Innate immune responses to allergens by airway epithelial cells (AECs) help initiate and propagate the adaptive immune response associated with allergic airway inflammation in asthma. Activation of the transcription factor NF-κB in AECs by allergens or secondary mediators via G protein–coupled receptors (GPCRs) is an important component of this multifaceted inflammatory cascade. Members of the caspase recruitment domain family of proteins display tissue-specific expression and help mediate NF-κB activity in response to numerous stimuli. We have previously shown that caspase recruitment domain–containing membrane-associated guanylate kinase protein (CARMA)3 is specifically expressed in AECs and mediates NF-κB activation in these cells in response to stimulation with the GPCR agonist lysophosphatidic acid. In this study, we demonstrate that reduced levels of CARMA3 in normal human bronchial epithelial cells decreases the production of proasthmatic mediators in response to a panel of asthma-relevant GPCR ligands such as lysophosphatidic acid, adenosine triphosphate, and allergens that activate GPCRs such as Alternaria alternata and house dust mite. We then show that genetically modified mice with CARMA3-deficient AECs have reduced airway eosinophilia and proinflammatory cytokine production in a murine model of allergic airway inflammation. Additionally, we demonstrate that these mice have impaired dendritic cell maturation in the lung and that dendritic cells from mice with CARMA3-deficient AECs have impaired Ag processing. In conclusion, we show that AEC CARMA3 helps mediate allergic airway inflammation, and that CARMA3 is a critical signaling molecule bridging the innate and adaptive immune responses in the lung.
Caspase recruitment domain-containing membrane-associated guanylate kinase protein-1 (CARMA1), a member of the membrane associated guanylate kinase (MAGUK) family of kinases, is essential for T lymphocyte activation and proliferation via T cell receptor (TCR) mediated NF-κB activation. Recent studies suggest a broader role for CARMA1 regulating other T cell functions as well as a role in non-TCR mediated signaling pathways important for lymphocyte development and functions. In addition, CARMA1 has been shown to be an important component in the pathogenesis of several human diseases. Thus, comprehensively defining its mechanisms of action and regulation could reveal novel therapeutic targets for T cell-mediated diseases and lymphoproliferative disorders.
CARMA1; T cell; T cell receptor; NF-κB
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.
Influenza is a major cause of morbidity and mortality in the United States. Studies have shown that excessive T cell activity can mediate pneumonitis in the setting of influenza infection, and data from the 2009 H1N1 pandemic indicate that critical illness and respiratory failure following infection was associated with greater infiltration of the lungs with CD8+ T cells. T cell immunoglobulin and mucin domain 3 (Tim3) is a negative regulator of Th1/Tc1-type immune responses. Activation of Tim3 on effector T cells has been shown to down-regulate proliferation, cell-mediated cytotoxicity, and IFNγ production as well as induce apoptosis. Here, we demonstrate that deletion of the terminal cytoplasmic domain of the Tim3 gene potentiates its ability to down-regulate Tc1 inflammation and that this enhanced Tim3 activity is associated with decreased phosphorylation of the TCR-CD3ζ chain. We then show that mice with this Tim3 mutation infected with influenza are protected from morbidity and mortality without impairment in viral clearance or functional heterotypic immunity. This protection is associated with decreased CD8+ T cell proliferation and decreased production of inflammatory cytokines, including IFNγ. Furthermore, the Tim3 mutation was protective against mortality in a CD8+ T cell-specific model of pneumonitis. These data suggest that Tim3 could be targeted to prevent immunopathology during influenza infection and demonstrate a potentially novel signaling mechanism utilized by Tim3 to down-regulate the Tc1 response.
