bluuebunny/crossref_metadata_2025 huggingface dataset:
Biochemical Purification of Various Receptor Molecules Involved in Human T Lymphocyte Activation
Authors: LECA G. , BENICHOU G. , BENSUSSAN A. , KANELLOPOULOS J.
Published: June 2006

The now classical major histocompatibility complex (MHC)‐restricted receptor for antigen on human T lymphocytes has been identified as a 90 kDa disulphide‐linked heterodimer composed of two glycoproteins termed α and β. More recently, another type of T cell receptor for antigen has been described. which seems to mediate killing of target cells without any obvious requirement for MHC recognition. This T cell receptor for antigen is also a heterodimer composed of γ, λ chains non‐covalently associated with the three mon morphic CD3 subunits. Another disulphide‐linked dimer capable of triggering T lymphocytes has been defined recently by a monoclonal antibody: the anti‐human 9.3 antigen. In order to generate monoclonal or polyclonal reagents against variable and constant regions of the T cell receptor chains and against new epitopes of the 9.3 antigen, we have developed a biochemical method of purification of T lymphocyte disulphide‐linked dimers. Our method relies on two biochemical properties of the 9.3 surface molecule and the T cell receptor for antigen. (1) They are disulphide‐linked dimers and thus can be separated from the vast majority of the cell surface molecules by two‐dimensional (non‐reduced versus reduced) sodium dodecyl sulphate–polyacrylamide gel electrophoresis (SDS–PAGE). (2) T cell receptor chains are less hydrophobic than the 9.3 antigen, and thus can be isolated from it on reverse‐phase high‐performance liquid chromatography (HPLC) at a lower concentration of acetonitrile. Microsomal preparations from T cell clones and leukaemia lines were prepared by nitrocavitation and lysed in sodium deoxycholate. After concentration, this lysate was electrophoresed on SDS–PAGE in non‐reducing conditions. The gel slice corresponding to the molecular weight of the T cell receptor was cut out and run in reducing conditions in the second dimension. The T cell receptor spots were easily located on the gel by autoradiography as the microsomal lysate had been mixed with iodinated glycoproteins. The T cell receptor was eluted from the gel with about 85% yield. At this stage, the T cell receptor preparations also contained the 9.3 antigen, another disulphide‐linked dimer. The separation of this antigen from the T cell receptor chains had been achieved on reverse‐phase HPLC. This procedure allows the purification and separation of two disulphide‐linked dimers which are both involved in T cell activation. The obtention of antibodies against new epitopes of these important molecules would be extremely useful for analysing their role in T cell function and ontogeny.
Inhibitor of Interleukin 1 in Normal Human Urine
Authors: SVENSON M. , BENDTZEN K.
Published: June 2006

We investigated whether urine from normal individuals contains inhibitors of interleukin 1 (IL‐1). Diafiltered urine from normal afebrile donors suppressed IL‐1‐induced interleukin 2 (IL‐2) activity of mouse thymocyte supernatants. These supernatants, however, strongly suppressed IL‐2‐induced [3H] thymidine incorporation into the IL‐2‐sensitive cell line CTLL.‐2, whereas the urine preparations did not. This phenomenon was caused by an increased amount of thymidine secreted by the urine‐treated thymocytes. Therefore, in order to prevent interference, experiments were carried out with excess [3H]thymidine. Under these circumstances, suppression of IL‐1‐ and to a lesser extent IL‐2‐induced DNA synthesis was still observed, whereas the synergistic effect of IL‐1 on IL‐2‐induced DNA synthesis was only marginally reduced. We conclude that suppression of IL‐1‐induced IL‐2 production by mouse thymocytes is a major effect of the IL‐1‐inhibitory factor(s)in normal urine. When the murine EL 4 cell line was used, the diafiltered urine failed to inhibit IL‐1‐induced IL‐2 production. The detection of an IL‐1 inhibitor in urine is therefore dependent on the target cells as well as the effects of IL‐1 on these cells.
T Cell‐Mediated Production of Tumour Necrosis Factor‐α by Monocytes
Authors: DEBETS J. M. H. , VAN DER LINDEN C. J. , SPRONKEN I. E. M. , BUURMAN W. A.
Published: June 2006

The aim of this study was to examine the tumour necrosis Factor (TNF)‐α production by peripheral blood mononuclear cells activated by mitogens. Considerable amounts of TNF‐α, ranging from 1.0 to 5.0 ng/ml, were present in the supernatants of cultures of human peripheral blood mononuclear cells (PBMC), stimulated with either the anti‐CD3 monoclonal antibodt OKT3 or the lectin phytohaemagglutinin (PHA). The amount of TNF‐α secreted in the supernatant was closely correlated to the degree of T cell proliferation in such cultures, as measured by [3H]TdR incorporation. In the absence of proliferating T cells the mitogens did not induce secretion of TNF‐α by monocytes. Supernatant of proliferating T cells were shown to induce TNF‐α production by monocytes. The macrophage‐activating factor gamma interferon (IFN‐γ) was also shown to induce, in the absence of endotoxin, TNF‐α secretion by monocytes in a dose‐dependent manner. The induction of TNF‐α production by supernatants of proliferating T cells could largely be abrogated by passaging the supernatants on an anti‐IFN‐γ column before adding them to the monocytes. It is therefore concluded from this study that the production of TNF‐α by monocytes can be induced by proliferating T cells and that this induction can largely be attributed to the T cell lymphokine IFN‐γ.
Regulation of Experimental Allergic Thyroiditis
Authors: EISHI* Y. , McCULLAGH P.
Published: June 2006

