Abstract
Glucocorticoids are first-line agents for the treatment of many eosinophil-associated disorders; however, their effects on human eosinophils remain poorly understood. To gain an unbiased, genome-wide view of the early transcriptional effects of glucocorticoids on human eosinophils in vivo, RNA sequencing was performed on purified blood eosinophils obtained before and 30, 60, and 120 minutes after administration of a single dose of oral prednisone (1 mg/kg) to three unrelated healthy subjects with hypereosinophilia of unknown significance. The resulting dataset is of high quality and suitable for differential expression analysis. Flow cytometry and qPCR were then performed on three additional cohorts of human subjects, to validate the key findings at the transcript and protein levels. The resulting datasets provide a resource for understanding the response of circulating human eosinophils to glucocorticoid administration.
Highlights
Background & SummaryGlucocorticoids effectively suppress eosinophilia and its clinical manifestations, and they are first-line agents in a variety of eosinophil-associated disorders, including hypereosinophilic syndromes[1] and eosinophilic granulomatosis with polyangiitis[2]
An initial analysis of the RNA-seq data revealed the induction of a pro-apoptotic transcriptional program and differential expression of key genes related to leukocyte migration[9]
By qPCR, we evaluated the transcriptional response of three key genes implicated in eosinophil migration (CXCR4, CCR1, CCR3) and seven genes involved in eosinophil apoptosis (BCL2L11, XIAP, CASP9, PAK1, TNFAIP3, NOTCH1, ZBTB16)
Summary
Glucocorticoids effectively suppress eosinophilia and its clinical manifestations, and they are first-line agents in a variety of eosinophil-associated disorders, including hypereosinophilic syndromes[1] and eosinophilic granulomatosis with polyangiitis[2]. To evaluate whether the induction of a pro-apoptotic transcriptional program led to eosinophil apoptosis in vivo prior to the egress of eosinophils from the peripheral circulation, which occurs between 60 and 120 minutes (min) after glucocorticoid administration[9], we studied a third cohort, comprised of three subjects: two unrelated donors with normal eosinophil counts received a single dose of IV methylprednisolone 250 mg and one patient with HES received a single dose of oral prednisone 1 mg/kg. To assess whether the observation of transcript-level changes on key eosinophil migration genes led to changes in protein abundance at the cell surface within the time frame of glucocorticoid-induced eosinopenia, we studied a fourth cohort, comprised of six unrelated donors with normal eosinophil counts. Human eosinophils and in studies of the mechanisms behind glucocorticoid-induced eosinopenia and glucocorticoid resistance
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