摘要:

AbstractWe have previously reported 2 cases of healthy men showingin vivomonoclonal expansion of mature CD4−CD8−αβ T cells. In the present study, an additional 3 adults were found to exhibit such an expansion, among a total 464 adult donors studied. These 5 individuals were otherwise physiologically normal, with no history of severe illness and autoimmune disease at the time of examination. To investigate the mechanisms of the clonal expansion, further characterization of the clonal cells was attempted. No apparent preference for usage of the Tcell receptor β chain variable region was observed in the clonal T cells. These clonal T cells showed lectin‐dependent or redirected antibody‐dependent cell‐mediated cytotoxicities, whereas they could not lyse autologous lymphoblastoid cell lines. Failure of Fas antigen expression was not observed for any of these clones. These results suggest that clonal expansion of CD4−CD8−αβ T cells frequently occurs in the periphery without any T

ISSN:0014-2980    

DOI:10.1002/eji.1830231102

出版商:WILEY‐VCH Verlag GmbH    

年代:1993

数据来源: WILEY

摘要:

AbstractThe enzyme‐linked immunospot (ELISPOT) assay has been proven to be an efficient and sensitive method for the enumeration of single cells secreting antibodies or cytokines. Here we have used this method to determine the number of interleukin‐4 (IL‐4)‐ and interferon‐γ (IFN‐γ)‐producing cells inin vitrosecondary responses to tetanus toxoid (TT) and the mycobacterial antigen (purified protein derivative; PPD) or the mitogen phytohemagglutinin (PHA). PHA‐induced IL‐4 and IFN‐γ secretion was well correlated suggesting polyclonal activation of cells. This was not the case with the specific antigens, where PPD preferentially induced IFN‐γ‐ and very few IL‐4‐producing cells, while TT‐induced both IL‐4 and IFN‐γ. These differences are probably a reflection of the types of immunity the two antigens induce, mycobacteria preferentially inducing a cell‐mediated T helper type 1 (Th 1) type of immunity, while immunity to tetanus is an antibody‐dependent, Th 2 type of response.In individuals recently boosted with TT, a significant increase in both IL‐4‐ and IFN‐γ‐producing cells in response to TT was seen at day 7 after boost, followed by decline. This was in contrast to what was seen in response to PPD where an increase of IFN‐γ‐producing cells after the TT boost at day 7 persisted for at least 14 days. These results suggest that after anin vivoboost both antigen‐specific and nonspecific T cells are activated and that antigen‐specific cells home to other organs and therefore may be difficult to demonstrate in the circulation. Our data show that the ELISPOT assay is a powerful tool for determining the frequency of cells secreting cytokines. The assay has several advantages over other assays since it is sensitive, measures the number of actually secreting cells, and avoids the problems of b

ISSN:0014-2980    

DOI:10.1002/eji.1830231103

出版商:WILEY‐VCH Verlag GmbH    

年代:1993

数据来源: WILEY

摘要:

AbstractMajor histocompatibility complex (MHC) class I and II products are specialized to present antigens via different intracellular processing routes. Peptides originating from proteins in the cytoplasm can gain access to class I peptide‐binding grooves, most likely in the rough endoplasmic reticulum. Peptides from proteins in acidic endocytic vacuoles gain access to class II. It has been proposed that MHC class I products also can capture peptides from “exogenous” or noninfectious sources, and this assumption underlies the use of intact proteins as vaccines for CD8+cytotoxic T lymphocytes. Here we describe quantitative information comparing the efficacy of peptide presentation from exogenous proteins by administering a class I‐ and II‐restricted peptide within the same context, the CDR3 loop of the VHdomain of a self immunoglobulin. Antigen‐presenting cells (APC), including primary dendritic cells, efficiently present an influenza hemagglutinin peptide from the immunoglobulin (Ig) carrier (50% maximal response at 10 nM Ig‐HA) to an MHC class II‐restricted T cell. In contrast, these same APC are unable to present an influenza nucleoprotein (NP) peptide from the same context (1 μM Ig‐NP) to an MHC class I‐restricted T cell. Ig‐NP DNA transfectants do present the nucleoprotein viral peptide on class I. Thus, peptides within the complementarity‐determining region loops of Ig carriers can be presented on class I or II MHC products, but the endocytic compartment, when offered MHC class I‐ and II‐restricted peptides within the same carrier protein context, favors presentation by cl

