bluuebunny/crossref_metadata_2025 huggingface dataset:
Natural Killer (NK) Cell Activity in Mice with Acute Graft‐Versus‐Host Reactions: Characterization of a Thy‐1+ NK‐Like Cell with a Broadened Spectrum of Lytic Activity in the Spleen and Lymph Nodes
Authors: MACDONALD G. C. , GARTNER J. G.
Published: June 2006

We have shown that acute (GVH) reactions produced in the parental Kt hybrid combination. A/J→(C57BL/6 × A/J) F1 result in the activation of two cytotoxic cell populations: a host‐derived Thy‐1+/‐ natural killer (NK) cell with a lytic spectrum confined to YAC‐1 targets, and a donor‐derived Thy‐1+ NK‐like cell that has the ability to lyse target cells that are normally insensitive to lysis by NK cells. Cold‐target inhibition (CTI) experiments have shown that the greater range of target cell killing seen in the NK‐like population is mediated by a single effector cell with a broadened lytic repertoire. Percoll density fractionation studies have revealed that NK and NK‐like cell co‐fractionate, suggesting that btith are large granular lymphocytes. We have also shown that NK‐like cells do not express either Lyt‐2 or L3T4 markers. We have also observed that there is a close temporal relationship between elevated levels of mterleukin‐2 (IL‐2) secretion by spleen cell cultures from mice with GVH disease and the subsequent emergence of splenic NK activity in both acute [A/J→(C57BL/6×A/J)F1] and chronic (A/J→ CBA×A/J)F1 GVH reactions. We have also noted that, despite high levels of IL‐2 secretion, mice with chronic GVH reactions do not generate NK‐like activity. Interferon (IFN) measurements have shown that, although increased IFN activity can be detected in both acute and chronic models, a preponderance of IFN‐α/β and some IFN‐γ is produced in the acute reaction, whereas only IFN‐γ can be detected in the chronic model. These results suggest that, although IL‐2 may participate in augmenting conventional NK activity. IL‐2 by itself does not generate NK‐like activity. We suggest that IFN‐α/β may be the cytokinc that, either alone or in concert with IL‐2, triggers the NK‐like cell response. The NK‐like cell described in our study resembles a phenotypically identical, donor‐derived large granular lymphocyte, identified by others, in close proximity to dead or dying epithelial cells in mice with GVH disease [14]. It has been suggested that these cells may mediate tissue injury. If in fact these graft‐derived NK‐like cells are involved in the pathogenesis of acute GVH disease, our present findings suggest that they must first be activated by an appropriate complement of cytokines that includes IFN‐α/β. We postulate that whether or not a GVH reaction pursues an acute, rapidly lethal, course depends not only on the cohort of effector cells in the graft, but also on the stimulatory cytokines released during the early, lymphoproliferative phase of the reaction.
Serum Amyloid A Isoforms in Inflammation
Authors: RAYNES J. G. , McADAM K. P. W. J.
Published: June 2006

Serum amyloid A protein (SAA) was extracted from serum using hydrophobic interaction chromatography and four or six isoforms were separated by isoelectric‐focusing. These represented three pairs of isoforms, each with and without an N‐terminal arginine. SAA I (pi 6.1). SAA l des‐arg (pl 5.9). SAA2α (pi 6.9) and SAAα: des‐arg (pl 6.6) were found to be present in all individuals from Europe and the USA. A minority of these individuals (11 of 56) expressed SAA2/β (pl 7. 1) and SAA2/β des‐arg (pi 6.8).

Serum from patients in Papua New Guinea and Malawi both showed a much higher frequency ofSAA2/β. There was no indication of altered isoforms in regions with high incidence of reactive AA amyloidosis. In sequential serum samples, concentrations of des‐arg isoforms were found to reach a maximum 0 24 h later than isoforms with an arginine. Concentrations of the isoform SAA1 decreased faster in five of six patients (16±:7.5 h to decrease 50%) than SAAl des‐arg (22± 11 h to decrease 50%). Variations in the handling of N‐terminal arginine may be important for the formation‐susceptibility of amyloid deposits.
Expression of CD26 (Dipeptidyl Peptidase IV) on Resting and Activated Human T‐Lymphocytes
Authors: MATTERN T. , SCHOLZ W. , FELLER A. C. , FLAD H.‐D. , ULMER A. J.
Published: June 2006

CD26 is an activation antigen which is expressed on the surface of human T‐lymphocytes. It has been characterized to be the dipeptidyl peptidase IV (DPP IV). Considerable amounts of CD26 are already present on resting T‐lymphocytes. The expression of CD26 is enhanced by T‐cell milogens or antigens. A correlation of CD26 expression and of enhanced enzymatic activity was observed after T‐cell activation. Our data indicate that not only the immunoreactivity, but also the enzymatic activity of CD26 are detectable on the cell surface. In addition, de novo expression of CD26 was demonstrated on CD26‐negative T‐cells after mitogenic or antigenic stimulation.

CD26 expression is initiated during the G1 phase of the cell cycle. The expression occurs nearly simultaneously with HLA‐DR, but later than CD25. Similar lo CD25 and HLA‐DR, CD26 is not a permanent marker on the surface of T‐lymphocytes, but is down‐regulated after 7 days of culture. When testing the influence of interleukin I, interleukin 2, tumour necrosis factor, and interferon‐γ on the expression of CD26, no effect was found on unstimulated or on mitogen‐stimulated T‐lymphocytes. The binding of two different monoclonal antibodies against CD26 (anti‐DPP IV and anti‐Tal) to resting and activated T‐lymphocytes revealed a different pattern of immunoreactivity. Resting T‐lymphocytes reacted stronger with anti‐DPP IV than with anti‐Ta I. However, binding of the two monoclonal antibodies to T‐cell blasts did not show significant differences. These data indicate that CD26 may be expressed in differently modulated configurations on the surface of T‐cells, which may be associated with a distinct status of activation and or function.
Subcellular Distribution of Monoclonal Antibody Defined Epitopes on Immunodominant Mycobacterium tuberculosis Proteins in the 30‐kDa Region: Identification and Localization of 29/33‐kDa Doublet Proteins on Mycobacterial Cell Wall
Authors: RAMBUKKANA A. , DAS P. K. , CHAND A. , BAAS J. G. , GROOTHUIS D. G. , KOLK A. H. J.
Published: June 2006

