Search PubMed⌕ Search

Biomedical subjects

V L Braciale

Publications and source records attributed to V L Braciale.

At least 37 records · Page 2Linked to original sources
PubMed1989

Class I major histocompatibility complex-restricted T lymphocyte recognition of the influenza hemagglutinin. Overlap between class I cytotoxic T lymphocytes and antibody sites.

The influenza hemagglutinin is a critical regulator of disease expression during influenza virus infection and serves as a major target for the host immune response to this pathogen. In this report, we have analyzed an immunodominant site on the hemagglutinin (residues 202-221) recognized by murine class I MHC-restricted T lymphocytes. This analysis has revealed evidence for the duplication of a T cell recognition site within the region 202-221. We have also identified critical amino acids necessary for class I-restricted T cell recognition within these two epitopes. In addition, we provide evidence that a site on the influenza hemagglutinin recognized by neutralizing antibody directly overlaps with an epitope recognized by class I MHC-restricted CTL.

Amino Acid Sequence↗
PubMed1989

Regulation of T lymphocyte proliferation. Interleukin 2-mediated induction of c-myb gene expression is dependent on T lymphocyte activation state.

We previously reported that with time, after antigenic stimulation of antigen-regulated murine T lymphocyte clones, total IL-2-R expression decayed 10-50-fold, commensurate with a decline in the ability of the cells to proliferate to IL-2. However, late after antigenic stimulation, when the cells were refractory to the IL-2-proliferative stimulus, high levels of high affinity IL-2-R remained. In this report we further explore the basis of unresponsiveness to IL-2 in the quiescent clones. We show that the proto-oncogene c-myc is induced in the late cell population by IL-2 to comparable levels observed early after antigen stimulation. IL-2-dependent c-myb induction, however, is seen only early after activation but not in the late-activated population. Analysis of the IL-2-dependent expression of c-myb mRNA with time after antigenic stimulation showed that steadystate c-myb expression declines dramatically with kinetics closely paralleling a decay in IL-2-dependent proliferative ability. In contrast, steadystate c-myc expression remains high throughout this period. Expression of c-myb is critical for proliferation of these cells since antisense oligodeoxy-nucleotide to c-myb can inhibit their IL-2-dependent proliferation. We present evidence for a pathway of c-myb induction via the TCR that is independent of the IL-2/IL-2-R interaction. In addition, the inhibition of IL-2-R-induced c-myb expression by 2-aminopurine and enhanced induction of c-myb via the TCR demonstrate that TCR activation and IL-2-R activation lead to induction of c-myb by different mechanisms.

Animals↗
PubMed1989

Class I major histocompatibility complex-restricted cytolytic T lymphocytes recognize a limited number of sites on the influenza hemagglutinin.

Two distinct regions of the influenza A/JAP/305/57 hemagglutinin molecule are identifiable as sites recognized by murine class I major histocompatibility complex (MHC) (H-2d)-restricted cytolytic T lymphocytes (CTL) generated in response to immunization with infectious type A influenza virus. Each of these sites can be mimicked by a synthetic oligopeptide of approximately 20 amino acids. Data presented herein indicate that these two sites define the dominant immunogenic epitopes on the hemagglutinin recognized by H-2Kd-restricted CTL. These same sites are not efficiently recognized by hemagglutinin-specific class I MHC-restricted CTL of several unrelated MHC haplotypes. These observations show that even for a large complex glycoprotein molecule like the influenza hemagglutinin, only a limited number of class I CTL recognition sites are generated in the infected cell and that the subset of immunogenic epitopes is dependent on the MHC haplotype of the responding individual. These parameters need to be considered in the design of synthetic and recombinant vaccines.

Amino Acid Sequence↗
PubMed1988

Class I MHC-restricted recognition of cells expressing a gene encoding a 41 amino acid product of the influenza hemagglutinin.

Class I H-2Kd-restricted influenza hemagglutinin (HA)-specific CTL recognize two immuno-dominant sites as represented by synthetic peptides spanning epitopes located in the HA1 and hydrophobic transmembrane domains of the influenza HA. Using a vaccinia virus recombinant expression system, we have examined CTL recognition of HA deletion mutants expressed in target cells. We have demonstrated that a truncated influenza HA gene encoding only the transmembrane anchor region containing a class I recognition site and a short segment of the cytoplasmic tail of the HA can be efficiently presented to class I CTL. These results set the stage for detailed analyses of the intracellular events associated with Ag presentation to class I CTL and offer novel possibilities for future vaccine design.

