytes were studied during graft‐versus‐host (GvH) reaction in immunodeficient C. B‐17 SCID (SCID) mice. After neonatal injection the donor cells primarily migrated to the spleen plus lymph nodes (SL) and the thymus of the recipients. Thirteen days post engraftment, CD8+ cells in SL had increased five times in cell number with an 18‐fold increase of CD8+ Vβ14+ cells, paralleled by clinical signs of GvH disease (GvHD). Donor lymphocytes from these mice were proliferative unresponsive to allogeneic Balb/c or C57B1/6 SL cells, whereas 8 weeks post injection the tolerance was confined to H‐2d specific donor cells. Here, spleens had a total cell content similar to untreated SCID mice but the average percentage of donor cells had reached 25%. Moreover, the CD4/CD8 cell ratio in the donor population in SL and thymus had changed to normal and the TCR Vβ repertoire was similar to that of the originally injected cells. Following secondary transfer into syngeneic CBA/Ca nu/nu recipients donor cells regained a significant but reduced response to H‐2d stimulators indicating that the antigen specific tolerance of allogeneic donor cells in the SCID mice was due, at least in part, to a reversible state of anergy.
Biological Implications of HLA‐DR Expression in Tumours
Authors: CABRERA T. , RUIZ‐CABELLO F. , GARRIDO F.
Published: July 2006

HLA‐DR antigens show restricted tissue distribution in comparison with the more extensive expression of HLA class I molecules. This constitutive expression is genetically controlled by well‐defined mechanisms. In addition, DR antigen expression can be induced by a variety of cytokines through different molecular genetic events that convert DR‐negative epithelia into positive cells. In this review we analyse the two major pathological situations in which abnormal DR expression occurs: autoimmune diseases and tumour development. We hypothesize that conversion to DR‐positivity may produce two opposite effects in both clinical situations: (1) a useful one in tumours associated with a good prognosis; and (2) a harmful one in autoimmune diseases with increased tissue damage.
Local T‐Cell Proliferation and Differentiation in Inflammatory Myopathies
Authors: LINDBERG C. , OLDFORS A. , TARKOWSKI A.
Published: July 2006

Our objective was to investigate the patterns of proliferation and differentiation of infiltrating cells in inflammatory myopathies. Immunohistochemical staining was performed on muscle biopsy specimens from 18 patients with inclusion body myositis, polymyositis and dermatomyositis using monoclonal and polyclonal antibodies.

An abundance of cells were TNF‐α+ (4–8%), ICAM‐1+ (7–65%). IFN‐γ+ (3–6%), and Ki‐67+ (4–8%). It was shown that 70% of the Ki‐67+ cells were Ki‐67+CD3+ cells. Very few mononuclear cells were IL‐2R+. MHC‐I expression was found on nearly all muscle fibres in all cases, while MHC‐II expression was found on occasional muscle fibres in 1/3 of cases. Analysis of repeated biopsies from four IBM patients after prednisolone treatment showed no change in the proportions of TNF‐α, ICAM‐1, IFN‐γ or Ki‐67 positive cells. In inflammatory myopathies there is an intense proliferation and differentiation of inflammatory cells in situ, indicating a local stimulation of the inflammatory process.
The Importance of Non‐Charged Amino Acids in Antibody Binding to Humicola lanuginosa Lipase
Authors: NAVER H. , LØVBORG U.
Published: July 2006

The antigenicity of 36 Humicola lanuginosa lipase (HL) variants, generated by site directed mutagenesis, was compared with that of the unchanged enzyme. Polyclonal antibodies raised against variant lipases were investigated and compared with the antibodies raised against the wild type lipase in an ELISA competition assay. The results showed that exchange of charged amino acids with polar residues in surface epitopes of HL, results in a tighter binding of the antibody to the epitope. Four amino acids (Trp at position 89, Asp at positions 96 and 254 and Phe at position 211) were found to be essential for antibody binding in each their epitope of the wild type enzyme.
S. Typhi Vaccine Strain Ty21a Can Cause a Generalized Infection in Whole Body‐Irradiated But Not in Hydrocortisone‐Treated Mice
Authors: VAN DISSEL J. T. , KWAPPENBERG K. , VAN FURTH R.
Published: July 2006