Two different groups of monoclonal antibodies (MAbs) directed to different epitopes on 30‐kDa region proteins of Mycobacterium tuberculosis were isolated; MAbs 5F9, 5D5 and 5D2 reacted with a single 33‐kDa protein band, whereas MAb 3A8 readied with A distinct 29,33‐kDa doublet when analysed by immunoblotting. The present paper describes the distribution of MAbs defined epitopes in the 29 33‐kDa region proteins in well‐characterized subcellular fractions: cytosol, plasma membrane, cell wall as well as culture filtrate of M. tuberculosis. MAbs 5F9, 5D5 and 5D2 reactive epitopes were found in cytosol, whereas 3AS epitope is distributed in all cellular compartments of the mycobacterium as well as in the culture filtrate. Localization of these epitopes by indirect immunoflourescence and immunogold‐labelling demonstrated that only 3A8 epitope is present on the cell surface of the mycobacterium. Both immunoblotting and EL[SA showed that only MAh 3A8. and not 5F9. 5D5 and 5D2, reacted with secreted BCG 85 antigen complex of Mycobacterium bin‐it BCG. Furthermore, using an MAb 3AS‐coupled affinity column, we purified antigen 3A8 from the cytosol fraction of M tuberculosis. All these MAhs reacted with antigen 3AK with varying degrees of intensity, thus suggesting that they are directed to a single protein. Absence of 5F9, 5D5 and 5D2 epitopes in the cell wall, culture filtrate and BCG‐85 complex suggests that these epitopes might have been lost during the processing of the same 33‐kDa protein on its way out from cytosol to the cell wall or when the protein is secreted out into the culture filtrate. Our results demonstrate, for the first time, direct evidence of the presence of a 30‐kDa region protein not only in secreted antigens but also in the cell wall and on the cell surface of the mycobacterium.
Lymphocyte Activation and Regulation of Three Adhesion Molecules with Supposed Function in Homing: LECAM‐1 (MEL‐14 Antigen), LPAM‐1/2 (α4‐Integrin) and CD44 (Pgp‐1)
Authors: BÜHRER C. , BERLIN C. , JABLONSKI‐WESTRICH D. , HOLZMANN B. , THIELE H.‐G. , HAMANN A.
Published: June 2006

Directed migration of lymphocytes from blood into lymph nodes and gut‐associated lymphatic tissue, also referred to as homing, is subject to change following activation. Lymphocyte migration into lymphoid organs in vivo and binding to high endothelial venules in vitro is largely suppressed after short‐term stimulation with phorbol esters. The observed functional alterations were correlated with changes in the expression of three putative homing receptors. LECAM‐1 (MEL‐14 antigen), LPAM‐1/2 (α4‐integrin) and the murine CD44 (Pgp‐1, H‐CAM, Hermes‐antigen equivalent) upon different modes of cellular activation. Expression of LECAM‐1 (gp90 MEL‐14), a lymphocyte adhesion molecule implicated in targeting extravasation into lymph nodes, was found to be lost almost completely within minutes after protein kinase C activation LECAM‐1 re‐expression occurred within less than 24 h. Rapid loss of LECAM‐1 was also observed after calcium ionophores whereas anti‐CD3 or concanavalin A elicited a gradual and heterogeneous loss of LECAM‐1 becoming delectable after several hours only. A number of cytokines tested were not able to induce alterations in LECAM‐1 expression. In contrast, expression of LPAM‐1/2 (α4‐integrin) and CD44 (Pgp‐1, H‐CAM), two adhesion molecules supposed to direct extravasation into Peyer's patches, remained stable for hours after every stimulus tested; CD44 expression gradually increased 24 h after mitogenic activation, whereas a small reduction only was observed for the expression of the α4‐chain under certain conditions. Thus, reduced extravasation of lymphocytes into Peyer's patches after activation is not due to a decline in the surface density of LPAM‐1/2 α‐chain or CD44 whereas alterations in migration into lymph nodes parallel the expression of LECAM‐1.
Tumour Necrosis Factor B Gene Polymorphism in Relation to Complotype in Couples with Spontaneous Abortions and in Control Families
Authors: LAITINEN T. , LOKKI M.‐L. , TULPPALA M. , YLIKORKALA O. , KOSKIMIES S.
Published: June 2006

Nco I restriction fragment length polymorphism (RFLP) of the tumour necrosis factor B (TNFB) gene gives two allelic fragments of 5.5 and 10.5 kb, corresponding to the TNFB*1 and TNFB*2 alleles, respectively.

The frequencies of these alleles were analysed in 121 healthy Finns and 56 Finnish couples suffering from recurrent spontaneous abortions (RSA). In the healthy Finns the frequency of TNFB*1 was 37% and that of TNFB*2 63%, of which the frequency of TNFB*1 was significantly increased compared with the Danish population. No deviation was seen between the RSA couples and the Finnish controls.

TNF genes are located in major histocompalibility complex class III region close to complement (BF, C4A and CAB) genes. Some complotypes associated most often with the TNFB*1 (S01, S30, S02) and some (S31, F320) with the TNFB*2 allele in the healthy Finns.

The combination of the TNFB and the C4 ‘null’ allele differed between the RSA couples and the Finnish controls.
Cyclosporine A in Fat Emulsion Carrier: Immunosuppressive Effect In Vitro
Authors: TIBELL A. , RINGDÉN O. , SUNDBERG B.
Published: June 2006

We have recently shown in a rut model that the acute decrease in glomerular filtration rate caused by intravenous Sandimmun® can be avoided if Cyclosporine A (CyA) is given in a fat emulsion carrier instead. The present study was undertaken to determine the immunosuppressive effect in vitro of this new formulation.

