[Show abstract][Hide abstract] ABSTRACT: Activation of innate immunity through Toll-like receptors (TLR) can abrogate transplantation tolerance by revealing hidden T cell alloreactivity. Separately, the cholinergic anti-inflammatory pathway has the capacity to dampen macrophage activation and cytokine release during endotoxemia and ischemia reperfusion injury. However, the relevance of the α7 nicotinic acetylcholine receptor (α7nAChR)-dependent anti-inflammatory pathway in the process of allograft rejection or maintenance of tolerance remains unknown. The aim of our study is to investigate whether the cholinergic pathway could impact T cell alloreactivity and transplant outcome in mice. For this purpose, we performed minor-mismatched skin allografts using donor/recipient combinations genetically deficient for the α7nAChR. Minor-mismatched skin grafts were not rejected unless the mice were housed in an environment with endogenous pathogen exposure or the graft was treated with direct application of imiquimod (a TLR7 ligand). The α7nAChR-deficient recipient mice showed accelerated rejection compared to wild type recipient mice under these conditions of TLR activation. The accelerated rejection was associated with enhanced IL-17 and IFN-γ production by alloreactive T cells. An α7nAChR-deficiency in the donor tissue facilitated allograft rejection but not in recipient mice. In addition, adoptive T cell transfer experiments in skin-grafted lymphopenic animals revealed a direct regulatory role for the α7nAChR on T cells. Taken together, our data demonstrate that the cholinergic pathway regulates alloreactivity and transplantation tolerance at multiple levels. One implication suggested by our work is that, in an organ transplant setting, deliberate α7nAChR stimulation of brain dead donors might be a valuable approach for preventing donor tissue inflammation prior to transplant.
PLoS ONE 11/2013; 8(11):e79984. DOI:10.1371/journal.pone.0079984 · 3.23 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The goal of this health-psychology study was to investigate the range of motives that make someone consent or refuse participation in a clinical transplant trial.
The study involved (i) preparatory interviews with five transplant patients who took part in clinical trials and five persons from the general public; (ii) we created a questionnaire with scaled responses; (iii) we selected 468 patients, divided into two groups: patients waiting for a transplant and patients who already had a transplant; (iv) we obtained patient consents, sent out the questionnaire, and recorded responses; (v) data were analysed using descriptive statistics, exploratory factorial analyses, correlations and regressions.
Two hundred and ten patients answered the questionnaire. Motives were classified into the following: (i) Motives to consent participating in a clinical trial (pride in participating, hope for better quality of life, sufficient and clear information, discussion with others participating in a trial, altruism); or (ii) Motives to refuse participating (no information on medical teams, lack of explanation, fear of additional expenses, being considered a "guinea pig").
This study contributes to our understanding of the motivations of patients who accepted or refused participation in a clinical transplantation trial.
[Show abstract][Hide abstract] ABSTRACT: Primary graft dysfunction (PGD) and bronchiolitis obliterans (BO) are the leading causes of morbidity and mortality after lung transplantation. Reports from clinical and rodent models suggest the implication of IL-17A in either PGD or BO. We took advantage of the heterotopic trachea transplantation model in mice to study the direct role of IL-17A in post-transplant airway lesions. Across full MHC barrier, early lesions were controlled in IL-17A(-/-) or anti-IL17 treated recipients. In contrast, IL-17A deficiency did not prevent subsequent obliterative airway disease (OAD). Interestingly, this early protection occurred also in syngeneic grafts and was accompanied by a decrease in cellular stress, as attested by lower HSP70 mRNA levels, suggesting the involvement of IL-17A in ischemia-reperfusion injury (IRI). Furthermore, persistence of multipotent CK14(+) epithelial stem cells underlined allograft protection afforded by IL-17A deficiency or neutralisation. Recipient-derived γδ(+) and CD4(+) T cells were the major source of IL-17A. However, lesions still occurred in the absence of each subset, suggesting a high redundancy between the innate and adaptive IL-17A producing cells. Notably, a double depletion significantly diminished lesions. In conclusion, this work implicated IL-17A as mediator of early post-transplant airway lesions and could be considered as a potential therapeutic target in clinical transplantation.
