The effect of age on mitogen responsive T cell precursors in human beings is completely restored by interleukin-2.

Abstract It is well known that the function of T lymphocytes is significantly impaired by advancing age. In the present study, attempts have been made to further characterize the T cell impairment of elderly subjects. Thus, we have performed limiting dilution microculture analysis to evaluate the precursor frequency of T lymphocytes responding to a mitogenic stimulus in old and young subjects. Furthermore we have evaluated the activity of recombinant interleukin-2 (rIL-2) on these cells. The results demonstrate that in older subjects the frequency of these precursors is significantly decreased. The in vitro treatment with rIL-2 increased the frequency of mitogen responsive T lymphocyte prec…

Frequency and subset distribution of human CD8 T cells specific for mycobacterial peptides in healthy contact individuals.

Ligand‐Specific αβ and γδ T Cell Responses in Childhood Tuberculosis

The alphabeta and gammadelta T cell responses were analyzed in the peripheral blood of children affected by active tuberculosis (TB) and in healthy children who tested positive (PPD+) or negative (PPD-) for purified protein derivative. PPD+ healthy and diseased children responded equally well to PPD in vitro. In contrast, only 18% of PPD+ TB patients responded to peptide p38G derived from the 38-kDa protein of Mycobacterium tuberculosis. Analysis of the whole gammadelta T cell population and of its Vgamma9/Vdelta2 subset showed similar frequencies in PPD+ children with TB and in healthy PPD+ and PPD- children. Vgamma9/Vdelta2 cells from children with TB responded to 5 different phosphoantig…

In Naive mice in vivo localization of WI specific CD8+CD28- induced cells

Differentiation of Effector/Memory Vδ2 T Cells and Migratory Routes in Lymph Nodes or Inflammatory Sites

Vδ2 T lymphocytes recognize nonpeptidic antigens without presentation by MHC molecules and mount both immediate effector functions and memory responses after microbial infection. However, how Vδ2 T cells mediate different facets of a memory response remains unknown. Here, we show that the expression of CD45RA and CD27 antigens defines four subsets of human Vδ2 T cells with distinctive compartmentalization routes. Naive CD45RA+CD27+ and memory CD45RA−CD27+ cells express lymph node homing receptors, abound in lymph nodes, and lack immediate effector functions. Conversely, memory CD45RA−CD27− and terminally differentiated CD45RA+CD27− cells, which express receptors for homing to inflamed tissu…

Immunoregulatory role of Jα281 T cells in aged mice developing lupus-like nephritis

Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by the emergence of autoreactive T cells. Humans and mice with SLE have reduced numbers of CD1d-restricted invariant natural killer T (iNKT) cells, suggesting a key role for these cells in its immunopathogenesis. This subset uses an invariant TCR constituted by Valpha14 Jalpha281 chains paired with some Vbeta domains. The regulatory role for iNKT cells in non-autoimmune mice was suggested by our previous results showing that aged Jalpha281 knockout (KO) mice produce anti-dsDNA. Here we show that old Jalpha281 KO mice have proteinuria and antibodies against dsDNA and cardiolipin. Histological analysis of Jalpha281 KO m…

Research on complement: old issues revisited and a novel sphere of influence

Immunology in recent years has taken a somewhat surprising turn, expressed by a renewed interest in innate immunity. Especially intriguing is the regulatory role exerted by the innate components on the adaptive response, with Toll receptors and complement components being the most investigated. This function has been firmly established for complement protein CR2 (CD21) as part of the BCR co-receptor CD19/CD21/CD81. New findings are now providing a broader picture of complement and its tuning of the immune response; for example, complement proteins have been implicated in the control of T-cell-mediated responses. We will review some of these data here and summarize new discoveries in areas o…

Differential Greek and northern African migrations to Sicily are supported by genetic evidence from the Y chromosome

The presence or absence of genetic heterogeneity in Sicily has long been debated. Through the analysis of the variation of Y-chromosome lineages, using the combination of haplogroups and short tandem repeats from several areas of Sicily, we show that traces of genetic flows occurred in the island, due to ancient Greek colonization and to northern African contributions, are still visible on the basis of the distribution of some lineages. The genetic contribution of Greek chromosomes to the Sicilian gene pool is estimated to be about 37% whereas the contribution of North African populations is estimated to be around 6%. In particular, the presence of a modal haplotype coming from the southern…

Cross-talk between gamma delta T cells and dendritic cells in anti-mycobacterial immune response

D6 is a decoy and scavenger receptor for inflammatory CC chemokines. D6-deficient mice were rapidly killed by intranasal administration of low doses of Mycobacterium tuberculosis. The death of D6(-/-) mice was associated with a dramatic local and systemic inflammatory response with levels of M. tuberculosis colony-forming units similar to control D6-proficient mice. D6-deficient mice showed an increased numbers of mononuclear cells (macrophages, dendritic cells, and CD4 and CD8 T lymphocytes) infiltrating inflamed tissues and lymph nodes, as well as abnormal increased concentrations of CC chemokines (CCL2, CCL3, CCL4, and CCL5) and proinflammatory cytokines (tumor necrosis factor alpha, int…

Method of antigenic peptide identification and relative use for the preparation of a vaccine anti-HIV-1.

Aspetti medico-sanitari, etici e socio economici dell'attività solfiera in Sicilia (1791-1964)

In vivo γδ T Cell Priming to Mycobacterial Antigens by Primary Mycobacterium tuberculosis Infection and Exposure to Nonpeptidic Ligands

The recognition of phosphorylated nonpeptidic microbial metabolites by Vγ9Vδ2 T cells does not appear to require the presence of MHC molecules or antigen processing, permitting rapid responses against microbial pathogens. These may constitute an important area of natural anti-infectious immunity. To provide evidence of their involvement in immune reactivities against mycobacteria, we measured the responsiveness of peripheral blood Vγ9Vδ2 T cells in children with primary Mycobacterium tuberculosis (MTB) infections. Peripheral blood mononuclear cells from 22 children with MTB infections and 16 positivity of tuberculin (PPD)-negative healthy children were exposed to nonpeptidic antigens in vit…

Production of T suppressor factor specific for the hapten picryl chloride requires both T suppressor cells and an antigen-specific, genetically restricted second signal

Summary We investigated the requirement for activation of T suppressor cells specific for the hapten picryl chloride and the release of hapten-specific T suppressor factor. Using an in vivo experimental system, we report that activation of T suppressor cells and the consequent release of T suppressor factor required two signals: one was provided by primed T suppressor cells, i.e. spleen cells from mice injected with the tolerogen picrylsulphonic acid, and the other was provided by the specific antigen in the context of H-2 gene products. Mechanisms by which the interaction between these two signals led to activation of T suppressor cells and the production of T suppressor factor, as well as…

Induction and tolerization of anti-male CD8+ cytotoxic T lymphocytes by in vivo immunization with an H-Y-derived peptide

Abstract We have analyzed the immune response induced by a 9mer synthetic peptide derived from the male histocompatibility antigen H-Y and containing D b -binding motifs in C57BL/6 mice. In this study we report that a single, subcutaneous injection of the peptide emulsified in IFA gave rise to the development of male-specific CD8 + T cells which displayed H-Y-specific proliferative response in vitro and showed a Tc1-type pattern of cytokine production (i.e. they secreted IFN-γ and IL-2, but not IL-4 and IL-10). Development of a strong cytotoxic activity required in vitro stimulation with specific peptide and IL-2: under these culture conditions, we were able to generate potent CD8 + CTLs th…

DIFFERENT ROLE OF HUMAN HLA-DR AND -DQ MOLECULES IN XENOGENEIC TRANSPLANTATION USING TRANSGENIC MICE1

Background. The role of T lymphocytes in graft rejection in xenotransplantation is still unclear. The ability of the human HLA class II molecules DR and DQ to function as xenoantigens was investigated in a murine model of skin grafting, using HLA-DR1 and -DQ6-transgenic mice. Methods, Skin from HLA-DR1- or -DQ6-transgenic mice was transplanted in control littermates. Spleen cells from donors or recipients were tested in mixed lymphocyte reaction and cytotoxic assay. Results. Skin from HLA-DR1-transgenic mice was rejected and spleen cells from rejecting mice were able to proliferate to donor cells, although no rejection was observed when the skin of HLA-DQ6-transgenic mice was engrafted in c…

Graffiti dello Steri di Palermo e conoscenze mediche

The graffiti left by prisoners in the Inquisition gaols of Palermo's represent a testimony of the historical period between 1600 to 1793. In that period, by order of the viceroy Caracciolo, all the testimonies were removed at the same time in which the Inquisition court was suppressed. In this work the historical subdivision between sacred and profane themes is analyzed with the purpose to study human body in an anthropological key as a language in condition of limited freedom and under torture. Many of the profane graffiti are devoted to medical knowledge suggesting that doctors were involved in the activities of this religious court likewise happened in civil courts. Giovanni Filippo Ingr…

LE REAZIONI IMMUNOPATOGENE

Prophylaxis of lipopolysaccharide-induced shock by α-galactosylceramide

AbstractThe NKT cell ligand α-galactosylceramide and its synthetic homologue KRN7000 stimulate rapid and copious secretion of IFN-γ and TNF-α release, both of which are key mediators of LPS-induced shock. We showed that KRN7000, injected before or within 2 h after LPS challenge, was able to prevent endotoxic shock. KRN7000 induced survival when the mice were injected 6, 9, or 12 days before the first injection of LPS, and this protective effect was associated with reduction upon subsequent challenge in the levels of IFN-γ, TNF-α, MCP-1, and an increase of IL-10. Further analysis showed that the animals treated with KRN7000 prior to LPS challenge had lower numbers of F4/80+, NKT, and NK cell…

Selective depression of interferon-γ and granulysin production with increase of proliferative response by Vγ9/Vδ2 T cells in children with tuberculosis

Vgamma9/Vdelta2 T cells can contribute to protective immune response against Mycobacterium tuberculosis, although the extent to which and mechanisms by which they could actually protect against human tuberculosis remain unclear. We have previously reported that Vgamma9/Vdelta2 T cells from tuberculin purified protein derivative (PPD)-positive children, either healthy or affected by different clinical forms of tuberculosis, strongly proliferate to different phosphoantigens in vitro, whereas Vgamma9/Vdelta2 T cells from PPD-negative healthy subjects proliferate very poorly. We report here that Vgamma9/Vdelta2 T cells from tuberculous children have an increased proliferative activity, but decr…

Anti-16-kilodalton mycobacterial protein immunoglobulin M levels in healthy but purified protein derivative-reactive children decrease after chemoprophylaxis.