The inhibitory effect on the lymphocyte responsiveness of CyA in fat emulsion carrier was compared with that of CyA dissolved in alcohol, using mitogens as well as alloantigens as stimuli. In the mixed lymphocyte culture and cell‐mediated lympholysis tests and in the interleukin‐2 assay, both formulations seemed equally effective. CyA in fat emulsion was somewhat less effective than CyA in alcohol in the mitogen tests. In clinical practice the fat emulsion would be rapidly metabolized, and to further evaluate our new formulation, studies in experimental Organ transplantation are needed.
Invertebrate Immunity: Another Viewpoint
Authors: Cooper Edwin L. , Rinkevich Baruch , Uhlenbruck Gerhard , Valembois Pierre
Published: June 2006

All vertebrates and invertebrates manifest self/non‐self recognition. Any attempt to answer the question of adaptive significance of recognition must take into account the universality of receptor‐mediated responses. These may lake two forms: (1) rearranging, clonally distributed antigen‐specific receptors that distinguish in the broadest sense between self and non‐self, and non‐self A from non‐self B, latecomers on the evolutionary scene; (2) pattern recognition receptors, the earliest to evolve and still around, necessitating the requirement for induced second signals in T‐ and B‐cell activation. Either strategy need not force upon invertebrates the organization, structure and adaptive functions of vertebrate immune systems. Thus, we can freely delve into the unique aspects of the primitive immune mechanisms of invertebrates. In contrast, using the opposite strategy which is still problematic, i.e. linking invertebrate and vertebrate defence, seems to give us an approach to universality that might eventually reveal homologous kinship.
Lymphocytes Bearing the γ/δ T‐Cell Receptors in Down's Syndrome
Authors: BERTOTTO A. , SCALISE F. , GERLI R. , CASTELLUCCI G. , FABIETTI G. M. , SPINOZZI F. , SENSI L. , VACCARO R.
Published: June 2006

Subjects with Down's syndrome (DS), or trisomy 21, have an increased susceptibility to infections, malignant diseases and autoimmune phenomena. Various arms of the immune system are severely impaired in trisomie patients. We found that the proportion of blood lymphocytes bearing the γ/δ T‐cell receptor (TCR) was significantly higher in adults with trisomy 21 than in age‐ and sex‐matched healthy controls. Interestingly, the increase was mainly due to an over‐expansion of cells which bear non‐covalently bound γ/δ chains on their surface. This contrasts with the normal blood picture, where the great majority of γ/δ T cells express the disulphide‐linked form of the TCR. The fact that trisomie γ/δ T cells are both numerically and phenotypically unbalanced provides further evidence that immunological abnormalities are integral features of DS.
The Natural Killer Cell‐Like Lytic Activity Expressed by Cytolytic T Lymphocytes is Associated with the Expression of a Novel Function‐Associated Molecule
Authors: HARRIS D. T. , JASO‐FRIEDMANN L. , DEVLIN R. B. , KOREN H. S. , EVANS D. L.
Published: June 2006

Experiments were performed to analyse the natural killer cell (NK)‐like cytotoxicity frequently expressed by human antigen‐specific cytolytic T lymphocytes (CTL). To this end, several monoclonal antibodies (MoAbs) previously shown to identify a novel function‐associated molecule (FAM) involved in human NK function were utilized. Flow cytometry revealed that these MoAbs reacted with the majority of human NK, but only with a subpopulation of CTL isolated from primary mixed lymphocyte cultures. Preincubation of CTL with the MoAbs inhibited the NK‐like lysis of K562 targets. Experiments with anti‐CD3 MoAb demonstrated that neither the NK‐like cytotoxicity of CTL nor the lytic activity of NK were mediated by the CD3 complex. Expression of the novel FAM was found to develop in T‐cell cultures at the time that NK‐like cytotoxicity was observed. Repeated in vitro antigenic stimulation of CTL was shown to result in loss of NK‐like cytotoxicity, as well as loss of the FAM on the CTL surface. Thus, NK‐like cytotoxicity displayed by antigen‐specific CTL appears to be mediated by a novel FAM that is identical to that structure found on NK.
Light Chain Distribution of Anti‐Borrelia burgdorferi IgG Antibodies in Lyme Arthritis
Authors: CRUZ M. , SIDÉN A.
Published: June 2006

Joint fluid (JF) and serum IgG from eight Lyme arthritis patients was investigated by isoelectric focusing, blotting to membranes of uncoated nitrocellulose (NC) as well as Borrelia (B.) burgdorferi antigen‐coated NC (immunoblot) and immunoenzymatic staining for visualization of the IgG isotype and characterization of the kappa/lambda light chain distribution. Oligoclonal bands of total IgG occurred in JF as well as in serum (3 cases) or in JF only (1 case); IgG lambda components were present in all four subjects und IgG kappa in three. IgG (Fe frag) reactive but kappa/lambda light chain negative oligoclonal bands were present in JF of one further patient. Additional investigations supported the interpretation that these components were free gamma chains. Oligoclonal anti‐B. burgdorferi IgG antibodies occurred in JF as well as in serum (6 cases) or in JF only (I case). There were complete identities, as defined by isoelectric points and light chain types, between JF and serum antibodies in an individual patient while identities between oligoclonal bands of total IgG and anti‐B. burgdorferi IgG were infrequent. Antibodies of IgG kappa identity were present in nil seven patients and six of them also had IgG lambda antibodies. The antibody response was, therefore, by definition oligo‐and not monoclonal in 6 out of 7 patients. Furthermore, the anti‐B. burgdorferi antibodies of IgG kappa identity only that were present in one case did not exhibit any completely characteristic monoclonal microheterogcneily.
Murine Intestinal Humoral Responses in Chronic Schistosoma mansoni Infections
Authors: CRABTREE J. E. , PULLAR C. E. , TREJDOSIEWICZ L. K. , WILSON R. A.
Published: June 2006