PLoS ONE 07/2013; 8(7):e70236. DOI:10.1371/journal.pone.0070236 · 3.23 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: We report on a pilot study investigating the feasibility of early immunosuppression withdrawal after liver transplantation (LT) using antithymocyte globulin (ATG) induction and rapamycin.
LT recipients received 3.75 mg/kg per day ATG from days 0 to 5 followed by rapamycin-based immunosuppression. In the absence of acute rejection (AR), rapamycin was withdrawn after month 4. Immunomonitoring included analysis of peripheral T-cell phenotypes and clonality, cytokine production in mixed lymphocyte reaction, and characterization of intragraft infiltrating cells.
Ten patients were enrolled between October 2009 and July 2010. In the first three patients, complete withdrawal of immunosuppression after month 4 led to AR. No further withdrawals of immunosuppressive were attempted. Two AR occurred in the remaining seven patients. ATG induced profound T-cell depletion followed by CD8 T-cell reexpansion exhibiting memory/effector-like phenotype associated with progressive oligoclonal T-cell expansion (Vβ/HPRT ratio) and gradually enhanced anti-cytomegalovirus and anti-Epstein-Barr virus T-cell frequencies. Patients developing AR were characterized by decreased TCAIM expression. AR were associated with increased donor-specific production of interferon (IFN)-γ and interleukin (IL)-17, increased intragraft expression of IFN-γ mRNA, and significant CD8 T-cell infiltrates colocalizing with IL-17 cells.
High-dose ATG followed by short-term rapamycin treatment failed to promote early operational tolerance to LT. AR correlates with expansion of memory-type CD8 T cells and increased levels of IFN-γ and IL-17 in mixed lymphocyte reaction and in the graft. This suggests that resistance and preferential expansion of effector memory T-cell in lymphopenic environment could represent the major barrier for establishment of tolerance to LT in approaches using T-cell-depleting induction.
[Show abstract][Hide abstract] ABSTRACT: Calcineurin-inhibitor refractory bronchiolitis obliterans (BO) represents the leading cause of late graft failure after lung transplantation. T helper (Th)2 and Th17 lymphocytes have been associated with BO development. Taking advantage of a fully allogeneic trachea transplantation model in mice, we addressed the pathogenicity of Th cells in obliterative airway disease (OAD) occurring in cyclosporine A (CsA)-treated recipients. We found that CsA prevented CD8(+) T cell infiltration into the graft and downregulated the Th1 response but affected neither Th2 nor Th17 responses in vivo. In secondary mixed lymphocyte cultures, CsA dramatically decreased donor-specific IFN-γ production, enhanced IL-17 production and did not affect IL-13. As CD4(+) depletion efficiently prevented OAD in CsA-treated recipients, we further explored the role of Th2 and Th17 immunity in vivo. Although IL-4 and IL-17 deficient untreated mice developed an OAD comparable to wild-type recipients, a single cytokine deficiency afforded significant protection in CsA-treated recipients. In conclusion, CsA treatment unbalances T helper alloreactivity and favors Th2 and Th17 as coexisting pathways mediating chronic rejection of heterotopic tracheal allografts.
