bluuebunny/crossref_metadata_2025 huggingface dataset:
Phenotypical and Functional Characterization of Double‐Negative (CD4‐CD8‐) αβ T‐Cell Receptor Positive Cells from an Immunodeficient Patient
Authors: ILLUM N. , RALFKIAER E. , PALLESEN G. , GEISLERt C.
Published: June 2006

We have characterized CD4‐CD8‐ double‐negative (DN) αβTCR+ T cells from a patient with immunodeficiency, lymphocytosis, lymphadenopathy, and hepatosplenomegaly. The majority of peripheral blood lymphocytes were DN αβTCR+ T cells as evalualed by FACS and biochemical analysis. The DNT cells showed the following phenotype: αβTCR+, γδTCR‐, CD2+, CD3+, CD57+, and HLA‐DR+. Both southern blot analysis of TCR genes and F ACS analysis applying a panel of Vβ and Vα monoclonal antibodies (MoAbs) indicated a polyclonal T‐cell expansion. Thymic biopsy showed normal histology, whereas lymph node biopsy samples showed altered histological and immunohistoiogical patterns with markedly expanded paracortical areas containing the DN T ceils of the same phenotype as found in peripheral blood T cells. In functional studies, the DN T cells showed a profoundly reduced proliferative response upon stimulation with mitogens as well as MoAbs against the TCR /CD3 complex. CD2. and CD28, respectively, Addition of exogenous interleukiri‐2 (IL‐2) only minimally augmented the proliferative response. In contrast, the addition of a combination of Ca2+ ionophore and phorbol 12‐myristate 13‐acetaie (PMA) restored the proliferative response of the DM T cells to almost normal levels. This observation strongly suggests that the protein kinase C activity of the DN T cells was intact, but that the normal mechanism for transmembrane signal transduction was impaired in these unusual DN T cells.
Different Conditions for Generation of Non‐Cytotoxic Autorestricted CD4‐ CD8+ Human Suppressor T Cells and Allospecific Cytotoxic T Lymphocytes in One‐Way Mixed Lymphocyte Cultures: the Role of Adherent Mononuclear Cells
Authors: BALASHOV K. E. , BRONDZ B. D.
Published: June 2006

The purpose of the present study was to investigate the role of adherent mononuclear cells (AMC) in generation of suppressor T lymphocytes (T‐Lph) in one‐way allogeneic mixed lymphocyte cultures (M LC). Alloantigen‐specific cytotoxic T‐Lph were generated in MLC predominantly in the presence of stimulator AMC, but not responder AMC. The presence of responder AMC and the absence of stimulator AMC were optimal conditions for generation of non‐cytotoxic (through the whole duration of MLC) suppressor T‐Lph. These CD4‐ CDS8+ suppressor T‐Lph were allononspecific but autorestricted (with responders in test MLC).
Kinetics and Regulation of NK Activity by Interleukin‐2 and Interferon in Acute Toxoplasmosis
Authors: DÍEZ B. , NICOLÁS R. , GALDEANO A. , CISTERNA R. , CAÑAVATE M. L.
Published: June 2006

Natural killer (NK) activity against Toxoplasma gondii tachyzoites and tumour cells during acute toxoplasmosis was investigated using a single‐cell NK assay. During the course of infection the percentage of lymphocytes binding tachyzoites and tumour cells did not vary significantly and NK activity was enhanced due to an increase in the specific cytolytic capacity per cell. To determine whether regulatory mechanisms mediated by cytokines might explain the increased NK activity, the kinetics of interleukin‐2 (IL‐2) and interferon (IFN) production and the correlation between their concentrations and the level of NK activity were analysed at the same time. As the infection progressed NK activity increased in spite of the fact that IL‐2 production decreased (except for a small increase during the first day of infection). However, IFN production increased gradually in close temporal and quantitative association with the overall increase in NK activity. These results suggest that T. gondii, via its ability to produce interferon, enhances NK activity against itself and other cells.
Stimulation of Peripheral Blood T Cells by an Activated T‐Cell Line: a Novel Human Autologous T‐T Lymphocyte Reaction
Authors: DITZIAN‐KADANOFF R. , PARKS L. , EVAVOLD B. , QUINTANS J. , SWARTZ T. J.
Published: June 2006

T lymphocyte interactions have generally been described between discrete functional subsets. In our investigation of murine T‐cell interactions we described a type of T‐T interaction termed the 'Syngeneic T‐T Lymphocyte Reaction’in which activated T‐cell clones stimulated the proliferation of resting T cells mainly through a mechanism involving cell to cell contact

To investigate whether similar reactions occur in the human immune system we used the human autoreactive T‐cell line C.1 to stimulate peripheral T cells. Line C! cells, which are not transformed and do not secrete IL2. consistently caused proliferation of purified freshly isolated autologous peripheral human T cells as measured by a [3H]‐thymidine incorporation assay. The proliferation was seen in both the CD4and CDS subsets and could be inhibited with anti‐DR and anti‐CD2 antibodies. The stimulation is not due to carryover of classical antigen‐presenting cells or to the C.1 line cells acting as antigen‐presenting cells.

We propose that some activated T cells, probably by expression of a surface molecule, can stimulate resting T cells thereby allowing for antigen‐non‐specific augmentation of the immune response.
Allelic Variation in the TNF‐β Gene Does Not Explain the Low TNF‐β Response in Patients With Primary Biliary Cirrhosis
Authors: MESSER G. , SPENGLER U. , JUNG M. C. , HONOLD G. , EISENBURG J. , SCHOLZ S. , ALBBRT E. D. , PAPE G. R. , RIETHMÜLLER G. , WEISS E. H.
Published: June 2006

Autoimmune disorders in humans are often associated with particular alleles of major histocompatibility genes. However, the chronic inflammatory liver disease primary biliary cirrhosis (PBC) has not been found to be correlated with certain haplotypes so far.

