Biomedical subjects

A Wack

Publications and source records attributed to A Wack.

At least 19 recordsLinked to original sources

PubMed2005

[Infectious complications and misuse of high-dose buprenorphine].

BACKGROUND: High-dose buprenorphine (HDB) treatment began in France in 1996 according to relatively unrestricted prescription rules. Continued heroin injection by patients on HDB maintenance treatment and even HDB injection remain underestimated and may lead to a variety of infectious diseases. OBJECTIVES: Description of infectious complications occurring in patients receiving HDB maintenance treatment. METHODS: Retrospective study of drug addicts receiving HDB maintenance treatment, injecting (or highly suspected of injecting) it, and hospitalized for infections (other than HIV or viral hepatitis) in the department of infectious and tropical diseases in Nancy University Hospital. Data collection covered 1998 through 2003. RESULTS: We identified 21 case reports, 9 concerning infectious endocarditis, 8 cutaneous abscesses, 2 osteoarticular infections, 1 meningitis and 1 Candida retinitis. The sex-ratio was of 1 woman for 2 men, and the patients' mean age was 29.8 years. Globally 13 patients had systemic infections. Nine patients admitted having injected HDB (and no other drugs) (including the case of Candida retinitis), while in the other 12 cases, the patients continued injecting heroin as well. The role of misused HDB was strongly suspected in those 12 infections, but was not clearly confirmed. All patients recovered from the infections. The long-term psychosocial outcome remains unknown. CONCLUSION: The cases analyzed illustrate the dual reality that HDB is often ineffective as a maintenance treatment, since some patients continue to inject heroin, and that its misuse can have infectious consequences. The results of HDB maintenance treatment substitution are mixed. The individual benefit/risk ratio must be improved. Networking is crucial, notably between physician and pharmacist, and the monitoring system must be reinforced.

Abscess↗

PubMed2001

Binding of the hepatitis C virus envelope protein E2 to CD81 provides a co-stimulatory signal for human T cells.

Chronic hepatitis C virus (HCV) infection frequently develops into liver disease and is accompanied by extra-hepatic autoimmune manifestations. The tetraspanin CD81 is a putative HCV receptor as it binds the E2 envelope glycoprotein of HCV and bona fide HCV particles. Here we show that HCV E2 binding to CD81 on human cells in vitro lowers the threshold for IL-2 receptor alpha expression and IL-2 production, resulting in strongly increased T cell proliferation. HCV E2-induced co-stimulation also enhances the production of IFN-gamma and IL-4 and causes increased TCR down-regulation. This suggests that binding of HCV particles to CD81 on T cells in vivo may lead to activation by otherwise suboptimal stimuli. Therefore, co-stimulation of autoreactive T cells by HCV may contribute to liver damage and autoimmune phenomena observed in HCV infection.

Antigens, CD↗

PubMed2001

Rationally designed strings of promiscuous CD4(+) T cell epitopes provide help to Haemophilus influenzae type b oligosaccharide: a model for new conjugate vaccines.

The age-related and T cell-independent immunological properties of most capsular polysaccharides limit their use as vaccines, especially in children under 2 years of age. To overcome these limitations, polysaccharide antigens have been successfully conjugated to a variety of carrier proteins, such as diphtheria toxoid or tetanus toxoid (TT) and the diphtheria mutant (CRM197) to produce very successful glycoconjugate vaccines. The increasing demand for new conjugate vaccines requires the availability of additional carriers providing high and long-lasting T helper cell immunity. Here we describe the design and construction of three recombinant carrier proteins (N6, N10, N19) constituted by strings of 6, 10 or 19 human CD4(+) T cell epitopes from various pathogen-derived antigens, including TT and proteins from Plasmodium falciparum, influenza virus and hepatitis B virus. Each of these epitopes is defined as universal in that it binds to many human MHC class II molecules. When conjugated to Haemophilus influenzae type b (Hib) oligosaccharide, these carriers elicit a potent anti-Hib antibody response in mice. In the case of the N19-Hib conjugate, this response is at least as good as that observed with CRM197-Hib, a conjugate vaccine currently used for mass immunization. We also show that some of the universal epitopes constituting the recombinant carriers are specifically recognized by two human in vitro systems, suggesting that T cell memory is provided by the selected epitopes. The data indicate that rationally designed recombinant polyepitope proteins represent excellent candidates for the development and clinical testing of new conjugate vaccines.

