Inhibiting CXCR3-dependent CD8+ T cell trafficking enhances tolerance induction in a mouse model of lung rejection

Lung transplantation remains the only effective therapy for patients with end-stage pulmonary diseases. Unfortunately, acute rejection of the lung remains a frequent complication and is an important cause of morbidity and mortality. The induction of transplant tolerance is thought to be dependent, in part, on the balance between allograft effector mechanisms mediated by effector T lymphocytes (Teff), and regulatory mechanisms mediated by FOXP3(+) regulatory T cells (Treg). In this study, we explored an approach to tip the balance in favor of regulatory mechanisms by modulating chemokine activity. We demonstrate in an adoptive transfer model of lung rejection that CXCR3-deficient CD8(+) Teff have impaired migration into the lungs compared with wild-type Teff, which results in a dramatic reduction in fatal pulmonary inflammation. The lungs of surviving mice contained tolerized CXCR3-deficient Teff, as well as a large increase in Treg. We confirmed that Treg were needed for tolerance and that their ability to induce tolerance was dependent on their numbers in the lung relative to the numbers of Teff. These data suggest that transplantation tolerance can be achieved by reducing the recruitment of some, but not necessarily all, CD8(+) Teff into the target organ and suggest a novel approach to achieve transplant tolerance.     

A randomised, blinded, placebo-controlled trial in dementia patients continuing or stopping neuroleptics (the DART-AD trial)

Background

There have been increasing concerns regarding the safety and efficacy of neuroleptics in people with dementia, but there are very few long-term trials to inform clinical practice. The aim of this study was to determine the impact of long-term treatment with neuroleptic agents upon global cognitive decline and neuropsychiatric symptoms in patients with Alzheimer disease.

Methods and Findings

Design: Randomised, blinded, placebo-controlled parallel two-group treatment discontinuation trial.Setting: Oxfordshire, Newcastle and Gateshead, London and Edinburgh, United Kingdom.Participants: Patients currently prescribed the neuroleptics thioridazine, chlorpromazine, haloperidol trifluoperazine or risperidone for behavioural or psychiatric disturbance in dementia for at least 3 mo.Interventions: Continue neuroleptic treatment for 12 mo or switch to an identical placebo.Outcome measures: Primary outcome was total Severe Impairment Battery (SIB) score. Neuropsychiatric symptoms were evaluated with the Neuropsychiatric Inventory (NPI).Results: 165 patients were randomised (83 to continue treatment and 82 to placebo, i.e., discontinue treatment), of whom 128 (78%) commenced treatment (64 continue/64 placebo). Of those, 26 were lost to follow-up (13 per arm), resulting in 51 patients per arm analysed for the primary outcome. There was no significant difference between the continue treatment and placebo groups in the estimated mean change in SIB scores between baseline and 6 mo; estimated mean difference in deterioration (favouring placebo) −0.4 (95% confidence interval [CI] −6.4 to 5.5), adjusted for baseline value (p = 0.9). For neuropsychiatric symptoms, there was no significant difference between the continue treatment and placebo groups (n = 56 and 53, respectively) in the estimated mean change in NPI scores between baseline and 6 mo; estimated mean difference in deterioration (favouring continue treatment) −2.4 (95% CI −8.2 to 3.5), adjusted for baseline value (p = 0.4). Both results became more pronounced at 12 mo. There was some evidence to suggest that those patients with initial NPI ≥ 15 benefited on neuropsychiatric symptoms from continuing treatment.

Conclusions

For most patients with AD, withdrawal of neuroleptics had no overall detrimental effect on functional and cognitive status. Neuroleptics may have some value in the maintenance treatment of more severe neuropsychiatric symptoms, but this benefit must be weighed against the side effects of therapy.Trial registration: Cochrane Central Registry of Controlled Trials/National Research Register (#ISRCTN33368770).     

The lysophosphatidic acid receptor LPA1 links pulmonary fibrosis to lung injury by mediating fibroblast recruitment and vascular leak

Aberrant wound-healing responses to injury have been implicated in the development of pulmonary fibrosis, but the mediators directing these pathologic responses have yet to be fully identified. We show that lysophosphatidic acid levels increase in bronchoalveolar lavage fluid following lung injury in the bleomycin model of pulmonary fibrosis, and that mice lacking one of its receptors, LPA1, are markedly protected from fibrosis and mortality in this model. The absence of LPA1 led to reduced fibroblast recruitment and vascular leak, two responses that may be excessive when injury leads to fibrosis rather than to repair, whereas leukocyte recruitment was preserved during the first week after injury. In persons with idiopathic pulmonary fibrosis, lysophosphatidic acid levels in bronchoalveolar lavage fluid were also increased, and inhibition of LPA1 markedly reduced fibroblast responses to the chemotactic activity of this fluid. LPA1 therefore represents a new therapeutic target for diseases in which aberrant responses to injury contribute to fibrosis, such as idiopathic pulmonary fibrosis.     