The susceptibility to experimental allergic thryoiditis (EAT) of rats that had previously been immunized against thyroglobulin was examined. Challenge with thryoglobulin 5 weeks previously conferred strong resistance to the induction of EAT after exposure to a sensitization protocol that was otherwise highly effective. Resistance to induction of EAT could also be conferred by the inoculation of spleen cells from thryoglobulin‐sensitized donors on the day of birth. Heavily irradiated spleen cells from sensitized donors were able to confer resistance on neonatal recipients, indicating that prolonged survival of the transferred cells was not required.
Cytolysis by Cloned Helper T Cells: Induction by Specific Antigen or by Anti‐CD3 Hybrid Antibodies
Authors: VYAKARNAM A. , STRANGEWAYS A. L. , GLOVER R. E. , LACHMANN P. J.
Published: June 2006

We have explored the use of hybrid antibodies—prepared by covalently linking anti‐CD3 to an antibody Specific for a monomorphic major histocompatibility complex (MHC) class II determinant using N‐succinimudyl 3‐(2‐pyridyldithio)proprionate/succinimidyl 4‐(N‐malcimidomethyl)cyclohexane‐1‐carhoxylate (SPDP/SMCC) as coupling reagent—in inducing cytolysis in human tuberculin (PPD)‐specific T helper (TH) clones, These clones have been shown to lyse PPD‐bound Epstein‐Barr virus (EBV)‐transformed B‐cell lines (B‐EBV) in an MHC class II‐restricted manner. In this paper anti‐CD4‐induced cytolysis is compared with antigen/MHC‐induced cytolysis with the same clones. Cytolysis induced by the hybrid antibodies was highly efficient, with killing of both syngeneic and allogeneic tumour cells positive for MHC class II. Conjugate‐induced cytolysis was maximal within 4 h; that of antigen‐positive targets at 16 h. Kilting of bystander cells was seen only when cytolysis was triggered by antigen/MHC, suggesting that the mechanism of Cytolysis in the two systems may be distinct. Targets treated simultaneously with hybrid antibody and with antigen, thereby providing both activation signals to the clones, are lysed less efficiently than those treated with either PPD or hybrid antibody alone. Evidence is presented showing that this inhibition is most marked against syngeneic PPD‐coated cells treated with hybrid antibody, suggesting that two signals independently capable of activating cytolytic function in the clones, when presented simultaneously, interfere with the induction of the cytolytic process.
Heterogeneity in the Specificities of an Alloreactive T Helper Cell Line
Authors: RAJASEKAR R. , ANDERSSON J. , LEANDERSON T.
Published: June 2006

Heterogeneity in antigen recognition of an alloreactive helper T‐cell line was studied by repeated limiting dilution cloning. Analysis of the line specificities of these clones showed that the gain of reactivity to new antigens by the T cell line included the generation of non‐cross‐reactive T cells specific to new major histocompatibility (MHC) antigens. Analysis of function revealed that all the T‐cell clones studied had similar cell surface antigens and secreted interleukin 4(IL‐4) and IL‐5/IL‐3 but not IL‐2 or gamma interferon (IFN‐γ). The T‐cell clones served as helper cells for B cells expressing the appropriate MHC antigens to both clonal proliferation and differentiation into antibody‐producing cells.
Modulation of Natural Killer Cell Activity in Peripheral Blood by Physical Exercise
Authors: PEDERSEN B. K. , TVEDE N. , HANSEN F. R. , ANDERSEN V. , BENDIX T. , BENDIXEN G. , BENDTZEN K. , GALBO H. , HAAHR P. M. , KLARLUND K. , SYLVEST J. , THOMSEN B. S. , HALKJÆR‐KRISTENSEN J.
Published: June 2006

The present study was designed to examine the effect of physical exercise on human natural killer (NK) cells. Six healthy volunteers underwent two different acute physical exercise tests with an interval of at least 1 week: (1) 60min bicycle exercise at 80% of maximal oxygen uptake (VO2max) and (2) 60 min back‐muscle training at up to 29% of VO2max; blood samples were collected before and during the last few minutes of exercise, as well as 2 h and 24 h afterwards. The NK cell activity (lysis/fixed number of mononuclear cells) increased during bicycle exercise, dropped to a minimum 2 h later and returned to pre‐exercise levels within 24 h. Back‐muscle exercise did not significantly influence NK cell activity. Plasma levels of adrenaline, noradrenaline, and cortisol were elevated during bicycling, but not during back‐muscle exercise, indicating that exercise intensity is a determinant of NK cell activity. During bicycle exercise the NK cell subset (CD 16+ cells) of mononuclear cells increased significantly. Furthermore an improved interleukin 2 (IL‐2) boosting of the NK cell activity was found during work as compared to IFN‐α and indomethacin‐enhanced NK cell activity. These results indicate that NK cells with a high IL‐2 response capacity are recruited to the peripheral blood during exercise. The decreased NK cell activity demonstrated 2 h after work was probably not due to fluctuations in size of the NK cell pool, since the proportion of CD16+ cells was normal. The finding that indomethacin fully restored the suppressed NK cell activity in vitro and the demonstration of a twofold increase in monocyte (CD20+ cells) proportions 2 h after work, strongly indicate that prostaglandins released by monocytes during the heavy physical exercise are responsible for the down‐regulation of the NK cells.
Interleukin 1‐Induced Down‐Regulation of Antibody Binding to CD4 Molecules on Human Lymphocytes
Authors: TVEDE N. , CHRISTENSEN L. D. , ØDUM N. , WIIK A. , BENDTZEN K.
Published: June 2006

