ABSTRACT
Microenvironment-based alterations in phenotypes of mast cells influence the susceptibility to anaphylaxis, yet the mechanisms underlying proper maturation of mast cells toward an anaphylaxis-sensitive phenotype are incompletely understood. Here we report that PLA2G3, a mammalian homolog of anaphylactic bee venom phospholipase A2, regulates this process. PLA2G3 secreted from mast cells is coupled with fibroblastic lipocalin-type PGD2 synthase (L-PGDS) to provide PGD2, which facilitates mast-cell maturation via PGD2 receptor DP1. Mice lacking PLA2G3, L-PGDS or DP1, mast cell-deficient mice reconstituted with PLA2G3-null or DP1-null mast cells, or mast cells cultured with L-PGDS-ablated fibroblasts exhibited impaired maturation and anaphylaxis of mast cells. Thus, we describe a lipid-driven PLA2G3-L-PGDS-DP1 loop that drives mast cell maturation.
Subject(s)
Group III Phospholipases A2/immunology , Mast Cells/immunology , Paracrine Communication/immunology , Prostaglandin D2/immunology , Receptors, Prostaglandin/immunology , Animals , Blotting, Western , Bone Marrow Cells/immunology , Bone Marrow Cells/metabolism , Cell Differentiation/genetics , Cell Differentiation/immunology , Cells, Cultured , Fibroblasts/cytology , Fibroblasts/immunology , Fibroblasts/metabolism , Gene Expression Profiling , Group III Phospholipases A2/genetics , Group III Phospholipases A2/metabolism , Humans , Intramolecular Oxidoreductases/genetics , Intramolecular Oxidoreductases/immunology , Intramolecular Oxidoreductases/metabolism , Lipocalins/genetics , Lipocalins/immunology , Lipocalins/metabolism , Mast Cells/metabolism , Mast Cells/ultrastructure , Mice , Mice, 129 Strain , Mice, Inbred C57BL , Mice, Knockout , Microscopy, Electron, Transmission , Oligonucleotide Array Sequence Analysis , Paracrine Communication/genetics , Prostaglandin D2/metabolism , Receptors, Prostaglandin/genetics , Receptors, Prostaglandin/metabolism , Reverse Transcriptase Polymerase Chain ReactionABSTRACT
Ceramides are the predominant lipids in the stratum corneum (SC) and are crucial components for normal skin barrier function. Although the composition of various ceramide classes in the human SC has been reported, that in mice is still unknown, despite mice being widely used as animal models of skin barrier function. Here, we performed LC/MS/MS analyses using recently available ceramide class standards to measure 25 classes of free ceramides and 5 classes of protein-bound ceramides from human and mouse SC. Phytosphingosine- and 6-hydroxy sphingosine-type ceramides, which both contain an additional hydroxyl group, were abundant in the human SC (35% and 45% of total ceramides, respectively). In contrast, in mice, phytosph-ingosine- and 6-hydroxy sphingosine-type ceramides were present at â¼1% and undetectable levels, respectively, and sphingosine-type ceramides accounted for â¼90%. In humans, ceramides containing α-hydroxy FA were abundant, whereas ceramides containing ß-hydroxy or ω-hydroxy FA were abundant in mice. The hydroxylated ß-carbon in ß-hydroxy ceramides was in the (R) configuration. Genetic knockout of ß-hydroxy acyl-CoA dehydratases in HAP1 cells increased ß-hydroxy ceramide levels, suggesting that ß-hydroxy acyl-CoA, an FA-elongation cycle intermediate in the ER, is a substrate for ß-hydroxy ceramide synthesis. We anticipate that our methods and findings will help to elucidate the role of each ceramide class in skin barrier formation and in the pathogenesis of skin disorders.