Amino Acid Sequence↗

PubMed2000

Early onset of CD8 transgene expression inhibits the transition from DN3 to DP thymocytes.

In this paper we show that the effects of transgenic coreceptor expression on thymocyte development depend on the onset of transgene expression. Thus, a CD8 transgene expressed on CD44+CD25+ (DN2) and CD44-CD25+ (DN3) cells causes a partial block at the stage when TCRbeta selection takes place and diminishes expansion at the subsequent developmental stages, resulting in increased DN3 and markedly reduced double-positive (DP) thymocyte numbers. This effect is evident on a polyclonal TCR repertoire as well as in TCR-transgenic mice (F5). By contrast, a CD8 transgene that leads to the same degree of overexpression on DP thymocytes, but is not expressed on double-negative subsets, has no effect on thymus size or composition. Therefore, the reduction of DP thymocyte numbers in CD8 TCRtg mice can be attributed to interferences at early developmental stages rather than to increased negative selection of DP cells.

Animals↗

PubMed2000

Life, activation and death of intrahepatic lymphocytes in chronic hepatitis C.

The healthy liver of adult humans has little or no lymphocyte component and the histological finding of intrahepatic lymphocytes (IHL) is evidence of liver pathology. In a liver injured by chronic hepatitis C, the most common chronic liver disease, most IHL are activated/pro-inflammatory cells, which are particularly enriched for effectors of innate immunity (natural killer (NK), natural T, and other NK-like T cells). IHL do not undergo clonal expansion in the liver but migrate from extrahepatic sites to the chronically infected liver, where they display effector function and subsequently die, suggesting that maintenance of the IHL pool depends on continuous lymphocyte migration. The cytotoxic and inflammatory functions of these IHL have three potential outcomes: 1) they could be helpful in clearing the virus (a rare case in hepatitis C virus (HCV) infection); 2) they could be useless and have no effect on the infection; or 3) they could be harmful, whereby overaggressive lymphocyte responses destroy the liver in a continuous and unsuccessful attempt to clear the virus. Unfortunately, we do not know as of yet which of these possibilities is the case and, therefore, a more complete picture of the intrahepatic immune response will be relevant to the development of new therapeutic strategies against HCV. Additionally and from a more general perspective, due to the availability of biopsied material and the high prevalence (approximately 3%) of HCV infection worldwide, studying the chronically inflamed liver of hepatitis C patients is an ideal model to investigate the poorly understood processes of lymphocyte trafficking, activation and death to non-lymphoid sites of chronic inflammation in man.

Adult↗

PubMed1999

Th cells and Th2 responses can develop in the absence of MHC class II-CD4 interactions.

In this paper, we address the question whether CD4 and MHC class II expression are necessary for the development of the T helper lineage during thymocyte maturation and for activation-induced Th2 responses. To bypass the CD4-MHC class II interaction requirements for positive selection and activation, we used mice that are doubly transgenic for CD8 and for the MHC class I-restricted TCR F5. This transgene combination leads to MHC class I-dependent maturation of CD4 lineage cells. Upon activation, these CD4 lineage T cells secrete IL-4 and give help to B cells but show no cytotoxic activity. Remarkably, neither MHC class II nor CD4 expression are necessary for the generation and helper functions of these cells. This suggests that under normal conditions, coreceptor-MHC interactions are necessary to ensure the canonical combinations of coreceptor and function in developing thymocytes, but that they do not determine functional commitment. Our results also imply that expression of the CD4 gene does not influence, but is merely associated with the decision to establish the T helper program. In addition, we show that activation through TCR-MHC class I interactions can induce Th2 responses independently of CD4 and MHC class II expression.