Cytokine polymorphisms in silicosis and other pneumoconioses

Silicosis and coal workers' pneumoconiosis are complex multifactorial lung diseases whose etiopathogenesis are not well defined. It is generally accepted that fibrotic lung disorders are mediated by macrophage-derived cytokines and growth factors. There is evidence showing a crucial role for tumor necrosis factor- (TNF-) and interleukin-1 (IL-1) in inflammation caused by silica dust and in the transition from simple to progressive massive fibrosis. In this review we discuss genetic polymorphisms responsible for regulating the production of these proinflammatory cytokines and their role in modifying silicosis severity.     

Donor T cell activation initiates small bowel allograft rejection through an IFN-gamma-inducible protein-10-dependent mechanism

The poor success in controlling small bowel (SB) allograft rejection is partially attributed to the unique immune environment in the donor intestine. We hypothesized that Ag-induced activation of donor-derived T cells contributes to the initiation of SB allograft rejection. To address the role of donor T cell activation in SB transplantation, SB grafts from DO11.10 TCR transgenic mice (BALB/c, H-2L(d+)) were transplanted into BALB/c (isografts), or single class I MHC-mismatched (L(d)-deficient) BALB/c H-2(dm2) (dm2, H-2L(d-)) mutant mice (allografts). Graft survival was followed after injection of control or antigenic OVA(323-339) peptide. Eighty percent of SB allografts developed severe rejection in mice treated with antigenic peptide, whereas <20% of allografts were rejected in mice treated with control peptide (p < 0.05). Isografts survived >30 days regardless of OVA(323-339) administration. Activation of donor T cells increased intragraft expression of proinflammatory cytokine (IFN-gamma) and CXC chemokine IFN-gamma-inducible protein-10 mRNA and enhanced activation and accumulation of host NK and T cells in SB allografts. Treatment of mice with neutralizing anti-IFN-gamma-inducible protein-10 mAb increased SB allograft survival in Ag-treated mice (67%; p < 0.05) and reduced accumulation of host T cells and NK cells in the lamina propria but not mesenteric lymph nodes. These results suggest that activation of donor T cells after SB allotransplantation induces production of a Th1-like profile of cytokines and CXC chemokines that enhance infiltration of host T cells and NK cells in SB allografts. Blocking this pathway may be of therapeutic value in controlling SB allograft rejection.     

IFN-gamma-inducible protein 10 (CXCL10) contributes to airway hyperreactivity and airway inflammation in a mouse model of asthma

Allergic asthma is an inflammatory disease of the airways characterized by eosinophilic inflammation and airway hyper-reactivity. Cytokines and chemokines specific for Th2-type inflammation predominate in asthma and in animal models of this disease. The role of Th1-type inflammatory mediators in asthma remains controversial. IFN-gamma-inducible protein 10 (IP-10; CXCL10) is an IFN-gamma-inducible chemokine that preferentially attracts activated Th1 lymphocytes. IP-10 is up-regulated in the airways of asthmatics, but its function in asthma is unclear. To investigate the role of IP-10 in allergic airway disease, we examined the expression of IP-10 in a murine model of asthma and the effects of overexpression and deletion of IP-10 in this model using IP-10-transgenic and IP-10-deficient mice. Our experiments demonstrate that IP-10 is up-regulated in the lung after allergen challenge. Mice that overexpress IP-10 in the lung exhibited significantly increased airway hyperreactivity, eosinophilia, IL-4 levels, and CD8(+) lymphocyte recruitment compared with wild-type controls. In addition, there was an increase in the percentage of IL-4-secreting T lymphocytes in the lungs of IP-10-transgenic mice. In contrast, mice deficient in IP-10 demonstrated the opposite results compared with wild-type controls, with a significant reduction in these measures of Th2-type allergic airway inflammation. Our results demonstrate that IP-10, a Th1-type chemokine, is up-regulated in allergic pulmonary inflammation and that this contributes to the airway hyperreactivity and Th2-type inflammation seen in this model of asthma.     

Cutting edge: Th2 cell trafficking into the allergic lung is dependent on chemoattractant receptor signaling

Th2 cells are recruited to the lung where they mediate the asthma phenotype. Since the molecular mechanisms regulating Th2 cell trafficking remain unknown, we sought to determine whether trafficking of Th2 cells into the lung is mediated by G alpha i-coupled chemoattractant receptors. We show here that in contrast to untreated Th2 cells, pertussis toxin-treated Th2 cells were unable to traffic into the lung, airways, or lymph nodes following Ag challenge and therefore were unable to induce allergic inflammation in vivo. Pertussis toxin-treated Th2 cells were functional cells, however, and when directly instilled into the airways of mice, bypassing their need to traffic to the lung, were able to induce airway eosinophilic inflammation. These studies conclusively demonstrate that trafficking of Th2 cells into the lung is an active process dependent on chemoattractant receptors.     

