Effect of Thymoquinone on Cytosolic pH and Na+/H+ Exchanger Activity in Mouse Dendritic Cells

Posted by Admin on Wednesday, August 15, 2012
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Yang W, Bhandaru M, Pasham V, Bobbala D, Zelenak C, Jilani K, Rotte A, Lang F. Effect of thymoquinone on cytosolic pH and Na+/H+ exchanger activity in mouse dendritic cells. 1. Cell Physiol Biochem. 2012;29(1-2):21-30. Epub 2012 Mar 1.
The anti-inflammatory nigella sativa component thymoquinone compromises the function of dendritic cells (DCs), key players in the regulation of innate and adaptive immunity. DC function is regulated by the Na(+)/H(+) exchanger (NHE), which is stimulated by lipopolysaccharides (LPS) and required for LPS-induced cell swelling, reactive oxygen species (ROS) production, TNF-? release and migration. Here we explored, whether thymoquinone influences NHE activity in DCs. To this end, bone marrow derived mouse DCs were treated with LPS in the absence and presence of thymoquinone (10 ?M). Cytosolic pH (pH(i border=0> was determined from 2',7'-bis-(2-carboxyethyl)-5-(and-6)-carboxyfluorescein (BCECF) fluorescence, NHE activity from the Na(+)-dependent realkalinization following an ammonium pulse, cell volume from forward scatter in FACS analysis, ROS production from 2',7'-dichlorodihydrofluorescein diacetate (DCFDA) fluorescence, TNF-? production utilizing ELISA and DC migration with transwell migration assays. As a result, exposure of DCs to LPS (1 ?g/ml) led within 4 hours to transient increase of NHE activity. Thymoquinone did not significantly modify cytosolic pH or cellular NHE activity in the absence of LPS, but abrogated the effect of LPS on NHE activity. Accordingly, in the presence of thymoquinone LPS-treatment resulted in cytosolic acidification. LPS further increased forward scatter and ROS formation, effects similarly abrogated by thymoquinone. Again, in the absence of LPS, thymoquinone did not significantly modify ROS formation and cell volume. LPS further triggered TNF-? release and migration, effects again blunted in the presence of thymoquinone. NHE1 inhibitor cariporide (10 ?M) blunted LPS induced TNF-? release and migration. The effects of thymoquinone on NHE activity and migration were reversed upon treatment of the cells with t-butyl hydroperoxide (TBOOH, 5 ?M). In conclusion, thymoquinone blunts LPS induced NHE activity, cell swelling, oxidative burst, cytokine release and migration of bone marrow derived murine dendritic cells. NHE inhibition may thus contribute to the antiinflammatory action of thymoquinone.

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