ABSTRACT Serum responses against Mycobacterium tuberculosis HSP16 were determined for children with tuberculosis (TB) and for healthy purified protein derivative (PPD)-positive and PPD-negative children. Immunoglobulin G (IgG) and IgM responses were higher for TB patients than for other groups. After chemotherapy, IgM and IgG responses decreased for TB patients and PPD-positive subjects. Monitoring of anti- M. tuberculosis HSP16 responses could assist in the management of pediatric TB.

A SUBSET OF VGAMMA9 VDELTA2 T CELLS HELP B CELLS FOR ANTIBODY PRODUCTION.

Analysis of the immune response induced by a single xenoantigen in vivo

Transgenic mice expressing human major histocompatibility complex (MHC) class II molecules would provide a valuable model system for studying murine anti-human MHC immune response. We have previously shown that skin from HLA-DR1 transgenic mice was rejected by control littermates and spleen cells from rejecting mice were able to proliferate to donor cells. The aim of this paper is to analyze the mechanism of recognition of this xenoantigen and the possible involvement of antibody response in anti-HLA-DR1 immune response. Control littermates were immunized with spleen cells from HLA-DR1 transgenic (TG) mice; at indicated times, xenoantigen-specific proliferation and IFNgamma production was a…

Cytokine production pathway in the elderly.

It is well known that aging is associated with various alterations in lymphoid cell functions, particularly with a progressive decline in immune responsiveness to exogenous antigens and increasing incidence of autoimmune phenomena. Many studies have been focused on the mechanisms of the immunologic features of aging. this review describes our results of studies performed to determine the influence of age on the capacity to produce interleukin-2 (IL-2), interferon-gamma (IFN-gamma), interleukin-4 (IL-4), interleukin-t (IL-5), interleukin-6 (IL-6) and tumor necrosis factor (TNF). Mitogen-stimulated cultures of mononuclear cells (MNC) from human beings were assessed for cytokine-producing capa…

Impaired contact hypersensitivity to trinitrochlorobenzene in interleukin-4-deficient mice

We have examined the role of endogenously produced interleukin-4 (IL-4) in the contact hypersensitivity (CH) reaction to the haptene trinitrochlorobenzene (TNCB). The CH reaction was abolished in IL-4 genetically deficient mice (IL-4 KO), when compared to wild-type (wt) mice. The CH reaction was restored by treatment with IL-4 and further analysis revealed that IL-4 exerted its action both at the induction and effector stages of the CH reaction. Despite failure to develop a CH reaction, IL-4 KO mice developed a T helper type 1 (Th1) response to TNCB, in terms of lymphokine production in vitro. Furthermore, the number of Vgamma3+ cells accumulating in the lymph nodes of TNCB-immune IL-4 KO m…

Role of Gamma-Delta T Cells in Cutaneous Hypersensitivity Reactions

In vitro effects of aminobisphosphonates on Vgamma9Vdelta2 T cell activation and differentiation.

A methodological strategy for PAH genotyping in populations with a marked molecular heterogeneity of hyperphenylalaninemia.

Abstract The elucidation of the molecular basis of hyperphenylalaninemia in various world populations (PKU Consortium Database: http://www.mcgill.ca/pahdb/) has revealed a remarkable molecular heterogeneity at the locus encoding for phenylalanine hydroxylase. As a consequence, genotyping of HPA patients has prompted the establishment of an impressive number of mutation detection protocols. In spite of the large variety of methods proposed so far, no comprehensive strategy has been yet developed for the detection of PAH gene mutations. Therefore, new approaches, combining the advantages of individual methods are required, especially in populations with a high number of PAH gene mutations. In…

Selection of microbial epitopes for immune recognition. North-South transfer in Biotechnology of Tuberculosis and AIDS. University of Rome “Tor Vergata”. UNESCO Interdisciplinary Chair in Biotechnology. Vittorio Colizzi, Luc Mantagnier, 2004

Development of hapten-induced IL-4-producing CD4+ T lymphocytes requires early IL-4 production by alphabeta T lymphocytes carrying invariant V(alpha)14 TCR alpha chains

This paper investigates the mechanisms responsible for the generation of IL-4-producing CD4+ T cells during contact sensitization with the hapten trinitrochlorobenzene (TNCB). Lymph node cells taken 1 day after immunization spontaneously released IL-4 while lymph node cells taken 2 and 3 days after immunization did not produce IL-4. A second wave of IL-4 production that was both antigen-specific and MHC class II (I-A)-restricted was observed 4 days after immunization. The spontaneous release of IL-4 at day 1 was due to the alphabeta+ double-negative (CD4- CD8-) T lymphocytes that also expressed NK1.1 and showed V(alpha)14 rearrangement, while alphabeta+ CD4+ T lymphocytes were the source of…

Role of γδ T lymphocytes in immune response in humans and mice

T lymphocytes recognize antigen through the T cell receptor. T cells expressing the gamma delta T cell receptor have been found in many species. Whereas murine alpha beta T cells are concentrated in the lymphoid organs, gamma delta T cells represent only a minor population in the adult thymus and peripheral lymphoid organs (less than 5% of the population). However, murine gamma delta cells predominate in epidermis, in epithelial layers of small intestine, in lung, and in female reproductive organs. In contrast, human gamma delta cells predominate in lymphoid organs. Despite extensive progress in the molecular characterization of the gamma delta T cell receptor and its genes, the physiologic…

"L'informatione de Pestifero et Contagioso Morbo" di G. F. Ingrassia: Percorso diagnostico.

Towards a genetic history of Sicily

γδ cells involved in contact sensitivity preferentially rearrange the Vγ3 region and require interleukin-7

Ptak and Askenase showed that both alphabeta and gammadelta cells are required for transfer of contact sensitivity (CS). This study confirms that day 4 immune cells depleted of gammadelta cells fail to transfer CS to trinitrochlorobenzene (TNP-Cl) systemically and demonstrates that administration of anti-gammadelta monoclonal antibodies (mAb) in vivo abolishes the CS reaction. Moreover, gammadelta cells accumulate at the antigen challenge site: these cells have the unusual phenotype CD8alpha+, CD8beta-, IL-4 R+ which we suggest is due to their state of activation. Following immunization with contact sensitizer on the skin, the absolute number of gammadelta cells increases in the regional ly…

IL-5 Enhances in Vitro and in Vivo Antigen-Specific IgA Production in MHC Genetically Determined Low IL-5 Responder Mice

Lymphonode cells from BALB/k mice, but not from BALB/c mice, immunized with picryl chloride (PCl) produce IL-5 when stimulated with the specific antigen in vitro and this correlates with picryl-specific IgA levels in vivo, which are 6 to 10 times higher in BALB/k mice. B lymphocytes from BALB/k mice cultured with PCl-immune T cells from BALB/k produce in vivo anti-PCl-IgA, while B lymphocytes from BALB/c mice, cultured with T cells from BALB/c mice, fail to produce appreciable amounts of anti-PCl IgA, unless IL-5 is added to cultures. B lymphocytes from both strains of mice produce similar amounts of total IgA antibodies when stimulated in vitro with lipopolysaccharide. In vivo administrati…

Delayed-type skin reactions in bursectomized or thymectomized chickens.

Chickens can easily be induced to develop delayed-type skin reactions to oxazolone when animals are sensitized 7 days before the challenge. The reaction is quantitated by assessing the increase in wattle thickness: maximum reactions occur 24 h after challenge. The reaction is inhibited by neonatal thymectomy or bursectomy; these findings therefore suggest also an important B-derived component in delayed hypersensitivity to oxazolone.

In vitro effects of aminobisphosphonates on Vgamma9Vdelta2 T cell activation and differentiation.

In this study we have evaluated the in vitro effects of four different aminobisphosphonates, alendronate, risedronate, neridronate and zoledronate, on Vγ9Vδ2 T cell activation and differentiation. All tested aminobisphosphonates induce an IL-2-dependent activation and expansion of Vγ9Vδ2 T lymphocytes in primary PBMC cultures of healthy donors. Most notably, they also determine a different distribution of Vγ9Vδ2 T cell subsets, with decrease of Tnaive and TCM cells and increase of TEM and TEMRA Vγ9Vδ2 cells, indicating that in vitro treatment with aminobisphosphonates induces Vγ9Vδ2 T lymphocytes to differentiate towards an effector/cytotoxic phenotype. Accordingly, Vγ9Vδ2 T lymphocytes cu…

Reciprocal stimulation of gammadelta T cells and dendritic cells during the anti-mycobacterial immune response.