American Journal of Transplantation 01/2013; 13(3). DOI:10.1111/ajt.12067 · 6.19 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Interferon regulatory factor 3 (IRF3) is an important transcription factor in Toll‐like receptor 4 (TLR4) signaling, a pathway that is known to play a critical role in liver ischemia‐reperfusion injury. In order to decipher the involvement of IRF3 in this setting, we first compared the intensity of hepatic lesions in IRF3‐deficient versus wildtype mice. We found increased levels of blood transaminases, enhanced liver necrosis, and more pronounced neutrophil infiltrates in IRF3‐deficient mice. Neutrophil depletion by administration of anti‐Ly6G monoclonal antibody indicated that neutrophils play a dominant role in the development of severe liver necrosis in IRF3‐deficient mice. Quantification of cytokine genes expression revealed increased liver expression of interleukin (IL)‐12/IL‐23p40, IL‐23p19 messenger RNA (mRNA), and IL‐17A mRNA in IRF3‐deficient versus wildtype (WT) mice, whereas IL‐27p28 mRNA expression was diminished in the absence of IRF3. The increased IL‐17 production in IRF3‐deficient mice was functionally relevant, as IL‐17 neutralization prevented the enhanced hepatocellular damages and liver inflammation in these animals. Evidence for enhanced production of IL‐23 and decreased accumulation of IL‐27 cytokine in M1 type macrophage from IRF3‐deficient mice was also observed after treatment with lipopolysaccharide, a setting in which liver gamma‐delta T cells and invariant natural killer T cells were found to be involved in IL‐17A hyperproduction. Conclusion: IRF3‐dependent events downstream of TLR4 control the IL‐23/IL‐17 axis in the liver and this regulatory role of IRF3 is relevant to liver ischemia‐reperfusion injury. (HEPATOLOGY 2013)
[Show abstract][Hide abstract] ABSTRACT: Immune responses in newborn mice are known to be biased toward the helper type 2 phenotype. This may account for their propensity to develop tolerance. Herein, we evaluated the effects of IL-4 deprivation on CD4(+) T-cell activities elicited by neonatal exposure to allogeneic spleen cells. We showed that chimerism, Th2-type polarization and pathology, as well as skin allograft acceptance were inhibited in BALB/c mice immunized at birth with (A/J x BALB/c) F(1) spleen cells upon in vivo IL-4 neutralization. While IL-4 neutralization inhibited the development of Th2 cells in this model, it led to the accumulation of IL-17A, IL-17F, IL-22, IL-6 and RORγt mRNA in the spleen or graft tissues. Moreover, IL-4 deprivation led to the differentiation of donor-specific Th17 cells with a concomitant Th1 response characterized by IFN-γ production. The Th17-type response emerging in IL-4-deprived mice was found to mediate both intragraft neutrophil infiltration and the abrogation of B-cell chimerism. Neutralization of this Th17 response failed however to restore functional skin graft acceptance. Collectively, our observations indicate that the neonatal Th2 response opposes the development of Th17 cells, and that Th17 cells are responsible for controlling lymphoid chimerism in mice neonatally injected with semiallogeneic cells.
American Journal of Transplantation 01/2012; 12(1):81-9. DOI:10.1111/j.1600-6143.2011.03778.x · 6.19 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Protein kinase C (PKC) isoforms play a critical role in the regulation of innate immune responses. We have previously demonstrated that conventional PKC (cPKC) α is involved in interferon regulatory factor 3 (IRF-3) activation and IFN-β synthesis. Herein, we investigated the role of cPKCs in the regulation of IL-12 family members expression mediated by the Toll-like receptor 3 (TLR3) and TLR4. First, inhibition of cPKCs activity in human DCs by a cPKC-specific inhibitor, Gö6976 downregulated the expression of IL-12p70 and IL-27p28 but not IL-12/IL-23p40, IL-23, IL-27EBI3 induced by LPS or poly(I:C). Furthermore, reporter gene assays in RAW 264.7 macrophages showed that cPKCs regulate IL-12p35 and IL-27p28 promoter activities since Gö6976 repressed LPS and poly(I:C)-mediated transcriptional activities of IL-12p35 and IL-27p28. In contrast, no effect was observed with IL-12/IL-23p40 and IL-23p19 reporter constructs. These results prompted us to study the role of IRF-3 on IL-23 expression. Bone marrow-derived DC (BMDCs) from IRF-3(-/-) mice produced comparable levels of IL-23 induced by both LPS and poly(I:C) as compared to wild type BMDCs, indicating that IRF-3 is not involved in IL-23 production. Finally, BMDCs from PKCα(-/-) mice displayed a reduced synthesis of IL-27 induced by poly(I:C). Collectively, these data identify cPKCs as critical components that control IRF-3-dependent IL-12p35 and IL-27p28 gene expression downstream of TLR3 and TLR4.