Specific and non‐specific production of immunoglobulins (Ig) by the intestinal mucosa was examined in mice infected with the human blood fluke Schistosoma mansoni. Ileal and colonic mucosal tissue samples were cultured for 2 days, the medium replaced and the culture continued for a further 2 days. Ig concentrations and specific antibodies to soluble schistosome egg antigens in culture supernatants were estimated by isotype‐specific ELISA. Cultured mucosae from control mice produced little IgG, but significant amounts of IgA and IgM on prolonged culture, IgG concentrations were increased in infected animals, mainly in the initial culture period, indicative of systemic, rather than local origins. By contrast, significantly increased local production of IgA and IgM occurred after the start of egg deposition in the intestinal mucosae. Although specific anti‐egg antibodies of the IgG and IgM class were detected, none of the local IgA response was specific for schistosome eggs. We conclude that specific intestinal immune responses to schistosome eggs reflect systemic responses, whereas locally increased IgA production is largely non‐specific. This pattern of response is likely to be related to the prior systemic exposure to schistosome eggs, which results in polyclonal local B‐cell activation, but fails to trigger an antigen‐specific IgA mucosal response.
Accessory Function of Endothelial Cells in Anti‐CD3‐Induced T‐Cell Proliferation: Synergism with Monocytes
Authors: WESTPHAL J. R. , TAX W. J. M. , WILLEMS H. W. , KOENE R. A. P. , RUITER D. J. , DE WAAL R. M. W.
Published: June 2006

Monoclonal antibodies to CD3 can induce proliferation of resting T cells. In vitro this effect is dependent on the presence of monocytes. They serve as accessory cells providing a co‐stimulatory signal after cross‐linking of the antibody‐coated TcR/CD3 complex by the Fc receptor on the monocytes. We have studied whether endothelial cells can replace monocytes with regard to (his function. Highly purified T‐cell preparations were cultured in the presence of anti‐CD3 antibody, purified monocytes, and human umbilical vein endothelial cells. Anti‐CD3 and endothelial cells alone were unable in support T ‐cell proliferation, due to lack of FcR expression. Addition, however, of as few as 10110 FcR+ monocytes (0.8% of the number of T cells present) to a co‐culture of T cells and endothelial cells (EC) in the presence of soluble anti‐CD3 resulted in a strong proliferation of T cells. When anti‐CD3 was presented in an immobilized form (coated to the culture well or to Sepharose beads), or when phytohaemagglutinin was added to the culture as a cross‐linking agent, EC could support T‐cell proliferation in the absence of any monocytes. We conclude that EC by themselves cannot support the proliferation of pure T cells induced by soluble anti‐CD3, but are potent generators of the co‐stimulatory signal(s). They provide a suitable starling material to further define this co‐stimulatory activity.
Phenotypical and Functional Characterization of Small Intestinal TcRγδ+ T Cells in Coeliac Disease
Authors: RUST C. , KOOY Y. , PEÑA S. , MEARIN M.‐L. , KLUIN P. , KONING F.
Published: June 2006

Increased numbers of TcRγδ+ T cells are present in the small intestinal epithelium of patients with coeliac disease (CoD). Their function, however, is unknown. In order lo facilitate detailed functional studies, intestinal γα T cells have been isolated from small intestinal biopsies of patients with CoD (n= 18) and controls (n= 14). As expected, increased numbers of Vδ1+ TcRγδ+ T cells were detected in freshly isolated intraepithelial cell suspensions (IEL) from CoD patients. Also, in the in vitro expanded IEL T‐cell populations from CoD patients the numbers of Vδ1+ TcRγδ+ T cells were increased compared with similar cell cultures from control patients. From IEL cultures derived from six CoD patients, 107 T‐cell clones were generated by limiting dilution and analysed. Sixty of these clones were either CD4 or CD8 positive TcRαβ+ clones. The remaining 47 clones expressed the TcRγδ. Further phenotypical analysis of the γδ T‐cell clones indicated that the TcRγδ+ T‐cell population in the small intestinal epithelium of CoD patients is heterogeneous: four TcRγδ phenotypes could be detected and, although the majority of the TcRγδ+ T cells were CD4‐CD8‐, γδ T‐cell clones expressing either a CD8αα homodimer, a CD8αβ+ heterodimer or CD4 were also identified. In contrast to the TCRαβ+ IEL, most TcRγδ+ IEL were CD5 negative. Furthermore, biochemical analysis indicated that the increase in Vδ1+γδ T cells in the small intestinal epithelium of CoD patients was not the result of a monoclonal expansion. The small intestinal epithelium‐derived γδ T‐cell clones were functional in vitro since the majority of these clones were able to lyse target cell lines such as K562, Molt4 and Daudi. These novel findings therefore indicate that the γδ T cells in the small intestine of CoD patients represent a heterogeneous population and that such cells are functional in vitro. The isolation and the in vitro propagation and cloning of these cells may open new avenues for the study of the putative immune mechanisms leading lo coeliac disease.
The Implications of the Failure to Generate Autoantibody‐Producing Hybridomas from Rat Erythrocyte‐Immunized Mice
Authors: ELLIOTT J. I. , COOKE A.
Published: June 2006

Injection of mice with rat erythrocytes (RRBC) has long been thought to provide an experimental model in which suppressor T cells (Ts) control autoimmunity. The basis of this is that whilst mice immunized with RRBC produce an antibody response, of which a proportion cross‐reacts with autologous red cells, the RRBC‐immunized recipients of RRBC‐primed spleen cells make no, or little, autoantibody, and secondly because the transfer of this autoantibody‐specific suppression can be abrogated by T‐cell depletion of transferred spleen cells. Here an alternative explanation of these phenomena is described.
Adjuvant Composition Determines the Induction of Type II Collagen‐Induced Arthritis
Authors: ELLIS J. S. , CHAIN B. M. , COOKE A. , IBRAHIM M. A. , KATZ D. R.
Published: June 2006

