bluuebunny/crossref_metadata_2025 huggingface dataset:
Vaccination with the major secretory protein of Legionella pneumophila induces cell-mediated and protective immunity in a guinea pig model of Legionnaires' disease.
Authors: Blander S J , Horwitz M A
Published: June 2004

We have examined the capacity of the major secretory protein (MSP) of Legionella pneumophila to induce humoral, cell-mediated, and protective immunity in a guinea pig model of Legionnaires' disease. MSP was purified to homogeneity by ammonium sulfate precipitation, molecular sieve chromatography, and ion-exchange chromatography. The purified MSP was nonlethal and nontoxic to guinea pigs upon subcutaneous administration. Guinea pigs immunized with a sublethal dose of aerosolized L. pneumophila or a subcutaneous dose of MSP developed a strong cell-mediated immune response to MSP. Such guinea pigs exhibited marked splenic lymphocyte proliferation and cutaneous delayed-type hypersensitivity to MSP in comparison with control animals. Guinea pigs immunized with MSP also developed a strong humoral immune response to MSP, as assayed by an ELISA. The median reciprocal antibody titer was 362 (range 45 to greater than 2,048) for immunized animals compared with less than 8 for controls. In contrast, guinea pigs immunized with a sublethal dose of L. pneumophila failed to develop anti-MSP antibody. Guinea pigs immunized with MSP and then challenged with a lethal aerosol dose of L. pneumophila exhibited highly significant protective immunity in each of five consecutive experiments. MSP induced protective immunity in dose-dependent fashion (40 greater than 10 greater than 2.5 greater than 0.6 micrograms MSP); vaccination with two doses of as little as 2.5 micrograms MSP induced significant protective immunity (p = 0.01, Fisher's Exact Test, two-tailed). Altogether, 21 (81%) of 26 animals immunized with 40 micrograms MSP survived challenge compared with 0 (0%) of 26 sham-immunized control animals (p = 7 x 10(-10), Fisher's Exact Test, two-tailed). MSP-immunized but not control guinea pigs were able to limit L. pneumophila multiplication in their lungs. This study demonstrates that (a) guinea pigs sublethally infected with L. pneumophila develop a strong cell-mediated immune response to MSP; (b) guinea pigs immunized with MSP develop a strong humoral and cell-mediated immune response to MSP; (c) guinea pigs immunized with MSP develop a very high level of protective immunity to lethal aerosol challenge with L. pneumophila; and (d) MSP-immunized animals are able to limit L. pneumophila multiplication in their lungs. MSP, an extracellular protein of an intracellular pathogen, has potential as a vaccine for the prevention of Legionnaires' disease. Secretory molecules of other intracellular pathogens may also have vaccine potential.
Studies of lymphokine-activated killer (LAK) cells. I. Evidence using novel monoclonal antibodies that most human LAK precursor cells share a common surface marker.
Authors: Morris D G , Pross H F
Published: June 2004

Separation of LAK precursor (LAKp) cells (as defined by LAK effector generation after incubation with IL-2 for 7 d) from cells with NK activity/LGL morphology was achieved on Percoll gradients using a longer, slower centrifugation than that used for optimal NK enrichment. mAb were generated using the various Percoll fractions as the immunizing cells and used for separation and depletion studies. Two mAbs DM-1 (IgM,k) and DM-2 (IgM,k) recognizing 2-15% and 15-30% of PBL, respectively, abrogated a large proportion of LAK generative potential after complement depletion, but had little effect on NK or LAK effector activity. Cell sorting experiments indicated that the majority of LAKp cells are found within the DM-1+ population and that DM-1+ cells are not simply an accessory cell required for LAKp generation. Further, these two mAbs do not recognize cells that are responsible for generating cytotoxicity during MLC or co-culture with the PR-1 EBV lymphoblastoid cell line. Western blot analysis indicated that DM-1 and DM-2 recognize a 38,000 and 44,000 dalton moiety, respectively. The frequency of cells bearing these antigens and the intensity of cell surface staining decreased during the 7-d culture period, suggesting that these antibodies recognize determinants found only at the precursor level. These findings indicate that cells other than NK effectors or mature T cells are capable of generating a LAK cell response. These LAK precursor cells share a common differentiation surface antigen and are different from AK or antigen-specific CTL precursors. The possibility exists that these cells are identical to, or include, the NK precursor cell.
Potent leukocidal action of Escherichia coli hemolysin mediated by permeabilization of target cell membranes.
Authors: Bhakdi S , Greulich S , Muhly M , Eberspächer B , Becker H , Thiele A , Hugo F
Published: June 2004

The contribution of Escherichia coli hemolysin (ECH) to bacterial virulence has been considered mainly in context with its hemolytic properties. We here report that this prevalent bacterial cytolysin is the most potent leukocidin known to date. Very low concentrations (approximately 1 ng/ml) of ECH evoke membrane permeability defects in PMN (2-10 x 10(6) cells/ml) leading to an efflux of cellular ATP and influx of propidium iodide. The attacked cells do not appear to repair the membrane lesions. Human serum albumin, high density and low density lipoprotein, and IgG together protect erythrocytes and platelets against attack by even high doses (5-25 micrograms/ml) of ECH. In contrast, PMN are still permeabilized by ECH at low doses (50-250 ng/ml) in the presence of these plasma inactivators. Thus, PMN become preferred targets for attack by ECH in human blood and protein-rich body fluids. Kinetic studies demonstrate that membrane permeabilization is a rapid process, ATP-release commencing within seconds after application of toxin to leukocytes. It is estimated that membrane permeabilization ensues upon binding of approximately 300 molecules ECH/PMN. This process is paralleled by granule exocytosis, and by loss of phagocytic killing capacity of the cells. The recognition that ECH directly counteracts a major immune defence mechanism of the human organism through its attack on granulocytes under physiological conditions sheds new light on its possible role and potential importance as a virulence factor of E. coli.
Induction of classical transplantation tolerance in the adult.
Authors: Qin S X , Cobbold S , Benjamin R , Waldmann H
Published: June 2004