CARMA1 is a lymphocyte-specific scaffold protein necessary for T cell activation. Deletion of CARMA1 prevents the development of allergic airway inflammation in a mouse model of asthma due to a defect in naïve T cell activation. However, it is unknown if CARMA1 is important for effector and memory T cell responses after the initial establishment of inflammation, findings which would be more relevant to asthma therapies targeted to CARMA1. In the current study, we sought to elucidate the role of CARMA1 in T cells that have been previously activated. Using mice in which floxed CARMA1 exons can be selectively deleted in T cells by OX40 driven Cre recombinase (OX40+/CreCARMA1F/F), we report that CD4+ T cells from these mice have impaired T cell reactivation responses and NF-κB signaling in vitro. Furthermore, in an in vivo recall model of allergic airway inflammation that is dependent on memory T cell function, OX40+/CreCARMA1F/F mice have attenuated eosinophilic airway inflammation, T cell activation and Th2 cytokine production. Using MHC class II tetramers, we demonstrate that the development and maintenance of antigen-specific memory T cells is not affected in OX40+/CreCARMA1F/F mice. In addition, adoptive transfer of Th2-polarized OX40+/CreCARMA1F/F antigen-specific CD4+ T cells into wild-type mice induces markedly less airway inflammation in response to antigen challenge than transfer of wild-type Th2 cells. These data demonstrate a novel role for CARMA1 in effector and memory T cell responses and suggests that therapeutic strategies targeting CARMA1 could help treat chronic inflammatory disorders such as asthma.
Remodeling of the pulmonary arteries is a common feature among the heterogeneous disorders that cause pulmonary hypertension. In these disorders, the remodeled pulmonary arteries often demonstrate inflammation and an accumulation of pulmonary artery smooth muscle cells (PASMCs) within the vessels. Adipose tissue secretes multiple bioactive mediators (adipokines) that can influence both inflammation and remodeling, suggesting that adipokines may contribute to the development of pulmonary hypertension. We recently reported on a model of pulmonary hypertension induced by vascular inflammation, in which a deficiency of the adipokine adiponectin (APN) was associated with the extensive proliferation of PASMCs and increased pulmonary artery pressures. Based on these data, we hypothesize that APN can suppress pulmonary hypertension by directly inhibiting the proliferation of PASMCs. Here, we tested the effects of APN overexpression on pulmonary arterial remodeling by using APN-overexpressing mice in a model of pulmonary hypertension induced by inflammation. Consistent with our hypothesis, mice that overexpressed APN manfiested reduced pulmonary hypertension and remodeling compared with wild-type mice, despite developing similar levels of pulmonary vascular inflammation in the model. The overexpression of APN was also protective in a hypoxic model of pulmonary hypertension. Furthermore, APN suppressed the proliferation of PASMCs, and reduced the activity of the serum response factor–serum response element pathway, which is a critical signaling pathway for smooth muscle cell proliferation. Overall, these data suggest that APN can regulate pulmonary hypertension and pulmonary arterial remodeling through its direct effects on PASMCs. Hence, the activation of APN-like activity in the pulmonary vasculature may be beneficial in pulmonary hypertension.
pulmonary hypertension; pulmonary artery smooth muscle cells; metabolism; adiponectin
The balance between Th17 and T regulatory (Treg) cells critically modulates immune homeostasis, with an inadequate Treg response contributing to inflammatory disease. Using an unbiased chemical biology approach, we identified a novel role for the dual specificity tyrosine-phosphorylation-regulated kinase DYRK1A in regulating this balance. Inhibition of DYRK1A enhances Treg differentiation and impairs Th17 differentiation without affecting known pathways of Treg/Th17 differentiation. Thus, DYRK1A represents a novel mechanistic node at the branch point between commitment to either Treg or Th17 lineages. Importantly, both Treg cells generated using the DYRK1A inhibitor harmine and direct administration of harmine itself potently attenuate inflammation in multiple experimental models of systemic autoimmunity and mucosal inflammation. Our results identify DYRK1A as a physiologically relevant regulator of Treg cell differentiation and suggest a broader role for other DYRK family members in immune homeostasis. These results are discussed in the context of human diseases associated with dysregulated DYRK activity.