Interleukin 1 (IL‐1) is involved in the early activation of T lymphocytes. The CD4 antigen, described as a phenotypic marker of helper T cells, is also important in early T‐cell activation by its ability to hind to MHC class II molecules on antigen‐presenting cells, and to transmit positive (and negative) signals to the cells. We observed that purified human monocyte IL‐1 as well as recombinant IL‐1α and IL‐1β selectively decreased the binding of monoclonal antibodies to CD4 on the surface of otherwise unstimulated blood T cells, in contrast to prestimulated and continuously grown CD4+ cells. Under optimal growth conditions, the initial reduction in antibody binding to CD4 was followed by an apparent re‐expression of the CD4 antigen even in the presence of high concentrations of IL‐1. This re‐expression did not occur if the cells were cultured at 4°C, or after treatment with actinomycin D or cytochalasin B, indicating that protein synthesis and intact microfilament function were essential for re‐expression of CD4 binding. The mechanism by which CD4 molecules are physically and/or functionally modulated by IL‐1 is unclear.
Characterization and Expression of the Human T Cell Receptor‐T3 Complex by Monoclonal Antibody F101.01
Authors: GEISLER C. , PLESNER T. , PALLESEN G. , SKJØDT K. , ØDUM N. , LARSEN J. K.
Published: June 2006

A murine monoclonal antibody (MoAb) F101 01 reacting with the T cell receptor (TCR)‐T3 complex is presented. Immunohistological studies showed that FH101.01 specifically stains T‐zone lymphocytes in lymph nodes, tonsils, and splenic tissue. Two‐colour immunofluorescence and flow cytometry demonstrated co‐expression of the antigen defined by F101.01 and the pan‐T cell antigens defined by CD2, CD3, CD5 and CD7 antibodies. Cells Stained with CD4 and CDS antibodies were both included in the F101.01‐positive population, whereas CD16‐positive natural killer cells (NK). B cells (CD19 and CD20), and myeloid cells (CD13 and CD33) were excluded. The target antigen of F101.01 comodulated with the CD3‐defined antigen (T3) and the TCR recognized by the MoAb WT‐31. CD3 antibody and WT‐31 both blocked binding or F101.01. F101.10 precipitated the TCR‐T3 complex from lysates of 125‐labelled peripheral blood mononuclear cells (PBMC) and HPB‐ALL, when the lysate was prepared with a detergent (digitonin) that conserves the TCR‐T3 complex. FACS analysis of T cells from a patient with a T cell immunodeficiency demonstrated that δ‐TCS‐1‐CD3‐CD4‐ and δ‐TCS‐1‐CD3+CD8+ cells were brightly F101.01+, whereas a large subpopulation of δ‐TCS‐1+CD3+CD4− CD8− cells were weakly F101.01+. We conclude that F101.01 recognizes a conformational epitope of theTCR‐T3 complex and that it reacts with the αβ TCR‐T3 and the γδ TCR‐T3 complexes with different intensities.
Activation of Human T Cells by Neuraminidase‐Galactose Oxidase‐Treated Erythrocytes Involving CD2 (T11) and its Complementary Structure
Authors: OCKLIND G. , TALTS J. , LINDAHL‐KIESSLING K.
Published: June 2006

Human T lymphocyte proliferation induced by neuraminidase‐galactose oxidase (NAGO)‐treated autologous erythrocytes (HENAGO) plus polyethylene glycol (PEG) has previously been shown to be independent of accessory cells. Here, we show that the response to HENAGO+PEG was accompanied by interleukin 2 (IL‐2) release and was inhibited by anti‐IL‐2 and anti‐IL‐2 receptor antibodies. HENAGO alone initiated DNA synthesis together with phorbol ester (12‐O‐tetradecanoyl‐phorbol‐13‐acetate; TPA). To elucidate the nature of the stimulatory signals NAGO‐treated sheep erythrocytes (SENAGO) were used in additional experiments. In parallel to the superior rosetting capacity of SE compared to HE, SENAGO were by themselves stimulatory, and the response was further enhanced by PEG or TPA. Antibody L180/1, specific for the T11 (CD2) target structure (T11TS) on SE, homologous to the human CD2 ligand LFA‐3, abolished the response to SENAGO alone or when combined with PEG or TPA. The results suggest that ENAGO induce T‐cell response through CD2‐LFA‐3‐T11TS interaction, and via other surface antigens bound by the oxidatively induced aldehyde groups on ENAGO.
Biological Characterization of T Cell‐Replacing Factor in the Synovial Fluid of Rheumatoid Arthritis Patients
Authors: MÖLLER E. , STRÖM H.
Published: June 2006

The synovial fluid of patients with rheumatoid arthritis (RA) contains a biologically active factor which has the ability to replace T cells for the induction of antibody secretion by human blood lymphoid cells stimulated by pokeweed mitogen (PWM) in vitro. This factor, which will be referred to as RA‐SF (synovial fluid), also has the capacity to act as a B cell‐stimulatory factor of mouse splenic lymphocytes in the presence of lipopolysaccharide (LPS). Using a test system developed for the definition of interleukin 4 (IL‐4), which is a B cell‐stimulating lymphokine which preferentially activates the synthesis of selected Ig classes in mouse lymphoid cells, we have shown that RA‐SF has properties similar to IL‐4 in that it induces differentiation of antibody secretion in the LPS‐pretreated mouse cell, but unlike IL‐4, which gives IgG1 and IgE, it selectively induces IgG2b synthesis. The present study demonstrates that RA‐SF has a biological activity that is reminiscent of other B cell‐stimulating mouse lymphokines, but it is biologically distinct from IL‐2, IL‐4, and IL‐5. Recent data also indicate that it is distinct from gamma interferon (IFN‐γ). Therefore, we conclude that the biological activity of RA‐SF has properties in common with a T‐cell replacing (TRF) and B‐cell differentiation factor (BCDF) and probably represents yet another biological activity which so far lacks an experimental counterpart. The relevance of this factor for autoantibody synthesis is discussed.
Characterization of Bovine Amyloid Proteins SAA and AA
Authors: HUSEBEKK A. , HUSBY G. , SLETTEN K. , SKOGEN B. , NORDSTOGA K.
Published: June 2006