Transplantation tolerance across histoincompatibilities in multiple non-H-2 minors (B10.BR into CBA/Ca) and "minor" plus H-2D (B10.A into CBA/Ca) antigens has been achieved successfully by combined adult bone marrow transplantation and treatment with CD4 and CD8 mAbs. The tolerant state was confirmed by permanent acceptance of donor strain skin grafts, and in vitro unresponsiveness to donor cells. Tolerance was associated with partial donor chimerism to various degrees. Tolerance to minor transplantation antigens induced in this manner was restricted to recipient-type MHC. The possibility was raised that tolerance resulted, at least in part, from clonal anergy rather than deletion.
Endotoxin and tumor necrosis factor challenges in dogs simulate the cardiovascular profile of human septic shock.
Authors: Natanson C , Eichenholz P W , Danner R L , Eichacker P Q , Hoffman W D , Kuo G C , Banks S M , MacVittie T J , Parrillo J E
Published: June 2004

Survivors of both human and animal bacterial shock develop a characteristic pattern of progressive changes in cardiovascular function over a period of 7-10 d. In this present study, we examined whether endotoxin (a product of Gram-negative bacteria) or TNF (a cytokine released from macrophages) could reproduce the same complex cardiovascular changes observed in septic shock over a period of 7-10 d. To test this hypothesis, we implanted a thrombin-fibrin clot containing purified endotoxin from E. coli into the peritoneal cavity of eight dogs, and infused TNF into eight different dogs. Over the next 10 d, serial simultaneous heart scans and thermodilution cardiac outputs were performed in these awake nonsedated animals. By day 2 after challenge with either endotoxin or TNF, animals developed a decrease (p less than 0.05) in both mean arterial pressure and left ventricular ejection fraction. With fluid resuscitation, animals manifested left ventricular dilatation (increased [p less than 0.05] end diastolic volume index), increased or normal cardiac index, and decreased or normal systemic vascular resistance index. In surviving animals, these changes returned to normal with 7-10 d. The time course of these changes was concordant (p less than 0.05) with that previously described in a canine model of septic shock using viable bacteria. During the 10-d study, control animals receiving sterile clots or heat-inactivated TNF had not significant changes in hemodynamics. The results from this canine model demonstrate that either endotoxin or TNF alone can produce many of the same hemodynamic abnormalities seen in human septic shock and in a canine septic shock model induced by live bacteria. These findings support the hypothesis that the action of endogenous mediators (TNF) responding to bacterial products (endotoxin) is the common pathway that produces the serial cardiovascular changes found in septic shock.
Amastigotes of Trypanosoma cruzi escape destruction by the terminal complement components.
Authors: Iida K , Whitlow M B , Nussenzweig V
Published: June 2004

We studied the effect of complement on two life cycle stages of the protozoan parasite Trypanosoma cruzi: epimastigotes, found in the insect vector, and amastigotes, found in the mammalian host. We found that while both stages activate vigorously the alternative pathway, only epimastigotes are destroyed. The amounts of C3 and C5b-7 deposited on the amastigotes were similar to those bound to the much larger epimastigotes. Binding of C9 to amastigotes was four to six times less than binding to epimastigotes, resulting in a lower C9/C5b-7 ratio. Although a fairly large amount of C9 bound stably to amastigotes, no functional channels were formed as measured by release of incorporated 86Rb. The bound C9 had the characteristic properties of poly-C9, that is, it expressed a neo-antigen unique to poly-C9, and migrated in SDS-PAGE with an apparent Mr greater than 10(5). The poly-C9 was removed from the surface of amastigotes by treatment with trypsin, indicating that it was not inserted in the lipid bilayer. Modification of amastigote surface by pronase treatment rendered the parasites susceptible to complement attack. These results suggest that amastigotes have a surface protein that binds to the C5b-9 complex and inhibits membrane insertion, thus protecting the parasites from complement-mediated lysis.
The generation of antigen-specific, major histocompatibility complex-restricted cytotoxic T lymphocytes of the CD4+ phenotype. Enhancement by the cutaneous administration of interleukin 2.
Authors: Hancock G E , Cohn Z A , Kaplan G
Published: June 2004

We have examined an in vitro system in which PBMC from purified protein derivative (PPD)-sensitized patients generate CTL after in vitro activation with antigen. These cells selectively destroy mycobacterial antigen PPD-pulsed monocyte targets. These CTL are of the CD4+ phenotype and exhibit MHC class II restriction. After exposure to antigen these cells require 5-7 d for maximal development, whereas, a separate antigen-independent population is generated within 3-4 d. CD8+ cells are poorly, if not at all, cytotoxic under similar conditions. Cells with properties of the NK and LAK lineage are also present in these cultures and kill other specific targets. Human rIL-2 was injected into the skin of lepromatous patients at 10-micrograms doses, given at 48-h intervals, for three doses. Peripheral blood cells obtained 8-14 d after the initiation of IL-2 injection demonstrated enhanced antigen-dependent destruction of monocyte targets. The efficacy of antigen-dependent and -independent populations and their amplification by IL-2-dependent mechanisms is discussed in terms of the local destruction of parasitized macrophages and the subsequent disposal of M. leprae.
Interferon alpha associated with systemic lupus erythematosus is not intrinsically acid labile.
Authors: Yee A M , Buyon J P , Yip Y K
Published: June 2004