The Toll-like receptor 5 stimulus bacterial flagellin induces maturation and chemokine production in human dendritic cells

Toll-like receptors (TLRs) are pattern recognition receptors that serve an important function in detecting pathogens and initiating inflammatory responses. Upon encounter with foreign Ag, dendritic cells (DCs) go through a maturation process characterized by an increase in surface expression of MHC class II and costimulatory molecules, which leads to initiation of an effective immune response in naive T cells. The innate immune response to bacterial flagellin is mediated by TLR5, which is expressed on human DCs. Therefore, we sought to investigate whether flagellin could induce DC maturation. Immature DCs were cultured in the absence or presence of flagellin and monitored for expression of cell surface maturation markers. Stimulation with flagellin induced increased surface expression of CD83, CD80, CD86, MHC class II, and the lymph node-homing chemokine receptor CCR7. Flagellin stimulated the expression of chemokines active on neutrophils (IL-8/CXC chemokine ligand (CXCL)8, GRO-alpha/CXCL1, GRO-beta/CXCL2, GRO-gamma/CXCL3), monocytes (monocyte chemoattractant protein-1/CC chemokine ligand (CCL)2), and immature DCs (macrophage-inflammatory protein-1 alpha/CCL3, macrophage-inflammatory protein-1 beta/CCL4), but not chemokines active on effector T cells (IFN-inducible protein-10 kDa/CXCL10, monokine induced by IFN-gamma/CXCL9, IFN-inducible T cell alpha chemoattractant/CXCL11). However, stimulating DCs with both flagellin and IFN-inducible protein-10 kDa, monokine induced by IFN-gamma, and IFN-inducible T cell alpha chemoattractant expression, whereas stimulation with IFN-beta or flagellin alone failed to induce these chemokines. In functional assays, flagellin-matured DCs displayed enhanced T cell stimulatory activity with a concomitant decrease in endocytic activity. Finally, DCs isolated from mouse spleens or bone marrows were shown to not express TLR5 and were not responsive to flagellin stimulation. These results demonstrate that flagellin can directly stimulate human but not murine DC maturation, providing an additional mechanism by which motile bacteria can initiate an acquired immune response.     

CD1d-restricted NKT cells express a chemokine receptor profile indicative of Th1-type inflammatory homing cells

CD1d-restricted T cells (NKT cells) are innate memory cells activated by lipid Ags and play important roles in the initiation and regulation of the immune response. However, little is known about the trafficking patterns of these cells or the tissue compartment in which they exert their regulatory activity. In this study, we determined the chemokine receptor profile expressed by CD1d-restricted T cells found in the peripheral blood of healthy volunteers as well as CD1d-restricted T cell clones. CD1d-restricted T cells were identified by Abs recognizing the invariant Valpha24 TCR rearrangement or by binding to CD1d-Fc fusion tetramers loaded with alpha-GalCer. CD1d-restricted T cells in the peripheral blood and CD1d-restricted T cell clones expressed high levels of CXCR3, CCR5, and CCR6; intermediate levels of CXCR4 and CXCR6; and low levels of CXCR1, CCR1, CCR2, and CX(3)CR1, a receptor pattern often associated with tissue-infiltrating effector Th1 cells and CD8+ T cells. Very few of these cells expressed the lymphoid-homing receptors CCR7 or CXCR5. CCR4 was expressed predominantly on CD4+, but not on double-negative CD1d-restricted T cells, which may indicate differential trafficking patterns for these two functionally distinct subsets. CD1d-restricted T cell clones responded to chemokine ligands for CXCR1/2, CXCR3, CXCR4, CXCR6, CCR4, and CCR5 in calcium flux and/or chemotaxis assays. These data indicate that CD1d-restricted T cells express a chemokine receptor profile most similar to Th1 inflammatory homing cells and suggest that these cells perform their function in peripheral tissue sites rather than in secondary lymphoid organs.     

Peroxisome proliferator-activated receptor-gamma activators inhibit IFN-gamma-induced expression of the T cell-active CXC chemokines IP-10, Mig, and I-TAC in human endothelial cells

Peroxisome proliferator-activated receptor-gamma (PPARgamma), a member of the nuclear hormone receptor superfamily originally shown to play an important role in adipocyte differentiation and glucose homeostasis, is now known to regulate inflammatory responses. Given the importance of endothelial cell (EC)-derived chemokines in regulating leukocyte function and trafficking, we studied the effects of PPARgamma ligands on the expression of chemokines induced in ECs by the Th1 cytokine IFN-gamma. Treatment of ECs with PPARgamma activators significantly inhibited IFN-gamma-induced mRNA and protein expression of the CXC chemokines IFN-inducible protein of 10 kDa (IP-10), monokine induced by IFN-gamma (Mig), and IFN-inducible T-cell alpha-chemoattractant (I-TAC), whereas expression of the CC chemokine monocyte chemoattractant protein-1 was not altered. PPARgamma activators decreased IFN-inducible protein of 10 kDa promoter activity and inhibited protein binding to the two NF-kappaB sites but not to the IFN-stimulated response element ISRE site. Furthermore, PPARgamma ligands inhibited the release of chemotactic activity for CXC chemokine receptor 3 (CXCR3)-transfected lymphocytes from IFN-gamma-stimulated ECs. These data suggest that anti-diabetic PPARgamma activators might attenuate the recruitment of activated T cells at sites of Th1-mediated inflammation.