Various mutations including gal E in the S. typhi vaccine strain Ty21a are thought to prevent proliferation of these micro‐organisms in the host, and elimination of Ty21a would occur independent of the immune system of the host. To investigate this issue, we determined whether Ty21a can proliferate in immunosuppressed mice, and assessed the role of phagocytes in the eradication of Ty21a from tissues. Mice were rendered lymphocytopenic and monocytopenic by hydrocortisone s.c., or were made leucocytopenic by whole body irradiation. Bacteria were injected into a tail vene to evaluate eradication from the blood, liver and spleen, and into thigh muscle, i. e. a tissue that lacks resident macrophages. Ty21a were grown overnight in glucose [glu], or galactose and glucose [gal. glu]; only the Ty21a [gal. glu] expressed somatic O‐antigens. After i. v. injection of 104 to 106 micro‐organisms, Ty21a were rapidly eliminated from the liver and spleen of normal and immunosuppressed mice, i. e. within 1 day a 95% reduction of bacterial counts was observed. After i. m. injection of 104 to 106 bacteria, the number of viable Ty21a decreased in normal and hydrocortisone‐treated mice, but in irradiated mice the microorganisms proliferated and caused generalized infection. In all cases, Ty21a [glu] was eliminated more rapidly than Ty21a [gal. glu], confirming reports that killing of bacteria that lack O‐antigens is more rapid than that of smooth bacteria of the same species. These results indicate that elimination of the vaccin strain against typhoid fever, Ty21a, from host tissues is not due to an intrinsic property of the micro‐organisms that prevents proliferation but instead depends on the action of resident macrophages and exudate monocytes and granulocytes.
Impaired Antigen‐Specific B‐Cell Response and Altered Splenic Microstructure in Mice Following Continuous Administration of IL‐4 In Vivo
Authors: SCHILIZZI B. M. , SAVELKOUL H. F. J. , DE JONGE M. W. A. , THE T. H. , DE LEIJ L.
Published: July 2006

The effect of long term in vivo administration of IL‐4 on the induction of antigen‐specific B cells, the splenic microenvironment and the yield of antigen‐specific antibody producing hybridomas was studied. Immunization with DNP‐KLH, followed by 12 weeks continuous IL‐4 treatment resulted in increased numbers of total splenic (non‐DNP) IgM and IgG AFC (antibody forming cells) on day 5 after booster, whereas the DNP‐specific IgG and IgG1 AFC were reduced compared to age‐matched control animals not treated with IL‐4. In addition, an almost 300‐fold increase in non‐DNP IgE was found while the IgE anti‐DNP response was minimal.

When the splenic cells were used in a fusion protocol, a relative decrease in yield of antigen‐specific hybridomas was found in the long term IL‐4 treated mice. Immunohistological staining of spleen sections from mice treated with IL‐4 up until the time of booster revealed reduced B‐cell follicle area and germinal centre numbers. These results show that extensive IL‐4 treatment reduced antigen‐specific B‐cell formation and suggests a reduction in the number of B cells entering the memory B‐cell pathway in the spleen.
Anti‐GD 3 Antibodies are Potent Activators of Human γ/δ and α/β Positive T Cells
Authors: SCHLAAK J. F. , CLAUS C. , MEYER K.‐H. , BÜSCHENFELDE ZUM , DIPPOLD W.
Published: July 2006