In this sludy we have investigated the influence of adjuvant composition on the development of collagen‐induced arthritis and of anti‐collagen type II specific B‐ and T‐cell responses rollowing immunization with type II collagen. DBA/1 mice immunized with bovine collagen type II emulsified in complete Freund's adjuvant (CFA) containingMycobacterium tuberculosisstrain H37Ra developed footpad swelling indicative of arthritis. Animals immunized with collagen type 11 plus CIA containingMycobacterium butyricumor incomplete Freund's adjuvant showed no significant increase in footpad width. Induction of anti‐CII specific T‐cell proliferation was also dependent upon immunization with CM plus CFA containing M. tb H37RA. In contrast, ovalbumin‐reactive T‐cell proliferation was unaffected by the species of mycobacteria, indicating that the difference in adjuvant activity of the mycobacterial species is specific for anti‐collagen type II T‐cell responses. Antibody response to collagen type II, unlike T‐cell responses, was not significantly different using the two adjuvants. This study therefore demonstrates that murine collagen‐induced arthritis requires immunization with collagen type II together with complete Freund's adjuvant containingMycobaterium tuberculosisH37RA. Since only this combination of antigen and adjuvant induces detectable arthritis and T‐cell responses against collagen type II, while antibody synthesis does not have such stringent adjuvant requirements, this suggests that the development of the full pattern of the collagen‐induced arthritis disease requires synergistic activation of both humoral and cell‐mediated responses.
Nearly Identical T‐Cell Receptor V‐Gene Usage at Birth in Two Cohorts of Distinctly Different Ethnic Origin: Influence of Environment in the Final Maturation in the Adult
Authors: RAMAKRISHNAN N. S. , GRUNEWALD J. , JANSON C. H. , WIGZELL H.
Published: June 2006

We have previously analysed the T‐cell receptor (TCR) V‐gene usage in peripheral blood T lymphocytes from a group of healthy Scandinavians, and described a biased representation (i.e. a statistically significant higher median representation) for some of the TCR V genes towards the CD4+ subpopulation. In a subsequent study the usage of the same V genes was analysed in single positive (CD4+CD8− and CD4−CD8+) human thymocytes, and a similar type of skewness was noted. These observations might be explained by an influence of the specificity of the TCR of thymocytes on the maturation into the CD4+ or the CD8+ lineage. Such a model would assume an interaction between a common determinant on the major histocompatibility complex (MHC) class I or class II molecules, or with a peptide that is preferentially presented by either of the two molecules, and the TCR on the maturing thymocyte. To investigate the possible influence of a different genetic background and environment on skewed TCR V‐gene representation, we have in this study analysed the TCR V‐gene usage in peripheral blood and umbilical cord blood lymphocytes obtained from Asians, with a different ethnic and environmental background from our previous Scandinavian subjects. In the umbilical cord blood lymphocytes the TCR V‐gene usage was close to identical between the two different ethnic groups in both CD4+ and CD8+ subpopulations. Analysing the peripheral blood lymphocyte (PBL) TCR V‐gene usage, we found that three of the four monoclonal antibodies (MoAb) with a biased reactivity towards the CD4+ subpopulation in the Scandinavian group also showed a similar skewed reactivity in this study. Thus, the majority of the TCR V genes were used in a similar way. Some minor but definite disrepancies could be detected when comparing ICR V‐gene usage in adult individuals from these two different ethnic groups. These differences could be inferred to be due to selective peripheral expansion through environmental pressure of T cells utilizing a specific Vβ gene segment.

We conclude that a striking preservation of biased TCR V‐gene usage does exist in humans of distinctly different ethnic origin.
Recombinant Vaccine Vector‐Induced Protection of Athymic, Nude Mice from Influenza A Virus Infection. Analysis of Protective Mechanisms
Authors: KARUPIAH G. , RAMSAY A. J. , RAMSHAW I. A. , BLANDEN R. V.
Published: June 2006

Athymic, nude mice, which normally succumb to virus infection, can resolve infection with recombinant vaccinia virus (rVV) engineered to express IL‐2. We have demonstrated that interferon‐γ (IFN‐γ) produced by natural killer (NK) cells and other immunocytes in response to the virus‐encoded interleukin‐2 (IL‐2) is crucial to recovery. Here, we extend this work to show that nude mice, when primed intravenously with rVV co‐expressing both IL‐2 and an influenza virus haemagglutinin (HA) gene, are also protected following challenge with a lethal dose of homologous influenza virus. A substantial increase in the number of influenza virus‐reactive antibody‐secreting cells producing antibody of the IgM isotype, but not of the IgG or IgA isotypes, was found in spleens and lungs of the protected mice. Treatment with monoclonal antibodies to IFN‐γ or to the NK marker, as GM1, at challenge and thereafter, led to their death however, though the specific IgM antibody response was unaffected. These data suggest that both specific antibody and non‐specific antiviral reactivity are important elements of the protective response and show that this immunization strategy may be used to protect severely immunocompromised individuals.
Over‐Estimation of the Number of Complement Receptor Type 1 (CR1) on Erythrocytes
Authors: QUADRI R. A. , SCHIFFERLI J. A.
Published: June 2006

Quadri RA, Schifferli JA. Over‐Estimation of the Number of Complement Receptor Type I (CR1) on Erythrocytes. Scand J Immunol 1992;36:125‐30