Interestingly, an impaired production of tumour necrosis factor β (TNF‐β) upon mitogen stimulation was observed for PBC patients, especially in the immunologically active stages of the disease. Furthermore, the identification of alleles of the TNF‐β gene which differ in one unique amino acid, and in the production of TNF‐β after phytohaemagglutinin stimulation, has prompted the idea of a possible linkage between the impaired TNF‐β response in PBC and the genetic prevalence of a certain TNF haplotype.

We report here a rapid method for typing the TNFB*1 and TNFB*2 genes by a standard polymerase chain reaction, PBC patients(n = 60)as well as randumized healthy controls (n = 179) of the Munich area were studied for the occurrence of the TNF alleles. No deviation was found in the PBC collective (0.7) for the TNFB*2 distribution when compared with the control (0.67).
Cell‐to‐Cell Mediated Inhibition of Natural Killer Cell Proliferation by Monocytes and its Regulation by Histamine H2‐Receptors
Authors: HELLSTRAND K. , HERMODSSON S.
Published: June 2006

Human peripheral blood monocytes. recovered by counter‐current centrifugal elutriation, suppressed interleukin‐2 (IL‐2)‐induced proliferation of autologous lymphocytes, recovered from low density Percoll fractions. Cell sorting experiments, analysis of phenotype of proliferating cells, and removal of defined cellular subsets by complement cytotoxicity revealed that IL‐2‐induced proliferation was confined to CD3 /16+/56+ natural killer (NK) cells. Monocyte‐induced suppression of NK‐cell proliferation was completed within 1 h of incubation with monocytes and unrelated to the formation of prostaglandins or other intermediary factors. The biogenic amine histamine. acting via H2‐type histamine receptors (H;R) on monocytes. completely counteracted the monocyte‐mediated suppression of IL‐2‐induced NK‐cell proliferation. Our data are suggestive of a H2R‐regulated, cell‐cell‐mediated mechanism by which monocytes down‐modulate NK‐cell proliferation.
Reversal of T‐Cell Anergy in Autoimmune (NZB X NZW)F1 Mice by Transfer with Non‐Antigen Specific CD4+ T Cells from Young, Syngeneic Donors
Authors: CARLSTEN H. , TARKOWSKI A
Published: June 2006

The F1 hybrid of NZB and NZW mice is a well‐known model of ds systemic lupus erythematosus characterized by B‐cell hyperactivity, production of autoantibodies and immune‐complex formation. These phenomena have been regarded as pathogenetic for the development of glomerulonephritis and early death in renal failure. U has been previously reported that aged and overt diseased NZB/W mice also display impaired T‐cell responses. In the present report we demonstrate an age‐dependent decline in the capacity of NZB/W mice to mount in vivo cutaneous delayed type hypersensitivity (DTH) and antibody responses to a hapten. oxazolone (OXA). Moreover, in vitro proliferative response lo Concanavalin A (Con A) and production of interleukin 2 (lL‐2) were severely depressed in aged NZB/W mice, In transfer experiments we show that a single i. p. injection of non‐immune mononuclear spleen cells from young NZB/W mice to old diseased syngeneic recipients restores DTH reactivity and increases the antibody response to OXA, This effect is a CD4+‐ dependent process since the restoration of T‐cell responses was abrogated upon elimination of CD4+ donor T cells. Overall, our results indicate that limited numbers ofCD4* T cells display substantial immunoregulatory properties reversing the state of anergy in autoimmune NZB/W mice.
B Lymphocyte Fibronectin Receptors: Expression and Utilization
Authors: STUPACK D. G. , STEWART S. , CARTER W. G. , WAYNER E. A. , WILKINS J. A.
Published: June 2006

Lymphocytes adhere to fibronectin (FN) via multiple receptors of the VLA β1 CD29 integrin) family. The cellular requirement for the variety of FN receptors (FNR) which have been described is unclear, but they may be associated with differential signalling processes, cooperative effects which may stabilize cellular attachment, or cell homing and retention processes. The present study was undertaken to examine the FN adherence properties and receptor utilization patterns of human B cells. Often B‐cell culture lines which were studied, six demonstrated a significant adherence to FN. Among these. Tour employed α4β1. (CD49d/29) and two employed α4β1/α5β1 (CD49d/29, CD49e/29), There was no apparent correlation between the differentiation status of the lilies and their FNR utilization patterns. Furthermore, expression of FNR alone was not sufficient to confer FN binding potential.

Freshly isolated tonsil B cells did not display significant adherence to FN, Following stimulation, a marked increase in VLA antigens was observed, and the capacity to attach to FN‐coated surfaces was co‐acquired. Analysis of the induced bulk B‐cell population demonstrated that both α4β1 and aα5β1 were used for adherence. These results clearly indicate that activated B cells, similar to T cells, may express and utilize α5β1 as a FNR.
Presence of Human Chromosome 1 with Expression of Human Decay‐Accelerating Factor (DAF) Prevents Lysis of Mouse/Human Hybrid Cells by Human Complement
Authors: WANG M.‐W. , WRIGHT L. J. , SIMS M. J. , WHITE D. J. G.
Published: June 2006

Xenogeneic organs transplanted to phylogenetically distant species are subject lo rapid destruction mediated by complement. In humans, the complement activation is regulated by several proteins encoded by a series of closely linked genes (RCA locus) located on chromosome I. The mouse/human hybrid cell line BIO was found to have retained human chromosome I, FACS analysis confirmed that RCA products such as decay‐accelerating factor (DAF) were expressed on the membrane surface of B10 cells. When exposed to human or rabbit complement in the presence of naturally occurring’human anti‐mouse antibodies these cells were not lysed by human complement but were killed by rabbit complement. This effect could be abrogated by addition of anti‐DAF monoclonal antibody (IC6). The results offer potential for genetic manipulation of the human complement regulatory products in animals lo overcome xenograft hyperacute rejection.
IL‐2‐Dependent Murine T‐Cell Lines and Clones Expressing Gamma/Delta T‐Cell Antigen Receptors. I, Functional and Biochemical Characterization
Authors: MORKOWSKI J. J. , NANNO M. , CHEN P.‐F. , ITOH K. , IOANNIDES C. G. , KRUZEL E. , BECKER F. F. , PLATSOUCAS C. D.
Published: June 2006

