Biomedical subjects

Thomas R Malek

Publications and source records attributed to Thomas R Malek.

17 recordsLinked to original sources

PubMed2007

Cytokine-dependent Blimp-1 expression in activated T cells inhibits IL-2 production.

After Ag activation of naive T cells in vitro, extensive growth and differentiation into effector cells depend upon IL-2. DNA microarray analysis was used to identify IL-2-dependent molecules regulating this process. In this study, we show that the transcriptional repressor B lymphocyte-induced maturation protein 1 (Blimp-1) is expressed by a cytokine-dependent pathway in activated T lymphocytes. IL-2 production by activated CD4(+) and CD8(+) T cells inversely correlated with Blimp-1 levels as higher IL-2 production was associated with lower Blimp-1 expression. Furthermore, ectopic expression of Blimp-1 by activated T cells inhibited IL-2 production but enhanced granzyme B and CD25 expression. Collectively, these findings indicate that there is a negative feedback regulatory loop in activated T cells such that IL-2 inhibits its own production through induction of Blimp-1 while promoting an effector cell phenotype.

Animals↗

PubMed2006

Selective availability of IL-2 is a major determinant controlling the production of CD4+CD25+Foxp3+ T regulatory cells.

The development and maintenance of T regulatory (Treg) cells critically depend on IL-2. This requirement for IL-2 might be due to specificity associated with IL-2R signal transduction or because IL-2 was uniquely present in the niche in which Treg cells reside. To address this issue, we examined the capacity of IL-7R-dependent signaling to support Treg cell production and prevent autoimmunity in IL-2Rbeta(-/-) mice. Expression of transgenic wild-type IL-7R or a chimeric receptor that consisted of the extracytoplasmic domain of the IL-7R alpha-chain and the cytoplasmic domain of IL-2R beta-chain in IL-2Rbeta(-/-) mice did not prevent autoimmunity. Importantly, expression of a chimeric receptor that consisted of the extracytoplasmic domain of the IL-2R beta-chain and the cytoplasmic domain of IL-7R alpha-chain in IL-2Rbeta(-/-) mice led to Treg cells production in the thymus and periphery and prevented autoimmunity. Signaling through the IL-2R or chimeric IL-2Rbeta/IL-7Ralpha in vivo or the culture of thymocytes from IL-2Rbeta(-/-) mice with IL-7 led to up-regulation of Foxp3 and CD25 on Treg cells. These findings indicate that IL-7R signal transduction is competent to promote Treg cell production, but this signaling requires triggering through IL-2 by binding to the extracytoplasmic portion of the IL-2R via this chimeric receptor. Thus, a major factor controlling the nonredundant activity of the IL-2R is selective compartmentalization of IL-2-producing cells with Treg cells in vivo.

Animals↗

PubMed2006

Redundant and unique regulation of activated mouse B lymphocytes by IL-4 and IL-21.

IL-21 distinctively regulates B cell growth and death, and it redundantly functions with IL-4 for IgG production. B cells likely encounter IL-4 and IL-21 in vivo, as both are secreted by activated T cells. Therefore, the action of both these cytokines was investigated during activation of B cells. IL-21 or the combination of IL-4 and IL-21 inhibited proliferation by purified mouse B cells to LPS or CpG DNA, whereas these cytokines enhanced proliferation after engaging the BCR or CD40. Although B cell subsets expressed somewhat varied levels of the IL-21 receptor, LPS-stimulated follicular and marginal B cell subsets were also dominantly susceptible to IL-21-induced growth arrest and cell death. After activation of B cells with CD40 and LPS, IL-4 and IL-21 distinctively regulated the expression of CD23, CD44, and CD138, and they cooperatively promoted IgG1 class-switching and synthesis. These findings support a model in which the presence of IL-4 and IL-21 inhibits B cells activated by polyclonal innate signals, and they promote B cell expansion and differentiation during T cell-dependent antibody responses, although the individual responses to IL-4 and IL-21 do not always overlap.

Animals↗

PubMed2006

Cutting edge: allogeneic CD4+CD25+Foxp3+ T regulatory cells suppress autoimmunity while establishing transplantation tolerance.

