Ex vivo Stimulation of Lymphocytes with IL-10 Mimics Sepsis-Induced Intrinsic T-Cell Alterations

Profound T-cell alterations are observed in septic patients in association with increased risk of secondary infection and mortality. The pathophysiological mechanisms leading to such dysfunctions are not completely understood and direct and indirect mechanisms have been described.

In this study we evaluated whether ex vivo stimulation of lymphocytes with IL-10, an immunosuppressive cytokine released at the systemic level during sepsis, could mimic sepsis-induced intrinsic T-cell alterations.

We showed that recombinant human IL-10 priming of T cells altered their proliferative response to anti-CD2/CD3/CD28 antibody-coated beads and PHA stimulations, in a dose-dependent manner independently of accessory cells. This priming also significantly decreased T-cell secretion of IL-2 and IFNγ following stimulation. Furthermore, we demonstrated that IL-10 reduction of T-cell functionality was associated with increased FOXP3 expression in CD4⁺CD25⁺CD127^? regulatory T cells as observed in sepsis. Finally, we found that blocking the increased IL-10 concentration in plasma from septic shock patients increased the proliferative response of responding T cells from healthy controls.

We describe here an ex vivo model recapitulating features of sepsis-induced intrinsic T-cell alterations. This should help, in further studies, to decipher the pathophysiological mechanisms of T-cell alterations induced after septic shock.     

Interleukin 4-producing CD4 T cells arise from different precursors depending on the conditions of antigen exposure in vivo

The precursor origin of T helper (Th) cell subsets in vivo has been difficult to study and remains poorly investigated. We have previously shown that chronic administration of soluble protein antigen induces selective development of antigen-specific CD4 Th2 cells in genetically predisposed mouse strains. To analyze the origin of effector T cells in this model, we designed a competitive polymerase chain reaction-based approach to track public BV-J rearrangement expressed by CD4 T cells specific for hen egg white lysozyme (HEL) in BALB/c mice. We show that public T cell clones are predominantly associated with type 1 or 2 effector Th cells recovered after primary immunization in complete or incomplete Freund's adjuvant, respectively. Conversely, continuous administration of soluble antigen, which induces strong memory Th2 response, is associated with a dose-dependent reduction of public clone size by a mechanism resembling clonal anergy. Thus, soluble HEL-induced Th2 cells do not express the public complementarity determining region 3 motifs characteristic of immunogenic challenge in the presence of adjuvant. These results demonstrate that there are multiple pathways of induction of Th2 responses depending on the condition of antigen exposure in vivo, i.e., clonal immune deviation versus recruitment of a different pool of precursor cells.     

Induction and treatment of anergy in murine leprosy

Leprosy is a disease consisting of a spectrum of clinical, bacteriological, histopathological and immunological manifestations. Tuberculoid leprosy is frequently recognized as the benign polar form of the disease, while lepromatous leprosy is regarded as the malignant form. The different forms of leprosy depend on the genetic and immunological characteristics of the patient and on the characteristics of the leprosy bacillus. The malignant manifestations of lepromatous leprosy result from the mycobacterial‐specific anergy that develops in this form of the disease. Using murine leprosy as a model of anergy in this study, we first induced the development of anergy to Mycobacterium lepraemurium (MLM) in mice and then attempted to reverse it by the administration of dialysable leucocyte extracts (DLE) prepared from healthy (HLT), BCG‐inoculated and MLM‐inoculated mice. Mice inoculated with either MLM or BCG developed a robust cell‐mediated immune response (CMI) that was temporary in the MLM‐inoculated group and long‐lasting in the BCG‐inoculated group. DLE were prepared from the spleens of MLM‐ and BCG‐inoculated mice at the peak of CMI. Independent MLM intradermally‐inoculated groups were treated every other day with HLT‐DLE, BCG‐DLE or MLM‐DLE, and the effect was documented for 98 days. DLE administered at a dose of 1.0 U (1 × 10⁶ splenocytes) did not affect the evolution of leprosy, while DLE given at a dose of 0.1 U showed beneficial effects regardless of the DLE source. The dose but not the specificity of DLE was the determining factor for reversing anergy.     