We have developed two stable IL‐2‐dependenl T‐cell lines designated AKV‐I and AKV‐N from the enlarged spleens, respectively, of an AKV l and an NFS mouse, Immunofluorescence staining with the appropriate monoclonal antibodies revealed that cells of the AKV‐I cell line were αβJTCR‐CD3+CD4‐ CD5‐ CD8+CD25+. Where as cells of the AKV‐N cell line were αβTCR‐CD3+CD4‐ CD5+ CD8‐CD25+. A number of T‐cell clones were developed from the AKV‐1 or AKV‐N T‐cell lines by limiting dilution and analysed by immunofluorescence. All clones tested were αβTCR‐CD3+CD4‐CD25+. Certain T‐cell clones expressed the CD5 antigen, whereas others expressed the CD8 antigen. The AKV‐I cell line responded by proliferation lo rIL2, rlL4, phorbol myristate acetate (PMA), PMA plus lL‐4 and PMA plus PHA or Con A, In contrast, the AKV‐N cell line did not respond to rIL‐4orrIL‐4plus PMA and exhibited only a modest prolifrative response lo PMA alone. Both AKV‐1 and AKV‐N T‐cell lines as well as a large number of T‐cell clones examined were able to T‐cells of the PU5‐1R murine cell line in the presence of the anti‐CD3 (clone 145‐2C11) MoAb, demonstrating their ability lo mediate cytotoxicity in this system. Biochemical analysis of both AKV tines and a number of clones by immunoprecipitation with the anti‐CD3 MoAb, followed by one‐dimensional (either non‐reducing or reducing) or two‐dimensional (non‐reducing/reducing) SDS‐PAGE, revealed that the AKV lines and clones expressed a disulphide‐linked dimer. Under non‐reducing conditions, a band in the range of 75 85 kDa was observed and upon reduction it was resolved into two discrete polypeptide chains of43‐44 kDa and 48 kDa in certain AKV‐I cells or 38 kDa and 42 kDa in certain AKV‐N cells. In other T‐cell clones or lines a broad band of42 47 kDa was observed in AKV‐I cells or 38‐45 kDa in AKV‐N cells. These results suggest the presence of different terms of disulphide‐linked dimers on these cells. Northern blotting analysis using probes specific for the constant regions of the α, βγ‐ and lD‐chains of the T‐cell antigen receptor revealed that all the AKV cell lines or clones tested expressed full‐length α, γ‐ and lD‐chain mRNA. whereas β‐chain mRNA was absent. These results suggest that the AKV T‐cell lines and clones express the; γlDTCR, These cells will be useful in investigating the properties of the γlD TCR and studying its diversity.
Immune Responses to 6 and 30‐kDa Mycobacterial Antigens in Rheumatoid Patients, and Vβ Usage by Specific Synovial T‐Cell Lines and Fresh T Cells
Authors: SIOUD M. , KJELDSEN‐KRAGH J. , QUAYLE A. J , WTKER H. G , SØRSKAAR D. , NATVIG J. B. , FØRRE Ø.
Published: June 2006

We have investigated both the humoral and the cellular immune responses of patients with juvenile rheumatoid arthritis (JRA) and rheumatoid arthritis (RA) to mycobacterial antigens. The JRA group was not Bacillus Calmette Guerin (BCG) vaccinated whilst the majority of the RA group was.

As determined by immunoblotting, 79% of sera from patients with JRA reacted mainly with a 18.6‐kDa protein (P18,6), whilst 70% of sera from patients with RA reacted mainly with a 30‐kDa protein (P30) of BCG, M. tuberculosis and M, kansasii. In contrast, only a moderate proportion of the control sera (25% of adult and 20% of children) showed reactivity to P30, and none of the samples had significant reactivity with the P18,6 antigen. Furthermore, T‐cell proliferation to the P18,6 and P30 antigens was detected in the majority of JRA and RA patients, and was nearly always higher in synovial fluid (SF) than in the peripheral blood (PB).

We also investigated the usage of Vvβ family genes in P18,6, and P30 antigen‐specific T‐cell lines established from the SF of one patient with active RA, We showed that Vβ‐2‐4,‐5,‐6,‐7,‐14,‐17,‐18 and Vβ19 were over‐represented compared with other known Vβ families. We also noted that the proportion of Vβ14 was higher in freshly isolated SF mononuclear cells compared with the blood in this patient and in 2 out of 4 other RA patients examined. Other Vβ families such as Vβ6. Vβ8 Vβ16 Vβ18 and Vβ19 were also over‐represented in the SF compared with the blood in some patients. Taken together our results provide more information concerning the role of mycobacterial antigens in RA and suggest that there may be an in vivo clonal expansion of T lymphocytes in the synovium.
Activated CD3−CD16+ Natural Killer Cells Express a Subset of the Lymphokine Genes Induced in Activated αβ+ and γω+ T cells
Authors: BIASSONI R. , FERRINI S. , PRIGIONE I. , PELAK V. S. , SEKALY R. P. , LONG E. O.
Published: June 2006

In this study we analysed the potential of highly purified polyclonal TcR αβ+ TcR γω+ and CD3− NK cells, to produce lymphokines in response to mitogenic stimulation. RNA hybridizations were performed to detect with high sensitivity the induction of multiple lymphokine genes. Upon stimulation with lectin and phorbol ester TcR γω+ lymphocytes expressed the same sei of lymphokine genes as the TcR αβ+ lymphocytes, which included IL‐2. ‐3. ‐4. ‐5. GM‐CSF. TNF α and β, IFN γ. In contrast, a more limited set of lymphokine genes (GM‐CSF. TNF α and β. IFN γ) was induced in activated CD3− NK cells, thus indicating that this subpopulation of cells may display different regulatory functions, with respect to CD3+ T lymphocytes.
Mechanism of T‐Cell Activation by Mycobacterial Antigens in Inflammatory Synovitis
Authors: BURFORD‐FOGGS A. M. , SAMBOL S. P. , POPE R. M.
Published: June 2006