The bovine serum amyloid A (SAA) and tissue amyloid A (AA) proteins were isolated and characterized. SAA was isolated from acute phase high density lipoprotein (HDL) of a cow suffering from acute mastitis, and was identified by amino acid sequence analysis. No AA‐like protein was found in complex with HDL in serum. Amyloid fibrils isolated from a bovine kidney contained a 9 kDa AA protein and a considerable amount of a 14 kDa protein. Amino acid sequence analysis showed that the largest protein probably represents undegraded SAA. This is an interesting observation which confirms previous works indicating that SAA can be incorporated in the amyloid fibrils without a prior degradation to AA. The partial amino acid sequences of bovine SAA and AA were strikingly homologous to the sequences of corresponding proteins in man and other species.
Leukaemic B Cells from Patients with Chronic Lymphocytic Leukaemia Suppress Immunoglobulin Production by Lymphocytes from Normal Donors
Authors: KUNICKA J. , PLATSOUCAS C. D.
Published: June 2006

We observed that highly purified E‐rosette‐negative largely leukaemic B cells from 9 out of 15 patients with chronic lymphocytic leukaemia (CLL) significantly suppressed immunoglobulin production by mixtures of T4 and B cells from normal donors in the presence of pokeweed mitogen (PWM). This suppression by leukaemic B cells was concentration‐dependent. Addition of equal numbers of B cells from normal donor to the mixtures of normal T4 and B cells increased, or had no effect on the production of IgM, IgA, and IgG Treatment of purified largely leukaemic B cells from patterns with CLL with either the anti‐B1 or anti‐Leu 1 monoclonal antibody plus complement abolished their ability to suppress immunoglobulin production. In contrast, treatment with either the anti‐Leu 5 or the OKMI monoclonal antibody plus complement had no effect on the suppression. These results tugged that leukaemic B cells from certain patients with CLL may exhibit, or can be induced to exhibit, immunosuppressive properties.
Altered Natural Killer Cell Activity in Childhood Acute Non‐lymphoid Leukaemia
Authors: SØRSKAAR D. , FØRRE Ø. , TJØNNELAND S. , LIE S. O.
Published: June 2006

Peripheral blood and bone marrow mononuclear cells from 25 children with acute non‐lymphoid leukaemia were analysed for natural killer cell activity and for cells with the Leu‐7 and Leu‐11b(CD 16) markers. Significantly reduced spontaneous cytotoxicity was detected in peripheral blood from children with untreated and active acute non‐lymphoid leukaemia compared with that of the controls (P=0.01 and P<0.05). Patients to remission, however, had normal natural cytotoxicity and normal number of Leu‐7 and Leu‐11b(CD 16)‐positive cells. The natural killer cell activity in hone marrow from patients with untreated acute non‐lymphoid leukaemia was also significantly reduced (P=0.025). On die other hand, patients in remission had both an increased percentage of Leu‐7 and Leu‐11b (CD 16)‐positive cells (P<0.05) and an increased natural killer cell activity (P<0.0005) in their bone marrow cells in comparison with the control group. This augmented natural killer cell activity is most probably a result of anti‐leukaemic treatment. Stimulation with recombinant alpha interferon and recombinant interleukin 2 caused an increase in natural killer cell activity that was both significant and normal in both peripheral blood and bone marrow from children with acute non‐lymphoid leukaemia.
Characteristics of Soluble Tumour‐Derived Proteins That Inhibit Natural Killer Activity
Authors: HEISKALA M. K. , STENMAN U. H. , KOIVUNEN E. , CARPÉN O. , SAKSELA E. , TIMONEN T.
Published: June 2006

We have previously shown that various benign and malignant natural killer (NK)‐resistant monolayer cells inhibit endogenous human NK activity, probably by reducing the secretion of cytotoxic factors from the effector cells. The nature of she molecules responsible for the inhibition has been unclear. In this study we show that phosphate‐buffered saline (PBS) extracts of ovarian cystadenocarcinoma tissue and normal uterine smooth muscle strongly inhibit NK activity. Fractionation of tumour extracts by gel chromatography revealed major inhibitory activity in the Mr range 160,000–180,000, and other weaker Inhibiting activities in the Mr ranges 50,000–70,000 and 20,000. The active material of Mr range 160,000–180,000 was adsorbed on anion exchange chromatography column at neutral pH and physiologic NaCl concentration, and it was eluted by 0.31–0.34 m NaCl. The inhibitory molecule was sensitive to proteolysis. No relation of this compound to immunoglobulins or trypsin and urokinase inhibitors was detected. The unfractionated extract inhibited NK activity apparently by the same mechanism as the monolayer target cells, i.e. by reducing the secretory capacity of effector cells. The data strongly suggest that the NK‐inhibiting compounds described in this work lire involved in the inactivation of NK cells by intact monolayer cells
Monoclonal Autoantibodies Specific for Kidney Proximal Tubular Brush Border from Mice with Experimentally Induced Chronic Graft‐versus‐Host Disease
Authors: ROLINK A. G. , RADASZKIEWICZ T. , MELCHERS F.
Published: June 2006