Animals↗

PubMed1999

T-cell clonality in immune responses.

Recent methodological advances allow the analysis of clonal composition within T-cell subsets. Here, Mala Maini and colleagues review the available data on clonality in acute immune responses and steady-state situations. They highlight and explore reasons for the striking differences in clonality between the CD4+ and CD8+ T-cell subsets.

Animals↗

PubMed1998

Susceptibility and resistance to antigen-induced apoptosis in the thymus of transgenic mice.

Injection of TCR transgenic mice with antigenic peptide results in the deletion of immature thymocytes expressing the transgenic TCR. We have analyzed this process in mice transgenic for a TCR (F5) that recognizes a peptide from the influenza nucleoprotein (NP68). To determine whether deletion of immature thymocytes is the result of specific recognition of the antigenic peptide by the thymocytes or mature T cell activation, bone marrow chimeric mice were generated using a mixture of cells from F5 transgenic and nontransgenic mice. Injection of these mice with antigenic peptide leads to the preferential depletion of F5 transgenic thymocytes, whereas nontransgenic thymocytes remain largely unaffected. Furthermore, exposure of F5 fetal thymic lobes to peptide leads to thymocyte deletion even though no mature single positive T cells are present at this stage. These data suggest that Ag-induced death of immature thymocytes is due to peptide-specific recognition, although activated mature T cells appear to potentiate such deletion. Further administration of antigenic peptide to F5 mice results in the appearance of double-positive thymocytes that are resistant to Ag or anti-CD3-induced apoptosis. These data suggest a change in the ability of the cells to signal through the TCR-CD3 complex, resembling the state of anergy induced in peripheral T cells following chronic exposure to Ag.

Animals↗

PubMed1998

Interactions with multiple peptide ligands determine the fate of developing thymocytes.

Thymocytes are positively or negatively selected depending on interactions between their T cell receptors (TCR) and peptides presented by major histocompatibility complex molecules. We have previously shown that apoptosis of thymocytes from an alpha beta TCR-transgenic mouse (F5), induced by antigenic peptide, can be inhibited specifically by an antagonist peptide variant in an in vitro culture model. We have now extended these experiments by demonstrating that the antagonist peptide can inhibit natural negative selection of maturing thymocytes, induced by endogenously expressed antigen, in fetal thymic organ cultures (FTOC). This inhibition resulted in the rescue and maturation of thymocytes that would otherwise have been deleted. Mature T cells generated in these cultures were able to respond to antigen by producing limited quantities of interferon-gamma, but unlike T cells from control FTOC, they required exogenous interleukin-2 to generate cytolytic effector cells. Interestingly, the antagonist peptide also accelerated the development of F5 thymocytes in the absence of the negatively selecting ligand. These data suggest that the developmental fate of a thymocyte may be determined by the recognition of multiple distinct peptide ligands during thymic selection. Alterations in the profiles of selecting peptides presented in the thymus would thus have profound effects on the size and autoreactive potential of the T cell repertoire generated.

Animals↗

PubMed1998

A comparison of two techniques for the molecular tracking of specific T-cell responses; CD4+ human T-cell clones persist in a stable hierarchy but at a lower frequency than clones in the CD8+ population.

Oligoclonal or clonal T-cell expansions, presumed to be antigen driven, are frequently sought and followed for diagnostic and prognostic purposes, as well as to understand more about their natural history. Techniques based on conservation of T-cell receptor CDR3 length are increasingly widely used, often without assessment of sensitivity or specificity. We present a comparative evaluation of a novel modified heteroduplex technique and a CDR3-length-based assay. Dilution of a known clone in a mixed T-cell population shows that in our hands the heteroduplex technique is at least 10-fold more sensitive than the CDR3-length-based assay. However, even with this level of sensitivity, we do not detect clonal expansions in unstimulated CD4+ T cells. This contrasts with the frequent detection of CD8+ clones in fresh samples and suggests different mechanisms of clonal homeostasis in the two subsets. We show that both techniques detect functional expansions after in vitro stimulation with a recall antigen. The distinct molecular footprint seen with the heteroduplex technique allows reproducible follow up of specific clonal expansions. We have exploited this to demonstrate that the repertoire of clones expanded by in vitro tetanus toxoid stimulation shows stability within an individual, implying long-term maintenance of multiple CD4+ clones.