Clinical Trials Report: Central & Peripheral Nervous Systems: Drugs in development for the treatment of multiple sclerosis: antigen-specific therapies

In recent years, several new terms and concepts have been added to the lexicon of immune regulation, including: peripheral tolerance, anergy, activation-induced apoptosis, altered peptide ligand, and bystander suppression. At the same time, great strides were made in the identification of autoantigenic peptides and T-cell receptors (TCRs) involved in the pathology of multiple sclerosis (MS). The convergence of these discoveries and limitations of current MS treatments sparked the inception of several novel therapeutic approaches that were applied to animal models and T-cell clones. This review summarises the research that has led to the development of various proposed antigen-specific therapies for MS. Included are approaches that target specific components of the TCR:peptide: MHC II triad involved in the activation of autoreactive T-cells. Also included are approaches which purport to induce specific cells to inactivate cells in their vicinity, a form of ‘specific non-specificity’ generally termed bystander suppression. The trial status of these antigen-specific therapies and results to date are summarised.     

Autoreactive T-cell responses in primary biliary cirrhosis are proinflammatory whereas those of controls are regulatory

Background:  Autoreactive T cells that proliferate in response to autoantigens are found in both autoimmune diseases and controls but have important qualitative differences in relative activation states, costimulation signal requirements and pathogenetic significance.
Methods:  To understand the differences between autoreactive T cells in PBC versus controls, we have developed autoreactive T-cell clones (TCC) from patients with PBC and healthy controls, and have used a peptide corresponding to the CD4 major autoantigen (Ag) to define the relative proliferative response. Peripheral blood mononuclear cells (PBMC) from PBC respond to the Ag in a costimulation-independent manner, but PBMC from controls respond to the Ag in a costimulation-dependent manner. Next, we established nine autoreactive TCC from patients with PBC and eight from healthy controls.
Results:  Among 17 TCC, eight were the costimulation-dependent type and nine were independent. In addition, costimulation-dependent autoreactive TCC became anergic after stimulation in the presence of APC that did not provide costimulatory signals. Finally, we observed that anergic TCC exhibit regulatory functions.
Conclusions:  In the case of regulatory dendritic cells, we could not induce TCC anergy. On the other hand, when using peptide analog in a costimulation-deficient manner, we could induce TCC anergy, even though these TCC were costimulation independent.     

MHC Restricted Recognition by Cloned T Cells

The critical role of major histocompatibility complex (MHC) gene products in T cell activation was appreciated and extensively studied well before the availability of monoclonal T cell populations. However, the availability of cloned T cells has dramatically enhanced the ability to characterize the nature of MHC-restricted recognition by T cells. In certain areas, the use of monoclonal T cells has allowed substantial extension of principles already established through the use of heterogeneous T cells. In other cases, studies employing T cell clones have led to findings which were inaccessible to approaches using only heterogeneous T cell populations. On balance, it should also be pointed out that a significant number of critical questions concerning the MHC-restricted T cell repertoire remain best approached by studying the development of interactions of functionally heterogeneous T cell populations. The present review will focus upon recent progress in several areas in which the use of cloned T cells has been particularly important in characterizing the nature of MHC-restricted recognition. In particular, the restriction of T cell recognition by class II MHC products (Ia) is examined in detail.     

Multiple paths to loss of anergy and gain of autoimmunity

B cells and autoimmunity: cells of the immune system have the capacity to recognize/neutralize a myriad array of disease-causing pathogens, while simultaneously minimizing damage to self tissue. Obvious breakdowns in this ability to distinguish between self and non-self are evident in multiple forms of autoimmune disease, where B and T cells mount damaging attacks on cells and organs. B cells may directly damage tissue by producing pathogenic antibodies that bind self antigen, fix complement or form immune complexes. Recent evidence also suggests B cells indirectly induce autoimmunity by concentrating low avidity self antigen through the B cell receptor and presenting self-peptides to autoreactive T cells. B cells may also initiate autoimmunity when provided sufficient help from autoreactive T cells that have escaped deletion in the thymus. Here, we will review the role of anergy in maintenance of tolerance and how alterations in the normal balance of positive and negative signals may contribute to the development of autoimmune disease in mouse models and humans.     