Inflammation is used by the immune system to protect and repair tissues after an injury or infection. However, if inflammation is too strong, or goes on for too long, it can damage tissues. This is seen in autoimmune diseases such as inflammatory bowel disease and type 1 diabetes. Therefore, precise regulation of the inflammatory response is essential for maintaining human health.
White blood cells known as T cells are central regulators of tissue inflammation. To achieve this goal, they develop into subtypes with specialized roles. For example, some T helper cells release chemical signals that trigger inflammation and other immune responses. Regulatory T (Treg) cells then shut down these immune responses once they are no longer needed. Many autoimmune and other inflammatory diseases are thought to arise—at least partially—because Treg cells fail to stop the inflammatory response. Boosting the number or the activity of Treg cells could therefore help to treat these diseases. However, technical difficulties have made it difficult to investigate the genes and molecular pathways that control how this subtype of white blood cells develops.
Khor et al. thought that discovering new chemicals that increase the number of Treg cells without harming them could help to identify the pathways that control their development. Khor et al. screened over 3000 chemicals, many of which are drugs currently approved for use in humans, for their effect on immature T cells that were taken from mice and grown in the laboratory. This ‘unbiased chemical biology’ approach identified several chemicals that both encouraged the T cells to develop into Treg cells and reduced the numbers that became inflammation-promoting T helper cells.
Khor et al. then focused on one of these chemicals, called harmine. Tests in mice showed that harmine reduces the extent of experimentally induced inflammatory reactions. Treg cells generated by treating immature T cells with harmine had the same effect. Further experiments showed that harmine exerts these effects, at least in part, by inhibiting the activity of a protein called DYRK1A. When DYRK1A was removed from maturing mouse T cells grown in the laboratory, the T cells tended to develop into anti-inflammatory Treg cells.
These findings therefore identify DYRK1A as part of a pathway that suppresses the development of Treg cells. It remains to be discovered how it does this, and whether other DYRK protein family members have similar roles.
T cell differentiation; inflammation; dual-specificity tyrosine-regulated kinase signaling; human; mouse
Pulmonary arterial hypertension (PAH) is a condition of unknown etiology whose pathological features include increased vascular resistance, perivascular inflammatory cell infiltration and pulmonary arteriolar remodeling. Although risk factors for PAH are poorly defined, recent studies indicate that obesity may be an important risk factor for this condition. The mechanisms leading to this association are largely unknown, but bioactive mediators secreted from adipose tissue have been implicated in this process. One of the most important mediators released from adipose tissue is the adipokine adiponectin. Adiponectin is highly abundant in the circulation of lean healthy individuals, and possesses well-described metabolic and antiinflammatory actions. Levels of adiponectin decrease with increasing body mass, and low levels are directly linked to the development of PAH in mice. Moreover, overexpression of adiponectin has been shown to protect mice from developing PAH in response to inflammation and hypoxia. Based on the findings from these studies, it is suggested that the effects of adiponectin are mediated, in part, through its antiinflammatory and antiproliferative properties. In this review, we discuss the emerging evidence demonstrating a role for adiponectin in lung vascular homeostasis and discuss how deficiency in this adipocyte-derived hormone might explain the recent association between obesity and PAH.