Nine hybridomas producing monoclonal autoantibodies specific for kidney proximal tubular brush border were Found in 600 hybridomas derived from (C57BL/6J×DBA/2)F1 mice injected with DBA/2 T cells. None of the 1100 hybridomas derived from nonautoimmane (C57BL/6J×DBA/2)F1 mice produced antibodies with a similar specificity. Four of these nine monoclonal antibodies were characterized further. They did not bind to cryosections of liver, lung, stomach, or intestine. Three bound to kidney proximal tubular brush border of mouse, cattle, sheep, pigs, rabbits, rats, and humans, whereas the fourth was specific only for murine brush border. All four precipitated from mouse kidney microvilli, a protein with an apparent molecular weight of 160,000 under reducing as well as nonreducing conditions. Removal of asparagine‐linked carbohydrate with EndoF reduced the molecular weight of the 160,000 protein by about 20,000. One of the three multi‐species‐specific antibodies bound to pig kidney aminopeptidase, a glycosylated enzyme located on the microvilli of kidney proximal tubular brush border. Three antibodies have a heavy‐chain variable region encoded by VH genes of the J558 family, whereas the heavy‐chain variable region of the fourth is encoded by a VH gene of the 7183 family. Attempts to passively transfer immune complex glomerulonephritis to normal mice by injection of the purified monoclonal antibodies or by growth of the corresponding hybridoma cells in mice have so far been unsuccessful. However, antibodies recognizing the 160,000 molecule are present in the serum of mice with chronic graft‐versus‐host disease and can be eluted from kidneys with immune complex glomerulonephritis.
Dynamics of Cytotoxic T Lymphocyte Precursors in Vivo Assessed by Change in the Radiation Sensitivity
Authors: HASEGAWA Y. , NAKASHIMA I. , ANDO K. , MIZOGUCHI K. , NAGASE F. , ISOBE K. , KAWASHIMA K. , SHIMOKATA K. , YOSHIDA T. , IWAMOTO T.
Published: June 2006

The dynamics of cytotoxic T lymphocyte precursors (CTL‐p) in mice injected with allogeneic spleen cells (SC) was studied with special reference to changes in their radiation sensitivity. Whole‐body 400 rad X‐ray irradiation of allo‐SC‐primed and unprimed mice virtually abolished the capacity of their SC to proliferate and to generate CTL in primary or secondary mixed leucocyte culture (MLC). However, the impaired ability of SC to generate CTL in the primary MLC was restored by interleukin 2 (IL‐2). This showed that helper cells whose activity was replaceable with IL‐2 (IL‐2‐producing cells) were functionally more radiation‐sensitive than CTL‐p in unprimed mice. In contrast, the radiation‐impaired activity in secondary MLC was not restored by IL‐2, suggesting that memory CTL‐p in allo‐SC‐primed mice were unexpectedly sensitive to radiation. The D37 values determined from the percentage of residual CTL‐p activity of SC in bulk cultures 1 day after irradiation were 525 rad for virgin CTL‐p and 75 rad for memory CTL‐p. Further studies demonstrated that the radiation‐sensitive memory CTL‐p were generated from relatively radiation‐resistant precursors, largely independent of radiation‐sensitive IL‐2‐producing cells and of cellular proliferation. The mean frequency of CTL‐p in SC measured by limiting dilution assay was not significantly increased by the priming. This supports our conclusion that the development of the memory CTL‐p activity in allo‐SC‐primed mice did not depend on clonal expansion. Whole‐body 400 rad‐irradiation reduced the frequency of CTL‐p in SC from unprimed‐mice 1/2–1/3 and that in SC from allo‐SC‐primed mice to 1/8–1/15. This supports the view that the majority of radiation‐resistant virgin CTL‐p functionally mature to radiation‐sensitive memory CTL‐p without cellular proliferation in allo‐SC‐primed mice.
Ovalbumin‐Specific Human B‐Cell Activation and Maturation
Authors: HART L. A.'t , ZIJLSTRA J. , HEIJNEN J. J. , BALLIEUX R. E.
Published: June 2006

By means of a panel of monoclonal antibodies it is demonstrated that, in cultures of human peripheral blood mononuclear cells (PBMC) with the T‐cell dependent (TD) antigen ovalbumin (OA), responding B ceils are activated from the resting Mate The differentiation of the activated B cells to high rate‐secreting plasma blasts, however, is attested in an early activation phase, in which they can be detected at low rate‐secreting plaque‐forming cell. The arrest does not occur when stimulation with OA occurs in the presence of antigen‐nonspecific activation and maturation factors, which are provided in the culture by the anamnestic response to the TD antigen tetanus toxoid.
Expression of B220 Antigen on an Interleukin 4‐ and Interleukin 5‐Producing T‐Cell Line
Authors: SANDER B. , SIDERAS P. , MÖLLER E.
Published: June 2006

The marine T‐cell line 2.19, orginally used for cloning of interleukin 4 (IL‐4) and IL‐5, was analysed for surface marker characteristics using monoclonal antibodies and a FACS‐4analyser and found to be positive for the B220 antigen. Thus, this lymphokine‐producing cell with charactistic T‐cell markers expresses an antigen earlier thought to be specific for B cells. With two‐colour fluorescence it was found that a small fraction (2–4%) of Thyl+ spleen cells from normal CBA mice were also positive for B220.
The Vβ8 Gene Family is Preferentially Used by‘Naturally’Activated T Cells
Authors: FORNI L. , COUTINHO A.
Published: June 2006

Receptors encoded by the Vβ8 gene family detected by the monoclonal antibody F23.1 are expressed among ‘naturally’ activated T cells in normal spleen at frequencies significantly higher than in the total CD4+ and CD8+ cell populations. The positive selection of these clones into ‘natural’ T‐cell activity could be the reason for the high frequencies of cells expressing Vβ8 genes. This phenotype is strain‐dependent.
Role of CD8 (Lyt‐2) in Cytotoxic T‐Cell Function
Authors: MUNAKATA T. , SCHMIDT‐ULLRICH R. , BERTSCHMANN M. , EICHMANN K.
Published: June 2006

The role of CD8 (Lyt‐2) in the function of a long‐term cytotoxic T‐cell (CTL) clone (C196) was analysed. Previous studies had shown that C196 cells utilize the αβT‐cell receptor and depend on Lyt‐2 to recognize and lyse P815 mastocytoma target cells. Recognition is H‐2Kd restricted, presumably involving a P815 specific antigenic structure. Here we analyse a number of variants selected from C196 that have reduced or virtually abolished expression of Lyt‐2, alone or in combination with other deficiencies. We studied the ability of these variant cells to lyse either P185, a process requiring both specific antigen recognition and triggering of cytolytic function, or to lyse the anti‐CD3 hybridoma 145‐2C11, a process that requires the triggering of cytolytic function only. The results shows that in the case or a permanently activated CTL, effector cell such as C196, Lyt‐2 is required for antigen recognition but is not essential for the triggering of cytotoxicity.
Expression of Deoxyadenosine and Deoxyguanosine Toxicity at Different Stages of Lymphocyte Activation
Authors: SCHARENBERG J. G. M. , RIJKERS G. T. , TOEBES E. A. H. , STAAL G. E. J. , ZEGERS B. J. M.
Published: June 2006

