Gao Xiugong, Ray Radharaman, Xiao Yan, Barker Peter E, Ray Prabhati
Section of Molecular Biology, Department of Biology, Division of Experimental Therapeutics, Walter Reed Army Institute of Research, Silver Spring, MD 20910, USA.
BMC Cell Biol. 2007 May 24;8:17. doi: 10.1186/1471-2121-8-17.
Sulfur mustard (SM) is a potent chemical vesicant warfare agent that remains a significant military and civilian threat. Inhalation of SM gas causes airway inflammation and injury. In recent years, there has been increasing evidence of the effectiveness of macrolide antibiotics in treating chronic airway inflammatory diseases. In this study, the anti-cytotoxic and anti-inflammatory effects of a representative macrolide antibiotic, roxithromycin, were tested in vitro using SM-exposed normal human small airway epithelial (SAE) cells and bronchial/tracheal epithelial (BTE) cells. Cell viability, expression of proinflammatory cytokines including interleukin (IL)-1beta, IL-6, IL-8 and tumor necrosis factor (TNF), and expression of inducible nitric oxide synthase (iNOS) were examined, since these proinflammatory cytokines/mediators are import indicators of tissue inflammatory responses. We suggest that the influence of roxithromycin on SM-induced inflammatory reaction could play an important therapeutic role in the cytotoxicity exerted by this toxicant.
MTS assay and Calcein AM/ethidium homodimer (EthD-1) fluorescence staining showed that roxithromycin decreased SM cytotoxicity in both SAE and BTE cells. Also, roxithromycin inhibited the SM-stimulated overproduction of the proinflammatory cytokines IL-1beta, IL-6, IL-8 and TNF at both the protein level and the mRNA level, as measured by either enzyme-linked immunosorbent assay (ELISA) or real-time RT-PCR. In addition, roxithromycin inhibited the SM-induced overexpression of iNOS, as revealed by immunocytochemical analysis using quantum dots as the fluorophore.
The present study demonstrates that roxithromycin has inhibitory effects on the cytotoxicity and inflammation provoked by SM in human respiratory epithelial cells. The decreased cytotoxicity in roxithromycin-treated cells likely depends on the ability of the macrolide to down-regulate the production of proinflammatory cytokines and/or mediators. The results obtained in this study suggest that macrolide antibiotics may serve as potential vesicant respiratory therapeutics through mechanisms independent of their antibacterial activity.
硫芥（SM）是一种强效化学糜烂性战剂，仍然是重大的军事和民用威胁。吸入SM气体可导致气道炎症和损伤。近年来，越来越多的证据表明大环内酯类抗生素在治疗慢性气道炎症性疾病方面具有有效性。在本研究中，使用暴露于SM的正常人小气道上皮（SAE）细胞和支气管/气管上皮（BTE）细胞，在体外测试了一种代表性大环内酯类抗生素罗红霉素的抗细胞毒性和抗炎作用。检测了细胞活力、包括白细胞介素（IL）-1β、IL-6、IL-8和肿瘤坏死因子（TNF）在内的促炎细胞因子的表达以及诱导型一氧化氮合酶（iNOS）的表达，因为这些促炎细胞因子/介质是组织炎症反应的重要指标。我们认为罗红霉素对SM诱导的炎症反应的影响可能在这种毒物所施加的细胞毒性中发挥重要的治疗作用。
MTS法和钙黄绿素AM/乙锭同二聚体（EthD-1）荧光染色显示，罗红霉素降低了SAE和BTE细胞中的SM细胞毒性。此外，通过酶联免疫吸附测定（ELISA）或实时RT-PCR检测，罗红霉素在蛋白质水平和mRNA水平均抑制了SM刺激的促炎细胞因子IL-1β、IL-6、IL-8和TNF的过量产生。此外，使用量子点作为荧光团的免疫细胞化学分析显示，罗红霉素抑制了SM诱导的iNOS过表达。
本研究表明罗红霉素对SM在人呼吸道上皮细胞中引发的细胞毒性和炎症具有抑制作用。罗红霉素处理的细胞中细胞毒性降低可能取决于大环内酯类药物下调促炎细胞因子和/或介质产生的能力。本研究获得的结果表明，大环内酯类抗生素可能通过与其抗菌活性无关的机制作为潜在的糜烂性呼吸道治疗药物。