Amino Acid Sequence↗
PubMed1988

Antigen form influences induction and frequency of influenza-specific class I and class II MHC-restricted cytolytic T lymphocytes.

The induction of class I and class II MHC-restricted CTL in response to different forms of A/JAP/57 influenza virus was compared. Splenocytes removed from influenza-immune BALB/c mice and stimulated in vitro with infected syngeneic splenocytes are mainly CD8+ (Lyt-2+) and specifically lyse infected Ia- and Ia+ target cells. To a lesser extent they also lyse non-infectious virus-pulsed Ia+ but not Ia- target cells. In contrast, syngeneic stimulators pulsed with non-infectious virus (exogenous Ag) induce effector T cells that specifically lyse both infected and non-infectious virus-pulsed Ia+ target cells. The cells present in this heterogeneous culture predominantly express the CD4 (L3T4) cell surface marker. Frequency analysis by limiting dilution of splenocytes derived directly from influenza-immune mice revealed a similar pattern of precursor induction: In vitro stimulation with infected splenocytes yielded primarily class I MHC-restricted CTL, whereas stimulation with non-infectious virus reciprocally induced primarily class II MHC-restricted CTL. Thus, the Ag form and consequently the intracellular route of viral Ag presentation profoundly influence the MHC restriction of CTL precursors induced.

Animals↗
PubMed1987

On the role of the transmembrane anchor sequence of influenza hemagglutinin in target cell recognition by class I MHC-restricted, hemagglutinin-specific cytolytic T lymphocytes.

We have examined the requirement for the transmembrane hydrophobic anchor sequence of the influenza hemagglutinin (HA) in the formation of the antigenic moiety on the surface of target cells recognized by class I MHC-restricted murine CTL. For this analysis we have used a line of CV-1 monkey epithelial cells that express the transfected murine H-2Kd gene product as target cells and have used recombinant SV40-based late replacement vectors to achieve expression of genes encoding wild-type and mutant forms of HA. We have found that the majority of Kd-restricted HA-specific CTL clones recognize target cells that express a secreted HA molecule that lacks the transmembrane and cytoplasmic domains of the parent glycoprotein. Several Kd-restricted CTL clones that recognize subtype-specific and crossreactive epitopes on HA fail to recognize the anchor-negative, secreted HA or chimeric HA molecules containing the transmembrane and cytoplasmic domains of unrelated glycoproteins. These CTL clones appear to be directed to antigenic epitopes located within the transmembrane domain of HA, as defined by their capacity to recognize target cells sensitized with a synthetic 23-amino-acid peptide corresponding to sequences within this domain. The implications of these results for class I MHC-restricted CTL recognition are discussed.

Amino Acid Sequence↗
PubMed1987

Lack of IL 2-dependent proliferation despite significant expression of high-affinity IL 2 receptor on murine cytolytic T lymphocyte clones late after antigenic stimulation.

In this study, we examine the expression of IL 2 receptors on class I and class II MHC-restricted, influenza-specific murine T lymphocyte clones at early (day 3) and late (day 8 to day 12) times after antigenic stimulation. IL 2 receptor expression on the three clones examined increases to peak levels early and subsequently decays 10-fold to 50-fold during this time period, as evidenced by monoclonal anti-IL 2 receptor antibody binding. However, in IL 2 binding site studies these clones retain high levels of high-affinity IL 2 receptors (46 to 97% of day 3 levels) at the later time points, despite their inability to proliferate in response to IL 2 in the form of supernatant from Con A-stimulated rat splenocytes. Considerable clone to clone variation is seen in binding site affinity for IL 2, whereas no significant change in binding affinity for IL 2 is exhibited for a particular clone as a function of time after activation, suggesting little structural change in the IL 2 receptor with time. Contrary to this finding, Con A-stimulated murine splenocytes exhibit a sharp decay in IL 2 receptor expression reaching 25% of peak (day 3) levels by day 10 after stimulation by lectin, with a significant decrease in the average binding site affinity for IL 2 in the population. This change in affinity in the Con A-stimulated population may, however, reflect a selection with time for lymphocytes with lower affinity for IL 2. To elucidate where the potential block may be that prevents these CTL clones from proliferating in an IL 2-dependent manner, the ability of the cells to internalize bound IL 2 at late times after activation is examined. All three clones late after activation are able to internalize bound IL 2 with efficiencies equivalent to that seen at day 3. Additionally, two of the three clones late after activation are able to upregulate expression of IL 2 receptors in response to picamolar concentrations of IL 2, indicating that the receptors on these clones are able to transmit a signal, although insufficient to induce proliferation of the cells. These observations strongly suggest that for the CTL clones examined, at late times after antigenic stimulation, engagement of IL 2 by the high-affinity receptor is not a sufficient signal to induce cells to transit through the cell cycle.

