Satoko Miyahara

Effector CD8+ T cells mediate inflammation and airway hyper-responsiveness

Allergic asthma is a complex syndrome characterized by airway obstruction, airway inflammation and airway hyper-responsiveness (AHR). Using a mouse model of allergen-induced AHR, we previously demonstrated that CD8-deficient mice develop significantly lower AHR, eosinophilic inflammation and interleukin (IL)-13 levels in bronchoalveolar lavage fluid compared with wild-type mice. These responses were restored by adoptive transfer of antigen-primed CD8+ T cells. Previously, two distinct populations of antigen-experienced CD8+ T cells, termed effector (TEFF) and central memory (TCM) cells, have been described. After adoptive transfer into CD8-deficient mice, TEFF, but not TCM, cells restored AHR, eosinophilic inflammation and IL-13 levels. TEFF, but not TCM, cells accumulated in the lungs, and intracellular cytokine staining showed that the transferred TEFF cells were a source of IL-13. These data suggest an important role for effector CD8+ T cells in the development of AHR and airway inflammation, which may be associated with their Tc2-type cytokine production and their capacity to migrate into the lung.

Contribution of Antigen-Primed CD8+ T Cells to the Development of Airway Hyperresponsiveness and Inflammation Is Associated with IL-13

The role of Th2/CD4 T cells, which secrete IL-4, IL-5, and IL-13, in allergic disease is well established; however, the role of CD8+ T cells (allergen-induced airway hyperresponsiveness (AHR) and inflammation) is less clear. This study was conducted to define the role of Ag-primed CD8+ T cells in the development of these allergen-induced responses. CD8-deficient (CD8−/−) mice and wild-type mice were sensitized to OVA by i.p. injection and then challenged with OVA via the airways. Compared with wild-type mice, CD8−/− mice developed significantly lower airway responsiveness to inhaled methacholine and lung eosinophilia, and exhibited decreased IL-13 production both in vivo, in the bronchoalveolar lavage (BAL) fluid, and in vitro, following Ag stimulation of peribronchial lymph node (PBLN) cells in culture. Reconstitution of sensitized and challenged CD8−/− mice with allergen-sensitized CD8+ T cells fully restored the development of AHR, BAL eosinophilia, and IL-13 levels in BAL and in culture supernatants from PBLN cells. In contrast, transfer of naive CD8+ T cells or allergen-sensitized CD8+ T cells from IL-13-deficient donor mice failed to do so. Intracellular cytokine staining of lung as well as PBLN T cells revealed that CD8+ T cells were a source of IL-13. These data suggest that Ag-primed CD8+ T cells are required for the full development of AHR and airway inflammation, which appears to be associated with IL-13 production from these primed T cells.

Leukotriene B4 Receptor-1 Is Essential for Allergen-Mediated Recruitment of CD8 + T Cells and Airway Hyperresponsiveness

Recent studies in both human and rodents have indicated that in addition to CD4+ T cells, CD8+ T cells play an important role in allergic inflammation. We previously demonstrated that allergen-sensitized and -challenged CD8-deficient (CD8−/−) mice develop significantly lower airway hyperresponsiveness (AHR), eosinophilic inflammation, and IL-13 levels in bronchoalveolar lavage fluid compared with wild-type mice, and that all these responses were restored by adoptive transfer of in vivo-primed CD8+ T cells or in vitro-generated effector CD8+ T cells (TEFF). Recently, leukotriene B4 and its high affinity receptor, BLT1, have been shown to mediate in vitro-generated TEFF recruitment into inflamed tissues. In this study we investigated whether BLT1 is essential for the development of CD8+ T cell-mediated allergic AHR and inflammation. Adoptive transfer of in vivo-primed BLT1+/+, but not BLT1−/−, CD8+ T cells into sensitized and challenged CD8−/− mice restored AHR, eosinophilic inflammation, and IL-13 levels. Moreover, when adoptively transferred into sensitized CD8−/− mice, in vitro-generated BLT1+/+, but not BLT1−/−, TEFF accumulated in the lung and mediated these altered airway responses to allergen challenge. These data are the first to show both a functional and an essential role for BLT1 in allergen-mediated CD8+ TEFF recruitment into the lung and development of AHR and airway inflammation.

