Effect of Calcium Channel Blockers on Endothelin-1 Production by HDL in Cultured Human Proximal Tubular Cells

Ahn, Seon Ho; Song, Ju Hung

Abstract

BACKGROUND: Human high density lipoprotein (HDL) is known to stimulate endothelin-1 (ET-1) production through the phospholipase C (PLC)/Ca2+/ protein kinase C (PKC) pathway. Calcium channel blockers may be involved in the decrease of HDL- induced ET-1 production. This study was designed to evaluate whether HDL-induced ET-1 production was affected by Ca2+ channel blockers in cultured human proximal tubular cells (PTC).
METHODS
The human PTC were obtained from human nephrectomized tissues, and cultured in six different media, which were bovine serum free (SF) DMEM/F12 medium alone, and five other SF DMEM/F12 media with 200 microgram/ml of HDL, with 200 microgram/ml of HDL and each 10 micrometer of diltiazem, nifedipine, and verapamil solved in 100% ethanol 0.1 volume%, and with 200 microgram/ml of HDL and 0.1 volume% of 100% ethanol as a control. After 24 hours of exposure, ET-1 in the supernatant was measured by radioimmunoassay, and ET-1 level in each well were marked as pg ET-1/mg cell protein/ 24 hr in consideration of cell count.
RESULTS
In SF medium, ET-1 production was 1.803+/-0.295pg/mg cell protein/24 hr. In SF medium with 200 microgram/ml of HDL, ET-1 production significantly increased from 1.803+/-0.295 to 10.860+/-0.476 pg/mg cell protein/24 hr (P<0.05). In SF medium with 200 microgram/ml of HDL and 100% ethanol 0.1 volume%, ET-1 production significantly decreased from 10.860+/-0.476 to 6.700+/-1.273pg/mg cell protein/ 24 hr (P<0.05). In SF media with 200 microgram/ml of HDL and each 10 micrometer of diltiazem, nifedipine, and verapamil solved in 100% ethanol 0.1 volume%, ET- 1 production was decreased from 6.700+/-1.273 to 4.043+/-1.550 by diltiazem (P<0.05), to 3.260+/-0.752pg/ mg cell protein/24 hr by verapamil (P<0.05), and to 4.414+/-1.567pg/mg cell protein/24 hr by nifedipine (P=0.067).
CONCLUSION
These results suggest that the HDL- induced ET-1 production in cultured human PTC was significantly decreased by diltiazem and verapamil, and it seemed to be decreased by nifedipine.