Expression pattern and signalling pathways in neutrophil like HL-60 cells after treatment with estrogen receptor selective ligands

Division for Reproductive Endocrinology and The Pediatric Endocrinology Unit, Department of Women's and Children's Health, Karolinska Institutet, Stockholm, Sweden.
Molecular and Cellular Endocrinology (Impact Factor: 4.41). 04/2012; 361(1-2):179-90. DOI: 10.1016/j.mce.2012.04.006
Source: PubMed


Estrogens play a role in the regulation of genes associated with inflammation and immunity in neutrophils. Estrogen signalling is mediated by estrogen receptor (ER)α, ERβ, and G-protein-coupled estrogen receptor-1 (GPER). The mechanisms by which estrogen regulate genes in neutrophils are poorly understood. Our aim was to identify the presence of ERs and to characterize estrogen responsive genes in terminally differentiated neutrophil like HL-60 (nHL-60) cells using estradiol and selective ER agonists. ERs were identified by Western blotting and immunocytochemistry. Microarray technique was used to screen for differentially expressed genes and the selected genes were verified by quantitative PCR. We show the presence of functional ERα, ERβ and GPER. Microarray analysis showed the presence of genes that are uniquely regulated by a single ligand and also genes that are regulated by multiple ligands. We conclude that ERs are functionally active in nHL-60 cells regulating genes involved in key physiological functions.

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Available from: Chellakkan Selvanesan Blesson, Apr 25, 2014
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    ABSTRACT: Neutrophils are major participants in innate host responses. It is well known that estrogens have an immune-modulatory role, and some evidence exists that neutrophil physiology can be altered by these molecules. Traditionally, estrogens act via classical nuclear estrogen receptors, but the identification of a G protein-coupled estrogen receptor (GPER), a membrane estrogen receptor that binds estradiol and other estrogens, has opened up the possibility of exploring additional estrogen-mediated effects. However, information on the importance of GPER for immunity, especially, in neutrophils is scant. In this study, we report that gilthead seabream (Sparus aurata L.) acidophilic granulocytes, which are the functional equivalent of mammalian neutrophils, express GPER at both mRNA and protein levels. By using a GPER selective agonist, G1, it was found that GPER activation in vitro slightly reduced the respiratory burst of acidophilic granulocytes and drastically altered the expression profile of several genes encoding major pro- and anti-inflammatory mediators. In addition, GPER signaling in vivo modulated adaptive immunity. Finally, a cAMP analog mimicked the effects of G1 in the induction of the gene coding for PG-endoperoxide synthase 2 and in the induction of CREB phosphorylation, whereas pharmacological inhibition of protein kinase A superinduced PG-endoperoxide synthase 2. Taken together, our results demonstrate for the first time, to our knowledge, that estrogens are able to modulate vertebrate granulocyte functions through a GPER/cAMP/protein kinase A/CREB signaling pathway and could establish therapeutic targets for several immune disorders in which estrogens play a prominent role.
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    ABSTRACT: Studying the effect of estrogenic chemicals, especially estrogenic endocrine disruptors, needs mechanism-based understanding of toxicity pathways, especially at the level of cell signaling. We first summarized how estrogen action can be monitored through gene expression profiles by means of estrogen-responsive genes, which are associated with or mediate various types of cell signaling through pathways, such as mitogen-activated protein kinase (MAPK), angiogenesis, nuclear receptor, ErbB/HER and ubiquitin/proteasome signaling pathways, and the regulation of cell functions, such as chromatin/epigenesis, apoptosis, autophagy, cellular metabolism, translational control, cell cycle/DNA damage/cytoskeletal formation, immunology/inflammation response, neurological diseases and development/differentiation. The cell signals induced by estrogenic chemicals can be monitored by appropriate sets of estrogen-responsive genes, where the above-mentioned signaling pathways are involved. The association of estrogenic endocrine disruptors and environmental estrogens, such as flavonoids, zearalenone, bisphenol A, perfluorooctane sulfonate and di(2-ethylhexyl) phthalate, with cell signaling is discussed along with a comprehensive list of signaling pathways induced by these chemicals. The signaling pathways identified could be used as candidate toxicity pathways to monitor and evaluate endocrine disruptor action.