The number of CR1 on erythrocytes, as measured by monoclonal antibodies, remains undefined because of the repetitive structure of CR1 and the presence of different types of structural CR1 alleles. We studied the number of CR1 per erythrocyte using two monoclonal antibodies. E11 and 3D9, which recognize different sites on CR1. The number of binding sites was higher for E11 than for 3D9 (ratio E11/3D9: 1.9 ± 0.4, n=17); however, this ratio was not affected by CR1 numbers or alleles. Partial digestion with papain of CR1 on erythrocytes abolished the binding of 3D9. It reduced the binding of E11 to one‐third of its initial value (0.35 ± 0.03; n= 13) using cells with different CR1 numbers or alleles. By immunoblotting, a unique 75‐kDa stump of CR1 remained attached to the erythrocytes for every allele studied. Taken together, these results suggest that the number of CR1 has been over‐estimated in the past using E11 by a factor of 2, or more probably 3. However, the over‐estimation of CR1 number has been identical for erythrocytes bearing different CR1 numbers or alleles.
Antibodies against Secreted and Non‐Secreted Antigens in Mice after Infection with Live Mycobacterium tuberculosis
Authors: VERBON A. , KUIJPER S. , JANSEN H. M. , SPEELMAN P. , KOLK A. H. J.
Published: June 2006

Mice from four different inbred strains were infected with live Mycobacterium tuberculosis and the immune response to M. tuberculosis was followed for 24 weeks, using Western blotting. Nearly all mice, irrespective of H‐2 type, reacted with the 3S‐kDa protein band. Antibodies against this secreted 38‐kDa protein were ihe first to appear, 4 weeks after infection. Thereafter the secreted 19‐kDa protein and non‐secreted antigens, such as the 65‐kDa and 33‐kDa proteins, were recognized. The immune response against the non‐secreted antigens was influenced by the mouse strain. However, the 33‐kDa protein band was recognized by all mouse strains after a second injection wilh live M. tuberculosis.

The specificity of the antibodies was analysed in Western blot using sonicates of M. tuberculosis, M. kansasii, M avium. M. terrae, M. gordonae and Escherichia coli. Antibodies against the 38‐kDa and 33‐kDa protein bands seemed to be specific for M. tuberculosis, while antibodies against the 19‐kDa protein band showed limited cross‐reactivity. Antibodies against the 65‐kDa prolein were strongly cross‐reactive.

These results suggest that the 38‐kDa protein is secreted in vivo and, therefore, may be available to the humoral immune system at an early stage of infection. The non‐secreted 33‐kDa protein is only recognized by all mouse strains afler prolonged contact with M, tuberculosis.
Expression of CD11b (Leu15) Antigen on CD3+, CD8+, CD16+ Peripheral Lymphocytes, Estimation of CD3+8+11b+ and CD3+4‐8‐11b‐ T‐Cell Subsets using a Single Laser Flow Cytometer
Authors: GANE P. , FAIN O. , MANSOUR I. , ROQUIN H. , ROUGER P.
Published: June 2006

CD11b (Leu15) epitope is expressed on 20‐30% of peripheral blood lymphocytes, including CD16+ large granular lytnphocytes and CD8+ cells. This study confirms that 30% of CD8+ lymphocytes and virtually all CD16+ NK cells from healthy subjects express this determinant. In parallel, our data show that various proportions of CD3+ 4‐8‐, TCR‐δ cytotoxic T lymphoeytes and occasionally CD4+ lymphocytes subsets could also express this epitope.

The CD8+ 11b+ phenotype is associated with suppression of T‐cell proliferative response and has been extensively used to characterize suppressor T lymphocytes. Since about 25% of CD8 lymphocytes are non‐T (CD3) and express the CD16 NK antigen (CD8+ 16+ 3 ‐), the expression of CD11b was also studied on CD8+ 3+ T‐cell and CD8+ 16+ NK‐cell subsets. To this end, we developed three methods using a flow cytometer equipped with a single laser and two fluorescence detectors. Results showed that T CD8+ 3+ 11b+ and NK CD8+ 16+ 11b+ lymphocytes account for 3O% and 70% of CD8+ 11b+ cells respectively. Consequently, the CD8+ 3+ 11b+ phenotype would be more specific for suppressor T lymphocytes than the total CD8+ 11b+ phenotype which includes high proportions of CD16+ NK cells.
The Effects of Deoxyspergualin on the Development of Diabetes in Diabetes‐Prone BB Rats
Authors: NICOLETTI F. , MERONI P. L. , DIMARCO R. , GRASSO S. , BARCELLINI W. , BORGHI M. O. , LUNETTA M. , MUGHINl L. , MENTA R. , SCHORLEMMER H. U. , ZANUSSI C.
Published: June 2006

The effects of the administration of the recently discovered immunosuppressant 15‐Deoxysper gualin (DSP) on the development of insulin‐dependent diabetes mellitus (IDDM) in diabetes prone BB rats were studied. The data show that 2 mg/kg body weight DSP, administered six times a week from the 30th day up to the 105th day of age, significantly reduced the incidence of diabetes in diabetes‐prone BB rats as compared with the PBS‐injectcd controls. The drug was also able to reduce the signs of pancreatic insulitis and the percentages of W3/25+ and OX6+ splenocytes. Interruption of the treatment resulted in a later onset of diabetes in a high percentage of animals within 41 days. These findings suggest that 15‐DSP may temporarily reverse the pathogenic mechanisms leading to beta cell destruction and autoimmune diabetes in a well‐known experimental model of human insulin‐dependent diabetes mellitus.
Activation of HLA‐DR and Interleukin‐6 Gene Transcription in Resting T Cells via the CD2 Molecule: Relevance to Chronic Immune‐Mediated Inflammation
Authors: LINARDOPOULOS S. , CORRIGALL V. , PANAYI G. S.
Published: June 2006