An important unresolved question with regard to T regulatory (Treg) cell specificity and suppressive activity is whether allogeneic Treg cells inhibit self-reactive T cells. In the present study, this issue was addressed using IL-2Rbeta-deficient mice that develop rapid lethal autoimmunity due to impaired production of Treg cells. We show that adoptive transfer of completely MHC-mismatched Treg cells into IL-2Rbeta(-/-) mice resulted in life-long engraftment of the donor cells, which exhibited skewed reactivity toward host alloantigens, and prevented autoimmunity. Thus, Treg cells that underwent thymic selection by peptide/MHC class II complexes distinct from those recognized by autoreactive T cells, still effectively suppress autoimmunity. Remarkably, when such animals were skin grafted, they exhibited dominant tolerance to those grafts bearing MHC molecules that were shared with donor Treg cells. Collectively, these data demonstrate that effective engraftment by allogeneic Treg cells controls autoimmunity and results in permissive conditions for long-term acceptance of allografts.

Adoptive Transfer↗

PubMed2006

Quantitative assessment concerning the contribution of IL-2Rbeta for superantigen-mediated T cell responses in vivo.

IL-2- and IL-2R-deficient mice readily develop T cell-dependent immune responses in vivo, but the relevance of this finding is complicated by severe concurrent autoimmunity. Furthermore, the detection of such responses does not address whether under normal circumstances IL-2 dominates T cell immunity. In the present report, we investigated the extent IL-2-independent T cell growth is mediated by other cytokines in the IL-2 family and compared such responses to those generated by IL-2/IL-2R-sufficient T cells. T cell expansion and contraction to the superantigen staphylococcal enterotoxin A (SEA) by autoimmune-free IL-2Rbeta-/- CD4 and CD8 T cells were comparable to normal control mice, although their CD8+ T cells did not optimally develop into IFNgamma-producing effector cells. The proliferation by these IL-2Rbeta-deficient T cells in vivo was independent of IL-2, IL-4 and IL-15 and not blocked by mAbs to the common gamma chain. However, in co-adoptive transfer experiments, wild-type T cells exhibited somewhat more extensive proliferation than IL-2Rbeta-deficient T cells to SEA and this difference was almost entirely accounted for by CD8+ T cells. Collectively, these data indicate that substantial T cell proliferation occurs in the absence of responsiveness to cytokines in the IL-2 family, although maximal T cell proliferation and development of IFNgamma-producing effector CD8+ T cells depend upon IL-2Rbeta.

Adoptive Transfer↗

PubMed2005

Essential role for interleukin-2 for CD4(+)CD25(+) T regulatory cell development during the neonatal period.

Although many aspects of CD4(+)CD25(+) T regulatory (T(reg)) cell development remain largely unknown, signaling through the IL-2R represents one feature for the production of T(reg) cells. Therefore, the present study was undertaken to further define early developmental steps in the production of T(reg) cells, including a more precise view on the role of interleukin (IL)-2 in this process. After adoptive transfer of wild-type T(reg) cells into neonatal IL-2Rbeta(-/-) mice, only a small fraction of donor T(reg) cells selectively seeded the lymph node (LN). These donor T(reg) cells underwent rapid and extensive IL-2-dependent proliferation, followed by subsequent trafficking to the spleen. Thus, IL-2 is essential for T(reg) cell proliferation in neonatal LN. The number and distribution of T(reg) cells in the periphery of normal neonatal mice closely paralleled that seen for IL-2Rbeta(-/-) mice that received T(reg) cells. However, for normal neonates, blockade of IL-2 decreased T(reg) cells in both the thymus and LN. Therefore, two steps of T(reg) cell development depend upon IL-2 in neonatal mice, thymus production, and subsequent expansion in the LN.

Adoptive Transfer↗

PubMed2004

Distinct activation signals determine whether IL-21 induces B cell costimulation, growth arrest, or Bim-dependent apoptosis.