Animals↗
PubMed1987

Anti-T cell receptor antibodies fail to inhibit specific lysis by CTL clones but activate lytic activity for irrelevant targets.

In this report, we describe the functional effects of anti-T cell receptor antibodies on a panel of MHC-restricted, influenza virus-specific CTL clones. Approximately 25 to 30% of these clones are recognized by KJ16-133, an anti-T cell receptor monoclonal antibody presumably specific for products of the V beta 8 gene family, and an antibody with similar specificity, F23.1. In contrast to most previous reports, both KJ16-133 and F23.1, over a wide range of antibody concentrations, fail to inhibit the antigen-specific effector function of these CTL. Instead, the antibodies activate the CTL to kill without regard for the MHC haplotype of the target cells or the presence of the appropriate viral antigen. This anti-T cell receptor antibody-induced cytolysis by our clones does not appear to be mediated by Fc receptors on target cells. Nuclear destruction of target cells as a result of antibody-induced lysis suggests that it occurs via a mechanism similar to antigen-specific lysis by CTL. In addition, both soluble bivalent F23.1 and F23.1 coupled-Sepharose beads are able to induce the secretion of interferon-gamma from these CTL clones.

Animals↗
PubMed1987

Expression of a murine polyclonal T cell receptor marker correlates with the use of specific members of the V beta 8 gene segment subfamily.

A series of murine T lymphocyte clones were examined for reactivity with the KJ16-133 and F23.1 mAbs. Clones that were KJ16-133+,F23.1+ and KJ16-133-,F23.1+ were identified, but no KJ16-133+,F23.1- clones were observed. Within our panel of clones, therefore, the KJ16-133 antibody identifies a subset of F23.1+ cells. All F23.1+ clones examined express members of the V beta 8 subfamily of beta chain variable region genes; clones expressing V beta 8.1 or V beta 8.2 reacted with both KJ16-133 and F23.1, while clones expressing V beta 8.3 reacted only with F23.1. Thus, the differential reactivity of the KJ16-133 and F23.1 antibodies with cloned T cells correlates with the V beta gene expression of each clone. Reactivity with these antibodies should therefore be of utility for predicting the V beta gene expression in some T cell clones.

Animals↗
PubMed1987

Antigen presentation pathways to class I and class II MHC-restricted T lymphocytes.

Our observations on the cellular immune response to type-A influenza suggest the existence of two distinct pathways of protein antigen presentation to T lymphocytes. One of these pathways is involved with presentation of antigens introduced into the presenting cell from without. This exogenous presentation pathway is the well-recognized route of presentation of soluble and particulate antigens to T lymphocytes. This pathway probably involves uptake of antigen into endocytic vesicles, alteration of antigen within an intracellular compartment, and subsequent display of antigen on the presenting cell surface (Unanue 1984). The second pathway is one which we have tentatively designated as an endogenous presentation pathway. The constraints on this pathway have yet to be fully defined. At a minimum, this pathway appears to involve the presentation of antigens which are synthesized de novo in the presenting cell utilizing the cell's biosynthetic machinery. This pathway may also handle preformed antigens located within the cytosolic compartment of the presenting cell. Perhaps the most striking feature of these two antigen presentation pathways is the close association between the MHC restriction of an antigen-specific T lymphocyte and the pathway of antigen presentation to that T lymphocyte. Our data suggest that this association holds both at the effector level and at the level of induction of T lymphocytes. Thus, presentation of a given antigen by the endogenous pathway preferentially triggers a response from class I MHC-restricted T lymphocytes directed to that antigen. The molecular basis for this link of class I MHC-restriction to the endogenous pathway and MHC class II restriction to the exogenous pathway is unknown. It seems likely that interactions between MHC molecules and antigen within the presenting cell may be critical for the demarcation of these pathways. Thus, for example, antigen presented by the endogenous route may only be able to associate intracellularly with newly synthesized or recycling class I MHC molecules. An understanding of the molecular basis of this phenomenon will require detailed information on the expression, intracellular trafficking, and transport of class I and class II MHC molecules in the antigen-presenting cell. An unresolved issue, at least in the case of viral antigens, is the nature and form of the antigenic moieties presented by the exogenous and endogenous pathways. In the case of viral antigen presentation to class II MHC-restricted T lymphocytes, there is strong, albeit indirect, evidence for processing of antigen and recognition of fragments of viral polypeptides (Lamb et al. 1982, Hackett et al. 1983).(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1987