ISSN:0014-2980    

DOI:10.1002/eji.1830231104

出版商:WILEY‐VCH Verlag GmbH    

年代:1993

数据来源: WILEY

摘要:

AbstractThe monoclonal antibodies (mAb) 15/7 and 24 recognize unique activation‐dependent, conformational epitopes on β1 and β 2‐integrins, respectively. The expression of both of these epitopes closely correlates with the ligand binding ability of their respective integrins, and thus serves as indicators of functional integrin “activation”. Here, we have used six‐parameter flow cytometry to examine the expression of these epitopes and conventional β1‐ and β2‐integrin epitopes during human T cell activation in secondary lymphoid tissuesin vivo, focusing particularly on the virgin to memory/effector cell transition. Fresh tonsil lymphocytes were stained with mAb against conventional or activation‐dependent integrin epitopes, followed by staining with mAb against CD3, CD45RA, and CD45RO, thus allowing the determination of integrin epitope expression on virgin (CD3+) T cells (CD45RA+/RO−to±), memory/effector (CD45RA−/RO++) T cells, and T cells undergoing the virgin to memory/effector transition: transition region‐1 (T1; CD45RA+to++/RO+); ‐2 (T2; CD45RA++/RO++); and ‐3 (T3; CD45RA+/RO++). Conventional β1‐ and β2‐integrin epitopes progressively increase during the virgin to T3 stages of the transition in tonsil, in keeping with the generally higher levels of these adhesion molecules on memory/effectorvs.virgin T cells. Expression of both the β1 (15/7)‐and β2 (24)‐integrin activation epitopes first appears on transitional T cells, and is maintained on a relatively constant number of cells (averaging 25‐30%) throughout the T1‐T3 stages. These epitopes are also noted on a subset of activated memory/effector T cells. Importantly, on both transitional and activated memory/effector T cell subsets, the expression patterns of the 15/7 and 24 epitopes vs. a variety of T cell activation antigens are identical, and the expression of these epitopes relative to each other is linearly correlated, findings strongly supporting the coordinate activation of β1 and β2 integrins duringT cell activationin vivo.These results provide the first evidence of integrin activation during anin vivoimmunologic response, and demonstrate the usefulness of mAb recognizing conformational epitopes and multiparameter flow cytometry in delineating the dynamic interplay of adhesio

ISSN:0014-2980    

DOI:10.1002/eji.1830231105

出版商:WILEY‐VCH Verlag GmbH    

年代:1993

数据来源: WILEY

摘要:

AbstractExpression of Vβ (β ‐chain variable region) gene segments was investigated in theMus m. domesticusDDO strain, which possesses a large genomic deletion encompassing 20 of the 29 Vβ gene segments known in BALB/c. Stainings using Vβ ‐specific monoclonal antibodies revealed that up to 60% of the peripheral T cells use 3 Vβ gene segments. Variable frequencies of Vβ 2 T cells were observed among DDO individuals. Segregation analyses of F2 crosses between Vβ 2‐deletor mice and mammary tumor virus (Mtv)‐free mice led to the identification of a new endogenous Mtv, named Mtv‐DDO, mediating Vβ 2 Tcell clonal deletion. Mtv‐DDO structure is conserved with the exception of the carboxy‐terminal region as compared to other Mtv. Comparison between Mtv sharing the same Vβ specificity and isolated from laboratory or wild mice confirms that a stretch of 11 amino acids, defined as the Vβ ‐specific region, is required for the Vβ ‐specific interaction. Limited substitutions in this region account for the shift of the Mtv spec