Earlier studies demonstrated enhanced proliferative responses to an acetone precipitable Mycobacterium tuberculosis (AP‐MT) antigenic complex by T lymphocytes from the synovial fluid, compared with the peripheral blood, of patients with inflammatory synovitis. including rheumatoid arthritis. In contrast. decreased proliferation and interleukin 2 (IL‐2) production in response to mitogens by synovial fluid lymphocytes from patients with rheumatoid arthritis has been demonstrated. In order to determine if IL‐2 was produced in response to AP‐MT, the peripheral blood and synovial fluid of patients with inflammatory arthritis were analysed by measuring proliferation and IL‐2 production in response to AP‐MT and tetanus toxoid. A reduction of IL‐2 production relative to proliferation was observed IN some, but not all, synovial fluids of patients who responded to the AP‐MT. Nevertheless, antibodies to IL‐2 as well as interleukin 4 (IL‐4). significantly inhibited proliferation of synovial fluid lymphocytes by AP‐MT. There was no inhibition by antibodies lo interleukin 6 (IL‐6). We conclude that AP‐MT induced proliferation by synovial fluid lymphocytes is mediated by both IL‐2 and lL‐4.
Studies on the Thymus in Chagas' Disease II. Thymocyte Subset Fluctuations in Trypanosoma cruzi‐infected mice: Relationship to Stress
Authors: LEITE DE MORAES M. C. , HONTEBEYRIE‐JOSKOWICZ M. , LEBOULENGER F. , SAVINO W. , DARDENNE M. , LEPAULT F.
Published: June 2006

Changes in thymic T‐cell subsets in mice acutely infected with Trypanosoma cruzi have been studied in both C3H/HeJ and C57BL/6 mice. The significant decrease in thymocyte number, observed in both mouse strains on day 14 post‐infection correlated with a drastic decrease in CD4+CD8+ cell number, whereas the number of CD4−CD8−, CD4+CD8− and CD4−CD8+ cells remained essentially unchanged. The important increase in CD3hi cell frequency confirmed that resistant thymocytes during Chagas' disease development were mostly medullary thymocytes, whereas the thymic cortex was largely depleted, as previously observed on thymus sections. This involution of the thymus could have been due to the increase of circulating glucocorticoid levels observed after infection. However, similar cell modifications were found in infected adrenalectomized mice whose serum corticosterone levels were only slightly augmented. Thus, the thymic alterations appear not lo be linked to stress responses, al least those dependent on high levels of circulating glucocorticoids.
Interleukin 4 and Interferon Gamma Production in Restimulated CD4+ and CD8+ Cells Indicates Memory Type Responsiveness
Authors: SANDER B. , CARDELL S. , MÖLLER E.
Published: June 2006

Interleukin 4 (IL‐4) and interferon gamma (IFN‐γ) production was analysed in murine spleen cells during primary and secondary mitogen Stimulation in vitro. The kinetics, frequency and phenotype of single lymphokine‐producing cells were studied by combining intracytoplasmatic immunofluorescence and surface staining. Both IL‐4 and IFN‐γ was produced by CD4+ as well as CD8+ cells, however 75–80% of IL‐4 producers were CD4+ and 90% or IFN‐γ+ cells were CD8+

In primary stimulations. concanavalin. A (Con A) activation or anti‐CD3 antibody together with phorbol 12‐myristale 13‐aeetate (PMA) induced different patterns of lymphokine production. Approximately the same frequency of IFN‐γ+ cells was induced by both stimulation procedures but the kinetics was different with a peak at 30 h using Con A and at 52 h using anti‐CD2and PMA. IL‐4 production peaked al 52 h, but the frequency of IL‐4+ cells was 8–10 times higher after stimulation by anii‐CD3 and PMA than after Con A stimulation.

During restimulation of the mitogen activated cells, lymphokines were rapidly produced; both IL‐4and IFN‐γ production peaked at 8–11 h. Only a small increase in the frequency of IL‐4+ cells was seen, at most two to three limes. No evidence for a major shift of lymphokine produced between primary and secondary stimulations could be found. Instead, the pattern of lymphokine production induced by the primary stimulus was dominant also in secondary cultures irrespective of stimulation condition.
The Influence of T Cells on the Immunoglobulin Repertoire and the Affinity Maturation of the Immune Response Against Dextran B512 in C57BL/6 Mice
Authors: FERNANDEZ C. , MÖLLER G.
Published: June 2006

A collection of hybridomas from C57BL/6 mice producing antibodies to dextran B512 was analysed and found to reflect the immune response in vivo with regard to immunoglobulin class expression. T cell dependency and antibody affinity. IgM‐. IgG‐3. and IgG‐2b. and IgA‐producing hybridomas were found. IgG3‐producing hybridomas were obtained from nude mice, indicating T cell independent IgG3 synthesis. All monoclonal antibodies were of light chain. A major anti‐dextran idiotype WU expressed in many monoclonals, Secondary immune responses to dextran were also suppressed at the hybridoma cell level. However. hybridomas from secondary responses produced antibodies expressing the major idiotype. suggesting that anti‐idiotype mediated suppression was not responsible for the reduced secondary response. Most monoclonals belonged to the VHJ558 family, but the IgG3‐producing hybridomas showed a preferential use of genes from the VHX24 family. All monoclonals were directed against internal structures of the dextran molecule. The affinity for dextran of the IgG antibodies produced in secondary immune responses was drastically increased, even when the mice were immunized with thymus‐independent forms of dextran. indicating that T helper cells need not be involved in affinity maturation of the immune response.
A Comprehensive Immunological Analysis in Chronic Fatigue Syndrome
Authors: GUPTA S. , VAYUVEGULA B.
Published: June 2006