Characterization and in vivo distribution of influenza-virus-specific T-lymphocytes in the murine respiratory tract.

Cloned populations of antigen-specific, murine T-lymphocytes, maintained in long-term continuous culture, have been reported to exhibit functional activity upon adoptive transfer. Few data are currently available on the in vivo distribution of these homogeneous, functional cell populations. Such information is important to understanding the mechanisms by which T-lymphocytes exert their effector activity. We examined 2 cloned populations of influenza virus-specific T-lymphocytes. One clone was a Lyt-2+, L3T4-, class I MHC-restricted, cytolytic T-lymphocyte subset. The other clone was a Lyt-2-, L3T4+, Class II MHC-restricted clone of the helper/amplifier T-cell subset. Both clones promote recovery from lethal pulmonary influenza infection upon adoptive transfer. Using cell populations labeled with [3H]thymidine, we examined the distribution of these cells in tissue sections from various organs. These cloned cell populations were preferentially retained in the lungs of uninfected and influenza-virus-infected animals. This retention is independent of the cell's viral antigenic specificity, but may be dependent on the phenotype of the clone. Once retained in the lungs, these cells migrated across the bronchial lamina propria and entered the epithelium and pulmonary lumen. The significance of these observations for in vivo T-lymphocyte functions is discussed.

Animals↗
PubMed1986

Features of target cell lysis by class I and class II MHC-restricted cytolytic T lymphocytes.

The lytic activity of influenza virus-specific murine cytolytic T lymphocyte (CTL) clones that are restricted by either H-2K/D (class I) or H-2I (class II) major histocompatibility (MHC) locus products was compared on an influenza virus-infected target cell expressing both K/D and I locus products. With the use of two in vitro measurements of cytotoxicity, conventional 51Cr release, and detergent-releasable radiolabeled DNA (as a measure of nuclear disintegration in the early post-lethal hit period), we found no difference between class I and class II MHC-restricted CTL in the kinetics of target cell destruction. In addition, class II MHC-restricted antiviral CTL failed to show any lysis of radiolabeled bystander cells. Killing of labeled specific targets by these class II MHC-restricted CTL was also efficiently inhibited by unlabeled specific competitor cells in a cold target inhibition assay. In sum, these data suggest that class I and class II MHC-restricted CTL mediate target cell destruction by an essentially similar direct mechanism.

Cell Nucleus↗
PubMed1986

Fine specificity and antigen receptor expression among influenza virus-specific cytolytic T lymphocyte clones.

Influenza virus stimulates a vigorous cytolytic T lymphocyte (CTL) response in the mouse that is directed to several virion polypeptides. This report examines the fine specificity of a panel of murine influenza-specific CTL clones restricted by MHC class I products of the H-2d haplotype. Ten of 22 A/JAPAN/305/57-specific CTL clones analyzed were directed to the A/JAPAN/305/57 hemagglutinin protein as detected by using target cells infected with a recombinant vaccinia virus containing hemagglutinin gene. Based on their fine specificity of hemagglutinin recognition, these clones defined four functional epitopes on the hemagglutinin. The remaining 12 cytolytic clones exhibited cross-reactivity for type A influenza viruses of the major human subtypes, and approximately 60% of these clones were directed to the nucleocapsid protein. KJ16-133 monoclonal antibody analysis of the utilization of the T cell receptor V beta 8 gene segment subfamily revealed that members of this V beta gene subfamily are expressed by both hemagglutinin- and nucleocapsid-specific MHC class I-restricted CTL (and by influenza-specific MHC class II-restricted T lymphocytes as well). These results suggest that CTL detect several distinct antigenic sites on the hemagglutinin. In addition, these results reveal no direct correlation between viral antigenic specificity and V beta gene expression by these virus-specific CLT clones.

