PHENOTYPIC REGULATION EICOSANOID PATHWAYS OF HUMAN MAST CELLS IN VITRO

皮肤肥大细胞的存在及其生物活性的影响

The presence of skin mast cells and the effects of their bioactive mediators has been reported in patients with a variety of dermatologic disorders, of which urticaria is a prototypic example. Specifically, mediators of mast cells include two families of lipid molecules known as eicosanoids, the cysteinyl leukotrienes and prostaglandins. The classic pruritic wheal and flare reaction is consistent with the effects of mast cell release of these prostaglandins and cysteinyl leukotrienes. Whereas prostaglandins cause pain and vasoconstriction, the leukotrienes have been well described in regard to their ability to cause pruritis and the vasodilation leading to the wheal and flare reaction of urticaria, regardless of its initiating event. Two phenotypes of mast cells exist: the connective tissue type and the mucosal type. Each of these phenotypes is responsible for a specific profile eicosanoids, either prostaglandins or leukotrienes, which may act as a phenotypic marker for the mast cell subtypes. The connective tissue type primarily produces prostaglandins via the prostaglandin GH synthase pathway, whereas the mucosal phenotypes of mast cells possess enzymes of the 5-lipoxygenase/leukotriene C4 synthase pathway leading to the formation of leukotrienes. Interestingly, although the connective tissue type mast cell has been identified in the skin, the effects of the cysteinyl leukotrienes, which are the major eicosanoids in the mucosal-type mast cell appear to predominate. This data regarding mast cell phenotype in the skin mast cell is conflicting and to address the problem, our laboratory has developed an in vitro culture system for the growth and development of an in vitro derived human mast cell progenitors. Use of the model system will allow insight into the development of the pathway of biosynthetic enzymes, and thus provide important knowledge about the phenotypic development of mast cells. We have proposed to investigate the balance between the biosynthetic enzymes of cysteinyl leukotrienes and prostaglandins as markers of phenotypic development in human mast cell progenitors. Specifically, activated human mast cell progenitors will be examined for eicosanoid generation to compare relative production of PGHS products to those of the 5 LO/LTC4-synthase pathway. Additionally, the effects of cytokines will be examined to determine their effects on activity, transcript and protein levels of the enzymes of these two eicosanoid pathways. Finally, the genomic regulation of the terminal enzyme in cysteinyl leukotriene production, leukotriene C4 synthase will be studied using DNAse hypersensitivity assays, followed by promoter-reporter constructs.