Gammadelta T cells and dendritic cells (DC) are two distinct cell types of innate immunity that participate in early phases of immune response against Mycobacterium tuberculosis infection. Here we show that a close functional relationship exists between these cell populations. Using an in vitro coculture system, Vgamma1 T cells from Tcrb(-/- )mice were found to be activated by DC infected in vitro with BCG, as indicated by the elevated CD69 expression, IFN-gamma secretion and cytotoxic activity. This activation process was due to a non-cognate mechanism since it required neither cell to cell contact nor interaction between the TCR and a specific antigen, but was mediated by DC-derived IL-12…

Sequestration of T lymphocytes to body fluids in tuberculosis: reversal of anergy following chemotherapy.

The specificity of CD4 T lymphocytes was investigated in 6 patients affected by tuberculosis who had negative tuberculin purified protein derivative (PPD) skin tests at diagnosis. Polyclonal CD4 T cell lines from the peripheral blood failed to proliferate to PPD and to the 16- or 38-kDa proteins of Mycobacterium tuberculosis, while CD4 cell lines from the disease site responded to PPD and to the 16- and 38-kDa proteins and derived epitopes in vitro. Four months after chemotherapy, the patients became responsive to PPD. The proliferative response to PPD and to the 16- or 38-kDa proteins and their derived peptides decreased in CD4 T cell lines from the disease site and increased in lines from…

Analysis of memory and effector CD8 T cell subsets in chronic-Graft-versus-Host disease.

CXCR5 identifies a subset of Vγ9Vδ2 T cells which secrete IL-4 and IL-10 and help B cells for antibody production

Abstract Vγ9Vδ2 T lymphocytes recognize nonpeptidic Ags and mount effector functions in cellular immune responses against microorganisms and tumors, but little is known about their role in Ab-mediated immune responses. We show here that expression of CXCR5 identifies a unique subset of Vγ9Vδ2 T cells which express the costimulatory molecules ICOS and CD40L, secrete IL-2, IL-4, and IL-10 and help B cells for Ab production. These properties portray CXCR5+Vγ9Vδ2 T cells as a distinct memory T cell subset with B cell helper function.

Partial and Ineffective Activation of Vγ9Vδ2 T Cells by Mycobacterium tuberculosis-Infected Dendritic Cells

Abstract γδ T cells and dendritic cells (DCs) participate in early phases of immune response against Mycobacterium tuberculosis. We investigated whether a close functional relationship exists between these two cell populations using an in vitro coculture in a human system. Vγ9Vδ2 T cells induce full maturation of M. tuberculosis-infected immature DCs, as demonstrated by upregulation of the costimulatory CD80, CD86, CD40, and HLA-DR molecules on infected DCs after 24 h of coculture. Reciprocally, infected DCs induced substantial activation of Vγ9Vδ2 T cells upon coculture, which was cell-to-cell contact and TCR dependent, as demonstrated in transwell experiments. However, infected DCs select…

Major histocompatibility complex regulation of cytokine production.

This review describes the phenomenon of the major histocompatibility complex (MHC) control of cytokine production both in experimental animals and in humans. H-2 (mouse MHC) regulates which type of cytokine is selectively produced in response to the hapten trinitrophenyl (TNP). T cells from TNP-immune H-2k mice produce interferon-gamma (IFN-gamma), interleukin-2 (IL-2), IL-3, IL-5, tumor necrosis factor-alpha (TNF-alpha), IL-10, and very low levels of IL-4 on reexposure to the specific antigen in vitro. By contrast, T cells from H-2d mice produce IL-3, TNF-alpha, IL-10, and IL-4 but very low levels of IL-2, IL-5 and IFN-gamma. As MHC-congenic matched strains (BALB/k and BALB/c) are used, th…

V gamma 9V delta 2 T lymphocytes efficiently recognize and kill zoledronate-sensitized, imatinib-sensitive, and imatinib-resistant chronic myelogenous leukemia cells.

Abstract Imatinib mesylate (imatinib), a competitive inhibitor of the BCR-ABL tyrosine kinase, is highly effective against chronic myelogenous leukemia (CML) cells. However, because 20–30% of patients affected by CML display either primary or secondary resistance to imatinib, intentional activation of Vγ9Vδ2 T cells by phosphoantigens or by agents that cause their accumulation within cells, such as zoledronate, may represent a promising strategy for the design of a novel and highly innovative immunotherapy capable to overcome imatinib resistance. In this study, we show that Vγ9Vδ2 T lymphocytes recognize, trogocytose, and efficiently kill imatinib-sensitive and -resistant CML cell lines pre…

Aminobisphosphonates as new weapons for gammadelta T Cell-based immunotherapy of cancer.

BACKGROUND: Activated V gamma 9 V delta 2 T cells are able to kill most tumour cells because of recognition by T cell receptor and natural killer receptors. OBJECTIVE: We discuss the possibility that the intentional activation of gammadelta T cells in vivo by aminobisphosphonates may represent a promising target for the design of novel and highly innovative immunotherapy in cancer patients. METHODS: The antitumoral effects of gammadelta T cells both in vitro and in vivo have been demonstrated suggesting a new therapeutic approach for translation into the clinical setting. RESULTS/CONCLUSION: V gamma 9 V delta 2 T lymphocytes represent a particularly interesting target for immunotherapeutic …

Differential activation of human γ δ cells by nonpeptide phosphoantigens

Human T cells expressing Vγ9/Vδ2-encoded TCR recognize several nonpeptide phosphoantigens in the absence of major histocompatibility complex restriction. As these cells respond differentially to increasing concentrations of structurally related phosphoantigens, such ligands constitute agonists of different strengths. By analyzing early cellular events and late effector responses of γ δ T cells, we compared their patterns of stimulation by weak, medium and strong phosphoantigen agonists. We found that, although the early metabolic activation as assessed by cytosensormicrophysiometry directly reflects the intensity of subsequent effector response by γ δ cells, TCR down-modulation is dissociat…

Characterization of lung γδ T cells following intranasal infection with Mycobacterium bovis bacillus Calmette-Guérin

The lungs are considered to have an impaired capacity to contain infection by pathogenic mycobacteria, even in the presence of effective systemic immunity. In an attempt to understand the underlying cellular mechanisms, we characterized the gammadelta T cell population following intranasal infection with Mycobacterium bovis bacillus Calmette-Guerin (BCG). The peak of gammadelta T cell expansion at 7 days postinfection preceded the 30 day peak of alphabeta T cell expansion and bacterial count. The expanded population of gammadelta T cells in the lungs of BCG-infected mice represents an expansion of the resident Vgamma2 T cell subset as well as an influx of Vgamma1 and of four different Vdelt…

Frequency, phenotype and function of Mycobacterium tuberculosis-specific CD8 T cells in patients with active tuberculosis and in individuals with latent infection

Homing and memory patterns of human gammadelta T cells in physiopathological situations.

Vgamma9Vdelta2 are a heterogeneous population of T cells and comprise distinct naive, memory and effector populations that can be distinguished on the basis of surface marker expression and effector functions. We review here these recently studied features of Vgamma9Vdelta2 T lymphocyte biology and the roles they play in infectious and autoimmune diseases.

Homing and memory patterns of human gamma delta T cells in physiopathological situations

An anti-inflammatory role for V alpha 14 NK T cells in Mycobacterium bovis bacillus Calmette-Guérin-infected mice.

Abstract The possible contribution of NKT cells to resistance to Mycobacterium tuberculosis infection remains unclear. In this paper we characterized the Vα14 NKT cell population following infection with Mycobacterium bovis bacillus Calmette-Guérin (BCG). BCG infection determined an early expansion of Vα14 NKT cells in liver, lungs, and spleen, which peaked on day 8 and was sustained until day 30. However, an NK1.1+ Vα14 NKT population preferentially producing IFN-γ predominated at an early stage (day 8), which was substituted by an NK1.1− population preferentially producing IL-4 at later stages (day 30). Despite the fact that Vα14 NKT cell-deficient mice eliminated BCG as did control mice…

Genetic control of C3 production by the S region of the mouse MHC.

SUMMARY The present paper reports evidence indicating that the level of the third complement component (C3) is regulated by the S region of the murine H-2 complex. In fact, using congenic strains of mice we demonstrate that mice with the k haplotype at the S region show high C3 levels, whereas mice with the d haplotype at the S region show low C3 levels.

Analysis of Mycobacterium tuberculosis-Specific CD8 T-Cells in Patients with Active Tuberculosis and in Individuals with Latent Infection

CD8 T-cells contribute to control of Mycobacterium tuberculosis infection, but little is known about the quality of the CD8 T-cell response in subjects with latent infection and in patients with active tuberculosis disease. CD8 T-cells recognizing epitopes from 6 different proteins of Mycobacterium tuberculosis were detected by tetramer staining. Intracellular cytokines staining for specific production of IFN-gamma and IL-2 was performed, complemented by phenotyping of memory markers on antigen-specific CD8 T-cells. The ex-vivo frequencies of tetramer-specific CD8 T-cells in tuberculous patients before therapy were lower than in subjects with latent infection, but increased at four months a…

Resistance of natural killer T cell-deficient mice to systemic Shwartzman reaction.