CD3 Complex↗

PubMed1998

Age-related modifications of the human alphabeta T cell repertoire due to different clonal expansions in the CD4+ and CD8+ subsets.

We have studied the effects of a life-long antigen stimulation on the clonal heterogeneity of human peripheral T cell subsets, as defined by their CD45 isoform expression. CD4+ or CD8+ T cells were obtained from healthy donors ranging in age from 20 to 100 years, and sorted into CD45RA+ and CD45RO+ populations. A modified PCR-heteroduplex analysis was then used to directly compare the TCR Vbeta clonal make up of either compartment pair. We find that the CD4+ T cell repertoire remains largely polyclonal throughout life, since CD4+ expanded clones are rare and accumulate predominantly in the CD45RO+ compartment of exceptionally old donors (100 years old). In contrast, the CD8+ T cell subset contains expanded clones which are already detectable in young adults and become very frequent in 70- to 75-year-old donors in both CD45RA+ and CD45RO+ compartments analyzed. Interestingly, some expanded clones are detectable in the CD45RA+ or in both CD45RA+ and CD45RO+ compartments of either CD4+ or CD8+ T cells. These results indicate that the age-dependent accumulation of expanded clones starts earlier and is more pronounced in the CD8+ than in the CD4+ T cell subset, reinforcing the concept that clonal expansion in the two subsets is controlled by substantially different mechanisms. Furthermore, whereas the finding of expanded CD45RO+ T cell clones is explained by antigen-driven proliferation, the detection of expanded clones in the CD45RA+ or in both CD45RA+ and CD45RO+ compartments would support the hypothesis of reversion from the CD45RO+ to the CD45RA+ phenotype after antigen encounter.

Adult↗

PubMed1997

Multiple sites of post-activation CD8+ T cell disposal.

Antigen-triggered activation of T cells leads to a sequence of differentiation steps including up-regulation of activation markers, blast formation, proliferation, delivery of effector functions, and ultimately apoptosis. It is still controversial in which anatomical site activation-induced apoptosis and elimination of T cells occur. To address this question, we used mice transgenic for a T cell receptor (F5) specific for an influenza virus nucleoprotein peptide (NP68) presented on the major histocompatibility complex H-2 Db molecule. Accumulation and apoptosis of T cells was studied using terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end-labeling in situ combined with immunohistology after intraperitoneal injection of the cognate peptide into F5 mice which are wild type or deficient for Rag-1. After 4 days of peptide treatment, large perivascular infiltrations of CD8+ cells were observed in liver, lung, and kidney of F5 mice. CD8+ cell numbers were also increased in skin and small intestine, but not in brain or heart muscle of peptide-treated animals. The infiltrating CD8+ cells show an increased percentage of apoptosis in liver, lung and, most strikingly, the kidney. These data suggest that in the F5 system, T cell disposal after activation occurs in a number of organs. Essentially identical findings were obtained in Rag-1(+/+) and Rag-1(-/-) F5 mice, suggesting that the deletion mechanism did not involve other T or B cells.

Animals↗

PubMed1997

In vitro positive selection of alpha beta TCR transgenic thymocytes by a conditionally immortalized cortical epithelial clone.