IL-21 costimulates B cell proliferation and cooperatively with IL-4 promotes T cell-dependent Ab responses. Somewhat paradoxically, IL-21 also induces apoptosis of B cells. The present study was undertaken to more precisely define the expression of the IL-21R, using a novel mAb, and the circumstances by which IL-21 promotes B cell growth vs death. The IL-21R was first detected during T and B cell development, such that this receptor is expressed by all mature lymphocytes. The IL-21R was further up-regulated after B and T activation, with the highest expression by activated B cells. Functional studies demonstrated that IL-21 substantially inhibited proliferation and induced Bim-dependent apoptosis for LPS or CpG DNA-activated B cells. In contrast, IL-21 induced both costimulation and apoptosis for anti-CD40-stimulated B cells, whereas IL-21 primarily costimulated B cells activated by anti-IgM or anti-IgM plus anti-CD40. Upon blocking apoptosis using C57BL/6 Bim-deficient or Bcl-2 transgenic B cells, IL-21 readily costimulated responses to anti-CD40 while proliferation to LPS was still inhibited. Engagement of CD40 or the BCR plus CD40 prevented the inhibitory effect by IL-21 for LPS-activated B cells. Collectively, these data indicate that there are three separable outcomes for IL-21-stimulated B cells: apoptosis, growth arrest, or costimulation. We favor a model in which IL-21 promotes B cell maturation during a productive T cell-dependent B cell response, while favoring growth arrest and apoptosis for nonspecifically or inappropriately activated B cells.

Animals↗

PubMed2004

Initial antigen encounter programs CD8+ T cells competent to develop into memory cells that are activated in an antigen-free, IL-7- and IL-15-rich environment.

Although much is known concerning the immunobiology of CD8+ T memory cells, the initial events favoring the generation of CD8+ T memory cells remain poorly defined. Using a culture system that yields memory-like CD8+ T cells, we show that 1 day after Ag encounter, Ag-activated T cells developed into memory-like T cells, but this optimally occurred 3 days after Ag encounter. Key phenotypic, functional, and molecular properties that typify central memory T cells were expressed within 48 h when the activated CD8+ T cells were cultured with IL-7 or IL-15 in the absence of Ag or following transfer into normal mice. These data support a model whereby Ag activation of naive CD8+ T cells not only programs effector cell expansion and contraction but the potential to develop into a memory cell which ensues in an Ag-free environment containing IL-7 or IL-15.

Adoptive Transfer↗

PubMed2004

Tolerance, not immunity, crucially depends on IL-2.

Interleukin-2 (IL-2) was identified based on its potent T-cell growth-factor activity and is widely considered to be a key cytokine in T-cell-dependent immune responses. However, the main non-redundant activity of this cytokine centres on the regulation of T-cell tolerance, and recent studies indicate that a failure in the production of CD4(+)CD25(+) regulatory T cells is the underlying cause of autoimmunity in the absence of IL-2. In marked contrast to the importance of IL-2 in peripheral T-cell tolerance, T-cell immunity is readily elicited to various agents in the absence of IL-2 in vivo. Here, we discuss these findings and, in particular, the action of IL-2 on regulatory T cells and effector cells, and the targeting of IL-2 and/or the IL-2 receptor in clinical settings.

Animals↗

PubMed2003

The main function of IL-2 is to promote the development of T regulatory cells.

Based primarily on vitro studies, interleukin (IL)-2 has been considered a key growth and death factor for antigen-activated T lymphocytes. IL-2 is also essential to maintain self-tolerance, as IL-2- and IL-2 receptor-deficient mice exhibit lethal autoimmunity. The intrinsic death-sensitizing activity of IL-2 was thought to be a key mediator for apoptosis of peripheral autoreactive T cells. However, recent in vivo studies strongly favor a model whereby IL-2 controls autoimmunity through the production of CD4+CD25+ T regulatory (Treg) cells. In this setting, IL-2 is essential for expansion of Treg cells within the thymus and in peripheral neonatal-immune tissue. Thus, from being considered the primary growth factor for antigen-activated T lymphocytes, these new findings redefine the pivotal role for IL-2 as the major inducer for the developmental production of suppressive Treg cells.

Animals↗

PubMed2003

Therapeutic limitations in tumor-specific CD8+ memory T cell engraftment.