A detailed analysis of cell‐mediated and antibody‐mediated immunity was performed in 20 CDC‐defined patients with chronic fatigue syndrome (CFS) and 20 age‐ and sex‐matched healthy controls. CD3 +, CD4 +, CD8 +, and CD20+ lymphocytes were comparable in two groups. Natural killer cells as defined by CD16, CD56 and CD57 antigens were significantly reduced in CFS. A significant increase in the proportions orCD4+ICAM 1+ T cells was observed in CFS. Monocytes from CFS displayed increased density (as determined by mean fluorescence channel numbers) of intercellular adhesion molecule 1 (ICAM‐1) and lymphocyte function associated antigen 1 (LFA‐1). but showed decreased enhancing response to recombinant interferon‐γ in vitro. The lymphocyte DNA synthesis in response lo phytohaemoglobulin (PHA). Concanavalin A (Con A) and pokeweed mitogen (PWM) was normal but the response to soluble antigens was significantly reduced. Serum IgM. IgG, IgA, and IgG sublclasses were normal. In vivo specific antibody response to pneumococcus vaccine was depressed in CFS. Forty percent of patients showed titres of anti‐human herpes virus 6 (anli‐HHV‐6) antibody higher than that in the controls (≤ 1/80). These data suggest immunological dysfunction in patients with chronic fatigue syndrome. The significance of these observations is discussed.
Auto‐ and Polyreactivity of IgM from CD5+ and CD5− Cord Blood B Cells
Authors: MACKENZIE L. E. , YOUINOU P. Y. , HICKS R. , YUKSEL B. , MAGEED R. A. , LYDYARD P. M.
Published: June 2006

The presence of the CD5 (67 kDa) molecule on the surface of B cells has been considered a marker for cells producing auto‐and polyreactive antibodies. Cord blood B lymphocytes (rich in CD5+ B cells) have been sorted into CD5 positive and negative populations by flow cytometry using monoclonal antibodies to CD20 and CD5. Clones of these populations were obtained by immortalization mill Epstein Barr virus. Clones derived from both CD5+ and CD5− B cells produced IGM which was auto‐ and polyreactive with a higher frequency of these specificities in the CD5+ population. These data indicate Ihm expression of surface CD5 on cord blood B cells Is not a definitive marker of an auto/polyreactive population.
A Monoclonal IgM Antibody to a Methylcholanthrene‐Induced Tumour
Authors: GIGLIOTTI D. , TENEBERG S. , ANDFRSSON R. , ÅNGSTRÖM J. , KARLSSON K.‐A. , WIGZELL H. , HANSSON M.
Published: June 2006

A monoclonal IgM antibody, H17, has been obtained from rats immunized with mouse fibrosarcoma cells from an in vitro established methylcholanthrene(MCA)induced tumour. H17 shows specific and very selective binding to α‐N‐acetylgalactosamine (GalNAcα) when tested for reactivity to a panel of glycolipids. It cross‐reacts with GalNAcα on the Forssman antigen extracted from dog small intestine, but not from the human blood group A determinant, a finding not commonly observed among antibodies with this specificity. Despite its specificity. H17 does not react with TA3‐Ha, a mouse mammary adenocarcinoma, known to express the Tn antigen (GalNAcα‐O‐Ser/Thr). The uniqueness of HI7 probably relates to the fact that it has been generated against an MCA‐induced tumour rather than against the pure saecharide itself, Minimum energy conformation structures of different GalNAcα containing saecharide molecules were computer modelled lo allow a plausible interpretation of the accessible site of GalNAcα for successful interaction with the H17 para tope as compared lo other GalNAcα binding antibodies.
The Effect of 20‐Trifluoromethyl Leukotriene B4 on Neutrophil Functional Responses
Authors: NILSSON E. , GYLLENHAMMAR H. , LERNER R. , PALMBLAD J. , RINGERTZ B.
Published: June 2006

20‐triHuoromelhyl‐lcLikotriene B4 (20CF3LTB4) is a stable derivative of leukotriene B4 (LTB4) that is not subjected to co‐oxidation to less active metabolites. 2OCK3LTB4 was as potent as LTB4 as a chemotactic, adhesion‐promoting and aggregatory agent for human neutrophils, but bad only 11 ± 3% of the ability to induce an Oxidative response. Nonetheless, both compounds were equally efficient in order to confer a rapid and monophasic increment of the concentration of cytosolic calcium. The kinetics of the calcium, aggregatory and chemiluminescent responses to 2OCF3‐LTB4 were similar to that of LTB4. These findings suggest that the insertion of the trifluoromethyl group into the LTB4 molecule causes a shift of the biological activity profile, suggesting that 20CF3LTB4 binds mainly to high affinity LTB4 receptors. Moreover, the similarity of the response kinetics of LTB4 and 20CF3‐ LTB4 suggests that the mechanism for the rapid and transient responses of LTB4 is not due to its w‐oxidation.
Differential Effects of Interferon‐γ and Low Molecular Weight BCGF on Growth of Human B Lymphocytes; Interferon‐γ Prolongs the Increased c‐MYC mRNA Levels After Activation
Authors: LØMO J. , SMELAND E. B. , STOKKE T. , HOLTE H. , FUNDERUD S. , BLOMHOFF H. K.
Published: June 2006

We have characterized the growth‐stimulating effect of Interferon‐γ (TFN‐γ) on various parameters of B cell growth, and compared the effects with those of low molecular weight B cell growth factor (lmw BCGF). We have found that IFN‐γ did not affect early changes induced by anti‐μ, like initial calcium‐flux and rise in mRNA‐ and protein levels of the proto‐oncogene c‐myc measured at 3 h. On the other hand, IFN‐γ enhanced the effect of anti‐μ on parameters measured later in the GI phase of the cell cycle, such as expression of the transferrin receptor and general transcriptional activity, measured as an increase in 7‐amitioactinoiTiycin D binding. In particular, whereas the c‐myc levels in anti‐μ‐treated cells peaked al 3 h and then gradually declined. IFN‐γ together with anti‐μ maintained the c‐myc levels at 24 h at approximately the same levels as seen at 3 h. Overall, lmw BCGF had a more potent effect on the parameters affected by IFN‐γ, correlating with stronger enhancement of DNA synthesis. However, in contrast to IFN‐γ lmw BCGF did not affect anti‐μ‐induced c‐myc mRNA levels. Thus this study has revealed differences between two B cell growth factors in effects on B cell cycle parameters.
Characterization of a Monoclonal Antibody Directed Against the Phosphotyrosine Kinase p56lck, in Human T Cells
Authors: ANSOTEGUI I. J. , CHOW S. C. , JEDDI‐TEHRANI M. , MELOCHE S. , PAWSON A. , SEKALY R.‐P. , MAKI T. W. , WIGZELL H.
Published: June 2006

