Modulation of nonselective cation current by oxidized LDL and lysophosphatidylcholine and its inhibitory contribution to endothelial damage.
OBJECTIVE
This study examined the effects of oxidized low-density lipoprotein (LDL) and its major lipid constituent lysophosphatidylcholine (LPC) on nonselective cation (NSC) current and its inhibitory contribution to LPC-induced cytotoxicity in cultured human umbilical endothelial cells (HUVECs).


METHODS
Patch-clamp technique and the resazurin-based cell viability assay were used.


RESULTS
In voltage-clamped cells, oxidized LDL or LPC slowly activated NSC current. NSC current was also activated by loading cells with Ca(2+) solution buffered at various concentrations using a patch pipette or by applying the sarcoplasmic reticulum Ca(2+) pump blocker 2,5-di-t-butyl-1,4-benzohydroquinone (BHQ), the metabolic inhibitor CN(-) or the hydroperoxide donor tert-butyl hydroperoxide (TBHP). On the contrary, when intracellular Ca(2+) was strongly buffered with 12mM BAPTA or cells were loaded with superoxide dismutase using a patch pipette, LPC or BHQ did not activate NSC current. Furthermore, NSC current activated by LPC, TBHP or CN(-) was inhibited by the antioxidant tempol or extracellular Ca(2+) depletion and NSC current activated by intracellular Ca(2+) was further augmented by oxidized LDL or LPC. LPC or oxidized LDL released Ca(2+) from intracellular stores and further enhanced store-operated Ca(2+) entry. LPC-induced cytotoxicity was augmented by inhibiting Ca(2+) influx and NO synthesis.


CONCLUSIONS
Oxidized LDL or its main component LPC activated Ca(2+)-permeable NSC current via releasing Ca(2+) from intracellular stores and producing ROS and thereby increased Ca(2+) influx. Ca(2+) influx through NSC channel might protect endothelial cells by producing NO.
Department of Physiology and Medical Research Institute, School of Medicine, Ewha Womans University, Seoul, Republic of Korea.
Guo Hua
GH
Liang
Seonghee
S
Park
Moon Young
MY
Kim
Ji Aee
JA
Kim
Shinkyu
S
Choi
Suk Hyo
SH
Suh
D002118:Calcium / Q000378:metabolism*; D002412:Cations; D002470:Cell Survival; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000378:metabolism; D006801:Humans; D008077:Lipoproteins, LDL / Q000378:metabolism*; D008244:Lysophosphatidylcholines / Q000378:metabolism*; D009569:Nitric Oxide / Q000096:biosynthesis*; D010078:Oxazines / Q000737:chemistry; D010084:Oxidation-Reduction; D018408:Patch-Clamp Techniques; D017382:Reactive Oxygen Species / Q000378:metabolism; D012519:Sarcoplasmic Reticulum / Q000378:metabolism; D014471:Umbilical Veins / Q000378:metabolism; D014966:Xanthenes / Q000737:chemistry
D002412:Cations; D008077:Lipoproteins, LDL; D008244:Lysophosphatidylcholines; D010078:Oxazines; D017382:Reactive Oxygen Species; D014966:Xanthenes; C099252:oxidized low density lipoprotein; C005843:resazurin; D009569:Nitric Oxide; D002118:Calcium