Lysosome-dependent Ca2+ release response to Fas activation in coronary arterial myocytes through NAADP: evidence from CD38 gene knockouts

Zhang F, Xia M, Li PL. Lysosome-dependent Ca2+ release response to Fas activation in coronary arterial myocytes through NAADP: evidence from CD38 gene knockouts. Am J Physiol Cell Physiol 298: C1209-C1216, 2010. First published March 3, 2010; doi:10.1152/ajpcell.00533.2009.-Activation of the death receptor Fas has been implicated in the development of vascular injury or disease, but most studies have focused on its role in the regulation of cell apoptosis and growth. The present study was designed to examine the early response of coronary artery to Fas activation by its ligand, FasL. The hypothesis being tested is that CD38 signaling pathway mediates FasL-induced intracellular Ca2+ release through nicotinic acid adenine dinucleotide phosphate (NAADP) in mouse coronary arterial myocytes (CAMs) and thereby produces vasoconstriction in coronary arteries. HPLC analysis demonstrated that FasL markedly increased NAADP production in CAMs from wild-type mice (CD38(+/+)) but not in cells from CD38 knockout (CD38(-/-)) mice. Using fluorescent Ca2+ imaging analysis, we found that FasL (10 ng/ml) significantly increased Ca2+ release from 142.5 +/- 22.5 nM at the basal level to 509.4 +/- 64.3 nM in CD38(+/+) CAMs but not in CD38(-/-) CAMs. However, direct delivery of NAADP, the CD38 metabolite, into CD38(-/-) CAMs still markedly increased Ca2+ release, which could be significantly attenuated by a lysosomal function inhibitor, bafilomycin A1 (Baf), or a NAADP antagonist, pyridoxalphosphate-6-azophenyl-2-disulfonic acid. Confocal microscopy further demonstrated that FasL produced a typical two-phase Ca2+ release with a local Ca2+ burst from lysosomes, followed by a global Ca2+ response in CD38(+/+) CAMs. In isolated perfused septal coronary arteries from CD38(+/+) mice, FasL was found to significantly increase U-46619-induced vasoconstriction from 29.2 +/- 7.3 to 63.2 +/- 10.3%, which was abolished by Baf (100 nM). These results strongly indicate that the early response of CAMs to FasL is to increase intracellular Ca2+ levels and enhance the vascular reactivity through stimulation of NAADP production and lysosome-associated two-phase Ca2+ release in coronary arteries.