A monoclonal antibody (anti‐p56lck) was generated against a fusion protein containing the residues 145–509 of the human p56lck, a lymphoeyte‐specific membrane‐associated protein tyrosine kinase. The involvement of this enzyme in T‐cell transmembrane signalling seems to be an early and crucial event during T‐cell receptor‐mediated activation. We have produced a monoclonal antibody which recognizes p56lck in free form and when associated with CD4. H functions in western blot analysis and is capable of selectively blocking auto‐phosphorylation of this kinase. This monoclonal antibody should be useful for investigating the role of p56lck in T‐cell activation.
Non‐immune VH‐binding Specificity of Human Protein Fv
Authors: BOUVET J.‐P. , PIRES R. , QUAN C. , ISCAKI S. , PILLOT J.
Published: June 2006

The specificity of human F(ab)‐binding Protein Fv (previously called Protein F), a sialoprotein released into the digestive tract mainly during hepatitis, was investigated with fragments or chains of monoclonal immunoglobulins. Protein Fv bound an unreduced H‐chain dimer of a monoclonal human IgA2m(I) molecule but neither the corresponding L‐chain dimer, nor several Bence‐Jones molecules. Using enzymatic subfragments of F(ab)μ(, or F(ab′)2γ, a significant binding was observed with Fv fragments (VH + VL), while Fb fragments (CHI+CL) were inactive. Taken altogether, these results prove that the structure recognized by Protein Fv is located in the VH domain. This structure probably involves discontinuous epitopes linked by a disulphide bond, which are destroyed by combined reduction and dissociation. Protein Fv does not interfere with the antigen‐binding site, since there was no reciprocal inhibition with the antigen‐antibody reaction.
CD5+ B‐Lymphocytes are as Dependent on T‐Helper Cells and as Responsive to T‐Suppressor Cells in Pokeweed Mitogen‐Stimulated Activation as Conventional CD5− B‐Lymphocytes
Authors: JONES B. M.
Published: June 2006

Pokeweed mitogen (PWM)‐stimulated cultures were established with varying numbers of CD4+ or CD8+ cells and CD5+ immunoglobulin‐secreting cells were subsequently identified using a combination of haemolytic plaque formation and rosetting wilh immunobeads coated with anti‐Leu‐1. Il was found that CD5+ and CD5+ B‐cells required similar numbers of CD4+ cells for half‐maximum plaque‐forming cell (PFC) induction and that 50% suppression of both CD5+ and CD5− PFC occurred with similar numbers of CD8+ cells. Suppression of both CD5+ and CD5− PFC could occur when T‐helper signals were supplied by a PWM‐stimulated culture supernatant, indicating that both subsets could be directly suppressed by CD8+ cells rather than by indirect suppression of T‐helper cells. It is concluded that CD5+ B‐cells are as responsive to T‐regulatory influences as conventional CD5− B‐cells.
Adenosine Deaminase and Thymocyte Maturation
Authors: DOHERTY P. J. , PAN S. , MULLOY J. C. , THOMPSON E. , THORNER P. , BARANKIEWTECZ J. , ROIEMAN C. M. , COHEN A.
Published: June 2006

The congenital absence of adenosine deaminase in humans results in severe combined immunodeficiency. To clarity the process whereby thymocytes are destroyed in the absence of adenosine deaminase activity, we induced a parallel condition in mice through the injection of an inhibitor of adenosine deaminase, deoxycoformycin. We have observed that deoxycoformycin, in addition to maintaining high levels of dATP in thymocytes, blocks the progression of thymocyte differentiation at two points. As a result of the first block, the corlex is depleted of immature cortical thymocyles while CD4+ CD8+ thymocytes with functionally rearranged T‐cell receptors survive. As a result of the second block, the CD4+ CD8− thymocytes are prevented from further differentiation lo mature CD4− CD8+ or CD4− CD8+ T lymphocytes and accumulate at the corticomedullar junction and in the medulla. These observations suggest that the maintenance of dNTP pools by adenosine deaminase is critical to at least two stages of thymocyte differentiation.
Oligopeptide Induction of a Secondary Cytotoxic T‐cell Response to Epstein‐Barr Virus In Vitro
Authors: SCHMIDT C. , BURROWS S. R. , MOSS D. J. , SCULLEY T. B. , MISKO I. S.
Published: June 2006

Three Epstein Barr virus (EBV) nuclear antigen (EBNA)‐encoded oligopeptide epitopes have been mapped, each capable of acting as a recognition determinant for class I‐restricted lysis by CDS+cytotoxic T lymphocytes (CTL). This report shows that each peptide, when presented on an appropriate autologous antigen‐presenting cell (APC), also stimulates EBV‐specific memory T cells present in peripheral blood mononuclear cell (PBMC) populations to develop in vitro into peptide‐specific CTL. These CTL specifically lysed autologous EBV‐infected lymphoblastoid cell lines (LCD and peptide‐sensitized uninfected targets. Identical viral oligopeptides could therefore function as recognition determinants for both the induction and commission of class I‐restricted specific cytotoxicity. A model system is described in which autologous phytohacm‐agglutinin (PHA) blasts present exogenous peptide during the stimulation phase. The magnitude of the peptide‐specific CTL response was dependent on the concentration of peptide added to the APC and specific lysis was inhibited by anti‐class I monoclonal antibody (MoAb) but not anti‐class II MoAb. Cultures depleted ofCD8+ T cells by cell separation with immunomagnetic beads prior to stimulation invariably failed to generate a peptide‐specific CTL response. However, the effect of CD4 depletion on CTL activity was equivocal and indicated that a need for CD4+ T cells as accessory helper cells may depend on the efficiency of the APC to elaborate their own help. This model has advantages in the analysis of events involved in the development of CTL activity in vitro.
Human Natural Killer Cells Express Different Integrins and Spread on Fibronectin
Authors: VIRTANEN I. , YLÄNNE J. , VARTIO T. , SAKSELA E.
Published: June 2006