[Show abstract][Hide abstract] ABSTRACT: Transplantation tolerance induced by neonatal injection of semi-allogeneic spleen cells is associated with a pathological syndrome caused by T helper type 2 (Th2) differentiation of donor-specific CD4(+) T lymphocytes. We have shown previously that this Th2-biased response is inhibited by host CD8(+) T cells. Herein, we demonstrate that upon neonatal immunization with (A/J × BALB/c)F(1) spleen cells, BALB/c mice expand a population of CD8(+) T cells expressing both CD25 and forkhead box P3 (FoxP3) markers. In this setting, CD8(+) CD25(+) T cells predominantly produce interferon (IFN)-γ and interleukin (IL)-10 and are efficient in controlling IL-4, IL-5 and IL-13 production by donor-specific CD4(+) T cells in vitro. CD8(+) FoxP3(-) T cells are single producers of IFN-γ or IL-10, whereas CD8(+) FoxP3(+) T cells are double producers of IFN-γ and IL-10. We further demonstrate that IFN-γ and IL-10 are two major cytokines produced by CD8(+) T cells involved in the in vivo regulation of Th2-type pathology. In this setting, we conclude that neonatal alloimmunization induces the expansion of several regulatory CD8(+) T cells which may control Th2 activities via IFN-γ and IL-10.
[Show abstract][Hide abstract] ABSTRACT: The immune system in early life is regarded as immature. However, the IL-12 family member IL-23 is highly produced upon TLR stimulation by neonatal DCs. Human adult Vγ9Vδ2 T cells can be stimulated specifically via their TCR by phosphoantigens (as the pathogen-derived HMB-PP) or agents and infections that lead to their endogenous accumulation (as the aminobisphosphonate zoledronate). As increasing evidence indicates that γδ T cells are especially important in early life, we investigated the effect of IL-23 on neonatal Vγ9Vδ2 T cells stimulated via their TCR. Zoledronate induced clear proliferation and IFN-γ production in neonatal Vγ9Vδ2 T cells. In contrast, HMB-PP did not elicit a distinct response unless at high concentrations. Addition of IL-23 to zoledronate enhanced the expression of IFN-γ and generated a distinct, IFN-γ-negative, neonatal Vγ9Vδ2 T cell population producing IL-17. Furthermore, IL-23 significantly enhanced the expression of a range of cytotoxic mediators (perforin, granzymes, granulysin). Although the costimulatory effect of IL-23 on IFN-γ and cytotoxic mediators was also observed within adult Vγ9Vδ2 T cells, the induction of an IL-17+IFN-γ- subset was unique to neonatal Vγ9Vδ2 T cells. In conclusion, neonatal DC-derived IL-23 combined with specific TCR signaling drives the generation of neonatal Vγ9Vδ2 T cells equipped with a range of cytotoxic mediators and distinct subpopulations producing IFN-γ and IL-17.
[Show abstract][Hide abstract] ABSTRACT: The long-term stability of renal grafts depends on the absence of chronic rejection. As T cells play a key role in rejection processes, analyzing the T-cell repertoire may be useful for understanding graft function outcomes. We have therefore investigated the power of a new statistical tool, used to analyze the peripheral blood TCR repertoire, for determining immunological differences in a group of 229 stable renal transplant patients undergoing immunosuppression. Despite selecting the patients according to stringent criteria, the patients displayed heterogeneous T-cell repertoire usage, ranging from unbiased to highly selected TCR repertoires; a skewed TCR repertoire correlating with an increase in the CD8(+) /CD4(+) T-cell ratio. T-cell repertoire patterns were compared in patients with clinically opposing outcomes i.e. stable drug-free operationally tolerant recipients and patients with the "suspicious" form of humoral chronic rejection and were found significantly different, from polyclonal to highly selected TCR repertoires, respectively. Moreover, a selected TCR repertoire was found to positively correlate with the Banff score grade. Collectively, these data suggest that TCR repertoire categorization might be included in the calculation of a composite score for the follow-up of patients after kidney transplantation.
