title: Non-genomic inhibitory effect of glucocorticoids on activated peripheral blood basophils through suppression of lipid raft formation.
abstract: We investigated the non-genomic effects of glucocorticoids (GCs) on inhibition of plasma membrane lipid raft formation in activated human basophils. Human basophils obtained from house dust mite (HDM)-sensitive volunteers were pretreated with hydrocortisone (CORT) or dexamethasone (Dex) for 30 min and then primed with phorbol 12-myristate 13-acetate (PMA, 10 ng/ml) or HDM (10 µg/ml). The expression of CD63, a basophil activation marker, was assessed by flow cytometry. Membrane-bound GC receptors (mGCRs) were analysed by flow cytometry and confocal laser microscopy. Lipid rafts were assessed using a GM1 ganglioside probe and visualization by confocal laser microscopy. Pretreatment of basophils with CORT (10(-4) M and 10(-5) M) and Dex (10(-7) M) significantly inhibited CD63 expression 20 min after addition of PMA or HDM. The inhibitory effects of GCs were not altered by the nuclear GC receptor (GCR) antagonist RU486 (10(-5) M) or the protein synthesis inhibitor cycloheximide (10(-4) M) (P < 0·05). CORT coupled to bovine serum albumin (BSA-CORT) mimicked the rapid inhibitory effects of CORT, suggesting the involvement of mGCRs. mGCRs were detectable on the plasma membrane of resting basophils and formed nanoclusters following treatment with PMA or HDM. Pretreatment of cells with BSA-CORT inhibited the expression of mGCRs and nanoclustering of ganglioside GM1 in lipid rafts. The study provides evidence that non-genomic mechanisms are involved in the rapid inhibitory effect of GCs on the formation of lipid raft nanoclusters, through binding to mGCRs on the plasma membrane of activated basophils.
authors.affiliation: Department of Respiratory Medicine and Allergology, Kinki University Faculty of Medicine, Osaka-Sayama, Japan.
authors.forename: S
authors.initials: S
authors.lastname: Yamagata
authors.forename: K
authors.initials: K
authors.lastname: Tomita
authors.forename: H
authors.initials: H
authors.lastname: Sano
authors.forename: Y
authors.initials: Y
authors.lastname: Itoh
authors.forename: Y
authors.initials: Y
authors.lastname: Fukai
authors.forename: N
authors.initials: N
authors.lastname: Okimoto
authors.forename: N
authors.initials: N
authors.lastname: Watatani
authors.forename: S
authors.initials: S
authors.lastname: Inbe
authors.forename: H
authors.initials: H
authors.lastname: Miyajima
authors.forename: K
authors.initials: K
authors.lastname: Tsukamoto
authors.forename: H
authors.initials: H
authors.lastname: Santoh
authors.forename: H
authors.initials: H
authors.lastname: Ichihashi
authors.forename: A
authors.initials: A
authors.lastname: Sano
authors.forename: R
authors.initials: R
authors.lastname: Sato
authors.forename: Y
authors.initials: Y
authors.lastname: Tohda
mesh_terms: D000818:Animals; D001491:Basophils / Q000187:drug effects* / Q000276:immunology / Q000378:metabolism; D002417:Cattle; D002462:Cell Membrane / Q000276:immunology / Q000378:metabolism; D002478:Cells, Cultured; D003513:Cycloheximide / Q000494:pharmacology; D003907:Dexamethasone / Q000276:immunology / Q000494:pharmacology; D005434:Flow Cytometry; D005677:G(M1) Ganglioside / Q000378:metabolism; D005786:Gene Expression Regulation; D005938:Glucocorticoids / Q000276:immunology / Q000494:pharmacology*; D006801:Humans; D006854:Hydrocortisone / Q000276:immunology / Q000494:pharmacology; D007963:Leukocytes, Mononuclear / Q000166:cytology; D021962:Membrane Microdomains / Q000187:drug effects* / Q000276:immunology / Q000378:metabolism; D018613:Microscopy, Confocal; D015735:Mifepristone / Q000494:pharmacology; D039981:Pyroglyphidae / Q000276:immunology / Q000378:metabolism*; D011965:Receptors, Glucocorticoid / Q000032:analysis / Q000037:antagonists & inhibitors / Q000378:metabolism*; D012710:Serum Albumin, Bovine / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000276:immunology / Q000494:pharmacology; D060149:Tetraspanin 30 / Q000032:analysis / Q000037:antagonists & inhibitors
chemical_list: D005938:Glucocorticoids; D011965:Receptors, Glucocorticoid; D060149:Tetraspanin 30; D012710:Serum Albumin, Bovine; D015735:Mifepristone; D005677:G(M1) Ganglioside; D003907:Dexamethasone; D003513:Cycloheximide; D013755:Tetradecanoylphorbol Acetate; D006854:Hydrocortisone