The ganglioside GD3 has a variety of biological functions. These include stimulatory effects is on proliferation, natural killer activity and cytokine production by freshly isolated peripheral T cells. In this study we have characterized anti‐GD3 antibody (MoAb Z21) mediated effects on T cell clones. Our data indicate that α/β TCR CD4+ and CD8+ as well as γ/δ TCR positive T cells can be stimulated resulting in proliferation and cytokine production. This effect could be blocked by cyclosporin A and did not involve the LFA‐3 or CD4 molecule. Apart from IFN‐γ and IL‐2 production by T helper I and T helper 0 cells we have observed production of IL‐4 and IL‐10 by T helper 2 cells indicating that the GD3 molecule is not a marker for a certain functional T cell subset. In contrast to anti‐CD3 mediated activation, the responsiveness of T cells to stimulation via GD3 was dependent on the cell surface expression of the molecule and could be enhanced by costimulation via CD2, CD3, CD26 or CD28. In addition, anti‐GD3 antibodies delivered a potent costimulatory signal for antigen‐induced proliferation of CD4+ T lymphocytes. In summary, our experiments illuminate the mechanisms of anti‐GD3 antibody induced T cell activation.
Identification of Two Distinct Function γδ TCR Complexes on the Surface of a Human T Cell Clone
Authors: ORSINI D. L. M. , KOOY Y. M. C. , STRUYK L. , OSSENDORP F. , VAN DEN ELSEN P. , KONING F.
Published: July 2006

In this study we describe the expression of two T cell receptor (TCR) γ chains on the surface of a human T cell clone isolated from the peripheral blood. Each γ chain was part of an independent and functional TCR. The dual receptor T cell clone (and all subclones derived from this clone) had stable expression of this phenotype. Immunoprecipitation studies revealed the expression of non‐disulfride linked TCRs by this Vγ4+Vγ9+Vγ1+ T cell clone, which was in agreement with the finding that both Vγ gene transcripts were rearranged to Cγ2‐associated joining elements. Both γ chains were derived from productive rearrangements of different (allelic) genes coding for a Vγ4+ and a Vγ9+Mγ‐chain, and both were coupled to a Vγ1+γ‐chain. Incubation of this Vγ4+Vγ9+Vγ1+ T cell clone with TCRγ‐chain‐specific MoAbs rapidly induced an increase in intracellular Ca++, indicating that both γ‐chains are functional. Furthermore, this clone responded to stimulation with S. aureus derived superantigens. We suggest therefore that exogenous (super)antigens can trigger dual receptor T cells resulting in activation of these T cells.
The Distribution and Abnormal Morphology of Plasma Cells in Rheumatoid Synovium
Authors: BROWN K. A. , PERRY M. E. , MUSTAFA Y. , WOOD S. K. , CRAWLEY M. , TAUB N. , DUMONDE D. C.
Published: July 2006

This study assessed the distribution and structural features of plasma cells in rheumatoid synovial tissue. Plasma cells were found to be the predominant infiltrating mononuclear cells (mean 40%) in relation to lymphocytes and monocytes, and there was a direct relationship between their number in the infiltrates and the total number of mononuclear leucocytes (P = 0.007). Plasma cells were also seen in intimate contact with macrophages intermixed with synovial lining cells, and closely associated with small blood vessels. They often surrounded these blood vessels and sometimes were seen lying within the vessel walls themselves. Ultrastructural analysis revealed that many synovial plasma cells were considerably larger than plasma cells of a normal size and possessed a marked distension of the cisternae of rough endoplasmic reticulum. Furthermore, plasma cells in close proximity to blood vessels often appeared to be undergoing migration. These observations imply that in rheumatoid synovium, plasma cells are metabolically very active and occupy a pivotal position for the secretion of antibodies into both the vascular and the extravascular compartments.
Natural Killer Suppressor Factors in Sera from HIV‐Infected Subjects
Authors: GOICOA M. A. , SEN L. , IANNITELLI P. S. , DIEZ R. A. , ESTEVEZ M. E.
Published: July 2006