The generalized Shwartzman reaction in mice which had been primed and challenged with lipopolysaccharide (LPS) depends on interleukin (IL)-12-induced interferon (IFN)-gamma production at the priming stage. We examined the involvement in the priming mechanism of the unique population of Valpha14 natural killer T (NKT) cells because they promptly produce IFN-gamma after IL-12 stimulation. We report here that LPS- or IL-12-primed NKT cell genetically deficient mice were found to be resistant to LPS-elicited mortality. This outcome can be attributed to the reduction of IFN-gamma production, because injection of recombinant mouse IFN-gamma, but not injection of IL-12, effectively primed the NKT …

Pivotal advance: alpha-galactosylceramide induces protection against lipopolysaccharide-induced shock.

Abstract α−galactosylceramide, a natural killer T cell ligand, and its synthetic homolog, KRN7000, consistently influence IFN-γ and TNF-α release, both mediators of LPS-induced shock. To modify the course of endotoxin shock, we injected KRN7000 at different time points of experimental systemic Shwartzman reaction. Mice treated with KRN7000 survived when it was injected within 2 h before and after LPS challenge. Mice survival was associated with low levels of T helper 1 (Th1) cytokines, such as IFN-γ and TNF-α. By contrast, protection from endotoxin shock was associated with an increase of T helper 2 (Th2) cytokines, like IL-4 and IL-10. A role of Th2 cytokines in counteracting LPS-induced s…

Efficient Killing of Human Colon Cancer Stem Cells by γδ T Lymphocytes

Abstract Colon cancer comprises a small population of cancer stem cells (CSC) that is responsible for tumor maintenance and resistant to cancer therapies, possibly allowing for tumor recapitulation once treatment stops. We previously demonstrated that such chemoresistance is mediated by autocrine production of IL-4 through the up-regulation of antiapoptotic proteins. Several innate and adaptive immune effector cells allow for the recognition and destruction of cancer precursors before they constitute the tumor mass. However, cellular immune-based therapies have not been experimented yet in the population of CSCs. Here, we show that the bisphosphonate zoledronate sensitizes colon CSCs to Vγ9…

A human leucocyte antigen-DR1 transgene confers susceptibility to experimental allergic encephalomyelitis elicited by an epitope of myelin basic protein.

Much evidence now indicates that human leucocyte antigen (HLA) class I and class II transgenic (Tg) mice can be of value in analysing HLA-restricted presentation of T-cell epitopes relevant to experimental models of autoimmune diseases. One area where this has been applied is the characterization of myelin epitopes presented by HLA class II molecules in experimental model of multiple sclerosis (experimental allergic encephalomyelitis (EAE)). As a first step towards humanized disease models in HLA Tg mice, we have analysed immune response of lymph node cells of HLA-DR1 Tg mice immunized with the human myelin basic protein (MBP) peptides 13–33, 87–106 and 139–154 bound by HLA-DR1. We report h…

Multifunctional CD4(+) T cells correlate with active Mycobacterium tuberculosis infection.

Th1 CD4(+) T cells and their derived cytokines are crucial for protection against Mycobacterium tuberculosis. Using multiparametic flow cytometry, we have evaluated the distribution of seven distinct functional states (IFN-gamma/IL-2/TNF-alpha triple expressors, IFN-gamma/IL-2, IFN-gamma/TNF-alpha or TNF-alpha/IL-2 double expressors or IFN-gamma, IL-2 or TNF-alpha single expressors) of CD4(+) T cells in individuals with latent M. tuberculosis infection (LTBI) and active tuberculosis (TB). We found that triple expressors, while detectable in 85-90%TB patients, were only present in 10-15% of LTBI subjects. On the contrary, LTBI subjects had significantly higher (12- to 15-fold) proportions of…

Analysis of memory and effector CD8+ T cell subsets in chronic graft-versus-host disease.

In humans, the selective depletion of CD8+ cells may prevent GVHD after allogeneic transplantation. These cells can infiltrate and damage target tissues. It is of interest to investigate the phenotypical characteristics and cytotoxic properties of the different CD8+ subsets in cGVHD patients. In a preliminary study we found that patients with cGVHD had a markedly elevated percentage of peripheral blood CCR7−/CD45RA+ cells compared to patients without cGVHD; conversely, the CCR7+/CD45RA+ subsets of CD8+ cells was significantly decreased. In this study, we report in depth on the phenotype of effector T cell subsets in cGVHD patients, as well as their proliferative capability, cytotoxic prope…

BASI PATOGENETICHE DELLE MALATTIE AUTOIMMUNI

Il "giuramento di G. F. Ingrassia nella peste di Palermo del 1575 nel suo 500° genetliaco

L'autore pone una riflessione sul giuramento nell'attività medica sia dal punto di vista etico che deontologico. Richiama il solenne impegno di Ingrassia nella tutela della città di Palermo colpita dalla peste e discute i punti di attualità.

Role of the chemokine decoy receptor D6 in balancing inflammation, immune activation, and antimicrobial resistance in Mycobacterium tuberculosis infection.

D6 is a decoy and scavenger receptor for inflammatory CC chemokines. D6-deficient mice were rapidly killed by intranasal administration of low doses of Mycobacterium tuberculosis. The death of D6(-/-) mice was associated with a dramatic local and systemic inflammatory response with levels of M. tuberculosis colony-forming units similar to control D6-proficient mice. D6-deficient mice showed an increased numbers of mononuclear cells (macrophages, dendritic cells, and CD4 and CD8 T lymphocytes) infiltrating inflamed tissues and lymph nodes, as well as abnormal increased concentrations of CC chemokines (CCL2, CCL3, CCL4, and CCL5) and proinflammatory cytokines (tumor necrosis factor alpha, int…

Impact of Human Leukocyte Antigen Polymorphisms in Human Immunodeficiency Virus Progression in a Paediatric Cohort Infected with a Mono-phyletic Human Immunodeficiency Virus-1 Strain

Objective: HLA polymorphisms within the peptide binding pocket have been associated with rapid and slowprogression to AIDS, suggesting that the capability to present efficiently HIV-1 epitopes is crucial for the infection control. To minimize the effects of genetic background due to population coming from different geographic area and viral strain variability in the cohort, an analysis of all the polymorphisms associated with the HLA-A, -B and -DR alleles has been performed in a cohort of children with a monophyletic HIV-1 infection (CRF02_AG) during an outbreak in Libya. Methods: High-resolution HLA-typing has been performed in 58 children infected with a monophyletic strain of HIV-1: 26 L…

Peptidi antigenici per la vaccinazione anti-HIV-1

Major histocompatibility complex regulation of interleukin-5 production in the mouse.

Lymph node cells of CBA (H-2k), but not of BALB/c (H-2d) mice immunized epicutaneously with picryl chloryde secrete interleukin (IL)-5 when stimulated with the specific antigen in vitro. The low IL-5 production in BALB/c mice persists when either picryl chloride or the unrelated antigen oxazolone are used, when the amount of antigen in vitro is varied and when a secondary response is studied. The difference in IL-5 production maps to the major histocompatibility complex (MHC) in the congenic BALB/b, BALB/c and BALB/k mice. Furthermore, lymph node cells from (k × d) F1 mice produce IL-5 when stimulated by antigen presented on H-2k but not on H-2d antigen-presenting cells. Finally, the low IL…

Cross-talk between human myeloid dendritic cells and VG9VD2 T cells during M. tuberculosis infection.

Basi patogenetiche delle malattie autoimmuni

Aminobisphosphonate-activated γδ T cells in immunotherapy of cancer: doubts no more

BACKGROUND: Activated V gamma 9 V delta 2 T cells are able to kill most tumour cells because of recognition by T cell receptor and natural killer receptors. OBJECTIVE: We discuss the possibility that the intentional activation of gammadelta T cells in vivo by aminobisphosphonates may represent a promising target for the design of novel and highly innovative immunotherapy in cancer patients. METHODS: The antitumoral effects of gammadelta T cells both in vitro and in vivo have been demonstrated suggesting a new therapeutic approach for translation into the clinical setting. RESULTS/CONCLUSION: V gamma 9 V delta 2 T lymphocytes represent a particularly interesting target for immunotherapeutic …

Mutations and polymorphisms of the PAH gene in Sicily: comparison with other DNA polymorphisms

In utero fetal liver hematopoietic stem cell transplantation: is there a role for alloreactive T lymphocytes?

The use of hematopoietic stem cells for in utero transplantation to create permanent hematochimerism represents a new concept in fetal therapy, although this approach has provided quite heterogeneous results. Flake and Zanjani have provided an excellent updated review of the current knowledge of in

A novel ELISA system for detecting serum granulysin in tuberculosis patients

lE IMMUNODEFICIENZE

Interleukin 4 suppresses primary interferon gamma response by T cells immunized in vivo and cultured in vitro with interleukin 2.

This paper describes a novel primary in vivo/in vitro culture system which allows analysis of the effect of IL-4 added to culture 1 day after immunization on the production of IFN-gamma. Mice are immunized epicutaneously with picryl chloride (TNP) and draining lymph node cells were harvested 1 day later. These cells (1 day lymph node cells), when cultured in vitro for 3 days in the presence of IL-2, either continuously or as a pulse, give an IFN-gamma response on reexposure to antigen 3 days later. This production of IFN-gamma is both antigen-specific and genetically (MHC)-restricted and is due to both CD8+ and CD4+ T cells. However, if 1 day lymph node cells are cultured with both IL-2 and…

IL COMPLESSO MAGGIORE DI ISTOCOMPATIBILITA' ED IL RIGETTO DEL TRAPIANTO

Reversible effect of MR and ELF magnetic fields (0.5 T and 0.5 mT) on human lymphocyte activation patterns.