Only a minority of T cells at cell‐mediated immune lesions are antigen specific. In the lesions of human autoimmune disease, such as the synovial membrane in rheumatoid arthritis, the T cells are activated as shown by a variety of phenotypic and functional changes including the expression of HLA‐DR and the production of interleukin‐6 (IL‐6). The stimulatory pathway involved is unknown but does not seem to involve the T‐cell receptor. Alternative pathways of activation which may be involved include the CD2 molecule. It is shown that the formation of sheep red blood cell (SRBC) rosettes with resting T cells from human peripheral blood, which is equivalent to CD2/LFA‐3 binding, leads to the de novo transcription ofthe HLA‐DR and IL‐6 genes and the expression of HLA‐DR on the surface of the T cells. There was no transcription of the interleukin‐2 (IL‐2) or the interleukin‐2 receptor (IL‐2R) genes and Tac expression was not seen. The rosetted T cells did not proliferate. These are all characteristics of T cells at chronic inflammatory sites. It is concluded that receptor‐ligand interactions between CD2/LFA‐3, which are expressed in increased amounts in the rheumatoid Joint, may be one pathway by which antigen non‐specific T cells are recruited as effector cells in lesions of human autoitnmune disease.
Immune Events in Skin. I. Spontaneous Cluster Formation of Dendritic (Veiled) Cells and Lymphocytes from Skin Lymph
Authors: GALKOWSKA H. , OLSZEWSKI W. L.
Published: June 2006

To investigate the mechanism of spontaneous attachment of afferent lymph lymphocytes to dendritic cells, cells from canine skin lymph were used. There were 3.3 ± 2.8% of veiled cells in clusters found in lymph flowing from the cannulated lymph vessel. The number of clusters forming ex vivo in the collected lymph samples increased as a function of time and was temperature dependent. Incubation of cells with proteolytic enzymes or monosaccharides did not alter cell interactions. The ability of veiled cells to bind lymphocytes was independent of divalent cations but reduced by xylocaine and retinoic acid. Among steroids only methylprednisolone showed an inhibitory effect on cluster formation, Indomethacin and acetylsalicylic acid had no blocking activity on cell binding. Also, no effect was seen after treatment with cycosporine A and azathioprine. An enhanced cluster formation after desialation with neuraminidase was observed The desialated cells were cultured in order to study their stimulatory and accessory cell functions. No enhancement of autologous mixed leucocyte reaction was seen, but a significantly higher responsiveness to a suboptimal dose of phytohaemagglutinin was observed. The N‐ase‐mediated non‐specific cell attachment could be abrogated by cell washing or treatment with EDTA or xylocaine.

This study indicates that cluster formation by skin lymph veiled cells and lymphocytes is a spontaneous process which cannot be controlled by means usually effective in regulating the in vitro induced clustering of antigen‐stimulated cells.
Enhancement of the Antibody Response to Protein Antigens by Specific IgG under Different Experimental Conditions
Authors: WIERSMA E. J.
Published: June 2006

Specific stimulation of the antibody response to protein iintigcns by IgG antibodies was studied in vivo‐The response to TNP‐coupled keyhole limpet haemocyanin (KLH‐TNP) was enhanced by a TNP‐spcciftc IgG monoclonal antibody, 7B4, as measured both in ELISA (enzyme‐linked immunosorbent assay) and by ELISPOT (enzyme‐linked immunospot) method. Enhancement was seen when using both high and low doses of antigen and antibody. The antibody response to TNP‐coupled bovine serum albumin (BSA‐TNP), bul not to TNP‐couplcd ovalbumin (OA‐TNP), tetanus toxoid (TT‐TNP) or diphtheria toxoid (DT‐TNP). was also etficiently augmented by 7B4, Enhancement was seen when using both high and low coupling ratio of TNP to KLH, The fact that IgG‐mediiited enhancement is seen under many ditVerent experimental conditions suggests that it is a physiologically relevant phenomenon. In order lo enhance the response to KLH‐TNP and BSA‐TNP, 7B4 had lo be injected while antigen was circulating in blood. This supports that igG‐mediated stimulation acts by concentrating circulating antigen into lymphoid centres.
Composition and Immunological Properties of the Protein Fraction of A36, a Major Antigen Complex of Mycobacterium paratuberculosis
Authors: DE KESEL M. , GILOT P. , COENE M. , COCITO C.
Published: June 2006

TMA (thermostable macromolecular antigens) are major mycobactcrial complexes present in all mycobticteria. We have purified A36, the TMA complex of M. paratuberculosis, the etiological agent of paratuberculosis (Johne's disease), and shown by the immune electron microscopy approach its presentation at the cell surface. The immunodominance of the A36 complex in Johne's disease was suggested by comparative ELISA analysis of infected bovine sera, using either A36 or M. paratuberculosis total soluble sonicate as antigens. The cross‐reactivity of TMA complexes from diifcrcnt tnycobacteria was evaluated by immunocn/ymometri c measurements. Percentage of shared epifopes was high tor the couple M. paratuberculosis‐ M. avium. and somewhat lower for the couple M. paratuberculosis and M. bovis Immunological kinship between M. paratuberculosis and M. leprae was suggested by the finding that oul of eleven anti‐M. lepraemonoclonals, four cross‐reacted with A36 proteins. The specificiiy missing al the level of the whole A36 complex was sought at the level of its protein components. Comparative immunoblot analysis of electrophoresed A36 proteins indicated three of them to contain epitopes not shared by M. bovis proteins, and one of them to contain epitopes specific wilh respect lo M. avium, M. bovis and M. phlei. The latter component, a 34‐kDa protein, could be an ideal reagent for a serological tesi for, Johne's disease, being immunodominant in infected cattle and endowed with species‐specific epitopes.
Involvement ofthe Avian μ Heavy Chain in Recolonization ofthe Bursa of Fabricius
Authors: PALOJOKI E. , LASSILA O. , JALKANEN S. , TOIVANEN P.
Published: June 2006