The physicochemical properties of apparently acid-labile IFN-alpha from patients with SLE have been studied. The antigenicity, apparent molecular size, and isoelectric point of SLE IFN-alpha are indistinguishable from those of conventional, previously characterized, acid-stable subspecies of IFN-alpha. However, after partial purification by anion-exchange chromatography, SLE IFN-alpha no longer exhibits acid lability, suggesting that other plasma factor(s) are responsible for the acid lability of SLE IFN-alpha. Addition of SLE plasma, but not normal plasma, to conventional acid-stable IFN-alpha renders the exogenous IFN-alpha acid labile. Preliminary results demonstrate that an acid-dependent IFN-inactivating activity can be partially purified from SLE plasma by anion-exchange chromatography.
Site-restricted persistent cytomegalovirus infection after selective long-term depletion of CD4+ T lymphocytes.
Authors: Jonjić S , Mutter W , Weiland F , Reddehase M J , Koszinowski U H
Published: June 2004

We have established a murine model system for exploring the ability of a CD4 subset-deficient host to cope with cytomegalovirus infection, and reported three findings. First, an antiviral response of the CD8 subset of T lymphocytes could be not only initiated but also maintained for a long period of time despite a continued absence of the CD4 subset, whereas the production of antiviral antibody proved strictly dependent upon help provided by the CD4 subset. Second, no function in the defense against infection could be ascribed as yet to CD4-CD8- T lymphocytes, which were seen to accumulate to a new subset as a result of depletion of the CD4 subset. This newly arising subset did not substitute for CD4+ T lymphocytes in providing help to B lymphocytes, and was also not effective in controlling the spread of virus in host tissues. As long as a function of these cells in the generation and maintenance of a CD8 subset-mediated response is not disproved, caution is indicated with concern to an autonomy of the CD8 subset. Third, even though with delay, the CD8+ effector cells raised in the CD4 subset-deficient host were able of clear vital tissues from productive infection and to restrict asymptomatic, persistent infection to acinar glandular epithelial cells in salivary gland tissue.
Lineage-specific expression of a T cell receptor variable gene promoter controlled by upstream sequences.
Authors: Diamond D J , Nelson F B , Reinherz E L
Published: June 2004

We have characterized the sequence contribution of DNA 5' of a functionally rearranged TCR promoter (V beta 8.1) on its T lineage-specific expression through the use of the chloramphenicol acetyl-transferase (CAT) reporter gene. A 230-bp fragment located 570 bp upstream of the determined transcription start site of the V beta 8.1 promoter confers a T lineage specificity of expression to a heterologous promoter. The inability of the V beta 8.1 promoter and its associated elements to function in B cells suggests the existence of a mechanism to prevent inappropriate V beta gene expression in B cells. Of considerable interest is the fact that both a B cell-specific and a nontissue-specific enhancer element were incapable of stimulating significant expression of this promoter in B cells. We discuss the implication of these results on the process of rearrangement of both Ig and TCR genes, and the differentiation of the lymphoid system.
Primary virus-induced lymphomas evade T cell immunity by failure to express viral antigens.
Authors: Vasmel W L , Sijts E J , Leupers C J , Matthews E A , Melief C J
Published: June 2004

T lymphoma induction by the mink cell focus-inducing murine leukemia virus MCF 1233 in C57BL/10 and C57BL/6 mice is influenced by a strongly Th-dependent, H-2I-A-restricted antiviral immune response (25). We compared the MHC class I as well as viral env and gag antigenic cell surface profiles of frequent T lymphomas of H-2I-A nonresponder-type mice to that of rare T lymphomas of H-2I-A responder-type mice. Membrane immunofluorescence studies, with a panel of anti-env mAbs (reactive with the highly conserved gp70f epitope, the p15Ec epitope, and the gp70-p15E complex), a polyclonal anti-p30 serum, and anti-H-2 class I mAbs, showed that all 17 nonresponder tumors tested expressed high levels of both env and gag viral proteins, and 15 of these 17 nonresponder tumors expressed high levels of H-2 class I K and D antigens. In contrast, 10 of 11 responder lymphomas lacked env and/or gag determinants. The only responder lymphoma with both strong env and gag expression failed to express H-2K and -D antigens. Preferential loss of env or gag expression did not correlate with H-2 class I allelic specificities. Both responder and nonresponder T lymphoma DNA contained multiple, predominantly MCF-like, newly acquired proviral integrations. Differences in viral antigen cell surface expression were confirmed at cytoplasmic and RNA levels. The amounts of 8.2- and 3.2-kb viral RNA were greatly reduced in two responder lymphomas when compared with four nonresponder lymphomas. In both responder lymphomas, aberrantly sized viral RNA species were found. Upon in vivo passage of these responder lymphomas in either immunocompetent or T cell-deficient nu/nu mice, it was found that various molecular mechanisms may underlie the lack of viral antigen expression at the cell surface of these lymphomas. One lymphoma re-expressed viral antigens when transplanted with nu/nu mice, whereas the other remained stably gag negative. The combined findings indicate that an H-2I-A-regulated antiviral immune response not only strongly reduces T lymphoma incidence, but also forces T lymphomas that still arise to poorly express viral antigens, thus explaining their escape from immunosurveillance.
Cloning of phenotypically different human lymphocytes originating from a single stem cell.
Authors: Hakoda M , Hirai Y , Shimba H , Kusunoki Y , Kyoizumi S , Kodama Y , Akiyama M
Published: June 2004