adipocyte; adipokine; inflammation; metabolism; treatment of pulmonary hypertension
Obesity is associated with an increased incidence and severity of asthma, as well as other lung disorders, such as pulmonary hypertension. Adiponectin (APN), an antiinflammatory adipocytokine, circulates at lower levels in the obese, which is thought to contribute to obesity-related inflammatory diseases. We sought to determine the effects of APN deficiency in a murine model of chronic asthma. Allergic airway inflammation was induced in APN-deficient mice (APN−/−) using sensitization without adjuvant followed by airway challenge with ovalbumin. The mice were then analyzed for changes in inflammation and lung remodeling. APN−/− mice in this model develop increased allergic airway inflammation compared with wild-type mice, with greater accumulation of eosinophils and monocytes in the airways associated with elevated lung chemokine levels. Surprisingly, APN−/− mice developed severe pulmonary arterial muscularization and pulmonary arterial hypertension in this model, whereas wild-type mice had only mild vascular remodeling and comparatively less pulmonary arterial hypertension. Our findings demonstrate that APN modulates allergic inflammation and pulmonary vascular remodeling in a model of chronic asthma. These data provide a possible mechanism for the association between obesity and asthma, and suggest a potential novel link between obesity, inflammatory lung disease, and pulmonary hypertension.
asthma; obesity; pulmonary hypertension
Caspase recruitment domain-containing membrane-associated guanylate kinase protein-1 (CARMA1) is a critical component of the nuclear factor κB (NF-κB) signaling cascade mediated by T cell receptor (TCR) engagement. In addition to activation of naïve T cells, TCR signaling is important for the development of agonist-selected T cell subsets such as regulatory T cells (Treg), natural killer T cells (NKT), and CD8ααT cells. However, little is known about the role of CARMA1 in the development of these lineages. Here we show that CARMA1-deficient mice (CARMA1−/−) have altered populations of specific subsets of agonist-selected T cells. Specifically, CARMA1−/− mice have impaired natural and adaptive Treg development, while NKT cell numbers are normal compared to wild-type mice. Interestingly, CD8αα T cells, which may also be able to develop through an extrathymic selection pathway, are enriched in the gut of CARMA1−/− mice, while memory-phenotype CD4+ T cells (CD62Llow/CD44high) are present at reduced numbers in the periphery. These results indicate that CARMA1 is essential for Treg development, but is not necessary for the development of other agonist-selected T cell subsets. Overall, these data reveal an important but differential role for CARMA1-mediated TCR signaling in T cell development.
CARMA1; regulatory T cell; NKT cell; CD8αα T cell
Induction of endogenous regulatory T cells (Tregs) represents an exciting new potential modality for treating allergic diseases such as asthma. Tregs have been implicated in the regulation of asthma but the anatomic location where they exert their regulatory function, and the mechanisms controlling their migration necessary for their suppressive function in asthma are not known. Understanding these aspects of Treg biology will be important for harnessing their power in the clinic.
To determine the anatomic location where Tregs exert their regulatory function in the sensitization and effector phases of allergic asthma, and to determine the chemokine receptors that control the migration of Tregs to these sites in vivo in mice and in humans.
The clinical efficacy and the anatomic location of adoptively transferred chemokine receptor-deficient CD4+CD25+ Foxp3+ Tregs was determined in the sensitization and effector phases of allergic airway inflammation in mice. The chemokine receptor expression profile was determined on Tregs recruited into the human airway following bronchoscopic segmental allergen challenge of subjects with asthma.
We show that CCR7, but not CCR4, is required on Tregs to suppress allergic airway inflammation during the sensitization phase. In contrast, CCR4, but not CCR7, is required on Tregs to suppress allergic airway inflammation during the effector phase. Consistent with our murine studies, humans with allergic asthma had an increase in CCR4 expressing functional Tregs in the lung following segmental allergen challenge.
The location of Treg function differs during allergic sensitization and during allergen-induced recall responses in the lung, and that this differential localization is critically dependent on differential chemokine function.
