Evidence of a role for cyclic ADP-ribose in calcium signaling and migration in human neutrophils

环状ADP-核糖(CADPR)具有强大的钙动员作用，是许多激动剂的第二信使，调节着广泛的钙离子介导的细胞过程。CD38是目前研究最充分的哺乳动物ADP-核糖环化酶，被认为是体内cADPR的重要来源。虽然[Ca^&lt；2&gt；]i的升高是中性粒细胞功能的关键信号，但[Ca^&lt；2&gt；]i的调节机制尚不清楚。本研究探讨了cADPR对人中性粒细胞钙离子动力学的调节及其生理作用。cADPR诱导洋地黄素通透性中性粒细胞释放钙离子，此作用可被cADPR、FK506和雷帕霉素的拮抗剂8BR-cADPR阻断。FMLP和PAF最初引起[Ca^&lt；2&gt；]i迅速升高，随后中性粒细胞持续升高。8BR-cADPR、抗β抗体、FK506、雷帕霉素、β…均能降低化学诱导剂和NAD引起的细胞内[Ca^-NAD]i升高更多的ASE和几种核苷转运体(NT)抑制剂。中性粒细胞表达ENT_1、ENT_2、CNT_2、CNT_3。这些结果表明，CD38合成的胞外cADPR通过NTS转运到细胞内，并通过FK506结合蛋白依赖的过程动员钙离子，这是中性粒细胞持续钙内流所必需的。EGTA或8BR-cADPR预处理人中性粒细胞可特异性地阻断fMLP或PAF刺激的趋化作用。同样，用FK506、抗CD38抗体、NADase和NT抑制剂处理中性粒细胞，可阻断中性粒细胞对fMLP或PAF的趋化。这些结果表明，中性粒细胞对fMLP和PAF的趋化依赖于cADPR介导的钙动员。因此，cADPR通过产生cADPR来控制中性粒细胞对趋化物质的趋化，是炎症和先天免疫反应的重要调节因子。由于cADPR调控的许多趋化受体与临床病理相关的配体结合，cADPR、CD38和NTS是治疗慢性炎症性疾病和神经退行性疾病的潜在药物靶点。较少

Cyclic ADP-ribose (cADPR) has a powerful Ca^<2+>-mobilizing action and behaves as a second messenger of many agonists, thereby modulating a wide number of Ca^<2+>-mediated cell processes. CD38, the best-characterized mammalian ADP-ribosyl cyclase, is postulated to be an important source of cADPR in vivo. Although an increase in [Ca^<2+>]i is a key signal neutrophil functions, the mechanisms for regulation of [Ca^<2+>]i is unclear. The present study examined the regulation by cADPR of Ca^<2+> dynamics and its physiological role in human neutrophils.cADPR induced Ca^<2+> release from digitonin-permeabilized neutrophils and the release was blocked by 8Br-cADPR, an antagonist of cADPR and FK506 and rapamycin. fMLP and PAF induced a initial rapid rise of [Ca^<2+>]i and the following sustained rise in intact neutrophils. β-NAD^+ added into the medium increased [Ca^<2+>]i. Chemoattractans- and β-NAD^+-induced [Ca^<2+>]i rise were reduced by 8Br-cADPR, anti-CD38 antibody, FK506, rapamycin, NAD … More ase and several nucleoside transporter (NT) inhibitors. ENT_1, ENT_2, CNT_2, CNT_3 are expressed in neutrophils. These results suggest that cADPR synthesized extracellulary by CD38 transported into the cells through NTs and mobilize Ca^<2+> by FK506-binding protein-dependent process and is required for sustained Ca^<2+> influx in neutrophils.Pretreatment of human neutrophils with either EGTA or 8Br-cADPR specifically blocked the chemotsxis stimulated with fMLP or PAF. Likewise, treatment of neutrophils with FK506, anti-CD38 antibody, NADase and NT inhibitors blocked the chemotaxis to fMLP or PAF. These results demonstrate that neutrophil chemotaxis to fMLP and PAF are dependent on Ca^<2+> mobilization mediated by cADPR.Thus, cADPR controls neutrophil chemotaxis to chemoattractants through its production of cADPR, and acts as a critical regulator of inflammation and innate immune responses. Since many of the chemoattractant receptors regulated by cADPR bind to ligands that are associated with clinical pathology, cADPR, CD38 and NTs represent novel drug targets with potential application in chronic inflammatory and neurodegenerative disease. Less