In Ihe chicken, the B eells develop in a specialized organ, the bursa of Fabricius. Earlier it was shown that neonatal bursal cells treated with polyclonal anti‐chicken immunoglobuiin antibodies are not able to recolonize tho bursa when transferred into cyclophosphamide‐treated chicks. In this study, 4‐day‐old bursal eells were treated with different polyclonal and monoclonal antiimmunoglobulin antibodies and transferred into 4‐day‐old eyelophosphamide‐treated chickens. Two monoclonal anti‐chicken IgM antibodies. CVI‐59.7 and 21‐2B2. recognizing distinct epitopes of the μ heavy chain, were inhibitory. Incubation of cells with 21‐2B2 antibody caused about 9O% inhibition of bursal recolonization. After incubation with CVI‐59.7 antibody ihe inhibition was 50%, The high inhibition by 21‐2B2 antibody was also seen when F(ab')2 fragments of the antibody were used. These results suggest that the entry of the cells needed for bursal reeolonization is inhibited almost totally by 21‐2B2 antibody, or that this antibody blocks further proliferation of the cells in bursal follicles. In eonelusion, we have shown that a μ heavy chain epitope is intimately involved in the recolonization of bursal follicles, and distinct epitopes of the (I heavy chain are not equally important in this proeess.
Hydrocortisone Treatment of BCG‐Infected Mice Impairs the Activation and Enhancement of Antimicrobial Activity of Peritoneal Macrophages
Authors: STOKVIS H. , LANGERMANS J. A. M. , DE BACKER‐VLEDDER E. , VAN DER HULST M. E. B. , VAN FURTH R.
Published: June 2006

The present study concerns the effect of hydrocortisone (HC) on the effector functions of Bacillus Calmette Guerin‐purified protein derivative (BCG‐PPD)‐activated macrophages. Such activated macrophages release greater amounts of H2O2 and NOi”, inhibit the intracellular proliferation of T. gondii and kill L, monocytogenes more efficiently than resident macrophages. This activation was not fully expressed by macrophages from BCG‐activated mice that had received a subcutaneous injection of HC 2 days before intraperitoneal injection of PPD, since the inhibition of the intracellular proliferation of T. gondii, the release of NO2’and the rate of intracellular killing of L. monocytogenes were lower than in macrophages from BCG‐PPD‐activated mice. However, treatment with HC did not impair the release of H2O2 by BCG‐PPD‐activated macrophages. The results show that the treatment of infected mice with HC inhibits their ability to develop adequate intracellular microbicidal mechanisms.
Analysis of HLA Genotypes and Susceptibility to Insulin‐Dependent Diabetes Mellitus: HLA‐DQα Complements HLA‐DQβ
Authors: BAISCH J. M. , O'BRIEN M. E. , HOOVER M. L. , CAPRA J. D.
Published: June 2006

It is well known that certain genes in the HLA‐D region confer increased susceptibility to insulin‐dependent diabetes mellitus (IDDM), Previous studies have documented an increased risk associated with the HLA‐DRβ chain alleles, DR3 and DR4, and the DQβ chain allele DQB1*0302 (formerly DQw8). Since DQα is also polymorphic and has been strongly implicated as the primary IDDM susceptibility locus in other races, we wanted to assess the contribution of DQα to IDDM in Caucasians, This information would enable us to define more precisely the class II association with IDDM as well as gain insight into issues of cu versus trans association of DQ heterodimers in this disease. To this end, the DQa genotype was determined for a large group of diabetic and normal Caucasian individuals who had been HLA‐DQβ and HLA‐DR typed previously. Using the polymerase chain reaction and a set of twelve oligonucleotide probes, we determined the DQα genotype of 323 patients with IDDM and 182 normal subjects.

We found that certain DQα alleles are decreased in the diabetic population compared with normal subjects (i.e. DQA1*102 and 0103), while others are significantly increased in patients with IDDM (i.e. DQA 1*0301 and *0501), In addition, certain combinations of DQα alleles are associated with increased susceptibility to disease (i.e. DQA 1*0301, *0501), These results parallel our findings at the DQβ locus; however, because of the various associations between DQα and DQβ chains, the risks conferred by DQα are generally lower than those at DQβ, Moreover, our data indicate that, in Caucasians, no single DQα allele accounts for the highest degree of susceptibility to IDD M as in other races, although DQα analysis may be informative in a few cases. When done in combination, however, oligonucleotide analyses at both DQβ and DQα complement each other and provide a more complete assessment of the HLA‐associated component of disease susceptibility in IDDM
Human Blood Dendritic Cells Exhibit a Distinct T‐Cell‐Stimulating Mechanism and Differentiation Pattern
Authors: XU H. , FRIEDRICHS U. , GIESELER R. K. H. , RUPPERT J. , OCKLIND G. , PETERS J. H.
Published: June 2006

In this study, the mechanisms underlying stimulation of T‐cell proliferation by human blood dendritic cells (BDC) and their differentiation have been defined with a panel of monoclonal antibodies (MoAbs)‐ It was found that the MoAbs against LFA‐I (CDlla), CD11c. LFA‐3 (CD58), ICAM‐1 (CD54) or HLA‐DR could significantly suppress T‐cell proliferation in an allogeneic mixed lymphocyte reaction (P<0.05), while being unable to inhibit clustering of BDC with T cells. Addition of anti‐CD18 or CD45 MoAbs increased the size of clusters after 18 h of culture, but had no effect on the proliferation of T cells (P<0.05). The suppressive effect of the MoAbs may be viewed not as an inhibition of contact between BDC and T cells, but rather as a blocking of co‐stimulatory signals for T‐cell activation, which are mediated by interaction of the adhesion molecules. After depleting the BDC preparations of monocytes. we used a double staining in FACS analysis to demonstrate that BDC do not express specific T (CD3), B (CD20 and CD2l)and myeloid cell markers (CDl1b, CDl3 and CD14), but abundant class II antigens. This pattern remained unaltered after 8 days of culture in the presence of 100 U/ml GM‐CSF, although a threefold increase of HLA‐DQ and ICAM‐1 molecules on the cultured cells was observed.