Asthma; regulatory T cells; chemokines; CCR4; CCR7; segmental allergen challenge
Cellular plasticity contributes to the regenerative capacity of plants, invertebrates, teleost fishes, and amphibians. In vertebrates, differentiated cells are known to revert into replicating progenitors, but these cells do not persist as stable stem cells. We now present evidence that differentiated airway epithelial cells can revert into stable and functional stem cells in vivo. Following the ablation of airway stem cells, we observed a surprising increase in the proliferation of committed secretory cells. Subsequent lineage tracing demonstrated that the luminal secretory cells had dedifferentiated into basal stem cells. Dedifferentiated cells were morphologically indistinguishable from stem cells and they functioned as well as their endogenous counterparts to repair epithelial injury. Indeed, single secretory cells clonally dedifferentiated into multipotent stem cells when they were cultured ex vivo without basal stem cells. In contrast, direct contact with a single basal stem cell was sufficient to prevent secretory cell dedifferentiation. In analogy to classical descriptions of amphibian nuclear reprogramming, the propensity of committed cells to dedifferentiate was inversely correlated to their state of maturity. This capacity of committed cells to dedifferentiate into stem cells may play a more general role in the regeneration of many tissues and in multiple disease states, notably cancer.
In asthma, the relationship among airway inflammation, airway hyperresponsiveness, and lung function is poorly understood. Methods to noninvasively assess these relationships in human subjects are needed. We sought to determine whether 18F-FDG uptake rate (Ki, min−1) could serve as a biomarker of eosinophilic inflammation and local lung function.
We used PET/CT to assess regional pulmonary perfusion (Q̇), specific ventilation per unit volume (sV̇A), fractional gas content (Fgas), airway wall thickness, and regional Ki 10 h after segmental allergen challenge to the right middle lobe in 6 asthmatic subjects with demonstrated atopy. Q̇, sV̇A, and Fgas in the allergen-challenged lobe were compared with the right upper lobe, where diluent was applied as a control. The airway wall thickness aspect ratio (ω) of the allergen-challenged airway was compared with those of similarly sized airways from unaffected areas of the lung. Differences in Ki between allergen and diluent segments were compared with those in cell counts obtained 24 h after the allergen challenge by a bronchoalveolar lavage of the respective segments.
We found systematic reductions in regional Q̇, sV̇A, and Fgas and increased ω in all subjects. The ratio of eosinophil count (allergen to diluent) was linearly related (R2 = 0.9917, P < 0.001) to the ratio of Ki.
Regional Ki measured with PET is a noninvasive and highly predictive biomarker of eosinophilic airway inflammation and its functional effects. This method may serve to help in the understanding of allergic inflammation and test the therapeutic effectiveness of novel drugs or treatments.
airway constriction; ventilation-perfusion ratio; pulmonary gas exchange; fluorine isotopes; emission-computed tomography; nitrogen isotopes
Acute rejection, a common complication of lung transplantation, may promote obliterative bronchiolitis leading to graft failure in lung transplant recipients. During acute rejection episodes, CD8+ T cells can contribute to lung epithelial injury but the mechanisms promoting and controlling CD8-mediated injury in the lung are not well understood. To study the mechanisms regulating CD8+ T cell–mediated lung rejection, we used a transgenic model in which adoptively transferred ovalbumin (OVA)-specific cytotoxic T lymphocytes (CTL) induce lung injury in mice expressing an ovalbumin transgene in the small airway epithelium of the lungs (CC10-OVA mice). The lung pathology is similar to findings in humans with acute lung transplant. In the presence of an intact immune response the inflammation resolves by day 30. Using CC10-OVA.RAG-/- mice, we found that CD4+ T cells and ICOS+/+ T cells were required for protection against lethal lung injury, while neutrophil depletion was not protective. In addition, CD4+Foxp3 + ICOS+ T cells were enriched in the lungs of animals surviving lung injury and ICOS+/+ Tregs promoted survival in animals that received ICOS-/- T cells. Direct comparison of ICOS-/- Tregs to ICOS+/+ Tregs found defects in vitro but no differences in the ability of ICOS-/- Tregs to protect from lethal lung injury. These data suggest that ICOS affects Treg development but is not necessarily required for Treg effector function.