Human natural killer (NK) cells adhered and most of them also actively spread on cellular fibronectin (cFn). plasma Fn (pFn) and its M, 120,000–140,000 or Mr 105,000 cell‐binding proteolytic Fn‐fragments as well as on heparin‐binding Fn‐fragments containing the alternative ceil binding site. The cells did not spread on vitronectin, laminin or collagens. Adhesion on M, 105,000 Fn fragment containing the cell binding site, could be prevented by the synthetic peptide GRGDS but not by an inactive peptide, whereas adhesion on heparin‐binding Fn fragments was unaffected by the peptide. Spreading of the NK cells led to a distinct reorganization of F‐actin. Immunoprecipitation with monoclonal antibodies (MoAb) against the β1integrin subunit of radioactively surface‐labelled cells revealed a broad polypeptide band of M, 140,000 under reducing conditions and a polypeptide doublet of Mr 160,000 and Mr110,000 under non‐reducing conditions. Identical polypeptides. corresponding to the α‐and β‐subunits of the Fn‐receptor complex, were bound to the Mr 105,000 chymotryptic Fn‐fragment coupled to Sepharose® Similar experiments with small lymphocytes did not reveal any polypeptides. Immunofluorescence results with McAbs suggested that among the α‐subunits of integrins the α3α4, and α5subunits are expressed in NK cells. The present results suggest that non‐activated NK cells, but not small lymphocytes, express β1‐integrins, and that at least the Fn‐receptors α4β1and α5β1may function in the adhesion and migration of NK cells.
Glycosylation is Required for Coronavirus TGEV to Induce an Efficient Production of IFNα by Blood Mononuclear Cells
Authors: CHARLEY B. , LAVENANT L. , DELMAS B.
Published: June 2006

Porcine peripheral blood mononuclear cells (PBMC) are induced to produce interferon alpha (IFNα) following in vitro exposure to coronavirus TGEV (transmissible gastroenteritis virus)‐infected glutaraldehyde‐fixed cell monolayers or lo TGEV virions. In the present report, we examined the possibility that glycosylation of viral proteins could play a major role in interactions with PBMC leading to the production of IFNα. Con A pretreatment of TGEV‐infected cell monolayers before fixation with glutaraldehyde and exposure to PBMC caused a dose‐dependent inhibition of IFNα induction, implying that masking of carbohydrates at the surface of infected cells lowered IFNα induction. Similarly, inhibition of N‐linked glycosylation by tunicamycin during viral infection of cell monolayers altered their ability to induce IFNα. In addition, complete cleavage of complex type' oligosaccharides by peptide‐N‐glycohydrolase Flowered the capacity of TGEV virions to induce IFNα. Thus, these findings strongly suggest that glycosylation of the viral proteins, and more precisely the presence of complex‐type oligosaccharides, is an important requirement for a completely efficient interaction with PBMC leading to the production of IFN‐α.
T Lymphocytes Degrade Fibronectin
Authors: BERGSTRÖM S. E. , HAUZENBERGER D. , SUNDQVIST K. G.
Published: June 2006

Mononuclear cells cause disappearance of fibronectin synthesized and released by fibroblasts. This disappearance of extracellular fibroblast fibronectin is accompanied by the appearance of components of “lower” molecular weight indicating that a fibronectin‐degrading enzymatic activity is responsible for the effect. Additional support for the existence of a fibronectin‐degrading enzyme is that mononuclear cells degrade iodinated fibronectin attached to a collagen matrix, Furthermore, lymphocytes seem to mediale the degradation of fibronectin whereas monocytes rather act inhibitory.
The Effect of Various Contexts of Stress on the Mouse Spleen Lymphocytes and Macrophage Co‐stimulatory Activity
Authors: AARSTAD H. J. , THIELE D. , SELJELID R.
Published: June 2006

Mice stressed daily by brief cold water immersions for 1, 8 or 14days showed changes in immune system function which were dependent on the number of mice per cage, frequency of stress exposures and total number of stress exposures. Changed percentages of spleen B AND C D4. but not of CPS cells were determined when the mice were stressed either once or twice daily. With CD4 cells, increased percentages were seen after stress once daily but a decreased percentage was seen after stress twice daily. furthermore, the Concanavalin A‐stimulated spleen cell mitogenesis was decreased after 1 day of stress in mice stressed once daily as opposed lo after H and 14 days of stress in mice stressed twice daily. After 14 days of stress, the lipopolysaccharide stimulated mitogenesis was increased if the mice were stressed once daily but decreased if the mice were stressed twice daily. With two mice per cage, we observed a decreased spleen cell mitogenesis after 14 days of stress. With four mice per cage, the spleen cell mitogenesis was decreased after 8 and 14 days of stress. If spleen cell populations from mice stressed twice daily for 8 days were depleted of macrophages and CD4 or CD8 cells. The effect of stress on the mitogenesis was removed from t he CDS cells. Spleen cells of mice stressed for 14 days showed a decreased mitogenesis when depleted HI adherent cells and reconstituted with adherent cells from control mice, Further more, the adherent cells from these mice had decreased ability to support mitogenesis of adherent cell‐depleted spleen cells from control mice as well as a decreased IL‐1 production.
Differential Kinetics of Various Subsets of Thymic Bone Marrow‐Derived Stromal Cells in Rat Chimeras
Authors: MURAWSKA M. B. , DUIJVESTIJN A. M. , KLATTER F. A. , AMMERLAAN W. , MEEDENDORP B. , NIEUWENHUIS P.
Published: June 2006