European Journal of Immunology 11/2010; 40(11):3280-90. DOI:10.1002/eji.201040301 · 4.52 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Mepolizumab, a monoclonal anti-IL-5 antibody, is an effective corticosteroid-sparing agent for patients with Fip1-like 1/platelet-derived growth factor receptor α fusion (F/P)-negative hypereosinophilic syndrome (HES). Lymphocytic variant hypereosinophilic syndrome (L-HES) is characterized by marked overproduction of IL-5 by dysregulated T cells.
To determine whether patients with L-HES respond to mepolizumab in terms of corticosteroid tapering and eosinophil depletion to the same extent as corticosteroid-responsive F/P-negative patients with HES and a normal T-cell profile.
Patients enrolled in the mepolizumab trial were evaluated for L-HES on the basis of T-cell phenotyping and T-cell receptor gene rearrangement patterns, and their serum thymus-and-activation-regulated chemokine (TARC) levels were measured. Response to treatment was compared in patient subgroups based on results of these analyses.
Lymphocytic variant HES was diagnosed in 13 of 63 patients with HES with complete T-cell assessments. The ability to taper corticosteroids on mepolizumab was similar in patients with L-HES and those with a normal T-cell profile, although a lower proportion of patients with L-HES maintained eosinophil levels below 600/μL. Increased serum TARC levels (>1000 pg/mL) had no significant impact on the ability to reduce corticosteroid doses, but a lower proportion of patients with elevated TARC achieved eosinophil control on mepolizumab.
Mepolizumab is an effective corticosteroid-sparing agent for patients with L-HES. In some cases however, eosinophil levels remain above 600/μL, suggesting incomplete neutralization of overproduced IL-5 or involvement of other eosinophilopoietic factors.
The Journal of allergy and clinical immunology 10/2010; 126(4):828-835.e3. DOI:10.1016/j.jaci.2010.06.049 · 11.25 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Pattern-recognition receptors (PRRs) are critically involved in the pathophysiology of airway allergy, yet most of the signaling pathways downstream of PRRs implicated in allergic airway sensitization remain unknown.
We sought to study the effects of genetic depletion of interferon response factor (IRF) 3 and IRF7, important transcription factors downstream of various PRRs, in a murine model of house dust mite (HDM)-induced allergic asthma.
We compared HDM-induced allergic immune responses in IRF3-deficient (IRF3(-/-)), IRF7(-/-), and wild-type mice.
Parameters of airway allergy caused by HDM exposure were strongly attenuated in IRF3(-/-), but not IRF7(-/-), mice compared with those in wild-type mice. Indeed, in HDM-exposed IRF3(-/-) mice HDM-specific T(H)2 cell responses did not develop. This correlated with impaired maturation and migration of IRF3(-/-) lung dendritic cells (DCs) on HDM treatment. Furthermore, adoptive transfer of HDM-loaded DCs indicated that IRF3(-/-) DCs had an intrinsic defect rendering them unable to migrate and to prime HDM-specific T(H)2 responses. Intriguingly, we also show that DC function and allergic airway sensitization in response to HDM were independent of signaling by type I interferons, the main target genes of IRF3.
Through its role in DC function, IRF3, mainly known as a central activator of antiviral immunity, is essential for the development of T(H)2-type responses to airway allergens.
The Journal of allergy and clinical immunology 10/2010; 126(4):836-844.e13. DOI:10.1016/j.jaci.2010.06.009 · 11.25 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Th17-mediated immune responses have been recently identified as novel pathogenic mechanisms in a variety of conditions; however, their importance in allograft rejection processes is still debated. In this paper, we searched for MHC or minor Ag disparate models of skin graft rejection in which Th17 immune responses might be involved. We found that T cell-derived IL-17 is critical for spontaneous rejection of minor but not major Ag-mismatched skin grafts. IL-17 neutralization was associated with a lack of neutrophil infiltration and neutrophil depletion delayed rejection, suggesting neutrophils as an effector mechanism downstream of Th17 cells. Regulatory T cells (Tregs) appeared to be involved in Th17 reactivity. We found that in vivo Treg depletion prevented IL-17 production by recipient T cells. An adoptive cotransfer of Tregs with naive monospecific antidonor T cells in lymphopenic hosts biased the immune response toward Th17. Finally, we observed that IL-6 was central for balancing Tregs and Th17 cells as demonstrated by the prevention of Th17 differentiation, the enhanced Treg/Th17 ratio, and a net impact of rejection blockade in the absence of IL-6. In conclusion, the ability of Tregs to promote the Th17/neutrophil-mediated pathway of rejection that we have described should be considered as a potential drawback of Treg-based cell therapy.
