This is a continuing, collaborative effort to understand the regulation of mast cell (MC)-associated effector functions and the relevance of these processes to asthma. MCs are unique among hematopoietic effector cells for their constitutive residence in the lung and other vascularized tissues, and their prominent contribution to both afferent and efferent phases of innate and adaptive immune responses. The central hypothesis of this Program Project is that MCs, by virtue of their constitutive and inducible effector pathways, form a functional bridge between innate and acquired immune responses and the clinical expression of asthma. The availability of purified recombinant MC tryptases and the development of mouse strains lacking the enzymes necessary to synthesize serglyin proteoglycans permits study of the role of proteoglycans in regulating the diverse functions of tryptases in vitro and in vivo. A novel in vitro approach will be used to define the mechanism by which GP49B1, a counterregulatory receptor bearing immunotyrosine-based inhibitory motifs (ITIMs), regulates MC activation through mechanistically diverse receptors to limit MC-dependent pathology in both allergic and innate immune esponses. The genes on mouse chromosomes 2 and 6 that interact to confer intrinsic MC-dependent AHR in A/J mice will be identified, and the MC-dependency of this phenotype will be confirmed with adoptive transfer of MCs into A/J mice rendered MC-deficient due to a mutation in the c-kit tyrosine kinase. Mouse strains lacking hematopoietic PGD2 synthase (PGDS) and LTC4synthase (LTC4S), respectively, as well as knockout strains lacking each receptor for PGD2 and LTC4, will be used to define the complementary and counterregulatory functions of these eicosanoids in vitro and in vivo. The role of the bronchoprotective eicosanoid, PGE2, and inducible PGE2 synthases in aspirin intolerant asthma (AIA) will be studied using a novel in vitro approach to the development of human MCs (hMCs) from well-characterized donors with AIA. Abnormalities in relevant synthases and PGE receptors will prompt resequencing for discovery of polymorphic variants. The receptors and mechanisms responsible for the inhibitory effects of PGE2 on hMC activation in vitro will be defined. These studies collectively provide information critical to understanding the biochemical and genetic regulation of key MC-associated effector functions, and defining their role in the control of intrinsic AHR, inflammation, and tissue repair subsequent to both allergic and innate immune responses that likely contribute to the pathophysiology of asthma.
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