Development of mature CD4 and CD8 single-positive T cells requires a process known as positive selection, which depends on the specific recognition of self-peptide-MHC complexes on thymic stromal cells by immature CD4+CD8+ thymocytes. We have used an in vitro reaggregate system to study the positive selection of thymocytes by conditionally immortalized thymic epithelial clones. Thymocytes from mice transgenic for the F5 alpha beta TCR, specific for a peptide from the influenza nucleoprotein in the context of H-2Db, are positively selected in the H-2b MHC background, but fail to mature in mice expressing the H-2q haplotype. Development of embryonic day 15 F5 H-2q transgenic thymocytes was followed in reaggregate cultures supplemented with H-2b-expressing epithelial clones. A conditionally immortalized cortical epithelial clone, derived from H-2Kb-tsA58 transgenic mice, was found to be as efficient as freshly isolated thymic stromal cells in positively selecting CD8 transgenic thymocytes. In contrast, an H-2b-expressing kidney epithelial clone did not augment positive selection above background levels, implying that the effect of the thymic epithelial clone was not merely the presentation of selecting MHC molecules. Mature transgenic thymocytes generated in reaggregate cultures were able to differentiate into functionally competent cytotoxic T cells. This model provides an important in vitro system for the detailed study of the specific molecular interactions leading to positive selection of developing thymocytes.

Animals↗

PubMed1996

Restricted TCR repertoire and long-term persistence of donor-derived antigen-experienced CD4+ T cells in allogeneic bone marrow transplantation recipients.

We investigated the contribution of transfer of Ag-experienced donor T cells to the immune reconstitution of allogeneic bone marrow transplantation (BMT) recipients. To this purpose, we used a combination of cell culture methods to isolate tetanus toxoid (TT)-specific T cell clones, and a sensitive and specific heteroduplex analysis to monitor the presence of a particular clonotype using TCR N region sequences. We document that patients after BMT display a small response to TT, entirely accounted for by few donor-derived clones. These patients show a strong polyclonal response to TT vaccination; however, the T cell clones transferred with the transplant can still be detected within the polyclonal T cell lines for up to at least 5 yr after BMT. We also demonstrate that vaccination of donors with TT before BMT results in a more relevant transfer of Ag-experienced T cells, allowing the recipients to mount a strong polyclonal response without need of vaccination. These findings provide a rationale for vaccinating donors to optimize adoptive transfer of protective T cell immunity into recipients, and suggest the possibility of using preventive T cell adoptive therapy in conjunction with marrow infusion.

Adult↗

PubMed1996

An improved PCR-heteroduplex method permits high-sensitivity detection of clonal expansions in complex T cell populations.

We have modified the PCR-heteroduplex technique to render it more suitable for the study of the clonal make up of complex T cell populations. This technique is based on separate PCR amplifications of all the TCR V beta genes expressed by a polyclonal T cell sample, followed by a heteroduplex reaction of the PCR products and a gel separation. Our modification involves performing each heteroduplex reaction in the presence of excess carrier DNA, which is the PCR product of a cloned TCR V beta cDNA having the same variable and constant region of the amplified V beta family, but a different N region. In this way, every clonotypic V beta chain that is amplified in the polyclonal mixture forms a unique and reproducible pair of heteroduplex bands with the carrier DNA. This molecular footprint permits the identification of a given T cell clone over time, or in different anatomical sites. The specificity and sensitivity of the detection of T cell clones can be further increased by hybridising the blotted heteroduplex gel with oligonucleotides specific for either a TCR V beta N region or the carrier DNA. In conclusion, we have developed a simple and reproducible technique that permits the simultaneous detection of the expanded T cell clones present in heterogeneous T cell populations in a very specific and sensitive manner.

Aged↗

PubMed1996

Direct visualization of thymocyte apoptosis in neglect, acute and steady-state negative selection.