By using hypoxanthine guanine phosphoribosyltransferase (hprt) gene alterations and chromosome aberrations as in vivo cellular markers, human T, NK, and B cells originating from a single stem cell have been successfully cloned from the peripheral blood of an atomic bomb survivor from Hiroshima. These mutant lymphocytes were selectively cloned, taking advantage of their resistance to a purine analogue, 6-thioguanine. The cloned lymphocytes possessed the same hprt gene alterations and the same chromosome aberration (20q-), but exhibited different surface or functional phenotypes and different rearrangements of TCR or Ig genes. The chromosome aberration patterns strongly suggested that the original stem cell initiated differentiation into each cell type after exposure to atomic bomb radiation. Since the person studied here was exposed to the bomb at 17 yr age, the results suggested that common stem cells exist in adults for at least T, NK, and B cells. The use of hprt gene alterations as specific cellular markers provides a novel method for identifying stem cells in the lymphocyte lineage and for studying lymphocyte differentiation in humans.
Human lymphocytes bearing T cell receptor gamma/delta are phenotypically diverse and evenly distributed throughout the lymphoid system.
Authors: Groh V , Porcelli S , Fabbi M , Lanier L L , Picker L J , Anderson T , Warnke R A , Bhan A K , Strominger J L , Brenner M B
Published: June 2004

A direct quantitative and phenotypic cytofluorographic analysis of TCR-gamma/delta+ lymphocytes as well as an immunohistologic study of their tissue distribution and microanatomy was made possible by the availability of two mAbs (anti-TCR-delta 1 and anti-C gamma M1) specific for framework determinants on human TCR gamma and delta chains, respectively. TCR-gamma/delta+ lymphocytes, ranging between greater than 0.5 and 16% of CD3+ cells, were found in fetal and postnatal thymus, fetal and adult peripheral lymphoid organs, and adult peripheral blood. While TCR-gamma/delta+ lymphocytes comprised a small subpopulation of T cells (mean, approximately 4%) occasionally greater than 10-16% of CD3+ cells expressed TCR-gamma/delta. Virtually all TCR-gamma/delta+ thymocytes/lymphocytes expressed CD7, CD2, and CD5 but were heterogeneous with respect to their expression of CD1, CD4, CD8, CD28, CD11b, CD16, and Leu-7. Human TCR-gamma/delta+ cells populate both organized lymphoid tissues (thymus, tonsil, lymphnode, and spleen) as well as the gut- and skin-associated lymphoid systems at similar frequencies without obvious tropism for epithelial microenvironments. TCR-gamma/delta+ lymphocytes tend to be located within a given organ wherever TCR-alpha/beta+ lymphocytes are found. This study shows that TCR-gamma/delta+ lymphocytes constitute a small but numerically important, phenotypically diverse T cell population distributed throughout the body. These results support the concept that TCR-gamma/delta+ cells comprise a distinct, functionally heterogeneous, mature T cell sublineage that may substantially broaden the T cell repertoire at all immunologically relevant sites.
Induction of human IgE synthesis requires interleukin 4 and T/B cell interactions involving the T cell receptor/CD3 complex and MHC class II antigens.
Authors: Vercelli D , Jabara H H , Arai K , Geha R S
Published: June 2004

The induction of IgE synthesis by IL-4 requires T cells and monocytes, as well as T cell- and monocyte-derived cytokines. Optimal cytokine combinations, however, fail to induce highly purified B cells to secrete IgE, indicating that additional signals are required. We show herein that the induction of human IgE synthesis by rIL-4 requires cognate interaction between the T cell receptor/CD3 complex on T cells and MHC class II antigens on B cells: mAbs directed against these molecules completely blocked IL-4-dependent IgE induction. mAbs against cell adhesion molecules (CD2, CD4, LFA-1) also inhibited IgE synthesis induced by IL-4, confirming that cell-cell contact is necessary for IgE induction. The requirement for cognate T/B cell interaction was further shown by comparing the IgE-inducing ability of two human IL-4-producing alloreactive T cell clones: F6, which recognizes MHC class II antigens on both B cells and monocytes, and A1, which recognizes an HLA-DP-associated epitope expressed on monocytes, but not on B cells. When incubated with B cells and monocytes from a normal donor bearing the appropriate alloantigen, clone F6, but not clone A1, induced vigorous IgE synthesis, although both clones proliferated and secreted IL-4. Taken together, our results suggest that at least two, possibly synergizing, signals are required for the T cell-dependent induction of IgE synthesis by B cells: one signal is delivered by cognate T/B cell interaction, the other by T cell-derived IL-4.
Monoclonal antibody defining a molecule possibly identical to the p75 subunit of interleukin 2 receptor.
Authors: Takeshita T , Goto Y , Tada K , Nagata K , Asao H , Sugamura K
Published: June 2004

A mouse hybridoma cell line, TU27, producing an mAb was established. TU27 mAb reacted with various human and Gibbon ape T cell lines bearing the IL-2R p75 (IL-2Rp75), but not with cell lines expressing only Tac antigen, IL-2Rp55, and numbers of its binding sites on cell surfaces were similar to those of high-affinity IL-2R. Radioimmunoprecipitation with TU27 mAb defined a molecule with a molecular mass of 75 kD on the surface of IL-2Rp75 bearing cells. TU27 mAb completely blocked IL-2 binding to IL-2Rp75 and to the high-affinity IL-2R but not to IL-2Rp55 composing the low-affinity IL-2R. The IL-2-dependent growth of a human T cell line, ILT-Mat, was significantly inhibited by TU27 mAb only at low concentrations of IL-2, and combination of TU27 mAb and H-31 mAb specific for IL-2Rp55 completely inhibited the cell growth even at high concentrations of IL-2. These data strongly suggest that TU27 mAb is specific for the human IL-2Rp75.
Mouse macrophage hemagglutinin (sheep erythrocyte receptor) with specificity for sialylated glycoconjugates characterized by a monoclonal antibody.
Authors: Crocker P R , Gordon S
Published: June 2004