Animals↗
PubMed1986

Differences in antigen presentation to MHC class I-and class II-restricted influenza virus-specific cytolytic T lymphocyte clones.

We have examined requirements for antigen presentation to a panel of MHC class I-and class II-restricted, influenza virus-specific CTL clones by controlling the form of virus presented on the target cell surface. Both H-2K/D- and I region-restricted CTL recognize target cells exposed to infectious virus, but only the I region-restricted clones efficiently lysed histocompatible target cells pulsed with inactivated virus preparations. The isolated influenza hemagglutinin (HA) polypeptide also could sensitize target cells for recognition by class II-restricted, HA-specific CTL, but not by class I-restricted, HA-specific CTL. Inhibition of nascent viral protein synthesis abrogated the ability of target cells to present viral antigen relevant for class I-restricted CTL recognition. Significantly, presentation for class II-restricted recognition was unaffected in target cells exposed to preparations of either inactivated or infectious virus. This differential sensitivity suggested that these H-2I region-restricted CTL recognized viral polypeptides derived from the exogenously introduced virions, rather than viral polypeptides newly synthesized in the infected cell. In support of this contention, treatment of the target cells with the lysosomotropic agent chloroquine abolished recognition of infected target cells by class II-restricted CTL without diminishing class I-restricted recognition of infected target cells. Furthermore, when the influenza HA gene was introduced into target cells without exogenous HA polypeptide, the target cells that expressed the newly synthesized protein product of the HA gene were recognized only by H-2K/D-restricted CTL. These observations suggest that important differences may exist in requirements for antigen presentation between H-2K/D and H-2I region-restricted CTL. These differences may reflect the nature of the antigenic epitopes recognized by these two CTL subsets.

Animals↗
PubMed1985

Expression of specific cytolytic activity by H-2I region-restricted, influenza virus-specific T lymphocyte clones.

Among murine class II major histocompatibility complex (MHC)-restricted cytotoxic T lymphocyte (CTL) clones specific for type A influenza virus, we have identified both noncytolytic clones and clones exhibiting H-2 I region-restricted cytolytic activity. After appropriate antigenic stimulation, both cytolytic and noncytolytic clones proliferated in the absence of exogenous interleukin 2. All of the clones possess the Thy-1.2+, Lyt-1+2-, L3T4+ phenotype. The class II MHC restriction of viral recognition by the CTL clones was mapped by proliferation using recombinant mouse strains and by inhibition of cytotoxic activity with monoclonal antibodies directed to class II MHC products and L3T4a. The restriction specificity of two CTL clones was unambiguously assigned to the E beta d chain by using L cell transfectant lines expressing E alpha kE beta d or E alpha kE beta k gene products. Analysis of the viral specificity of the cloned lines revealed subtype-specific and crossreactive patterns of viral antigen recognition; the pattern of viral antigen specificity exhibited by each clone in proliferation and cell-mediated cytotoxicity was identical. Each CTL clone also demonstrated antigen-dependent release of helper factor(s) that promoted in vitro primary anti-SRBC responses. Finally, the cytotoxic effector function of the class II MHC-restricted CTL clones was mediated by direct lysis of virus-infected cells, and not by secretion of a cytolytic lymphokine.

Animals↗
PubMed1985

Stable expression of cytolytic activity by influenza virus-specific cytotoxic T lymphocytes.

Influenza-specific immune cytotoxic T lymphocyte (CTL) populations maintain a constant level of in vitro cytolytic activity. This is demonstrable with both heterogeneous populations of anti-viral CTL from immune donors and long-term CTL clones derived from primed CTL precursors. Cytolytic machinery is stably expressed by these CTL populations under a variety of in vitro cultivation conditions. This finding is in contrast to results with alloreactive CTL generated by stimulation of primed CTL precursors that lose cytolytic activity on a per cell basis with time after stimulation. The results indicate that virus-specific, cloned CTL that stably express cytolytic activity are representative of the heterogeneous populations from which they are derived and further suggest a qualitative difference in the regulation and expression of cytolytic machinery between heterogeneous populations of influenza-specific CTL and alloreactive CTL.

Animals↗