Purpose: The aim of this study was to investigate the effects of magnetic fields (MF) of different intensity generated by a magnetic resonance (MR) unit (0.5 Tesla) and a double cylindrical coil (0.5 mTesla) on human CD4+ T cell lines. Materials and methods: CD4+ T cells were exposed for two hours under isothermal conditions (37 ± 0.5°C) to the above mentioned MF; a control group was provided for each exposed sample. After exposure, the samples were analysed in the laboratory for the following endpoints: Release of cytokines, expression of surface markers, cell proliferation and levels of cytosolic free-calcium. Results: Exposure to MF for 2 h and subsequent in vitro stimulation in the pres…

Predominance of Vγ9/Vδ2 T lymphocytes in the cerebrospinal fluid of children with tuberculous meningitis: Reversal after chemotherapy

We analyzed the γδ T cell composition and responses in the peripheral blood and cerebrospinal fluid (CSF) of children affected by tuberculous meningitis (TBM) and in control children. Peripheral blood and CSF samples were stimulated with different phosphoantigens and IL-2, and expansion of Vγ9/Vδ2 T cells assessed by FAC S analysis. Vγ9/Vδ2 lines were obtained by culturing CSF or peripheral blood mononuclear cells (PBMC) in vitro with phosphoantigens and IL-2 for 2 months, and tested for proliferation and cytokine production in response to phosphoantigens. Vδ2(D)Jδ junctional sequence length was assessed by PCR. The repertoire of γδ T cells from the CSF of TBM patients was characterized by …

Broad clonal heterogeneity of antigen-specific CD4+ T-cells localizing at the site of disease during tuberculosis

The repertoire of CD4+ T-lymphocytes was investigated in six patients affected by tuberculosis, who had a negative PPD skin test at diagnosis. Polyclonal CD4+ T-cell lines from the peripheral blood failed to proliferate to PPD and to the 16- or 38-kDa proteins of Mycobacterium tuberculosis, while CD4+ T-cell lines from the site of disease responded to PPD, and to the 16- and 38-kDa proteins, and derived epitopes in vitro. The repertoire of CD4+ T-cells accumulating at the site of disease was found to be widely heterogeneous as demonstrated by the finding that at least seven different peptides from the 16- and 38-kDa proteins were recognized by every patient. These results indicate that CD4+…

Role of HLA-B α-3 domain amino acid position 194 in HIV disease progression

HLA class I molecules play a role in the regulation of innate immune response. Therefore, the interaction of HLA class I molecules with different activating and inhibitory receptors leads to balancing the immune response. Among the different family of receptors, NK receptors KIR3DL1/S1 and LIR1, play a major role. Aim of this study was to evaluate the role of amino acid polymorphic positions of HLA class I molecules interacting with NK receptors in HIV progression. In order to minimize the influence of viral variability, a cohort of children with a nosocomial monophyletic HIV-1 infection from the Benghazi Children Hospital has been evaluated. To assess the role of single amino acid position…

Toxocara canis infection induces antigen-specific IL-10 and IFNgamma production in pregnant dogs and their puppies.

Identification of epitopes of Mycobacterium tuberculosis 16-kDa protein recognized by human leukocyte antigen-A*0201 CD8(+) T lymphocytes.

CD8(+) T cells could make an important contribution to protection against tuberculosis (TB), but the antigenic determinants recognized in the context of major histocompatibility complex class I molecules remain ill defined. Our aim was to identify nonamer peptides derived from the acr/16-kDa antigen. Two immunogenic peptides (p21-29 and p120-128) were identified by their ability to elicit cytotoxic CD8(+) T cells from juvenile patients recovering from TB. Epitope-specific recognition was demonstrated by the lysis of both Mycobacterium tuberculosis-infected and peptide-pulsed macrophages, the release of cytotoxic granules, and interferon-gamma and tumor necrosis factor-alpha production. CD8(…

Toxocara canis infection induces antigen-specific IL-10 and IFNγ production in pregnant dogs and their puppies

Toxocara canis (T. canis) is originally a parasite of canine bitches and their pups. The pathogenicity of T. canis infection is enhanced during pregnancy and puppyhood. The aim of this study was to investigate if modification of IFNgamma and IL-10 secretion occurs during infection in pregnant dogs and puppies. Analysis of cytokines secreted could let us hypothesize a role for IL-10 and/or IFNgamma in T. canis infection. We tested T. canis-specific production of IFNgamma and IL-10 by lymphocytes of pregnant dogs and their puppies after in vitro re-exposure to purified excretory/secretory antigen (ESAg) from T. canis. Blood mononuclear cells (BMC) isolated from pregnant dogs and their puppies…

TCR V alpha chain expression influences reactivity to the hapten TNP.

We have recently demonstrated a remarkable selection of in vitro cultivated, TNP-specific polyclonal T cell lines for the expression of a TCR beta chain encoded by the V beta 8.2 gene. The goal of the present study was to analyse V alpha usage in V beta 8.2 T cells responsive to TNP, using TNP-specific T cell lines derived from three common strains of mice, as well as from V beta 8.2 transgenic mice. Results indicate that in vitro TNP stimulation of T cells from TNP-immune mice results in significant skewing of V alpha usage among responding V beta 8.2+ T cells, with overexpression observed for V alpha 3.2 and V alpha 8. These results indicate that V alpha expression influences recognition …

γ δ T Cell Modulation in Anticancer Treatment

The broad antimicrobial and antitumoral reactivity of Vgamma9Vdelta2 T cells, their ability to produce inflammatory cytokines involved in protective immunity against intracellular pathogens and tumors and their strong cytolytic and bactericidal activities suggest their direct involvement in immune control of cancers and infections. gammadelta T cells can be selectively activated by naturally occurring or synthetic phosphoantigens, and drugs that enhance their accumulation into stressed cells, offering new avenues for the development of gammadelta T cell-based immunotherapies. The recent development of small drugs selectively activating Vgamma9Vdelta2 T lymphocytes, which upregulate endogeno…

Differentiation of human Vgamma9 Vdelta 2 T cell subsets by antigen or homeostatic cytokines

The evolution of didactic programs of Italian medical faculties towards an European standard

Damping excessive inflammation and tissue damage in Mycobacterium tuberculosis infection by Toll IL-1 receptor 8/single Ig IL-1-related receptor, a negative regulator of IL-1/TLR signaling.

Abstract Toll IL-1R 8/single Ig IL-1-related receptor (TIR8/SIGIRR) is a member of the IL-1R family, expressed by epithelial tissues and immature dendritic cells, and is regarded as a negative regulator of TLR/IL-1R signaling. Tir8-deficient mice were rapidly killed by intranasal administration of low doses of Mycobacterium tuberculosis, despite controlling efficiently the number of viable bacilli in different organs. Tir8−/−-infected mice showed an increased number of neutrophils and macrophages in the lungs; however, mycobacteria-specific CD4 and CD8 T cells were similar in Tir8−/− and Tir8+/+ mice. Exaggerated mortality of Tir8−/− mice was due to massive liver necrosis and was accompanie…

T Cells Recognize an Immunodominant Epitope of Heat Shock Protein 65 in Kawasaki Disease

Kawasaki disease (KD) is an acute systemic vasculitis of infancy and early childhood that is characterized by endothelial cell damage associated with T-cell activation. Lymphocytes infiltrating damaged tissues might be responsible for the disease through secretion of cytokines, such as tumor necrosis factor (TNF)-α, that could cause fever, as well as endothelial tissue damage. Debate is growing about the nature of antigen responsible for T-cell activation in KD. Bacillus Calmette Guerin (BCG) and purified protein derivative (PPD) hyper-responsiveness was observed in KD patients and this phenomenon was hypothetically ascribed to cross-reactivity between mycobacterial Heat Shock Protein (HSP)…

Efficient cytotoxic activity of zoledronate-activated gamma delta T cells against imatinib CML cell lines.

Differentiation of human CD8+ T cells in BCG vaccinated and tuberculous patients.

Differential requirements for antigen or homeostatic cytokines for proliferation and differentiation of human Vgamma9Vdelta2 naive, memory and effector T cell subsets.

We have compared four human subsets of Vgamma9Vdelta2 T cells, naive (T(naive), CD45RA(+)CD27(+)), central memory (T(CM), CD45RA(-)CD27(+)), effector memory (T(EM), CD45RA(-)CD27(-)) and terminally differentiated (T(EMRA), CD45RA(+)CD27(-)), for their capacity to proliferate and differentiate in response to antigen or homeostatic cytokines. Cytokine responsiveness and IL-15R expression were low in T(naive) cells and progressively increased from T(CM) to T(EM) and T(EMRA) cells. In contrast, the capacity to expand in response to antigen or cytokine stimulation showed a reciprocal pattern and was associated with resistance to cell death and Bcl-2 expression. Whereas antigen-stimulated cells a…

ESAT-6 Peptide Recognition by Bovine CD8 + Lymphocytes of Naturally Infected Cows in Herds from Southern Italy

ABSTRACT The aim of this study was to define epitopes of Mycobacterium bovis from ESAT-6 (early secretory antigen of 6 kDa) recognized by CD8 + T lymphocytes from cows naturally infected with Mycobacterium bovis . We found that bovine CD8 + T cells recognized 10 out of 11 ESAT-6 peptides tested.

Characterization of HLA-DR- and TCR-binding residues of an immunodominant and genetically permissive peptide of the 16-kDa protein of Mycobacterium tuberculosis.