The Journal of Immunology 09/2010; 185(6):3417-25. DOI:10.4049/jimmunol.0903961 · 5.36 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Th2-biased immune responses characterizing neonates may influence the later onset of allergic disease. The contribution of regulatory T cell populations in the prevention of Th2-driven pathologies in early life is poorly documented. We investigated the potential of CD8(+) T cells stimulated at birth with alloantigens to modulate the development of allergic airway inflammation. Newborn mice were immunized with semiallogeneic splenocytes or dendritic cells (DCs) and exposed at the adult stage to OVA aeroallergens. DC-immunized animals displayed a strong Th1 and Tc1/Tc2 alloantigen-specific response and were protected against the development of the allergic reaction with reduced airway hyperresponsiveness, mucus production, eosinophilia, allergen-specific IgE and IgG(1), and reduction of lung IL-4, IL-5, IL-10, and IL-13 mRNA levels. By contrast, splenocyte-immunized mice displayed a Th2 and a weak Tc2 alloantigen-specific response and were more sensitive to the development of the allergen-specific inflammation compared with mice unexposed at birth to alloantigens. DC-immunized animals displayed an important increase in the percentage of IFN-gamma-producing CD8(+)CD44(high), CD8(+)CD62L(high), and CD8(+)CD25(+) subsets. Adoptive transfers of CD8(+) T cells from semiallogeneic DC-immunized animals to adult beta(2)m-deficient animals prevented the development of allergic response, in particular IgE, IL-4, and IL-13 mRNA production in an IFN-gamma-dependent manner, whereas transfers of CD8(+) T cells from semiallogeneic splenocyte-immunized mice intensified the lung IL-4 and IL-10 mRNA level and the allergen-specific IgE. These findings demonstrated that neonatal induction of regulatory CD8(+) T cells was able to modulate key parameters of later allergic sensitization in a bystander manner, without recognition of MHC class I molecules.
The Journal of Immunology 07/2010; 185(2):884-91. DOI:10.4049/jimmunol.0903287 · 5.36 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Identifying transplant recipients in whom immunological tolerance is established or is developing would allow an individually tailored approach to their posttransplantation management. In this study, we aimed to develop reliable and reproducible in vitro assays capable of detecting tolerance in renal transplant recipients. Several biomarkers and bioassays were screened on a training set that included 11 operationally tolerant renal transplant recipients, recipient groups following different immunosuppressive regimes, recipients undergoing chronic rejection, and healthy controls. Highly predictive assays were repeated on an independent test set that included 24 tolerant renal transplant recipients. Tolerant patients displayed an expansion of peripheral blood B and NK lymphocytes, fewer activated CD4+ T cells, a lack of donor-specific antibodies, donor-specific hyporesponsiveness of CD4+ T cells, and a high ratio of forkhead box P3 to alpha-1,2-mannosidase gene expression. Microarray analysis further revealed in tolerant recipients a bias toward differential expression of B cell-related genes and their associated molecular pathways. By combining these indices of tolerance as a cross-platform biomarker signature, we were able to identify tolerant recipients in both the training set and the test set. This study provides an immunological profile of the tolerant state that, with further validation, should inform and shape drug-weaning protocols in renal transplant recipients.
The Journal of clinical investigation 06/2010; 120(6):1848-61. DOI:10.1172/JCI39922 · 13.77 Impact Factor