ISSN:0014-2980    

DOI:10.1002/eji.1830231106

出版商:WILEY‐VCH Verlag GmbH    

年代:1993

数据来源: WILEY

摘要:

AbstractWe have previously characterized an infectious mouse mammary tumor virus [(MMTV(SW)] which induces a strong superantigen responsein vivo. Here we describe the isolation and characterization of MMTV(C4) which was derived from milk of mice implanted with hyperplastic alveolar nodules. MMTV(C4) stimulates Vβ2 expressing T cells after local injectionin vivo. Comparison with known open reading frame (orf) sequences revealed high homology to Mtv‐6, an endogenous virus interacting with Vβ3‐expressing T cells. The carboxyl‐terminal amino acids were, however, altered. High homology including the carboxyl‐terminal orf amino acids were found with MMTV(C3H‐K). We show here that MMTV(C3H‐K) has lost its superantigen function. Sequence comparisons permitted the characterization of few key amino acids which could be important for T cell receptor interaction and superantig

ISSN:0014-2980    

DOI:10.1002/eji.1830231107

出版商:WILEY‐VCH Verlag GmbH    

年代:1993

数据来源: WILEY

摘要:

AbstractA murine solid tumor was transfected to express various levels of an allogeneic major histocompatibility complex class I gene (K216), in order to test the effect of the level of antigen expression on immunogenicity and sensitivity to lysis by cytotoxic T lymphocytes (CTL). The growth rates of clones of tumor cells expressing different levels of the transfected gene were similarin vitroand in nude mice. Although all tumor cells, including cells freshly isolated from growing tumors, were equally sensitive to lysis by specific CTL, only tumor cells expressing the highest level of the K216antigen stimulated CTL and were rejected by normal mice. In contrast, tumor cells expressing lower levels of antigen failed to immunize for CTL and grew progressively in normal mice, despite retaining expression of the transfected gene and remaining fully sensitive to CTL‐mediated lysis; thus, the threshold of antigen needed to stimulate CTL responses was considerably higher than that needed to lyse tumor cells. Reduction of K216antigen expression from 100‐fold to 40‐fold above background, impaired significantly the ability of the tumor cells to induce a K216‐specific immune response, while tumor cells expressing K216at levels 2‐fold above background were as susceptible to CTL‐mediated lysis as tumor cells expressing 50‐fold more antigen. The important implication of these findings is that some tumors occurring in nature may not be immunogenic but nevertheless express antigens which are potential targets for i

ISSN:0014-2980    

DOI:10.1002/eji.1830231108

出版商:WILEY‐VCH Verlag GmbH    

年代:1993

数据来源: WILEY

摘要:

AbstractThe immunodominant H‐2Dd‐restricted cytotoxic T lymphocyte (CTL) response to the HIV‐1 gp160 envelope glycoprotein maps to a single determinant in the V3 loop, designated p18. Using a series of peptides synthesized on pins we have determined that the minimal core sequence of this determinant required for CTL recognition comprises 8 amino acids (residues 320‐327). However, 9mer and l0mer peptides containing this core sequence were more effective than the 8mer peptide at sensitizing Dd‐expressing target cells. To analyze the antigenicity of endogenously synthesized p18, minigenes encoding a 10‐amino acid determinant (residues 318‐327) and a 67‐amino acid peptide (residues 281‐348; containing the V3 loop) were expressed using vaccinia virus (Vac) recombinants. Both peptides were as effective as wild‐type gpl60 in their ability to sensitize target cells for lysis by gpl60‐specific CTL. Immunization of BALB/c mice with Vac recombinants encoding both gp160 peptides elicited gp160‐specific CTL. These data demonstrate that both the V3 loop itself and a 10‐residue epitope are sufficient to prime CTLin vivoand strongly support the potential use of minigene‐encoded CTL epitopes for recombinant vaccines designed to induce protective T cell‐me

