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|Title:||Type I cell ROS kinetics under hypoxia in the intact mouse carotid body ex vivo: a FRET-based study|
|Authors:||A. Bernardini;U. Brockmeier;E. Metzen|
|subject:||ROS; carotid body; FRET-HSP33; tissue oxygen; membrane poten- tial; hypoxia; NADPH oxidase|
|Abstract:||Bernardini A, Brockmeier U, Metzen E, Berchner- Pfannschmidt U, Harde E, Acker-Palmer A, Papkovsky D, Acker H, Fandrey J. Type I cell ROS kinetics under hypoxia in the intact mouse carotid body ex vivo: a FRET-based study. Am J Physiol Cell Physiol 308: C61–C67, 2015. First published October 15, 2014; doi:10.1152/ajpcell.00370.2013.—Reactive oxygen species (ROS) mainly originating from NADPH oxidases have been shown to be involved in the carotid body (CB) oxygen-sensing cascade. For measuring ROS kinetics, type I cells of the mouse CB in an ex vivo preparation were transfected with the ROS sensor construct FRET- HSP33. After 2 days of tissue culture, type I cells expressed FRET- HSP33 as shown by immunohistochemistry. In one population of CBs, 5 min of hypoxia induced a significant and reversible decrease of type I cell ROS levels ( n 9 CBs; P 0.015), which could be inhibited by 4-(2-aminoethyl)benzensulfonylfluorid (AEBSF), a highly specific inhibitor of the NADPH oxidase subunits p47 phox and p67 phox . In another population of CBs, however, 5 min of hypoxia induced a significant and reversible increase of ROS levels in type I cells ( n 8 CBs; P 0.05), which was slightly enhanced by administration of 3 mM AEBSF. These different ROS kinetics seemed to coincide with different mice breeding conditions. Type I cells of both populations showed a typical hypoxia-induced membrane poten- tial (MP) depolarization, which could be inhibited by 3 mM AEBSF. ROS and MP closely followed the hypoxic decrease in CB tissue oxygen as measured with an O 2 -sensitive dye. We conclude that attenuated p47 phox subunit activity of the NADPH oxidase under hypoxia is the physiological trigger for type I cell MP depolarization probably due to ROS decrease, whereas the observed ROS increase has no influence on type I cell MP kinetics under hypoxia.|
|Appears in Collections:||American Journal of Physiology(Cell physiology) 2015|
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