title: Inhibition of sulfur mustard-induced cytotoxicity and inflammation by the macrolide antibiotic roxithromycin in human respiratory epithelial cells.
abstract: BACKGROUND
  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.
  
  
  RESULTS
  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.
  
  
  CONCLUSIONS
  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.
authors.affiliation: Section of Molecular Biology, Department of Biology, Division of Experimental Therapeutics, Walter Reed Army Institute of Research, Silver Spring, MD 20910, USA. xiugong.gao@na.amedd.army.mil <xiugong.gao@na.amedd.army.mil>
authors.forename: Xiugong
authors.initials: X
authors.lastname: Gao
authors.forename: Radharaman
authors.initials: R
authors.lastname: Ray
authors.forename: Yan
authors.initials: Y
authors.lastname: Xiao
authors.forename: Peter E
authors.initials: PE
authors.lastname: Barker
authors.forename: Prabhati
authors.initials: P
authors.lastname: Ray
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D002460:Cell Line; D002619:Chemical Warfare Agents / Q000633:toxicity; D016207:Cytokines / Q000235:genetics / Q000276:immunology; D003879:Dermatologic Agents / Q000633:toxicity*; D004305:Dose-Response Relationship, Drug; D004847:Epithelial Cells / Q000166:cytology / Q000187:drug effects* / Q000502:physiology; D005456:Fluorescent Dyes / Q000378:metabolism; D006801:Humans; D007249:Inflammation / Q000139:chemically induced*; D009151:Mustard Gas / Q000633:toxicity*; D052247:Nitric Oxide Synthase Type II / Q000235:genetics / Q000378:metabolism; D020545:Respiratory Mucosa / Q000166:cytology*; D015575:Roxithromycin / Q000494:pharmacology*
chemical_list: D000900:Anti-Bacterial Agents; D002619:Chemical Warfare Agents; D016207:Cytokines; D003879:Dermatologic Agents; D005456:Fluorescent Dyes; D015575:Roxithromycin; D052247:Nitric Oxide Synthase Type II; D009151:Mustard Gas