Close functional coupling between Ca2+ release-activated Ca2+ channels and reactive oxygen species production in murine macrophages.
OBJECTIVE
To investigate the role of Ca(2+) release-activated Ca(2+) (CRAC) channels in the ROS production in macrophages.


METHODS
The intracellular [Ca(2+)](i) was analyzed by confocal laser microscopy. The production of ROS was assayed by flow cytometry.


RESULTS
Both LPS and thapsigargin induced an increase in intracellular [Ca(2+)](i), either in the presence or absence of extracellular Ca(2+) in murine macrophages. The Ca(2+) signal was sustained in the presence of external Ca(2+) and only initiated a mild and transient rise in the absence of external Ca(2+). CRAC channel inhibitor 2-APB completely suppressed the Ca(2+) entry signal evoked by thapsigargin, and suppressed approximately 93% of the Ca(2+) entry signal evoked by LPS. The increase in intracellular [Ca(2+)](i) was associated with increased ROS production, which was completely abolished in the absence of extracellular Ca(2+) or in the presence of CRAC channel inhibitors 2-APB and Gd(3+). The mitochondrial uncoupler carbonyl cyanide p-trifluoromethoxy-phenylhydrazone and the inhibitor of the electron transport chain, antimycin, evoked a marked increase in ROS production and completely inhibited thapsigargin and LPS-evoked responses. Conclusions. These findings indicate that the LPS-induced intracellular [Ca(2+)](i) increase depends on the Ca(2+) entry through CRAC channels, and close functional coupling between CRAC and ROS production in murine macrophages.
Jin
Sheng-Wei
SW
Department of Anesthesiology, Union Hospital, Tongil Medical College, Huazhong University of Science and Technology, Wuhan, China.
Zhang
Li
L
Lian
Qin-Quan
QQ
Yao
Shang-Long
SL
Wu
Ping
P
Zhou
Xiao-Yan
XY
Xiong
Wei
W
Ye
Du-Yun
DY
D000818:Animals; D000968:Antimycin A / Q000031:analogs & derivatives / Q000494:pharmacology; D001896:Boron Compounds / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D015220:Calcium Channels / Q000187:drug effects / Q000378:metabolism*; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone / Q000494:pharmacology; D002460:Cell Line; D008070:Lipopolysaccharides / Q000494:pharmacology; D008264:Macrophages / Q000187:drug effects / Q000378:metabolism*; D051379:Mice; D017382:Reactive Oxygen Species / Q000378:metabolism*; D019284:Thapsigargin / Q000494:pharmacology; D014475:Uncoupling Agents / Q000494:pharmacology
D001896:Boron Compounds; D002121:Calcium Channel Blockers; D015220:Calcium Channels; D008070:Lipopolysaccharides; D017382:Reactive Oxygen Species; D014475:Uncoupling Agents; C032456:antimycin; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone; D000968:Antimycin A; D019284:Thapsigargin; C109986:2-aminoethoxydiphenyl borate; D002118:Calcium