BACKGROUND: Adoptive immunotherapy with cytotoxic T lymphocytes (CTL) represents an alternative approach to treating solid tumors. Ideally, this would confer long-term protection against tumor. We previously demonstrated that in vitro-generated tumor-specific CTL from the ovalbumin (OVA)-specific OT-I T cell receptor transgenic mouse persisted long after adoptive transfer as memory T cells. When recipient mice were challenged with the OVA-expressing E.G7 thymoma, tumor growth was delayed and sometimes prevented. The reasons for therapeutic failures were not clear. METHODS: OT-I CTL were adoptively transferred to C57BL/6 mice 21-28 days prior to tumor challenge. At this time, the donor cells had the phenotypical and functional characteristics of memory CD8+ T cells. Recipients which developed tumor despite adoptive immunotherapy were analyzed to evaluate the reason(s) for therapeutic failure. RESULTS: Dose-response studies demonstrated that the degree of tumor protection was directly proportional to the number of OT-I CTL adoptively transferred. At a low dose of OT-I CTL, therapeutic failure was attributed to insufficient numbers of OT-I T cells that persisted in vivo, rather than mechanisms that actively suppressed or anergized the OT-I T cells. In recipients of high numbers of OT-I CTL, the E.G7 tumor that developed was shown to be resistant to fresh OT-I CTL when examined ex vivo. Furthermore, these same tumor cells no longer secreted a detectable level of OVA. In this case, resistance to immunotherapy was secondary to selection of clones of E.G7 that expressed a lower level of tumor antigen. CONCLUSIONS: Memory engraftment with tumor-specific CTL provides long-term protection against tumor. However, there are several limitations to this immunotherapeutic strategy, especially when targeting a single antigen. This study illustrates the importance of administering large numbers of effectors to engraft sufficiently efficacious immunologic memory. It also demonstrates the importance of targeting several antigens when developing vaccine strategies for cancer.

Animals↗

PubMed2003

Long-term islet allograft survival in nonobese diabetic mice treated with tacrolimus, rapamycin, and anti-interleukin-2 antibody.

BACKGROUND: Nonobese diabetic (NOD) mice develop autoimmune diabetes with features similar to those observed in the human disease. The concurrence of allorecognition and recurrence of autoimmunity might explain why most of the treatments successful in preventing islet allograft destruction in other nonautoimmune combinations often fail in NOD recipients. To assess the value of the NOD mouse model for the evaluation of treatments relevant to clinical islet transplantation, the authors have tested the effect of a protocol closely resembling the one successfully used in the Edmonton clinical trial on the survival of islet allografts in NOD mice. METHODS: C57BL/6 islets were transplanted under the kidney capsule of spontaneously diabetic NOD mice. Treatment consisted of a combination of rapamycin, tacrolimus, and anti-interleukin (IL)-2 monoclonal antibody. Control groups received each treatment alone, a combination of two agents, or no treatment. RESULTS: Untreated animals invariably lost their graft within 13 days. Administration of rapamycin and tacrolimus significantly prolonged graft survival, with two of seven animals bearing a functional graft longer than 100 days. Addition of anti-IL-2 antibody therapy further improved graft survival, with six of eight grafts functioning longer than 100 days and two of eight grafts functioning longer than 200 days. CONCLUSIONS: In view of the limited success obtained with other treatments in this model, the dramatic prolongation of graft survival observed in the authors' study, by using a therapy that mimics one successfully used in clinical trials, seems to validate the NOD mouse as a meaningful model for the study of therapeutic interventions for the prevention of islet graft loss.

Animals↗

PubMed2003

Efficient induction of primary and secondary T cell-dependent immune responses in vivo in the absence of functional IL-2 and IL-15 receptors.

IL-2 and IL-15 are thought to be important cytokines for T cell-dependent immune responses. Mice deficient in IL-2, IL-2Ralpha, and IL-2Rbeta are each characterized by a rapid lethal autoimmune lymphoproliferative disorder that complicates their use in studies aimed at investigating the role of these cytokines and receptors for immune responses in vivo. We have previously characterized a novel transgenic (Tg) mouse on the IL-2Rbeta-/- genetic background (Tg-/- mice) that lacks autoimmune disease but still contains peripheral T cells that are nonresponsive to IL-2 and IL-15. In the present study, these mice were used to investigate the extent by which IL-2 and IL-15 are essential for T cell immunity in vivo. Tg-/- mice generated near normal primary and secondary Ab responses to OVA, readily mounted first and second set allogeneic skin graft rejection responses, and developed primary and recall CD8 T cell responses to vaccinia virus. However, Tg-/- mice generated a slightly lower level of IgG2a Abs to OVA, exhibited a somewhat delayed first set skin graft rejection response with lower allo-specific CTL, and developed a significantly lower number of IFN-gamma-producing vaccinia-specific CD8+ T cells. Thus, although T effector function is somewhat impaired, T cell immunity is largely functional in the absence of IL-2- and IL-15-induced signaling through IL-2Rbeta.

Animals↗

PubMed2002

On CD28/CD40 ligand costimulation, common gamma-chain signals, and the alloimmune response.