Interleukin (IL-) 36 cytokines (previously designated as novel IL-1 family member cytokines; IL-1F5– IL-1F10) constitute a novel cluster of cytokines structurally and functionally similar to members of the IL-1 cytokine cluster. The effects of IL-36 cytokines in inflammatory lung disorders remains poorly understood. The current study sought to investigate the effects of IL-36α (IL-1F6) and test the hypothesis that IL-36α acts as a pro-inflammatory cytokine in the lung in vivo. Intratracheal instillation of recombinant mouse IL-36α induced neutrophil influx in the lungs of wild-type C57BL/6 mice and IL-1αβ−/− mice in vivo. IL-36α induced neutrophil influx was also associated with increased mRNA expression of neutrophil-specific chemokines CXCL1 and CXCL2 in the lungs of C57BL/6 and IL-1αβ−/− mice in vivo. In addition, intratracheal instillation of IL-36α enhanced mRNA expression of its receptor IL-36R in the lungs of C57BL/6 as well as IL-1αβ−/− mice in vivo. Furthermore, in vitro incubation of CD11c+ cells with IL-36α resulted in the generation of neutrophil-specific chemokines CXCL1, CXCL2 as well as TNFα. IL-36α increased the expression of the co-stimulatory molecule CD40 and enhanced the ability of CD11c+ cells to induce CD4+ T cell proliferation in vitro. Furthermore, stimulation with IL-36α activated NF-κB in a mouse macrophage cell line. These results demonstrate that IL-36α acts as a pro-inflammatory cytokine in the lung without the contribution of IL-1α and IL-1β. The current study describes the pro-inflammatory effects of IL-36α in the lung, demonstrates the functional redundancy of IL-36α with other agonist cytokines in the IL-1 and IL-36 cytokine cluster, and suggests that therapeutic targeting of IL-36 cytokines could be beneficial in inflammatory lung diseases.
Interleukin-1 (IL-1) is a proinflammatory cytokine that signals through the Type I IL-1 receptor (IL-1RI). Novel IL-1–like cytokines were recently identified. Their functions in lung disease remain unclear. Interleukin-1 family member–9 (IL-1F9) is one such IL-1–like cytokine, expressed in the lungs of humans and mice. IL-1F9 signals through IL-1 receptor–related protein 2 (IL-1Rrp2/IL-1RL2), which is distinct from IL-1RI. We sought to determine if IL-1F9 acts as a proinflammatory cytokine in lung disease. IL-1F9 protein was increased in lung homogenates of house dust mite–challenged A/J mice compared with controls, and expression was seen in airway epithelial cells. The intratracheal administration of recombinant mouse IL-1F9 increased airway hyperresponsiveness and induced neutrophil influx and mucus production, but not eosinophilic infiltration in the lungs of mice. In addition, IL-1α protein levels in bronchoalveolar lavage fluid, chemokines, and chemokine-receptor mRNA expression in the lungs were increased after the instillation of intratracheal IL-1F9. Consistent with these changes, NF-κB transcription factor activity was increased in the lungs of mice challenged with IL-1F9 and in a macrophage cell line treated with IL-1F9. These data suggest that IL-1F9 is upregulated during inflammation, and acts as a proinflammatory cytokine in the lungs.
IL-1F9; chemokines; lung diseases; inflammation; neutrophil
Optical projection tomography is a new ex vivo imaging technique that allows imaging of whole organs in three dimensions at high spatial resolutions. In this Letter we demonstrate its capability to tomographically visualize molecular activity in whole organs of mice. In particular, eosinophil activity in asthmatic lungs is resolved using a Born-normalized fluorescence optical projection tomography and employing a near-IR molecular probe. The possibility to achieve molecularly sensitive imaging contrast in optical projection tomography by means of targeted and activatable imaging reporter agents adds a new range of capabilities for investigating molecular signatures of pathophysiological processes and a wide variety of diseases and their development.