The 16-kDa protein of Mycobacterium tuberculosis represents an important antigenic target during bacillary latency and, consequently, should be considered as candidate subunit vaccine component. In this study, we have used CD4 T cell clones that recognize the peptide p91-110, an immunodominant and genetically permissive epitope, in the context of five different HLA-DR molecules and truncated and substituted variants of this peptide, to identify the minimal binding sequence (HLA-DR-binding core) and the minimal stimulatory sequence (TCR-binding core), as well as the residues that contact HLA-DR molecules and the TCR. We have found a common 9-mer sequence, spanning amino acids 93-101, as the …

Decreased serum granulysin levels in childhood tuberculosis which reverse after therapy

Abstract Granulysin is a cytolytic protein of natural killer (NK) cells and cytotoxic T lymphocytes (CTLs). Serum levels of granulysin are related to host cellular immunity. We used an ELISA to quantify granulysin serum levels in children with tuberculosis (TB), before and after chemotherapy. The study involved children affected by different clinical forms of TB (n=72) and healthy control children (n=150) from the same geographical area and of similar socio-economic background. Serum granulysin levels before the initiation of TB therapy were significantly lower in children with TB compared to controls, with the lowest levels being found in TB patients who were PPD skin test negative. No sta…

Increase of CCR7- CD45RA+ CD8 T cells (TEMRA) in Chronic Graft-versus-host Disease

Among the late effects of hematopoietic stem cell transplantation (HSCT), chronic graft-vs-host disease (cGVHD) still remains as the major determinant of long-term outcome and quality of life. The disease typically appears between 3 months to 1.5 years following an allogeneic transplantation and is characterized by symptoms similar to those of autoimmune disease.

HLA Class I and Class II Polymorphism in Three Sicilian Populations

Two human leukocyte antigen (HLA) class I loci (HLA-A and HLA-B) and one class II locus (HLA-DR) were typed at the DNA level in the Sicilian population. Study participants were of Sicilian origin (183 for class I loci and 260 for class II loci) and live in three towns, chosen on the basis of geographic position and different historical events. These towns are Sciacca (southwest Sicily, located at sea level, conquered by Arabs in a.d. 814), Piana degli Albanesi (northwest Sicily, 720 m above sea level, has maintained religious, cultural, and linguistic peculiarities traced to Albanian settlement in 1488), and Troina (northeast Sicily, 1,120 m above sea level, known as the first settlement of…

Cross-talk between Vβ8+and γδ+T lymphocytes in contact sensitivity

We have previously reported that T lymphocytes proliferating in vitro to the hapten trinitrochlorobenzene (TNCB) exhibit a very restricted V beta gene usage and response to TNCB is limited to T-cell receptors (TCR) composed of V beta 8.2 in combination with V alpha 3.2, V alpha 8 and V alpha 10. This paper investigates the role played by T lymphocytes expressing the V beta 8.2 gene segment in the contact sensitivity (CS) reaction to TNCB in the intact mouse and in its passive transfer into naive recipient mice. Mice injected with monoclonal antibodies to V beta 8 are unable to develop CS upon immunization with TNCB and 4-day TNCB-immune lymph node cells from mice that had been depleted in v…

Characterization of HLA-DR- and TCR-binding residues of an immunodominant and genetically permissive peptide of the 16-kDa protein of Mycobacterium tuberculosis

Vγ9 / Vδ2 T lymphocytes reduce the viability of intracellularMycobacterium tuberculosis

An effective immune response against the intracellular pathogen Mycobacterium tuberculosis is strictly dependent on T cell activation. Although this protective response mainly depends on local release of pro-inflammatory cytokines by Th1 CD4(+) T cells, contribution of Vgamma9 / Vdelta2 T lymphocytes to immune protection against this pathogen is suggested by the antimycobacterial reactivity of this subset and its ability to produce large amounts of Th1 cytokines. Here we show that Vgamma9 / Vdelta2 T lymphocytes kill macrophages harboring live M. tuberculosis. The cytotoxic activity of Vgamma9 / Vdelta2 T lymphocytes was not MHC class I or class II restricted but was blocked by anti-TCR mon…

Immunomodulating role of bisphosphonates on human gamma delta T cells: an intriguing and promising aspect of their antitumour activity.

Vgamma9Vdelta2 T cells have the ability to produce inflammatory cytokines involved in protective immunity against intracellular pathogens and tumours and to display strong cytolytic as well as bactericidal activities. This suggests a direct involvement of Vgamma9Vdelta2 T lymphocytes in immune control of cancer and infections. These observations have recently aided development of novel immunotherapeutic approaches aimed at Vgamma9Vdelta2 T cell activation. Nitrogen-containing bisphosphonates (N-BPs) play a crucial role in Vgamma9Vdelta2 T lymphocyte activation and in the acquisition of effector functions. The preliminary results of these innovative strategies are encouraging. Moreover, comp…

IMMUNOLOGICAL TYPING AND ORGAN TRANSPLANTATION

Cytokine profile, HLA restriction and TCR sequence analysis of human CD4+ T clones specific for an immunodominant epitope of Mycobacterium tuberculosis 16-kDa protein

SUMMARY The identification of immunodominant and universal mycobacterial peptides could be applied to vaccine design and have an employment as diagnostic reagents. In this paper we have investigated the fine specificity, clonal composition and HLA class II restriction of CD4+ T cell clones specific for an immunodominant epitope spanning amino acids 91–110 of the 16-kDa protein of Mycobacterium tuberculosis. Twenty-one of the tested 28 clones had a Th1 profile, while seven clones had a Th0 profile. None of the clones had a Th2 profile. While the TCR AV gene usage of the clones was heterogeneous, a dominant TCR BV2 gene family was used by 18 of the 28 clones. The CDR3 regions of BV2+ T cell c…

Three cell subsets are required for the transfer of delayed-type hypersensitivity reaction by antigen-specific T cell lines.

Antigen (trinitrochlorobenzene)-specific T cell lines were obtained by repeated stimulation of lymph node cells from immune mice with antigen in vitro. These T cell lines, consisting of more than 90% CD4+ Vbeta8.2+ and 6 to 9% gammadelta+ T lymphocytes, transfer contact sensitivity (CS) locally when injected at the same site as the challenge antigen, but fail to mediate a systemic passive transfer when injected i.v. Injection of T cell lines together with spleen cells from mice immunized 1 day beforehand (1-day cells) allowed a successful, specific systemic transfer of CS. Phenotypic analysis showed that the 1-day immune cell was alphabeta+, gammadelta-, sIg-, CD3+, CD4-, CD8-, CD5+, B220 (…

Augmented Passive Transfer of Contact Sensitivity in Severe Combined Immunodeficiency Mice and Its Dependence of Vβ8<sup>+</sup> Cells in the Picryl System

The passive transfer of contact sensitivity using picryl chloride immune cells from H-2 syngenic BALB/c donors was analyzed in severe combined immunodeficiency (SCID) mice which lack functional T and B lymphocytes. H-2-restricted and antigen-specific contact sensitivity was transferred to SCID mice, and comparison between the level of contact sensitivity and the number of transferred cells showed a significantly more efficient transfer to SCID than to BALB/c mice. The cells passively transferring contact sensitivity were shown to carry the Vβ8 phenotype. Moreover, chromium-labeled cells from BALB/c PC1-primed donors localize normally in peripheral lymphoid organs, and an increased percentag…

Giuseppe Pitrè Quaderni di medicina e chirurgia

Th0 to Th1 switch of CD4 T cell clones specific from the 16-kDa antigen of Mycobacterium tuberculosis after successful therapy: lack of involvement of epitope repertoire and HLA-DR.

Abstract In this study, we have examined the influence of HLA-DR molecules and the structure of the epitope repertoire of the 16-kDa protein of Mycobacterium tuberculosis on the acquisition of the cytokine secretion pattern of CD4 T cell clones, obtained from tuberculous patients before and after anti-mycobacterial therapy. Our data indicate that TB patients have a predominant Th0 response against the 16-kDa protein and its epitopes and that healing, induced by anti-mycobacterial therapy, is associated with a shift toward a predominant Th1 phenotype. Moreover, both HLA-DR molecules restricting the clone specificity and the nature of the recognized epitope do not play any role in the generat…

The molecular basis of the low hemolytic activity of C4 molecules from low-C4 mice with IgM-coated erythrocytes.

This study investigated the origin of the different hemolytic activity of two allotypes of murine C4, C4H (C4-high) and C4L (C4-low) in the presence of IgM-coated erythrocytes. C4H displayed a threefold higher hemolytic titer (expressed in hemolytic units/microgram protein) than C4L. No difference was found between c4H and C4L either in stability at 37 degrees C at different pH values and in the rate of C4H and C4L hydrolysis by activated Cl. The major functional difference was found in the covalent binding capacity to IgM-coated erythrocytes, with the amount of C4H bound being about threefold higher than that of C4L. A marked difference in the reactivity of the C4b fragment of C4H and C4L …

Il Complesso Maggiore di Istocompatibilità ed il Rigetto dei Trapianti

Role of IL-4 in delayed type hypersensitivity

SUMMARY IL-4 plays a key role in the contact sensitivity skin reaction. This has several implications. First, the view that contact sensitivity (CS) is only mediated by cells with a Th1 profile of cytokine secretion needs modification, in the light of the essential role of IL-4 at the effector stage. Second, the concept of a single cell involved in the systemic transfer of CS is no longer tenable, as it is known that both αβ and γδ cells are required. Studies with the cell lines (which contain both αβ and a few γδ cells) suggest that this double requirement may involve the action of IL-4 on γδ cells, which bear receptors for IL-4. Finally, the view that T cell lines only transfer CS when in…

Memory and effector CD8 T cell subset in human tuberculosis.