Role of the LTB4/BLT1 Pathway in Allergen-induced Airway Hyperresponsiveness and Inflammation

LTB4, a proinflammatory lipid mediator generated from arachidonic acid through the action of 5-lipoxygenase, has been known for over two decades and is implicated in a wide variety of inflammatory disorders. BLT1, a G-protein-coupled receptor, has recently been identified as a high affinity receptor specific for LTB4. Recent studies in allergen-induced airway hyperresponsiveness and inflammation using mice lacking BLT1 have shown crucial new roles for leukotriene B4 and BLT1 in Th2 cytokine IL-13 production from lung T cells and recruitment of antigen-specific effector CD8+ T cells, suggesting novel mechanisms for their actions. The leukotriene B4-BLT1 pathway is an important target for the treatment of bronchial asthma.

Leukotriene B4 Receptor 1 Expression on Dendritic Cells Is Required for the Development of Th2 Responses and Allergen-Induced Airway Hyperresponsiveness

Dendritic cells (DC) are important APCs that control allergen-induced airway responses by interacting directly with T cells. Leukotriene B4 (LTB4), interacting with its high-affinity receptor, LTB4 receptor 1 (BLT1), is known to attract and activate leukocytes during inflammation. We have previously shown that BLT1 expression on Ag-primed T cells is required for the development of airway hyperresponsiveness (AHR; Miyahara et al. 2005. Am. J. Respir. Crit. Care Med. 172: 161–167). However, the role for the LTB4-BLT1 pathway in DC function in allergen-induced airway responses has not been defined. Bone marrow-derived DCs (BMDC) were generated. Naive BALB/c mice received OVA-pulsed BLT1-deficient (BLT1−/−) BMDCs or wild-type BMDCs intratracheally and were then challenged with OVA for 3 days. Airway responses were monitored 48 h after the last allergen challenge. BLT1−/− BMDCs showed normal maturation judged from surface expression of CD markers. Compared with recipients of wild-type BMDCs, mice that received BLT1−/− BMDCs developed significantly lower AHR to inhaled methacholine, lower goblet cell metaplasia, and eosinophilic infiltration in the airways and decreased levels of Th2 type cytokines in the bronchoalveolar lavage fluid. Migration of BLT1−/− BMDCs into peribronchial lymph nodes was significantly impaired compared with BLT1+/+ BMDCs after intratracheal instillation. These data suggest that BLT1 expression on DCs is required for migration of DCs to regional lymph nodes as well as in the development of AHR and airway inflammation.

Leukotriene B4 Release from Mast Cells in IgE-Mediated Airway Hyperresponsiveness and Inflammation

Previous studies have shown that leukotriene B4 (LTB4), a proinflammatory lipid mediator, is linked to the development of airway hyperresponsiveness through the accumulation of IL-13-producing CD8+ T cells, which express a high affinity receptor for LTB4, BLT1 (Miyahara et al., Am J Respir Crit Care Med 2005;172:161-167; J Immunol 2005;174:4979-4984). By using leukotriene A4 hydrolase-deficient (LTA4H-/-) mice, which fail to synthesize LTB4, we determined the role of this lipid mediator in allergen-induced airway responses. Two approaches were used. In the first, LTA4H-/- mice and wild-type (LTA4H+/+) mice were systemically sensitized and challenged via the airways to ovalbumin. In the second, mice were passively sensitized with anti-ovalbumin IgE and exposed to ovalbumin via the airways. Mast cells were generated from bone marrow of LTA4H+/+ mice or LTA4H-/- mice. After active sensitization and challenge, LTA4H-/- mice showed significantly lower airway hyperresponsiveness compared with LTA4H+/+ mice, and eosinophil numbers and IL-13 levels in the bronchoalveoloar lavage of LTA4H-/- mice were also significantly lower. LTA4H-/- mice also showed decreased airway reactivity after passive sensitization and challenge. After LTA4H+/+ mast cell transfer, LTA4H-/- mice showed increased airway reactivity after passive sensitization and challenge, but not after systemic sensitization and challenge. These data confirm the important role for LTB4 in the development of altered airway responses and suggest that LTB4 secretion from mast cells is critical to eliciting increased airway reactivity after passive sensitization with allergen-specific IgE.