We investigated the presence of NK suppressor factors in HIV+ sera. We further investigated if gp120 could be one of the substances responsible for the impairment of NKC regulation found in HIV+ asymptomatic patients. Our results indicate that HIV+ sera inhibit significantly normal NKC in a dose‐dependent way, even at concentrations as low as 1%. The inhibitory effect of HIV+ sera decreased, but was not completely removed, by adsorptions of IgG or by treatment with a MoAb against human FcIgG. Pretreatment of normal effector cells with anti‐CD 16 MoAb slightly reduced their cytotoxic capability, but did not modify the suppressor effect of HIV+ sera. The preincubation of normal PBMC with recombinant gp120 had also a suppressor effect even at 10ng/ml. Pretreatment of HIV+ sera with anti‐gp120 or anti‐FcIgG MoAb reduced, but not completely, their inhibitory effect. In conclusion, HIV+ serum has a dose‐dependent inhibitory effect on normal NKC. Most of this inhibition is caused by IgG, but other substances, such as gp120, can also contribute to it. Since the removal of IgG and further treatments of HIV+ sera were not able to abrogate completely the NK suppression, other serum factors still undetermined (TNF‐α, other cytokines), should be considered.
A Correlation Between the Expression of the bcr‐abl Chimeric Gene and Severity of the Clinical State of CML Patients with Time
Authors: AMIKAM D. , HENIG C. , CARTER A. , SHARON R. , BEN‐ISHAI Z.
Published: July 2006

Using the reverse polymerase chain reaction, four Ph positive CML patients were followed for 2 years; a correlation between the severity of the clinical state and the b3a2 expression was noted with time. Additionally, amplification of the c‐myc proto‐oncogene was observed, using Southern blot analysis, in one patient prior to his entry to the blast phase. No reorganization of the bcr‐abl rearrangement site was found in the latter patient. The data suggest that a routine follow‐up of CML patients using the Southern blot analysis and the reverse transcription‐polymerase chain reaction might be of importance in evaluating the progression of the disease.
Opposing Effects of Interleukin‐10 on Mouse Macrophage Functions
Authors: APPELBERG R.
Published: July 2006

Interleukin (IL)‐10 generally is considered a macrophage deactivating factor and thus inhibits the cell‐mediated responses against intracellular parasites. We evaluated the in vitro effects of IL‐10 on three different parameters of macrophage function. We found that IL‐10 inhibited gamma interferon (IFN‐γ) priming for enhanced O2− release of mouse bone marrow‐derived macrophages but had opposing effects on NO2− secretion according to the sequence of the treatment with IL‐10 and the agonists of NO2− secretion. Likewise, IL‐10 was able to induce a small but consistent degree of bacteriostasis of Mycobacterium avium and, also, to inhibit the bacteriostasis induced by IFN‐γ. Thus, we show that, according to the timing of exposure of macrophages to stimulating and inhibiting cytokines and agonists of their functions, IL‐10 shows different effects on macrophage function.
Chemospecificity and Cross‐Reactivity of Target Cell Recognition by Human CD56+ NK and LAK Cells
Authors: ZHU H.‐G. , VOETSCH W. , HAUER J. , ANDERER F. A.
Published: July 2006

Inhibition of specific cytotoxicity of highly purified (> 95%) human CD56+ NK and LAK cells against K562 tumour cells was studied with various sugar acetates. Maximum inhibitory specificity was obtained with 60%‐deacetylated penta‐acetates of mannose, galactose, glucose, or 80%‐deacetylated penta‐O‐acetate of N‐acetyt neuraminic acid. The inhibition was strictly dosedependent and 100% inhibition was achieved in the concentration range of 500 1000 nmoles/ml with all four sugar acetate samples. Enhancement of specific cytotoxicity in the presence of rhamnogalacturonan (RG; 500 ng/ml), acting as a bridging molecule, was also inhibited in a dose‐dependent manner with the same inhibitory specificity and within the same concentration range indicating involvement of the same number of sugar acetate‐specific receptors. Moreover, formation of lytic CD56+ effector cell/tumour cell (E/T) conjugates was equally well inhibited whereas formation of total E/T conjugates was only partially inhibited (NK: 44–73%; LAK: 46–50%). E/T conjugate formation in the presence of RG was enhanced. Inhibition of the enhancement of formation of lytic E/T conjugates in the presence of RG was again completely accomplished with the same inhibitory specificity and within the same concentration ranges as recorded for E/T conjugate formation in the absence of RG. However, inhibition of total E/T conjugate formation was again only partially achieved at the given concentrations. The data support the assumption of an NK cell receptor with specificity for acetylated carbohydrate moieties on target cells or on bridging molelcules such as RG.