Aspetti medico sanitari, etici e socio economici dell'attività solfifera in Sicilia (1791-1964)

Role of individual's T cell immunome in controlling HIV-1 progression

Viral and host factors can influence HIV-1 progression, among them human leucocyte antigen (HLA) has shown the strongest effect. However, studies on the functional contribution of HLA in controlling HIV progression toward AIDS are limited by multiple issues, including the viral strain variability within the study subjects. In this study, in a cohort of children infected with a monophyletic strain (CRF02_AG) during an outbreak, we evaluated the HIV-1 Gag, Vif, Vpr, Tat and hepatitis C virus E1/E2 (as control) proteins circulating in a cohort for the capability to be presented by the HLA molecules in the same population. A total of 70 Non-progressors and 37 Progressors to AIDS were evaluated.…

Change of Th0 to Th1 cell-cytokine profile following tuberculosis chemotherapy.

T cells mediate protection against tuberculosis, but little is known about their role during chemotherapy of patients with active disease. Here we examined the cytokine profile of CD4 T cells before and after four months of chemotherapy in six initial skin test anergic cases. Purified protein derivative (PPD) and 16-kDa antigen-reactive CD4 T-cell clones prior to therapy resided mostly in disease-associated body fluids and were of the Th0 (interferon (IFN)-gamma + interleukin (IL)-4) secreting profile. In contrast, the majority of postchemotherapy CD4 T-cell clones originated from blood and were of the IFN-gamma secreting Th1 type. However, the recognition of several peptides derived from t…

Aminobisphosphonates as New Weapons for γ δ T Cell-Based Immunotherapy of Cancer

Several observations in mice and in humans have collectively laid the foundation for examining the potential of γ δ T cells to exert tumor immunotherapy. Human γ δ T cells can be activated in a non-MHC dependent fashion either by low molecular mass phosphoantigens, or by agents that provoke the accumulation of endogenous pyrophosphates such as isopentenylpyrophosphate. Among the latter, aminobisphosphonates are well-established in the clinic, and extensive data are available on the compounds antiangiogenic, antiosteolytic and pro-apoptotic properties. In this review we discuss on the possibility that the intentional activation of γ δ T cells in vivo by aminobisphosphonates may represent a p…

Prevention of the post-chemotherapy relapse of tuberculous infection by combined immunotherapy

Summary We report that a recently developed combined immunotherapy (CIT) has the capacity to prevent a spontaneous relapse of replicating Mycobacterium tuberculosis bacilli in the lungs of BALB/c, C57Bl/6 or C3H/HeJ strains of mice, following 4 weeks of non-sterilising treatment with isoniazid and rifampicin. The CIT regimen, represented by recombinant IFNγ, anti-α crystalline monoclonal IgA antibody and IL-4 neutralizing polyclonal antibody, reduced the 8-week relapse of viable bacterial counts in the lungs most significantly, when CIT was inoculated during the 5th week post infection, i.e. during the 3rd week of chemotherapy. Although CIT enhanced lung granuloma area, nitric oxide, cytoki…

Natural killer and lymphokine-activated killer activity in HLA-B8,DR3-positive subjects.

Abstract The haplotype HLA-B8,DR3 is over-represented in several autoimmune diseases, implying that genes predisposing people to these disorders are linked to this haplotype. In these diseases, various dysfunctions reflecting an impairment of the immune system have been found. Several reports indicate also that in HLA-B8,DR3-positive healthy subjects similar disorders may be demonstrated. In the present work, we have evaluated NK and LAK activity in these subjects. The study has been performed on monocyte-depleted peripheral blood MNCs by using the K-562 cell line as a target for NK activity and the HL-60 cell line for as a target LAK activity. LAK cells were obtained by incubating MNCs for…

Novel insight into T cell immune response against Mycobacterium tuberculosis

Carcinoembryonic Antigen as a Monitor in Breast Cancer

: Carcinoembryonic antigen (CEA) levels were serially determined in 125 patients with breast cancer in order to study the diagnostic and prognostic use of serum CEA levels before and/or after surgery and during treatment with hormonal and chemotherapy. Serum CEA levels were elevated in 15.5% of nonmetastatic patients. Carcinoembryonic antigen increased according to stage (TNM classification); and a direct relationship between positive CEA levels and subsequent recurrence was found. After a three-year postoperative interval a 50% survival rate was exhibited in CEA-positive patients vs an 88% survival rate in those patients found to be CEA negative. There is a definite correlation between the…

gd T cells condition dendritic cells in vivo for priming pulmonary CD8 T cell responses against Mycobacterium tuberculosis

gammadelta T cells and dendritic cells are quickly recruited to the lungs shortly after intranasal vaccination with BCG, but the functional in vivo interplay between these two cell populations and its role in the induction of adaptive immune responses is unclear. Using TCR-deficient mice and bone marrow chimeras, we show here that gammadelta T cells provide a non-redundant early source of IFN-gamma in vivo, which enhances IL-12 production by lung dendritic cells. The in vivo-conditioned dendritic cells, in turn, prime a more efficient lung CD8 T cell response against Mycobacterium tuberculosis. Thus, strategies exploiting gammadelta T cell function and IFN-gamma production could be valuable…

Reversible effect of magnetic fields on human lymphocyte activation patterns: different sensitivity of naive and memory lymphocyte subsets.

The aim of this study was to investigate the influence of 50 Hz magnetic or static magnetic fields of 0.5 mT on subsets of human CD4(+) T cells in terms of cytokine release/content, cell proliferation and intracellular free calcium concentration. CD4(+) T cells can be divided into different subsets on the basis of surface marker expression, such as CD45, and T cells can be divided into naive (CD45RA(+)) and memory (CD45RA(-)) cells. In this study, the effects of magnetic fields after 24 and 48 h of cell culture were analyzed. We found that the CD4(+)CD45RA(-) T subset were more sensitive after 2 h of exposure. Decreases in the release/content of IFN-gamma, in cell proliferation and in intra…

A continuous infusion of a minor histocompatibility antigen-immunodominant peptide induces a delay of male skin graft rejection.

Abstract We previously reported that an inhibition of antigen-specific Interferon-γ release and cytotoxicity occurs after a continuous infusion of an HY immunodominant peptide although this treatment is not able to cause a significant delay of male skin grafts rejection. In vivo administration of high doses of an HY peptide, through mini-osmotic pumps, in naive female mice was used to study the effects on the male skin grafts rejection. A continuous infusion of 1 mg of an HY peptide induces a significant delay of male skin graft rejection. In vitro HY-specific Interferon-γ release was inhibited adding peptide-specific suppressor cells: the ability to inhibit Interferon-γ release was evident…

The effect of cyclosporin A, FK506 and rapamycin on the murine contact sensitivity reaction

We have evaluated the effects of three potent immunosuppressive agents, cyclosporin A (CsA), FK506 and rapamycin, on the murine contact sensitivity (CS) reaction to the hapten trinitrochlorobenzene. Development of CS reaction requires participation of three distinct T cell subsets: alphabeta+, CD4+ T lymphocytes, which are the classical effector cell of the CS reaction, gammadelta+ T lymphocytes, and alphabeta+, double-negative (CD4- CD8-) T lymphocytes that express the B220 molecule and produce IL-4. We found that all three drugs inhibit the development of the CS reaction, but they affect different target cells. In fact, rapamycin and FK-506 block both alphabeta+, CD4+ and gammadelta+ T ly…

Selection of distinct Valpha/beta T-cell receptor families during in vivo and in vitro T-cell maturation.

The experimental conditions influencing the use of Valphabeta TCR families were examined in lymph node (LN) cells from peptide-immunized C57BL/6 and Vbeta8.2 transgenic mice. Expanded proportions of Vbeta5, Vbeta8.2, Vbeta9, Vbeta12 and Vbeta14 positive cells and an association of Vbeta8.2 with Valpha11 was found in freshly harvested 8-day or 34-day immune LN cells. In contrast, peptide-specific T-cell lines generated in vitro from 8-day immune lymph node cells were found to be almost exclusively of the Valpha2/Vbeta12 family. However, T-cell lines originating from Vbeta8.2 transgenic mice did not show preferential Valpha usage. Anti-Vbeta8.2 antibody produced different effects: when added …

Development of a post-exposure paediatric anti HIV-AIDS vaccine based on the combined synthetic/recombinant HIV peptides and BCG for boosting innate and acquired immunity. North-South transfer in Biotechnology of Tuberculosis and AIDS. University of Rome “Tor Vergata”. UNESCO Interdisciplinary Chair in Biotechnology. Vittorio Colizzi, Luc Montagnier, 2004.