We have previously shown that deoxyguanosine (dGuo) is toxic to normal T and B lymphocytes, an effect mediated by intracellular accumulation of guanine ribonucleotides. In order to define the cellular processes that are sensitive to guanosine triphosphate (GTP) we have performed studies in which the effects of dGuo on normal T cells are compared with those of deoxyadenosine (dAdo) on adenosine deaminase (ADA)‐deficient T cells. Kinetic studies show that dAdo exerts its toxic effects on processes that precede the onset of DNA synthesis, like interleukin 2 receptor expression, whereas dGuo added as late as 24–48 h after initiation of the culture still inhibits mitogen‐induced proliferation. It can thus be concluded that dGuo toxicity as mediated through guauine ribonucleotides is manifested relatively late during the process of T‐cell activation, whereas dAdo acts early in T‐cell activation by a mechanism that cannot be explained by inhibition of ribonucleotide reductase.
Cell‐Mediated Immunity to Plasmodium falciparum Infection: Evidence against the Involvement of Cytotoxic Lymphocytes
Authors: THEANDER T. G. , ANDERSEN B. J. , PEDERSEN B. K. , JEPSEN S. , BYGBJERG I. C. , HVIID L. , LARSEN P. B. , KHARAZMI A.
Published: June 2006

Blood mononuclear cells (PBMC) recognizing soluble malaria antigens (SPag) are present in the peripheral blood of individuals clinically immune to malaria, and they proliferate after exposure to such antigens. To test whether these cells have effector activity against Plasmodium falciparum, we stimulated PBMC from malaria‐immune donors by SPag and purified protein derivative (PPD) in culture for 7 days. The PBMC were then co‐incubated with P. falciparum for 48 h, and parasitaemia was determined by microscopy. Parasite growth was only significantly impaired after incubation with PBMC stimulated by either SPag or PPD in the presence of immune serum. Studies on subpopulations of PBMC indicated that the inhibitory cells resided among the adherent cell fraction. Furthermore we tested PBMC for cytotoxic activity against P. falciparum‐infected autologous or heterologous erythrocytes. Experiments were done both in the absence and the presence of immune serum. Neither fresh PBMC nor PBMC activated by SPag or PPD for 7 days prior to assay were cytotoxic, indicating that cytotoxic T cells, natural killer (NK) cells, and K cells did not possess cytotoxic activity directed against parasitized erythrocytes. These data support the hypothesis that activated monocytes are the most important effector cells in the peripheral blood of malaria immune individuals.
Inefficient Binding of IgM Immune Complexes to Erythrocyte C3b–C4b Receptors (CR1) and Weak Incorporation of C3b–iC3b into the Complexes
Authors: KÁVAI M. , RASMUSSEN J. MØLLER , BAATRUP G. , ZSINDELY A. , SVEHAG S.‐E.
Published: June 2006

The binding of soluble complement‐reacted IgM immune complexes (IC) to erythrocyte (E) C3b–C4b receptors (CRI) and the incorporation of C3b–iC3b into solid phase IgM‐IC was investigated. The optimal binding of liquid phase IgM‐IC to E‐CRI was obtained with IC formed at moderate antibody excess, but the binding was low (2–3%) when compared to the binding of the corresponding IgG‐IC (50–60%). Solid phase IC were prepared by coming microwells with heat‐aggregated bovine serum albumin (BSA) followed by incubation with rabbit IgM anti‐BSA antibody. The IC were reacted with human serum at 37°C. The binding of C3b–iC3b was determined by use of biotinylated F(ab')2 antibodies to C3b‐C3c and avidin‐coupled alkaline phosphatase. The incorporation of C3b–iC3b into solid‐phase IgM‐IC increased when increasing amounts of IgM antibody were reacted with the antigen. The binding reaction was slow, reaching a maximum after about 2 h at 37°C. The binding of C3b–iC3b to the IgM‐IC was remarkably inefficient when compared to the incorporation into IgG‐IC reacted with the same amounts of BSA‐precipitating antibody.
Isolation and Characterization of Fc Receptors from Haemophilus somnus
Authors: YARNALL M. , WIDDERS P. R. , CORBEIL L. B.
Published: June 2006

Receptors that bind the Fc region of bovine immunoglobulin (Ig) have been isolated from the culture supernatant of Haemophilus somnus by chromatography on a Sepharose 4B column. One receptor with a relative molecular weight of 41,000 weakly binds both bovine IgG subclasses, IgA and IgM, while three high molecular weight receptors (350,000, 270,000, and 120,000) strongly bind bovine IgG2, FgA, and IgM. All four Fc receptors are antigenically related and the 41,000 receptor appears to be a subunit of the high molecular weight receptors. In addition to bovine Ig, the purified 270,000 Fc receptor strongly binds horse IgG, rabbit IgG, pig IgG, cat IgG, dog IgG, and sheep IgG. The receptor also reacts weakly with mouse, rat, chicken, human, and guinea pig IgG and does not bind goal IgG. Fc receptors from 19 H. somnus isolates were compared. Variations in the molecular weight of the 41,000 protein were demonstrated among preputial isolates from asymptomatic carriers, but ad other Isolates appeared to have identically migrating proteins.
Modulation of Human Monocyte Functions during Acute Bacterial Infection
Authors: MROWIETZ U. , CHRISTOPHERS E.
Published: June 2006