NF-κB activation in bronchial epithelial cells is important for the development of allergic airway inflammation, and may control the expression of critical mediators of allergic inflammation such as thymic stromal lymphopoietin (TSLP) and the chemokine CCL20. Members of the caspase recruitment domain (CARD) family of proteins are differentially expressed in tissue and help mediate NF-κB activity in response to numerous stimuli. Here we demonstrate that CARMA3 (CARD10) is specifically expressed in human airway epithelial cells, and that expression of CARMA3 in these cells leads to activation of NF-κB. CARMA3 has recently been shown to mediate NF-κB activation in embryonic fibroblasts after stimulation with lysophosphatidic acid (LPA), a bioactive lipid-mediator that is elevated in the lungs of individuals with asthma. Consistent with this, we demonstrate that stimulation of airway epithelial cells with LPA leads to increased expression of TSLP and CCL20. We then show that inhibition of CARMA3 activity in airway epithelial cells reduces LPA-mediated NF-κB activity and the production of TSLP and CCL20. In conclusion, these data demonstrate that LPA stimulates TSLP and CCL20 expression in bronchial epithelial cells via CARMA3-mediated NF-κB activation.
bronchial epithelial cells; asthma; lysophosphatidic acid; thymic stromal lymphopoietin; CARMA3
Recruitment of antigen-specific Th2 cells into the lung is critical for the development of allergic airway inflammation. Although CCR4 and CCR8 are preferentially expressed on Th2 cells and CCR4, CCR8 and CXCR3 ligands are increased in asthma, the specific relative contribution of these receptors to antigen-specific Th2 cell trafficking into the allergic lung is not known.
To determine the relative contribution of CCR4, CCR8 and CXCR3 to antigen-specific Th2 cell trafficking in a murine model of allergic pulmonary inflammation.
We used adoptive transfer experiments to compare the trafficking of wild type antigen-specific Th2 cells with antigen-specific Th2 cells deficient in CCR4, CCR8 or CXCR3.
CCR4-deficient antigen-specific Th2 cells failed to traffic efficiently into the lung and the airways. In contrast, CCR8-deficient antigen-specific Th2 cells accumulated in these sites. Trafficking of CXCR3-deficient antigen-specific Th2 cells and CCR4-deficient and CCR8-deficient antigen-specific Th1 cells were comparable to their wild type counterparts. Approximately 60% of IL-4 producing antigen-specific T cells expressed CCR4. Disruption of CCR4-mediated antigen-specific Th2 cell trafficking decreased the levels of Th2-type cytokines in the airways and reduced airway eosinophila and mucus production.
Our study demonstrates that CCR4 is required for the efficient entry of antigen-specific Th2 cells into the lung and the airways in murine model of allergic pulmonary inflammation.
Inhibition of CCR4-mediated Th2 cell trafficking may contribute to asthma therapy.
CCR4 is required for efficient entry of Th2 cells into the allergic lung. Disruption of CCR4-mediated antigen-specific Th2 cell trafficking decreases airway Th2-type cytokines, eosinophilia and mucus production,
CCR4; CCR8; CXCR3; asthma; chemokine; T cell trafficking
STAT6-mediated chemokine production in the lung is required for Th2 lymphocyte and eosinophil homing into the airways in allergic pulmonary inflammation, and thus is a potential therapeutic target in asthma. However, the critical cellular source of STAT6-mediated chemokine production has not been defined. Here we demonstrate that STAT6 in bone marrow-derived myeloid cells was sufficient for the production of CCL17, CCL22, CCL11, and CCL24 and for Th2 lymphocyte and eosinophil recruitment into the allergic airway. In contrast, STAT6 in airway lining cells did not mediate chemokine production or support cellular recruitment. Selective depletion of CD11b+ myeloid cells in the lung identified these cells as the critical cellular source for the chemokines CCL17 and CCL22. These data reveal that CD11b+ myeloid cells in the lung help orchestrate the adaptive immune response in asthma, in part, through the production of STAT6-inducible chemokines and the recruitment of Th2 lymphocytes into the airway.
