Macrolide Antibiotics Inhibit Mucus Secretion and Calcium Entry in Swine Airway Submucosal Mucous Gland Cells
Macrolide Antibiotics Inhibit Mucus Secretion and Calcium Entry in Swine Airway Submucosal Mucous Gland Cells
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DOI：
10.1124/jpet.110.171561
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发表时间：
2011-01-01
影响因子：
3.5
通讯作者：
Farley, J. M., Sr.
中科院分区：
文献类型：
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作者：
Lu, S.;Liu, H.;Farley, J. M., Sr.
大环内酯类抗生素如红霉素(EM)和阿奇霉素(Azm)在治疗炎症性肺部疾病的粘液高分泌方面是有益的。已经提出了几种间接和直接的作用机制。本研究探讨了大环内酯类化合物对分离的粘膜下粘液腺细胞(SMGCS)分泌功能的直接影响。我们假设大环内酯类抑制引起粘液分泌所必需的钙内流。为了验证这一点，我们用酶联免疫吸附试验、钙激活的K+(K-Ca)和钙激活的氯-(Cl-Ca)电流来量化粘蛋白蛋白的释放。用全细胞膜片钳技术测量非选择性阳离子电流(NSCC)；用Fura-2钙成像技术测量细胞内钙离子浓度([Ca~(2+)](I))。我们发现EM和Azm都是M受体的激动剂。EM(10 MU M)对乙酰胆碱(ACh)诱导的粘蛋白释放有明显的抑制作用，但对粘蛋白的释放有微弱的促进作用。EM和Azm(10mU M)均可引起钾-钙和氯-钙全细胞电流，阿托品可阻断这两种电流。EM和AZM在不改变峰值电流幅度的情况下，也加速了三磷酸肌醇诱发的钾-钙和氯-钙电流的衰减。同样，内施AZM(10mM)可增加ACh诱发的K-Ca和Cl-Ca电流的衰减率。EM(1-10 mU M)和Azm(0.1-10 mU M)在钙内流平台期应用时，缓慢地(超过25-30分钟)抑制thapsigargin(TG)诱导的Ca~(2+)内流，但在启动Ca~(2+)内流前5分钟处理后，可使TG诱导的Ca~(2+)内流减弱70%。EM可阻断TG诱导的NSCC。我们认为，大环内酯类抗生素是M受体的部分激动剂，但通过抑制SMGCS的钙内流来抑制刺激的粘液释放。
Macrolide antibiotics such as erythromycin (EM) and azithromycin (AZM) are beneficial in the treatment of mucus hypersecretion in inflammatory pulmonary diseases. Several indirect and direct mechanisms of action have been proposed. This study investigates the direct effect of macrolides on secretory function of isolated submucosal mucous gland cells (SMGCs). We hypothesize that macrolides inhibit the calcium influx necessary for evoked mucus secretion. To test this, we quantified mucin protein release using enzyme-linked immunosorbent assay, calcium-activated K+ (K-Ca), and calcium-activated Cl- (Cl-Ca) currents. We measured nonselective cation current (NSCC) using whole-cell patch-clamp techniques; intracellular calcium concentration ([Ca2+](i)) was measured using fura-2 Ca2+ imaging. We found that both EM and AZM are agonists at muscarinic receptors. EM (10 mu M) evoked a small but significant increase in mucin release but inhibited the mucin release induced by subsequent acetylcholine (ACh) treatment. Both EM and AZM (10 mu M) evoked K-Ca and Cl-Ca whole-cell currents, which were blocked by atropine. EM and AZM also accelerated the decay of inositol trisphosphate-induced K-Ca and Cl-Ca currents without changing the peak current amplitudes. Likewise, internal application of AZM (10 mu M) enhanced the decay rate of ACh-induced K-Ca and Cl-Ca currents. EM (1-10 mu M) and AZM (0.1-10 mu M) slowly (over 25-30 min) inhibited thapsigargin (TG)-induced Ca2+ entry when applied during the plateau phase of Ca2+ entry but blunted TG-induced Ca2+ entry by 70% after a 5-min pretreatment before initiating calcium entry. EM blocked TG-induced NSCC. We conclude that macrolide antibiotics are partial agonists at muscarinic receptors but inhibit stimulated mucus release by inhibiting calcium entry in SMGCs.