ISSN:0014-2980    

DOI:10.1002/eji.1830231109

出版商:WILEY‐VCH Verlag GmbH    

年代:1993

数据来源: WILEY

摘要:

AbstractThe present study investigated possible receptor‐like characteristics of glycosyl‐phosphatidylinositol (GPI)‐linked antigens on human monocytes and granulocytes by measuring cytoplasmic calcium fluxes and the oxidative burst in cells following cross‐linking of GPI‐linked antigens. Cross‐linking of cell‐bound anti‐CD14, ‐CDw52 and ‐CD55 induced cytoplasmic calcium fluxes and oxidative bursts in unprimed human monocytes similar to those observed following FcyR cross‐linking. In granulocytes primed with 200 nM N‐formyl‐Met‐Leu‐Phe (FMLP), cross‐linking of cell‐bound anti‐CD16, ‐CD24, ‐CD59 and ‐CD67 led to calcium fluxes and activation of the oxidative burst. The oxidative bursts mediated by GPI‐linked antigens were stronger than those induced by 200 nM FMLP, even though FMLP induced a larger increase in cytoplasmic calcium concentration. The responses were likely to be independent of FcyR interactions as F(ab′)2 fragments of IgG or IgM antibodies were used in the experiments. Activating effects of monoclonal antibody to GPI‐linked antigens were not observed in cells from patients with paroxysmal nocturnal hemoglobinuria, which are deficient in GPI‐linked antigens. In addition, treatment with GPI‐specific phospholipase C led to inhibition of cell activation through GPI‐linked antigens but not through transmembrane receptors. Cross‐linking of a number of non‐GPI‐linked antigens (CD11a, CD18, CD31, CD35, CD43, and CD45) neither induced calcium fluxes, nor activated the oxidative burst. The results indicate that most, if not all, GPI‐linked surface glycoproteins on myeloid cells are capable of mediating cell activation and suggest that t

ISSN:0014-2980    

DOI:10.1002/eji.1830231110

出版商:WILEY‐VCH Verlag GmbH    

年代:1993

数据来源: WILEY

摘要:

AbstractLeukocyte adhesion deficiency (LAD) is an autosomal recessive disease caused by heterogeneous mutations within the gene encoding the common β subunit (CD18) of the three leukocyte integrins LFA‐1 (CD11a/CD18), Mac‐1 (CD11b/CD18), and pl50,95 (CD11c/CD18). Based on the level of expression of CD18 on patient leukocytes, two phenotypes of LAD have been defined (severe and moderate) which correlate with the severity of the disease. We have investigated the molecular basis of the disease in two unrelated severe patients (HS and ZJO). Both patients share a complete absence of CD18 protein precursor and cell surface expression, but they differ in the level of CD18 mRNA, which is normal in HS and undetectable by Northern blot in ZJO. Determination of the primary structure of the patient HS CD18 mRNA revealed a 10‐base pair deletion between nucleotides 190‐200 (CD 18 exon 3), which eliminates residues 41–43 and causes a frameshift into a premature termination codon 17 base pairs downstream from the deleted region. The 10‐base pair frameshift deletion maps to a region of the CD18 gene where aberrant mRNA processing has been detected in HS and two other unrelated LAD patients. In the ZJO patient, amplification of lymphoblast CD 18 mRNA demonstrated the presence of a non‐sense mutation in the third nucleotide of the triplet encoding Cys534(TGC → TGA), within exon 12. Both genetic abnormalities were also detected at the genomic level, and affect the restriction pattern of their corresponding genes, thus enabling the detection of the mutant alleles among healthy heterozygous alleles in family studies. The identification of two new LAD CD18 alleles, either carrying a non‐sense mutation (ZJO) or a partial gene deletion (HS), further illustrates the heterogeneity of the genetic

ISSN:0014-2980    

DOI:10.1002/eji.1830231111

出版商:WILEY‐VCH Verlag GmbH    

年代:1993

数据来源: WILEY