An inhibitory rat mAb, SER-4, has been raised to the mouse macrophage (M phi)-restricted hemagglutinin, sheep erythrocyte receptor (SER), which binds unopsonized sheep erythrocytes through recognition of sialylated glycoconjugates. This receptor was originally defined on mouse resident bone marrow M phi where it was implicated in adhesive interactions of these cells with proliferating hematopoietic cells. In the present study using mouse serum-induced thioglycollate-elicited peritoneal M phi (TPM) as a model system for SER expression, mAb SER-4 IgG2a completely blocked rosette formation at 1 microgram/ml. The inhibition was likely to be via steric hindrance rather than through a direct interaction with the putative sialic acid binding site of SER because F(ab')2 and Fab fragments of mAb SER-4 gave a maximum inhibition of 50-60% and 0% respectively, despite binding effectively to the SER-4 antigen (Ag). Immunoprecipitation and Western blotting experiments with cultured M phi or tissue extracts demonstrated that the Ag recognized by SER-4 mAb is a single chain molecule with an apparent Mr by SDS-PAGE of 185 x 10(3) (reduced) or 170 x 10(3) (non-reduced) and is distinct from members of the leukocyte common Ag family. Expression of SER and SER-4 Ag in culture were closely correlated and depended on the presence of mouse serum for optimal induction. Further evidence that the SER-4 Ag is functionally equivalent to SER was provided by immunocytochemistry in which the overall pattern of staining in tissues was consistent with previous rosetting experiments. In the bone marrow, expression of the SER-4 Ag was restricted to the resident bone marrow M phi population with no expression on monocytes. High expression was also observed on stromal M phi within the subcapsular sinus and medullary cords in lymph nodes and on marginal metallophils in the spleen. These results therefore confirm that SER is a novel M phi-restricted receptor whose distribution and properties indicate a role in cellular interactions in hematopoietic and lymphoid tissues.
Human cytomegalovirus induces stage-specific embryonic antigen 1 in differentiating human teratocarcinoma cells and fibroblasts.
Authors: Andrews P W , Gönczöl E , Fenderson B A , Holmes E H , O'Malley G , Hakomori S , Plotkin S
Published: June 2004

Cell surface expression of stage specific embryonic antigen 1 (SSEA-1), or Lex (III3 FucnLC4), was induced in differentiated human teratocarcinoma cells and in human diploid fibroblasts 3-6 d after infection with human cytomegalovirus (HCMV). In parallel, fucosylated lactoseries glycolipids bearing the SSEA-1/Lex epitope were readily detected in the infected cells but not in the uninfected cells. HCMV infection also results in altered expression of several glycosyltransferases. SSEA-1/Lex induction is probably a consequence of both increased expression of beta 1----3N-acetylglucosaminyltransferase, which catalyzes the rate-limiting step in lactoseries core chain synthesis, and subtle alterations in the relative competition for common precursor structures at key points in the biosynthetic pathway. Since SSEA-1 has been suggested to play a role in some morphogenetic cell-cell interactions during embryonic development, the induction of this antigen at inappropriate times might provide one mechanism whereby intrauterine infection with HCMV can damage the developing fetal nervous system.
Active immunization against virus infections due to antigenic drift by induction of crossreactive cytotoxic T lymphocytes.
Authors: Kuwano K , Scott M , Young J F , Ennis F A
Published: June 2004

We have examined whether active immunization with c13 protein, a hybrid protein of the first 81 amino acids of the viral NS1 nonstructural protein and the HA2 subunit of A/PR/8 (H1N1) hemagglutinin, could protect BALB/c mice from challenge with A/PR/8 H1 subtype virus. Mice immunized with the c13 protein had a significant reduction of pulmonary virus titers with A/PR/8 (H1) virus, but failed to limit the replication of A/PC (H3) virus, which reflects the in vitro CTL activity of c13 immune spleen cells. We observed that the epitope recognized by HA2 specific CTL, which are induced by a derivative of c13 protein, is highly conserved among H1 and H2 subtype virus strains. This led us to test whether active immunization with c13 protein would also limit pulmonary virus replication in mice infected with the A/TW virus, a virus of the H1 subtype, which was isolated in 1986, and with a virus of the H2 subtype, A/Japan/305/57. Immunized mice had significantly lower lung virus titers than did control mice, and did not possess any neutralizing antibodies to the challenger viruses. These results indicate that active immunization with a fusion protein containing the cross-reactive CTL epitope protects mice from influenza infection by inducing CTL against influenza A H1 and H2 subtype virus strains, which markedly vary in their antibody binding sites on the HA1. The ability to induce active cross-reactive immunization with a fusion protein which contains a highly conserved CTL epitope offers a model for vaccine approaches against viruses which undergo significant variations in their antibody binding sites.
Early human IgH gene assembly in Epstein-Barr virus-transformed fetal B cell lines. Preferential utilization of the most JH-proximal D segment (DQ52) and two unusual VH-related rearrangements.
Authors: Nickerson K G , Berman J , Glickman E , Chess L , Alt F W
Published: June 2004