During thymocyte differentiation, the majority of the developing cells die in situ by apoptosis and are subsequently removed by macrophages. DNA fragmentation is one of the hallmarks of apoptosis and can be detected in situ by TdT-mediated dUTP-biotin nick end labeling (TUNEL). We used TUNEL combined with immunohistology to determine the sites of thymocyte apoptosis in mice transgenic for a TCR (F5) which recognizes a peptide (NP68) of the influenza virus nucleoprotein (NP) presented on the MHC class I H-2Db molecule. Apoptosis due to neglect was studied in F5 mice expressing a neutral MHC haplotype (F5/H-2q) and in beta 2-microglobulin-deficient F5 mice (F5/ beta 2m+). In both cases, the frequency of apoptotic cells was similar to that seen in F5/H-2b mice and non-transgenic C57BI/10 mice. Antigen-induced apoptosis was studied in F5 mice after i.p. Injection of the cognate NP68 peptide and in F5/NP double-transgenic mice. Three hours after peptide injection, apoptosis was high throughout the thymus cortex and clusters of apoptotic cells formed due to tissue macrophage uptake, whereas the thymic medulla remained unaffected. Massive recruitment of inflammatory cells into the thymus was seen as early as 1 h after peptide injection. Nine hours after peptide injection changes were apparent in the cortical epithelium and, by 4 days, the cortical network had collapsed to give scattered, compacted epithelial cells. In contrast, in F5/NP double-transgenic mice, thymocyte apoptosis induced by cognate self-peptide was localized at the cortico-medullary junction with little change seen in the epithelium of the cortex.

Animals↗

PubMed1996

The repertoire of T-lymphocytes recovered by bronchoalveolar lavage from healthy nonsmokers.

We reasoned that persistent exposure to a limited set of airborne antigens could drive the preferential expansion of single T-cell clones in the lower respiratory tract of normal individuals. To explore this issue, the normal human alpha/beta T-cell receptor repertoire was studied in lung lymphocytes obtained by bronchoalveolar lavage (BAL) from the humen of the lower respiratory tract. BAL T-cells obtained from five healthy volunteers were first analysed using polymerase chain reaction to amplify all known V alpha and V beta genes of the T-cell receptor. T-cells from peripheral blood were used as an internal control. Heteroduplex analysis of the amplified products was then performed, to assess the clonal composition of the repertoire of lung- versus blood-derived T-lymphocytes within each amplified variable gene family. In all subjects, the T-cell repertoire in the lung was largely as heterogeneous as peripheral blood in terms of clonal composition. This indicated lack of preferential expansion of single T-cell clones. A few T-cell clones were simultaneously expanded in blood and lung in all individuals within a limited number of V beta (mean 2.4; range 2-4) and V alpha (mean 1.6; range 1-3) genes. We also found that lung T-lymphocytes expressed all of the V gene families of the T-cell receptor that were expressed by peripheral blood T-cells. Our results indicate that T-cell clones in the lower respiratory tract of normal individuals are distributed according to a largely polyclonal pattern, which corresponds to that found in peripheral blood.

Adult↗

PubMed1994

Plasminogen activator inhibitor type 1 in rat spermatozoa: localisation in the tail, in the acrosome and on the surface of the head.

Isolated seminiferous tubules of rat testis contain considerable urokinase-inhibiting activity. An immunohistological analysis revealed the presence of plasminogen activator inhibitor type 1 (PAI-1) in the basement membrane as well as in the interior of the tubules. Distribution and intensity of the intratubular immunoreactivity depends on the stage of the seminiferous cycle. A relatively weak signal is present around elongated nuclei of spermatids at the beginning of chromatin condensation. The signal intensity increases in the course of differentiation until a maximum is reached at stages VII-VIII. In these stages PAI-1 immunoreactivity is localised around the nuclei of the late spermatids as well as along their tails. Spermatozoa in the ductus epididymis also strongly react with the PAI-1-specific antiserum, suggesting that the inhibitor remains associated with the germ cells after spermiation and during maturation in the epididymis. In intact mature spermatozoa isolated from epididymis cauda by "swimming-up' in non-capacitation medium, PAI-1 antigen is localised on the plasma membrane surrounding the head. In addition, in fixed and permeabilised cells the immunoreactivity is detectable in the acrosome and in the tail. Possible functions of PAI-1 in spermatogenesis, sperm motility and sperm-egg interaction are discussed.

Acrosome↗