Activation and robust expansion of naive T cells often require T cell costimulatory signals and T cell growth factors. However, the precise growth and costimulation requirements for activation and expansion of CD4(+) and CD8(+) T cells in vivo in allograft response are still not clearly defined. In the present study, we critically examined the role of CD28/CD40 ligand (CD40L) costimulation and the common gamma-chain (gamma(c)) signals, a shared signaling component by receptors for all known T cell growth factors (i.e., IL-2, IL-4, IL-7, IL-9, IL-15, IL-21), in activation and expansion of CD4(+) and CD8(+) T cells in the allogeneic hosts. We found that CD28/CD40L costimulation and the gamma(c) signals are differentially involved in proliferation and clonal expansion of CD4(+) and CD8(+) T cells in response to alloantigen stimulation. CD8(+) T cells are highly dependent on the gamma(c) signals for survival, expansion, and functional maturation, whereas in vivo expansion of alloreactive CD4(+) T cells is largely gamma(c) independent. T cell costimulation via CD28 and CD40L, however, is necessary and sufficient for activation and expansion of CD4(+) T cells in vivo. In a skin transplant model, blocking both CD28/CD40L and the gamma(c) pathways induced prolonged skin allograft survival. Our study provides critical insights that the CD4 and CD8 compartments are most likely governed by distinct mechanisms in vivo, and targeting both costimulatory and gamma(c) signals may be highly effective in certain cytopathic conditions involving activation of both CD4(+) and CD8(+) T cells.

Animals↗

PubMed2002

Structural basis for binding multiple ligands by the common cytokine receptor gamma-chain.

The common gamma-chain (gamma(c)) that functions both in ligand binding and signal transduction is a shared subunit of the multichain receptors for interleukin (IL)-2, IL-4, IL-7, IL-9, IL-15, and IL-21. The structural basis by which the ectodomain of gamma(c) contributes to binding six distinct cytokines is only partially defined. In the present study, epitope mapping of antagonistic anti-gamma(c) monoclonal antibodies led to the identification of Asn-128 of mouse gamma(c) that represents another potential contact residue that is required for binding IL-2, IL-7, and IL-15 but not IL-4. In addition, Tyr-103, Cys-161, Cys-210, and Cys-211, previously identified to contribute to binding IL-2 and IL-7, were also found to be involved in binding IL-4 and IL-15. Collectively, these data favor a model in which gamma(c) utilizes a common mechanism for its interactions with multiple cytokines, and the binding sites are largely overlapping but not identical. Asn-128 and Tyr-103 likely act as contact residues whereas Cys-161, Cys-210, and Gly-211 may stabilize the structure of the proposed ligand-interacting surface formed by the two extracytoplasmic domains.

Amino Acid Sequence↗

PubMed2002

CD4 regulatory T cells prevent lethal autoimmunity in IL-2Rbeta-deficient mice. Implications for the nonredundant function of IL-2.

Lethal autoimmunity associated with IL-2Rbeta-deficient mice is prevented after thymic transgenic expression of wild-type IL-2Rbeta in IL-2Rbeta(-/-) mice (Tg -/- mice). Here, we show that CD4(+)CD25(+) regulatory T cells were not readily detected in IL-2Rbeta(-/-) mice, but the production of functional CD4(+)CD25(+) T cells was reconstituted in Tg -/- mice. Adoptive transfer of normal CD4(+)CD25(+) T cells into neonatal IL-2Rbeta-deficient mice prevented this lethal autoimmune syndrome. The CD4(+)CD25(+) T cells in disease-free adult IL-2Rbeta-deficient recipient mice were present at a near normal frequency, were solely donor-derived, and depended on IL-2 for expansion. These observations indicate that the essential function of the IL-2/IL-2R system primarily lies at the level of the production of CD4(+)CD25(+) regulatory T cells.

Adoptive Transfer↗

PubMed2002

T helper cells, IL-2 and the generation of cytotoxic T-cell responses.

CD8 T-cell immunity is thought to require helper activity derived from CD4 T cells. Nevertheless, under some circumstances, effective CD8-dependent T-cell responses occur in vivo without CD4 T-cell help. Several recent papers help to explain this paradox and lead to a refined view concerning the role of T helper cells and interleukin-2 receptor signaling in the production of cytotoxic T lymphocytes.

Animals↗