Title: OSR1-sensitive regulation of Na<sup>+</sup>/H<sup>+</sup>exchanger activity in dendritic cells
Abstract: The oxidative stress-responsive kinase 1 (OSR1) is activated by WNK (with no K kinases) and in turn stimulates the thiazide-sensitive Na-Cl cotransporter (NCC) and the furosemide-sensitive Na-K-2Cl cotransporter (NKCC), thus contributing to transport and cell volume regulation. Little is known about extrarenal functions of OSR1. The present study analyzed the impact of decreased OSR1 activity on the function of dendritic cells (DCs), antigen-presenting cells linking innate and adaptive immunity. DCs were cultured from bone marrow of heterozygous WNK-resistant OSR1 knockin mice ( osr KI ) and wild-type mice ( osr WT ). Cell volume was estimated from forward scatter in FACS analysis, ROS production from 2′,7′-dichlorodihydrofluorescein-diacetate fluorescence, cytosolic pH (pH i ) from 2′,7′- bis-(2-carboxyethyl)-5-(and-6)-carboxyfluorescein fluorescence, and Na + /H + exchanger activity from Na + -dependent realkalinization following ammonium pulse and migration utilizing transwell chambers. DCs expressed WNK1, WNK3, NCC, NKCC1, and OSR1. Phosphorylated NKCC1 was reduced in osr KI DCs. Cell volume and pH i were similar in osr KI and osr WT DCs, but Na + /H + exchanger activity and ROS production were higher in osr KI than in osr WT DCs. Before LPS treatment, migration was similar in osr KI and osr WT DCs. LPS (1 μg/ml), however, increased migration of osr WT DCs but not of osr KI DCs. Na + /H + exchanger 1 inhibitor cariporide (10 μM) decreased cell volume, intracellular reactive oxygen species (ROS) formation, Na + /H + exchanger activity, and pH i to a greater extent in osr KI than in osr WT DCs. LPS increased cell volume, Na + /H + exchanger activity, and ROS formation in osr WT DCs but not in osr KI DCs and blunted the difference between osr KI and osr WT DCs. Na + /H + exchanger activity in osr WT DCs was increased by the NKCC1 inhibitor furosemide (100 nM) to values similar to those in osr KI DCs. Oxidative stress (10 μM tert-butyl-hydroperoxide) increased Na + /H + exchanger activity in osr WT DCs but not in osr KI DCs and reversed the difference between genotypes. Cariporide virtually abrogated Na + /H + exchanger activity in both genotypes and blunted LPS-induced cell swelling and ROS formation in osr WT mice. In conclusion, partial OSR1 deficiency influences Na + /H + exchanger activity, ROS formation, and migration of dendritic cells.
Publication Year: 2012
Publication Date: 2012-05-31
Language: en
Type: article
Indexed In: ['crossref', 'pubmed']
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Cited By Count: 12
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