We have analyzed the phenotypic characteristics and IgH gene rearrangements in a panel of EBV-transformed B lineage cell lines from human fetal liver and bone marrow. Some lines contained only populations of immature, Ig- Be cells, while others contained mixed populations of mature and immature B cells. The majority of identifiable IgH rearrangements involved joining of the most JH-proximal D segment, DQ52, to various JH segments, implying that DQ52 is a preferred target for initial DJH rearrangements. Three other rearrangements involving VH-related sequences were also characterized. Two involved VHDJH joining using VH3 genes, although one of these had a very unusual DJH structure. The third consisted of inverted 3' signal sequences and flanking regions of a VH4 gene appended to a JH. The mechanisms by which the later rearrangement could have occurred and its potential physiological significance are discussed.
The MHC molecule I-E is necessary but not sufficient for the clonal deletion of V beta 11-bearing T cells.
Authors: Bill J , Kanagawa O , Woodland D L , Palmer E
Published: June 2004

We have generated an mAb, RR3-15, that recognizes murine TCRs containing the V beta 11 domain. Using this antibody to stain peripheral T cells, we have demonstrated that V beta 11-bearing T cells are largely absent from strains of mice that express the class II MHC molecule, I-E. Studies with F1 mice demonstrate that this effect is dominant, consistent with tolerance. The clonal deletion of V beta 11-bearing T cells appears to occur intrathymically, as immature but not mature V beta 11+ T cells are present in the thymus of I-E-bearing mice. Examination of B6 x DBA/2 recombinant inbred strains demonstrates that the expression of I-E molecules is necessary for the clonal deletion of V beta 11-bearing T cells, but that other non-MHC genes control the clonal deletion process, as well. Paradoxically, only a small fraction of V beta 11+ T cell hybridomas are I-E reactive.
A cytotoxic T lymphocyte inhibits acquired immunodeficiency syndrome virus replication in peripheral blood lymphocytes.
Authors: Tsubota H , Lord C I , Watkins D I , Morimoto C , Letvin N L
Published: June 2004

CD8+ (suppressor/cytotoxic) lymphocytes block replication of HIV-1 or the simian immunodeficiency virus of macaques (SIVmac) in PBL of infected individuals. We now show that these CD8+ lymphocytes undergo clonal expansion in vivo after AIDS virus infection of the individual, suggesting they may be antigen-specific T cells. These CD8+ cells block replication of virus in autologous but not MHC class I-mismatched PBL. The inhibitory lymphocytes express the integrin family molecule 4B4 and the CTL-associated S6F1 epitope of LFA-1. Finally, physical contact is required for the CD8+ lymphocyte-mediated inhibition of AIDS virus replication, since this inhibitory function is blocked by anti-LFA-1 and anti-CD8 mAbs. These studies suggest that the cell that inhibits AIDS virus replication in PBL of infected individuals is a CTL.
Purification and amino acid analysis of two human glioma-derived monocyte chemoattractants.
Authors: Yoshimura T , Robinson E A , Tanaka S , Appella E , Kuratsu J , Leonard E J
Published: June 2004

Two chemoattractants for human monocytes were purified to apparent homogeneity from the culture supernatant of a glioma cell line (U-105MG) by sequential chromatography on Orange A-Sepharose, an HPLC cation exchanger, and a reverse phase HPLC column. On SDS-PAGE gels under reducing or nonreducing conditions, the molecular masses of the two peptides glioma-derived chemotactic factor 1 and 2 were 15 and 13 kD, respectively. Amino acid composition of these molecules was almost identical, and differed from other cytokines that have been reported. The NH2 terminus of each peptide was apparently blocked. When tested for chemotactic efficacy, the peptides attracted approximately 30% of the monocytes added to chemotaxis chambers, at the optimal concentration of 10(-9) M. Potency and efficacy were comparable with that of FMLP, which is often used as a reference attractant. The activity was chemotactic rather than chemokinetic. In contrast to their interaction with human monocytes, the pure peptides did not attract neutrophils. These pure tumor-derived chemoattractants can now be compared with attractants produced by normal cells and evaluated for their biological significance in human neoplastic disease.
A subset of memory CD4+ helper T lymphocytes identified by expression of Pgp-1.
Authors: Butterfield K , Fathman C G , Budd R C
Published: June 2004

The Pgp-1 glycoprotein (Ly-24 antigen) is acquired by mature murine T lymphocytes at the time of primary antigen stimulation Pgp-1 was previously shown to be a useful cell surface marker for distinguishing antigen-specific memory CD8+ T lymphocytes after immunization. Here we demonstrate that this observation extends to CD4+ T lymphocytes. Antigen-specific CD4+ T cells in mice immunized with sperm whale myoglobin or keyhole limpet hemocyanin were contained nearly exclusively in the minor Pgp-1+ subset.
L3T4 antigen expression by hemopoietic precursor cells.
Authors: Frederickson G G , Basch R S
Published: June 2004

L3T4 (CD4) is expressed on immature hematopoietic cells. Sorting bone marrow cells on the basis of their expression of this antigen produces populations of cells that are markedly enriched for multipotential stem cells (CFU-s) and for myeloid precursors (CFU-c). We believe that L3T4 is transiently expressed by most, if not all, hematopoietic precursors early in their maturation. We suggest that the expression of CD4 molecules on the surface of immature precursors is required for their interaction with Ia bearing cells within the hemopoietic inductive microenvironment(s) of the marrow and thymus.
Purification and characterization of a novel monocyte chemotactic and activating factor produced by a human myelomonocytic cell line.
Authors: Matsushima K , Larsen C G , DuBois G C , Oppenheim J J
Published: June 2004

A novel basic heparin-binding monocyte chemotactic factor (MCF) was purified to homogeneity from the conditioned media of human myelomonocytic cell line THP-1 based on its in vitro monocyte chemotactic activity. The purified MCF was homogenous and estimated to be 15 kD on SDS-PAGE. Purified MCF stimulated normal human monocytes to be growth inhibitory in vitro at 2-3 d for several human tumor cell lines. This represents the first report of the identification and purification of a chemoattractant cytokine that also activates monocytes but is distinct from interferons and other known cytokines.
H-2-linked genes influence the severity of herpes simplex virus infection of the peripheral nervous system.
Authors: Simmons A
Published: June 2004