Recurrent superantigen exposure in vivo leads to highly suppressive CD4+CD25+ and CD4+CD25‐ T cells with anergic and suppressive genetic signatures

Staphylococcal enterotoxin B (SEB) activates T cells via non‐canonical signalling through the T cell receptor and is an established model for T cell unresponsiveness in vivo. In this study, we sought to characterize the suppressive qualities of SEB‐exposed CD4⁺ T cells and correlate this with genetic signatures of anergy and suppression. SEB‐exposed CD25⁺ and CD25^‐Vβ8⁺CD4⁺ T cells expressed forkhead box P3 (FoxP3) at levels comparable to naive CD25⁺ T regulatory cells and were enriched after exposure in vivo. Gene related to anergy in lymphocytes (GRAIL), an anergy‐related E3 ubiquitin ligase, was up‐regulated in the SEB‐exposed CD25⁺ and CD25^‐FoxP3⁺Vβ8⁺CD4⁺ T cells and FoxP3^‐CD25^‐Vβ8⁺CD4⁺ T cells, suggesting that GRAIL may be important for dominant and recessive tolerance. The SEB‐exposed FoxP3⁺GRAIL⁺ T cells were highly suppressive and non‐proliferative independent of CD25 expression level and via a glucocorticoid‐induced tumour necrosis factor R‐related protein‐independent mechanism, whereas naive T regulatory cells were non‐suppressive and partially proliferative with SEB activation in vitro. Lastly, adoptive transfer of conventional T cells revealed that induction of FoxP3⁺ regulatory cells is not operational in this model system. These data provide a novel paradigm for chronic non‐canonical T cell receptor engagement leading to highly suppressive FoxP3⁺GRAIL⁺CD4⁺ T cells.     

Effect of factor VIII concentrate on antigen-presenting cell (APC)/T-cell interactions in vitro: relevance to inhibitor formation and tolerance induction

Inhibitor formation in patients with haemophilia receiving factor VIII (FVIII) concentrate is a common problem requiring tolerance induction therapy. Immune tolerance is dependent on defective T cell/antigen-presenting cell (APC) interactions and inhibitor antibody formation is associated with effective T-cell/B-cell interaction. We studied the expression of the cell-surface molecules involved with these interactions using multiparameter flow cytometry and a whole blood stimulation assay-phytohaemaglutinin (PHA) to activate T cells and Escherichia coli lipopolysaccharide (LPS) to activate monocytes and B cells. Up-regulation of T-cell co-stimulatory receptors CD11a, CD40 ligand (CD40L) and CTLA4 were inhibited in a dose-dependent manner by plasma-derived (pd)FVIII, but CD28 was unchanged. Up-regulation of monocyte and B-cell co-stimulatory ligands CD4O, B7-1 (CD80) and B7-2 (CD86) were also inhibited in a dose-dependent manner by pdFVIII, but LFA-3 (CD58) was unchanged. The combined inhibitory effect of prednisolone, an immunosuppressive agent used in several tolerance induction protocols, with pdFVIII on co-stimulatory molecules, was additive. There was no significant alteration in T-cell/APC adhesion or co-stimulatory molecules noted in the presence of recombinant (rh)FVIII concentrate. The inhibitory effect of pdFVIII on molecules involved in interaction between T cells and APCs may result in immune tolerance in recipients of pdFVIII concentrate. The inhibitory effect of pdFVIII on CD40/CD40L up-regulation may result in defective antibody formation. We now provide evidence that the use of pdFVIII, through interfering with APC/T-cell interactions, may be more appropriate than rhFVIII for tolerance induction.