New insight on immunological activation pathways of Langerhans cells, possible tolerogenic role

Langerhans cells are the prototype of antigen presenting cell, their role is to work as sentinel in the epidermis. Like every APC Langerhans cells act as bridge between innate and acquired immunity recognizing antigens into the epidermis and bringing them to drying lymph node, their work is well described by the Langerhans cell paradigm. Recently many works designed a new and amazing role of Langerhans cells in fact they often showed tolerogenic capacity, while in many cases, they seems not necessary to promote activation of acquired immunity. Anyway Langerhans cells remaining an interesting target for new vaccine strategies because of their localization in the epidermis and the ability to …

Activation of human V gamma9 V delta 2 T cells by plasmacytoid dendritic cells

gammadelta T Cell Modulation in Anticancer Treatment

The broad antimicrobial and antitumoral reactivity of Vgamma9Vdelta2 T cells, their ability to produce inflammatory cytokines involved in protective immunity against intracellular pathogens and tumors and their strong cytolytic and bactericidal activities suggest their direct involvement in immune control of cancers and infections. gammadelta T cells can be selectively activated by naturally occurring or synthetic phosphoantigens, and drugs that enhance their accumulation into stressed cells, offering new avenues for the development of gammadelta T cell-based immunotherapies. The recent development of small drugs selectively activating Vgamma9Vdelta2 T lymphocytes, which upregulate endogeno…

Granulysin‐Dependent Killing of Intracellular and ExtracellularMycobacterium tuberculosisby Vγ9/Vδ2 T Lymphocytes

Contribution of Vgamma9/Vdelta2 T lymphocytes to immune protection against Mycobacterium tuberculosis is still a matter of debate. It was reported earlier that Vgamma9/Vdelta2 T lymphocytes kill macrophages harboring live M. tuberculosis through a granule-dependent mechanism that results in killing of intracellular bacilli. This study found that Vgamma9/Vdelta2 T lymphocytes reduce the viability of both extracellular and intracellular M. tuberculosis. Granulysin and perforin, both detected in Vgamma9/Vdelta2 T lymphocytes, play a major role, which indicates that Vgamma9/Vdelta2 T lymphocytes directly contribute to a protective host response against M. tuberculosis infection.

CXCR5 identifies a subset of Vgamma9 Vdelta2 T cells which secrete IL-4 and IL-10 and help B cells for antibody production.

Vgamma9Vdelta2 T lymphocytes recognize nonpeptidic Ags and mount effector functions in cellular immune responses against microorganisms and tumors, but little is known about their role in Ab-mediated immune responses. We show here that expression of CXCR5 identifies a unique subset of Vgamma9Vdelta2 T cells which express the costimulatory molecules ICOS and CD40L, secrete IL-2, IL-4, and IL-10 and help B cells for Ab production. These properties portray CXCR5+ Vgamma9Vdelta2 T cells as a distinct memory T cell subset with B cell helper function.

Differential requirements for antigen or homeostatic cytokines for proliferation and differentiation of human Vgamma9Vdelta2 naive, memory and effector T cell subsets.

We have compared four human subsets of Vgamma9Vdelta2 T cells, naive (T(naive), CD45RA(+)CD27(+)), central memory (T(CM), CD45RA(-)CD27(+)), effector memory (T(EM), CD45RA(-)CD27(-)) and terminally differentiated (T(EMRA), CD45RA(+)CD27(-)), for their capacity to proliferate and differentiate in response to antigen or homeostatic cytokines. Cytokine responsiveness and IL-15R expression were low in T(naive) cells and progressively increased from T(CM) to T(EM) and T(EMRA) cells. In contrast, the capacity to expand in response to antigen or cytokine stimulation showed a reciprocal pattern and was associated with resistance to cell death and Bcl-2 expression. Whereas antigen-stimulated cells a…

Phenotypical and functional analysis of memory and effector human CD8 T cells specific for mycobacterial antigens

Abstract Mycobacterium tuberculosis infects one-third of the global population and claims two million lives every year. Because memory CD8 T cells exhibit a high heterogeneity in terms of phenotype and functional characteristic, we investigated the frequency, phenotype, and functional properties of Ag85A epitope-specific HLA-A*0201 CD8 T cells in children affected by tuberculosis (TB) before and 4 mo after chemotherapy and healthy contact children. Using Ag85A peptide/HLA-A*0201 pentamer, we found a low frequency of blood peptide-specific CD8 T cells in tuberculous children before therapy, which consistently increased after therapy to levels detected in healthy contacts. Ex vivo analysis of…

Tuning inflammation in tuberculosis: the role of decoy receptors

Decoy receptors are "silent scavengers" of CC chemokines and cytokines, which play a key role in damping inflammation and tissue damage. In this review we discuss on recent findings demonstrating that these receptors set the balance between antimicrobial resistance, immune activation and inflammatory response in Mycobacterium tuberculosis infection.

HUMAN MEMORY VGAMMA9 VDELTA2 T CELLS REQUIRE HOMEOSTATIC CYTOKINES FOR PROLIFERATION AND DIFFERENTIATION.

WHOLE BODY IRRADIATION INDUCES IFN-γ PRODUCTION IN BALB/c MICE BY PREVENTING THE APPEARANCE OF A Vα14+NK T DOWNREGULATORY POPULATION

Lymph node cells from TNCB-immune BALB/c mice fail to produce IFN-gamma when exposed to antigen in vitro. Conversely, lymph node cells of irradiated (550 rads) BALB/c mice produce IFN-gamma. Transfer experiments show that normal BALB/c mice contain cells which suppress IFN-gamma production. These downregulatory cells are CD4(+)alpha beta(+)and rearrange the invariant V alpha 14-J alpha 281 T cell receptor alpha chain, thus belonging to the NK T cell subset. Downregulatory cells probably act by producing IL-4 as their effect is blocked by mAb to IL-4.

Autosomal microsatellite and mtDNA genetic analysis in Sicily (Italy).

DNA samples from 465 blood donors living in 7 towns of Sicily, the largest island of Italy, have been collected according to well defined criteria, and their genetic heterogeneity tested on the basis of 9 autosomal microsatellite and mitochondrial DNA polymorphisms for a total of 85 microsatellite allele and 10 mtDNA haplogroup frequencies. A preliminary account of the results shows that: a) the samples are genetically heterogeneous; b) the first principal coordinates of the samples are correlated more with their longitude than with their latitude, and this result is even more remarkable when one outlier sample (Butera) is not considered; c) distances among samples calculated from allele an…

HLA-A, -B and -DRB1 allele frequencies in Cyrenaica population (Libya) and genetic relationships with other populations.

Abstract The frequencies of HLA-A, HLA-B and HLA-DRB1 alleles in 118 unrelated Libyans from Benghazi (Cyrenaica) were analysed using high resolution typing and compared with other populations. Their relatedness has been tested by correspondence analyses and principal component analysis. The most frequent HLA-A alleles were A ∗ 02:01:01:01 (15.7%), A ∗ 01:01:01:01 (11.4%) and A ∗ 03:01:01:01 (9.3%). For the HLA-B locus, the commonest allele was HLA-B ∗ 50:01:01 (14.4%) followed by B ∗ 51:01:01 (9.8%) and B ∗ 08:01:01 (6.4%). For the HLA-DRB1 locus, the commonest was HLA-DRB1 ∗ 07:01:01:01 (16.9%) followed by DRB1 ∗ 03:01:01:01 (13.6%) and DRB1 ∗ 13:02:01 (9.3%). The most frequent two-locus h…

mtDNA analysis of the human remains buried in the sarcophagus of Federico II

Abstract The sarcophagus containing the remains of Federico II, located in the Cathedral of Palermo (Sicily, Italy), was opened on 1998 to perform a multidisciplinary survey [1]. Next to the remains of Federico II and in close contact with them were laying two other skeletons belonging, according to historical records, to Pietro II di Aragona and to an anonymous person (“The Third Individual”), probably a woman. The bones appeared severely deteriorated. Chemical analysis performed on bone samples excluded that the bodies underwent some kind of embalming process. The analysis of mtDNA from bone samples taken from the three skeletons was successful in only one of the two labs involved. The HV…

IL-4 depletion enhances host resistance and passive IgA protection against tuberculosis infection in BALB/c mice.

The influence of Th2 cytokines in tuberculosis has been a matter of dispute. Here we report that IL-4 has a profound regulatory effect on the infection of BALB/c mice with Mycobacterium tuberculosis. Depletion of IL-4 with a neutralizing mAb caused only evanescent reduction of lung infection, but when combined with i.n. inoculations of IgA anti-mycobacterial alpha-crystallin mAb and mouse rIFN-gamma, we observed a 40-fold reduction of the bacterial counts in the lungs at 3 wks following i.n. infection (p<0.001). In genetically deficient IL-4-/- BALB/c mice, infection in both lung and spleen was substantially reduced for up to 8 wks without further treatment. Reconstitution of IL-4-/- mice w…

Biology of gama delta T Cells in Tuberculosis and Malaria

Tuberculosis and malaria remain the leading causes of mortality among human infectious diseases in the world. It is estimated that 3 to 5 million people die from tuberculosis and malaria each year. Although it is traditionally believed that CD4 and CD8 alphabeta T lymphocytes are mandatory for protective immune responses against Mycobacterium tuberculosis and Plasmodium falciparum (the ethiologic agents of tuberculosis and the most severe form of malaria, respectively), there is still incomplete understanding of the mechanisms of immune protection and of the causes of its failure in the affected patients. Several studies in humans and animal models have suggested that Vgamma9/Vdelta2 T cell…

Targeting human {gamma}delta} T cells with zoledronate and interleukin-2 for immunotherapy of hormone-refractory prostate cancer.

Abstract The increasing evidence that γδ T cells have potent antitumor activity suggests their value in immunotherapy, particularly in areas of unmet need such as metastatic carcinoma. To this end, we initiated a phase I clinical trial in metastatic hormone-refractory prostate cancer to examine the feasibility and consequences of using the γδ T-cell agonist zoledronate, either alone or in combination with low-dose interleukin 2 (IL-2), to activate peripheral blood γδ cells. Nine patients were enlisted to each arm. Neither treatment showed appreciable toxicity. Most patients were treated with zoledronate + IL-2, but conversely only two treated with zoledronate displayed a significant long-te…