Infection of the peripheral nervous system was studied after inoculation of HSV into the flank skin of H-2 congenic mice. The amount of virus recovered from the sensory ganglia varied significantly between the mouse strains tested. Differences became apparent 7 d after infection, at which time the severity of disease in H-2k mice was two to three orders of magnitude greater than that in H-2d animals. The association of the H-2k haplotype with impaired ability to clear HSV from the nervous system is the first clear demonstration that genes within the MHC can influence the severity of primary herpetic infection, in spite of numerous studies on genetic resistance to this disease.
Human monocytes can produce tissue-type plasminogen activator.
Authors: Hart P H , Vitti G F , Burgess D R , Singleton D K , Hamilton J A
Published: June 2004

Evidence has previously been presented that monocytes and macrophages produce urokinase-type plasminogen activator. We have shown for the first time that human monocytes, when stimulated appropriately in vitro, can produce tissue type-plasminogen activator (t-PA) of 70 kD. Detection of t-PA mRNA was consistent with the biochemical and immunological characterization of t-PA produced by human monocytes.
Human T cell clones define S1 subunit as the most immunogenic moiety of pertussis toxin and determine its epitope map.
Authors: De Magistris M T , Romano M , Bartoloni A , Rappuoli R , Tagliabue A
Published: June 2004

Human T lymphocyte clones specific for pertussis toxin (PT) were used to analyze the fine specificity of the response to PT, the basic component of new acellular vaccines against whooping cough. The majority (83%) of the clones specific for PT recognized S1, the subunit that in animal models has been shown to be highly immunogenic. To map T cell epitopes on S1, 18 S1-specific clones were tested for recognition of recombinant fragments representing NH2-terminal and COOH-terminal deletions of S1 and two recombinant S1 subunits containing amino acid substitutions. This approach led to the identification of three regions of the protein as the sequences containing T cell antigenic sites: 1-42, 181-211, and 212-235. Synthetic peptides were eventually used for a finer localization of the T cell epitopes. Two peptides, one of 13 residues (27-39) at the NH2 terminus and one of 24 residues (171-194) at the COOH terminus, stimulated proliferation of three and four clones, respectively. Both peptides are recognized in association with HLA DR1 molecules. These results stress the role of S1 in the immune response to PT and provide data useful for the development of a recombinant or synthetic antipertussis vaccine containing T cell epitopes from S1.
Analysis of V beta 17a expression in new mouse strains bearing the V beta a haplotype.
Authors: Kappler J W , Kushnir E , Marrack P
Published: June 2004

A set of new mouse strains were produced that carry the V beta a haplotype of the TCR-alpha/beta and any of a number of different H-2 haplotypes on backgrounds derived from related C57BL, C57L, and C57BR mice. Study of V beta 17a expression in these mice confirms the association between the presence of IE and the deletion of V beta 17a+ T cells. A second H-2 gene causing deletion of V beta 17a+ T cells was mapped in these mice to the K end of H-2k, and H-2 influences on the level of selection of CD4+ V beta 17a+ T cells were indicated.
Nitric oxide. A macrophage product responsible for cytostasis and respiratory inhibition in tumor target cells.
Authors: Stuehr D J , Nathan C F
Published: June 2004

A metabolic pathway of activated macrophages (M phi) involving oxidation of the guanido nitrogens of L-arginine is required for inhibition of growth and respiration of some target cells. The goal of this study was to identify the M phi metabolite(s) that induce these injuries. The stable products of the L-arginine pathway, NO2- and NO3-, were incapable of causing cytostasis under coculture conditions. However, NO2- became cytostatic upon mild acidification, which favors its transformation into nitrogen oxides of greater reactivity. This suggested that NO. (and/or NO2), recently identified as an M phi metabolite of L-arginine, could be a mediator. Authentic NO. caused cytostasis and respiratory inhibition in L1210 cells in a dose-dependent manner. The mitochondrial lesions caused by NO. were confined to complex 1 and 2, a pattern of injury identical to that seen after coculture with activated M phi. Inclusion of NO. scavenger systems prevented cytostasis from developing in M phi-L1210 cocultures. Thus, M phi-generated NO. can account for L-arginine-dependent cytostasis and respiratory inhibition.
Cytotoxic T lymphocytes specific for self tumor immunoglobulin express T cell receptor delta chain.
Authors: Wright A , Lee J E , Link M P , Smith S D , Carroll W , Levy R , Clayberger C , Krensky A M
Published: June 2004

CTL are thought to play a role in the elimination of transformed cells in vivo. The effectiveness of such CTL is in part dependent on recognition of tumor specific antigens. Among the best characterized tumor-specific antigens are the unique or idiotypic determinants on the Ig of B cell lymphomas. Here we describe the generation and properties of human CTL specific for the idiotype on autologous B cell tumors. These cells are CD3+,CD4-,CD8- and express the delta chain of the TCR. Such cells may prove useful in tumor-specific adoptive therapy.
Analysis of naturally occurring delayed-type hypersensitivity reactions in leprosy by in situ hybridization.
Authors: Cooper C L , Mueller C , Sinchaisri T A , Pirmez C , Chan J , Kaplan G , Young S M , Weissman I L , Bloom B R , Rea T H
Published: June 2004