Identification of those cells within the thymic stroma which are responsible for tolerance induction remains controversial. Evidence derived from studies of bone marrow chimeras or thymus transplants attributed this function to cells of haematopoietic origin, usually class II positive medullary dendritic cells (DC). Recent data suggest, however, that a stromal element located in the thymus cortex might be involved in negative selection. To further explore this issue we used immunohistology and immunocytology with a combination of allotype and cell type specific monoclonal antibodies (MoAb) to study the turnover of thymic stromal cells of haematopoietic origin to different rat models of allogeneic and congenic bone marrow (BM) radiation chimeras. Use of CFU‐GM cultured BM inoculum for congenic recipients allowed us to distinguish between direct homing of donor mycloid cells and the delayed migration of the donor stem cell progeny after the post‐irradiation recovery of the recipient. Our data indicate a heterogeneity in the turnover rate of thymic mobile stromal cells. While DC and a subset of macrophages located m the cortex as well as m the medulla (EDI+). within 4 weeks were virtually all of donor type, cortical macrophages detected by ED2 MoAbs were still incompletely replaced after a period as long as 20 weeks. Slow turnover, location and variable class II expression may imply a role for thymic cortical macrophages in (self‐) tolerance induction.
Idiotypic Profile of Natural Autoantibodies in Newborn and Young Adult BALB/c Mice
Authors: ZÖLLER M. , ACHTNICH M.
Published: June 2006

Idiotypic profiles of autoreactive monoclonal antibodies (MoAb) were evaluated by their reactivity with a panel of alkaline phosphatase (AP)‐coupled detector MoAb derived from the same fusions. Attention was given to the question of whether differences exist between MoAb derived from spleen cells (SC) or thymocytes (TC) and whether ID profiles would change during post‐natal development.

In the newborn, natural auioantibodies and MoAb which did not react with any one of eight autoantigens displayed different ID profiles, autoreactive MoAb being characterized by the expression of a restricted pattern of ID. During post‐natal development. changes of ID expression were only observed with autoreactive MoAb. Many ID which were detected on MoAb derived from 6‐day‐old mice were not detected on SC‐derived MoAb from young adults, while a few ID were significantly over‐represented. Furthermore, especially with TC‐derived MoAb, a clear linkage between certain idiotypes and autoantigen spectficities could be demonstrated.

Thus, in contract to non‐autoreactivc MoAb, natural autoantibodies in the young adult were characterised by expressing only a selected number of ID al high frequency. Furthermore, the B‐cell enivironment apparently played a role, since there were marked differences between ID profiles of TC‐ versus SC‐derived MoAb. The data are interpreted in the sense that expansion and maturation of naturally activated autoreactive B cells are controlled rather than being random processes.
Autoreactive Antibodies in Thymus and Spleen of Neonatal and Young Adult BALB/c Mice: Influence of Prenatal Tolerization
Authors: ACHTNICH M. , ZÖLLER M.
Published: June 2006

The repertoire of autoantibody‐producing B cells was evaluated in a collection of spleen‐ and thymus‐derived hybridomas from 6‐ and 2S‐day‐old BALB/c mice, which were untreated or prenatally tolerized with trinitrobenzenesulphonic acid (TNBS). MoAb were tested for their reactivity with TNP‐BSA and the autoantigens thyroglaobulin (TG), myoglobin (MG), actin (AC), cytochrome C (CY), collagen (CO), transferrin (TF), single‐stranded DNA (dsDNA), double‐stranded DNA (dsDNA), and bromelain‐lreated mouse red blood cells (BrMRBC).

More than 10% of spleen cell (SC)‐derived MoA b from 6‐and 28‐day‐old control mice did bind to AC, ssDNA, dsDNA, MY, and TG, the frequency of MoA b reacting with MY, TG, And BrMRBC increasing with age. Thymus cell (TC)‐derived hybridomas contained autoreactive clones Eoo. but only few of them produced multireactive MoAb.

MoAb from prenatally TNBS‐treated mice were more frequently autoreactive than MoAb from control mice, especially if derived from TC hybridomas. The most remarkable difference in the reactivity pattern as compared with MoAb from untreated mice consisted of a significant increase in the frequency of TG‐, My‐, ssDNA‐ and above all of dsDNA‐reactive MoAb, all TC‐derived multireactive MoAb binding to dsDNA.

The differences in autoreactivily between MoAb from prenatally untreated and TNBS‐treated mice as well as age‐ and organ‐related variations support the interpretation that part of the repertoire of naturally activated B cells is not random but is influenced by and responding to the available panel of self antigens.
Multiple Sclerosis: Cells Secreting Antibodies Against Myelin‐Associated Glycoprotein are Present in Cerebrospinal Fluid
Authors: BAIG S. , OLSSON T. , YU‐PING J. , HÖJEBERG B. , CRUZ M. , LINK H.
Published: June 2006

We evaluated the B‐cell response in cerebrospinal fluid (CSF) and blood by enumerating cells secreting antibodies to myelin‐associated glycoprotein (MAG) and, for reference, to myelin basic protein (MBP), two myelin components which may constitute targets for autoimmune attack in multiple sclerosis (MS). Among 25 untreated MS patients, 12 had cells in CSF secreting anti‐MAG IgG antibodies (mean value 1 per 1429 CSF cells) and three also had cells secreting anti‐MAG antibodies of the IgM Isotype but at lower levels. In CSF from 2 out of 10 MS patients examined, anti‐MAG and anti‐MBP IgG antibody‐secreting cells were present concurrently. Antibody‐secreting cells were less frequent in blood and bone marrow, reflecting compartmentalization to CSF. Anti‐MAG antibody‐secreting cells were found in CSF from only I out of 27 control patients. The intrathecal production of anti‐MAG and anti‐MBP antibodies may be important in the pathogenesis of MS.