The in vitro functions of highly purified blood monocytes were studied in 11 patients suffering from acute bacterial infections (e.g. erysipelas, appendicitis, abscesses). Chemotaxis, superoxide‐anion generation, and β‐glucuronidase release of the patients' monocytes in response to the receptor‐dependent stimuli formyl‐methionyl‐leucyl‐phenylalanine (FMLP), complement split product C5a, leukotriene B4 (LTB4), and opsonized zymosan particles were measured. All the patients were examined in a follow‐up study during the course of illness. A group of 33 healthy volunteers served as control. The patients revealed a transient decrease in monocyte chemotactic migration in response to all stimuli between days 3 and 5 after onset of clinical symptoms. Superoxide‐anion generation from patients' monocytes was found to he enhanced 3 days after impaired chemotaxis. Stimulated release of lysosomal β‐glucuronidase showed a decrease in the first days of the disease. However, spontaneous β‐glucuronidase release was enhanced between days 3 and 7 in the patients' monocytes. Serial measurements of monocyte functions carried out in six healthy subjects showed no significant alterations in monocyte responsiveness. These results indicate a distinct modulation of monocyte functions during the course of an acute bacterial infection. Changes in monocyte maturity and/or activation under inflammatory conditions may be responsible for these alterations in monocyte function.
Modulation of High‐Affinity Interleukin 2 Receptors on Activated Human T Lymphocytes by Activators of Protein Kinase C
Authors: LARSEN C. S. , CHRISTIANSEN N. O. , ESMANN V.
Published: June 2006

Phorbol myristate acetate (PMA) and 1‐oleoyl‐2‐acetyl‐rac‐glycerol (OAG) are shown to induce a rapid (within 30 min) down‐regulation of the capacity of activated human T lymphocytes to bind interleukin 2. This was associated with a manifold increase in membrane‐associated protein kinase C, whereas no change in free cytoplasmic calcium was observed. In contrast, a 10‐fold increase in free cytoplasmic calcium by ionomycin had no effect on interleukin 2 binding or sub‐cellular distribution of protein kinase C. The reduction of interleukin 2 binding was caused by a decreased number of high‐affinity interleukin 2 receptors, whereas the affinity of the remaining receptors was unchanged. However, PMA and OAG had no effect on the rate of initialization of the interleukin receptor. These data suggest that activation of protein kinase C. but not an increase in free cytoplasmic calcium, leads to a rapid decrease in the number of high‐affinity interleukin 2 receptors on activated human T lymphocytes. However, the mechanism and biological importance of this phenomenon have to be further elucidated.
Cytomegalovirus‐Infected Adherent Cells Interact Synergistically and Antagonistically with Staphylococcus aureus Protein A in vitro
Authors: PAULIN T. , RINGDÉN O. , WAHREN R.
Published: June 2006

Cytomegalovirus (CMV) has been shown to exert suppressive effects on the immune response but also to have mitogenic properties. A bacterial product, protein A from Staphylococcus aureus (SpA) was chosen to study possible interaction in vitro between bacterial products and adherent cells incubated with infectious CMV and ultraviolet light (UV)‐inactivated CMV. Small amounts of infectious CMV potentiated SpA‐induced DNA synthesis and Ig secretion measured by induction of plaque‐forming cells (PFC). The reason for this may be that CMV in small amounts may act in synergism with the non‐specific mitogen SpA. UV‐inactivated CMV did not influence these responses except for a markedly enhanced PFC induction with SpA in lymphocytes from seronegative individuals. This remarkable synergism with SpA was also seen in enriched B cells. No synergism was seen in lymphocytes from seropositive donors. Large amounts of infectious CMV markedly reduced SpA‐induced immune responses. Preliminary data suggest that the immunosuppressive effects are mediated by an interleukin l inhibitory factor. CMV was not shown to be a polyclonal B‐cell activator but may, possibly in small amounts, act as such together with bacterial products, which would explain certain autoimmune phenomena. To conclude, CMV could in interaction with a bacterial product generate both synergistic and suppressive effects on immune response.
Monoclonal Antibodies against the Major Internal Protein, p27, of a Psoriasis‐Associated Retrovirus‐Like Particle
Authors: ÅSBAKK K. , IVERSEN O.‐J.
Published: June 2006

Four murine monoclonal antibodies (MoAb) were raised against p27 isolated from psoriatic scale. The MoAb recognized the antigen on a subfraction (0.1–1%)of psoriatic lymphocytes as well as in skin biopsies from psoriatic patients. In contrast, the antigen could not be detected on lymphocytes or in skin biopsies from healthy controls. A sandwich ELISA assay using the MoAb for detection of p27 is described. This method, together with a competition antibody‐binding assay for distinction of epitopes, demonstrates that the MoAb recognize lour different antigenic determinants on p27.
Unidirectional Responses to Mls Determinants in Vivo
Authors: HOSONO M. , KINA T. , KATSURA Y. , HOSOKAWA T.
Published: June 2006

Polyclonal anti‐Mls responses of peripheral (mature) and thymic (immature) lymphocytes were studied in vivo in terms of local host‐versus‐graft and graft‐versus‐host reactions. The level of stimulatory activity differed between the various types of Mls antigens, with Mlsa and Mlsd having the highest levels, Mlsc a lower level, and Mlsb the lowest level. The immunogenicity of Mlsa and Mlsd may not be identical, since Mlsd mice responded to the Mlsd determinant, while Mlsa mice did not react with the Mlsd determinants. This suggests that the anti‐Mls response is unidirectional. The fact that Mlsb mice made tolerant at birth to Mlsd (Mlsa) antigens behave like Mlsd (Mlsa) mice when responding to Mls antigens supports this suggestion. Furthermore, thymus cells from the Mlsa‐tolerized BALB/c (Mlsb) mice were unresponsive to both Mlsa and Mlsd antigens, while those made tolerant to Mlsd were responsive to Mlsa. These results indicate that the polyclonal response to the strongly immunogenic Mlsad antigens is unidirectional, and that the immunogenicity of Mlsa and Mlsd is not identical. Based on the results of tolerance experiments and data on T‐cell clones, we suggest that the difference in immunogenicity between Mlsa. b. d antigens is due to different efficiencies of the responding T‐cell populations, probably because of a quantitative difference in a single common antigenic determinant expressed in each Mls haplotype, which results in different levels of stimulation of T cells according to the avidity for the determinant.
Accessory Cell Paradox: Monocytes Enhance or Inhibit Lectin‐Mediated Human T‐Lymphocyte Proliferation Depending on the Choice of Mitogen
Authors: KILPATRICK D. C.
Published: June 2006