Analysis of tissue lesions of the major reactional states of leprosy was undertaken to study the immune mechanisms underlying regulation of cell-mediated immunity and delayed-type hypersensitivity (DTH) in man. In situ hybridization hybridization of reversal reaction biopsy specimens for INF-gamma mRNA expression revealed a 10-fold increase in specific mRNA-containing cells over that observed in unresponsive lepromatous patients. Expression of huHF serine esterase, a marker for T cytotoxic cells, were fourfold increased in reversal reaction and tuberculoid lesions above that detected in unresponsive lepromatous individuals. Immunohistology of reversal reactions confirmed a selective increase of Th and T cytotoxic cells in the cellular immune response. Of interest, the microanatomic location of these serine esterase mRNA-containing cells was identical to the distribution of CD4+ cells. Analysis of erythema nodosum leprosum (ENL) lesions revealed differences in the underlying immune processes in comparison with reversal reaction lesions. Although phenotypic Th cells predominated in ENL lesions, IFN-gamma and serine esterase gene expression were markedly reduced. We suggest that reversal reactions represent a hyperimmune DTH response characterized by a selective increase of CD4+ IFN-gamma producing cells and T cytotoxic cells, which result in the clearing of bacilli and concomitant tissue damage. In contrast, ENL reactions may be viewed as a transient diminution of Ts cells and activity leading to a partial and transient augmentation in cell-mediated immunity, perhaps sufficient to result in antibody and immune complex formation, but insufficient to clear bacilli from lesions.
A shared idiotype among human anti-Ro/SSA autoantibodies.
Authors: Gaither K K , Harley J B
Published: June 2004

Idiotypes and antiidiotypes are thought to be important immune regulators and have provided clues for the origin and pathogenicity of autoantibodies. Many lupus and Sjögren's syndrome patients, as well as most neonatal lupus infants with congenital heart block or dermatitis, have antibodies to the ribonucleoprotein Ro/SSA, which is one of a group of RNA-protein autoantigens commonly found in human lupus sera. To characterize the fine specificity of anti-Ro/SSA antibodies, a rabbit antidiotypic serum was prepared against polyclonal affinity purified anti-Ro/SSA F(ab')2. The resulting antiidiotype, anti-Id-Rol, is specific for the F(ab')2 fraction of the anti-Ro/SSA immunogen and its binding to anti-Ro/SSA is inhibited by purified Ro/SSA. These data indicate that the Id-Rol epitope on anti-Ro/SSA is associated with the antigen binding site of these same antibodies. The Id-Rol idiotype was present by ELISA in 3 of 12 additional anti-Ro/SSA preparations from precipitin-positive donor sera and in anti-Ro/SSA from one normal donor with low level antibody. This is the first shared idiotype to be found in the human autoantibodies binding to this RNA-protein antigen. Idiotypic differences between anti-Ro/SSA autoantibodies have the potential to explain the variation in pathologic associations found in individuals who develop this autoantibody specificity.
A branched, synthetic octapeptide of ubiquitinated histone H2A as target of autoantibodies.
Authors: Plaué S , Muller S , van Regenmortel M H
Published: June 2004

Two peptides of eight (T2) and 10 (T1) residues corresponding to the branched moiety of ubiquitinated histone H2A have been synthesized and used for raising specific antibodies in rabbits. Antisera to peptide T1 reacted in ELISA with T1 and with H2A but not with ubiquitin; antisera to peptide T2 reacted with T2 but not with H2A or ubiquitin. When tested in immunoblotting, both peptide antisera reacted with ubiquitinated H2A but not with unconjugated H2A or with ubiquitin. Sera from patients with systemic lupus erythematosus (SLE) were shown previously to react with ubiquitin in ELISA and immunoblotting. When tested for their ability to react in ELISA with synthetic peptides T1 and T2, 96% of the SLE sera (diluted 1:500) that recognized ubiquitin also reacted with peptide T2. Of the SLE sera that did not react with ubiquitin, only 13% possessed antibodies able to bind peptide T2. Antibodies from seven SLE sera, purified on a T2-immunoadsorbent column, were also able to react either with H2A, and in three cases also with ubiquitin.
Relationship of variable region genes expressed by a human B cell lymphoma secreting pathologic anti-Pr2 erythrocyte autoantibodies.
Authors: Silberstein L E , Litwin S , Carmack C E
Published: June 2004

To study the biology of cold agglutinin disease we previously established EBV-transformed B cell clones isolated from a patient with splenic lymphoma of an early plasmacytic cell type and immune hemolysis due to an anti-Pr2 cold agglutinin. These clones had an aberrant chromosomal marker identical to the patient's B cell lymphoma and each secreted IgMk anti-Pr2 similar to the pathologic autoantibody in the serum of the patient. In this study, we have further investigated the Pr2-specific autoimmune response through nucleotide sequencing of VH and VL region genes. We have shown that the seven clones share the same VDJ/VJ gene segments and junctional elements confirming their clonal origin. The VH sequences were 88% homologous to a VHI germline gene while the VL sequences were 97% homologous to a VkIII germline gene. Only 4 somatic mutations (3 silent and 1 conservative) were found in greater than 5,000 bp sequenced, suggesting that a low mutation rate existed. Based on a tumor mass of 10(12) cells and a minimum of 40 divisions, we estimated the somatic mutation rate to be 4.45 x 10(-5) m/bp/d. This somatic mutation rate is similar to those estimated for acute lymphocytic leukemia (pre-B cell) and chronic lymphocytic leukemia (intermediate B cell), but significantly lower than the mutation frequency in follicular lymphomas (activated B cell). We propose that the difference in somatic mutation frequency of a B cell tumor may be related to the stage of B cell differentiation. In addition, the low mutation frequency observed in the Pr2-specific B cell tumor may also reflect, in part, selection by autoantigen to conserve sIg structure and specificity.