Th2 cells; allergy; chemokines; eosinophils
Mice lacking the chemokine receptor CCR5 are susceptible to mortality from a normally non-lethal influenza infection. Here we found that CXCR3-deficiency rescued CCR5-deficient mice from influenza-induced mortality. The number of mononuclear phagocytes in the airways was transiently increased in CCR5-deficient mice but not in CXCR3-CCR5 double-deficient mice. Antigen-specific CXCR3-CCR5 double-deficient CD8 effector cells were less efficient at entering the airways compared to wild-type or CCR5-deficient CD8 effector cells. The decrease in inflammatory cell infiltrates in CXCR3-CCR5 double-deficient infected mice correlated with a decrease in CCL2 and IFNγ production in the airways. Finally, CXCR3-CCR5 double-deficient mice that survived the primary viral challenge were protected from a lethal secondary challenge, indicating that T cell-mediated protective memory was not compromised in mice lacking these chemokine receptors. In conclusion, CXCR3 deficiency attenuated the lethal cellular immune response in CCR5-deficient influenza-infected mice without hindering viral clearance or long-term immunity.
T cells; Virology; Chemokines; Lung inflammation
Trafficking of leukocytes to sites of inflammation is an important step in the establishment of an immune response. Chemokines are critical regulators of leukocyte trafficking and are widely studied molecules for their important role in disease and for their potential as new therapeutic targets. The ability of chemokines to induce leukocyte recruitment has been mainly measured by in vitro chemotaxis assays, which lack many components of the complex biological process of leukocyte migration and therefore provide incomplete information about chemokine function in vivo. In vivo assays to study the activity of chemokines to induce leukocyte recruitment have been difficult to establish. We describe here the development of a robust in vivo recruitment assay for CD8+ and CD4+ T lymphocytes induced by the CXCR3 ligands IP-10 (CXCL10) and I-TAC (CXCL11). For this assay, in vitro activated T lymphocytes were adoptively transferred into the peritoneum of naïve mice. Homing of these transferred T lymphocytes into the airways was measured following intratracheal instillation of chemokines. High recruitment indices were achieved that were dependent on chemokine concentration and CXCR3 expression on the transferred lymphocytes. Recruitment was also inhibited by antibodies to the chemokine. The assay models the natural condition of chemokine-mediated lymphocyte migration into the airways as chemokines are expressed in the airways during inflammation. The nature of this model allows flexibility to study wildtype and mutant chemokines and chemokine receptors and the ability to evaluate chemokine antagonists and antibodies in vivo. This assay will therefore help elucidate a deeper understanding of the chemokine system in vivo.
Chemokines; cell trafficking; chemotaxis
Leukotriene B4 is a lipid mediator that recently has been shown to have potent chemotactic activity for effector T lymphocytes mediated through its receptor, BLT1. Here, we developed a novel murine model of acute lung rejection to demonstrate that BLT1 controls effector CD8+ T cell trafficking into the lung and that disruption of BLT1 signaling in CD8+ T cells reduces lung inflammation and mortality in the model. In addition, we used BLT1-deficient mice and a BLT1 antagonist in two tracheal transplant models of lung transplantation to demonstrate the importance of BLT1 for the recruitment of T cells into tracheal allografts. We also show that BLT1-mediated CD8+ T cell recruitment plays an important role in the development of airway fibroproliferation and obliteration. Finally, in human studies of lung transplant recipients, we found that BLT1 is up-regulated on T lymphocytes isolated from the airways of patients with obliterative bronchiolitis. These data demonstrate that BLT1 contributes to the development of lung rejection and obliterative bronchiolitis by mediating effector T lymphocyte trafficking into the lung. This is the first report that describes a pathologic role for BLT1-mediated T lymphocyte recruitment in disease and identifies BLT1 as a potential therapeutic target after lung transplantation.