Monocytes suppressed the mitogenic response of human T lymphocytes to the lectin from Datura stramonium but enhanced the mitogenic response of the same lymphocytes to wheat germ agglutinin under identical culture conditions. The inhibitory action was probably cell‐mediated, but was not the result of cytotoxicity. The stimulative activity could be replaced by cell‐free conditioned medium.
Intrinsic Cell Membrane Antigens Recognized by Antichromatin Autoantibodies
Authors: REKVIG O. P.
Published: June 2006

The main object of this study was to see whether or not chromatin constituents are present in cell membranes. The binding of antinuclear antibodies (ANA) to plasma membranes of leucocytes was studied by using autoantibodies and induced antibodies to different histones and histone peptides, and autoantibodies to double‐stranded DNA (dsDNA) in adsorption–elution experiments. Eluates were subsequently tested for antinuclear antibodies by a solid‐phase ELISA. No ANA activities in the eluates were observed, except when true cross‐reacting ANA were employed in the study. Futhermore, no binding of these antibodies to plasma membranes could be visualized by indirect membrane fluorescence tests. The conclusion of these studies was that freshly isolated viable leucocytes did not carry detectable amounts of ectopic chromatin components at the level of plasma membranes. Thus, it seems fairly unlikely that chromatin autoantibodies in general exert some tissue damage by binding to homologous nuclear antigens associated with plasma membranes in vivo.
S‐Protein Is Synthesized by Human Monocytes and Macrophages in Vitro
Authors: HETLAND G. , PETTERSEN H. B. , MOLLNES T. E. , JOHNSON E.
Published: June 2006

Human monocytes and alveolar and peritoneal macrophages were cultured in serum‐free medium with or without endotoxin (ET), agarose beads, or cycloheximide. The cell culture supernatants were collected after various intervals and examined by a monoclonal anti‐S‐protein antibody in Western blot and in a solid‐phase enzyme immunoassay. We found that the phagocytes synthesize and secrete S‐protein. ET stimulation or prolonged incubation of the cells did not favour S‐protein production, which was inhibited by cycloheximide. Agarose stimulation increased the S‐protein level in supernatants from monocyte but not from macrophage cultures.
The Relation between Cutaneous Blood Flow and Cell Content in the Tuberculin Reaction
Authors: BECK J. SWANSON , GIBBS J. H. , POTTS R. C. , JAWAD EMTITHAL S. , KARDJITO T. , SPENCE V. A. , GRANGE J. M.
Published: June 2006

The relationship of velocity of blood flow to density and microanatomical distribution of inflammatory cells in the dermis was studied in 20 human tuberculin tests. Must positive reactions showed maximal blood flow velocities (measured as red blood cell (RBC)flux) at the centre of the reaction, but the two most intense responses showed ‘central relative slowing’(CRS) with higher RBC flux at she periphery. Two of the four clinically negative reactions showed a considerable acceleration of blood flow, but the other two showed no such acceleration. The packing density of lymphocytes/monocytes in the perivascular zone was greater in the stronger positives than in the weaker reactions. The density of cells in the intervening dermis was markedly lower than in the foci: the lesions with CRS had the highest density of cells in the diffuse infiltrate of the reticular dermis. At the centre of the reaction, blood flow velocity was generally related to density of cellular infiltrate, except in those with CRS, which had a disproportionately lower blood flow velocity. The finding that the circulatory adaptation to a delayed hypersensitivity reaction can be inadequate may explain the dermal acidosis previously observed in intense skin test reactions, and may be the underlying mechanism of necrosis in hypersensitive reactions.
A Model System for the Analysis of B‐Cell Activation and Effector T‐Cell Functions
Authors: PEREIRA P. , BANDEIRA A. , KLEINE B. , MARQUEZ C. , COUTINHO A. , MARTINEZ‐A C.
Published: June 2006

We have attempted to develop an in vitro system where polyclonal B lymphocyte responses could be induced in ‘antigen‐like’ conditions, that is, where surface immunoglobulin‐dependant binding mediates interaction with a mitogen. Monoclonal anti‐μ and anti‐δ antibodies were covalently bound to lipopolysaccharide (LPS) and these complexes were shown to display mitogenic activity. Polyclonal plaque‐forming‐cell (PFC) responses, however, were diminished in cultures stimulated by anti‐μ‐LPS (but not by anti‐δ‐LPS) indicating that ‘anti‐μ inhibition’ of terminal B‐cell differentiation also applies to ‘specific’ antibody responses. Moreover, the analysis of the functional activity of monoclonal antibodies to major histocompatibility complex (MHC) class II molecules revealed a surprising synergy between low. non‐stimulatory concentrations of anti‐μ‐LPS (but not anti‐SdL‐LPS) with anti‐I‐A antibodies. These responses are T‐cell dependent and synergy with anti‐μ‐LPS conjugates can also be obtained with ‘naturally’ activated CD4+ cells isolated from normal donors. A model of molecular and cellular interactions was derived, which accounts for the present findings and is applicable in antigen‐dependent lymphocyte collaboration.