Scientific Report 2003-2004 - Cleveland Clinic Lerner Research ...

THE ERZURUMLABORATORYRESEARCH ASSOCIATESSuzy Comhair, Ph.D.Weiling Xu, M.D.CLINICAL FELLOWSTobias Piekert, M.D.Roberto Machado, M.D.VISITING SCIENTISTArnaud Chambellan, M.D.INSERM, Nantes, FrancePOSTDOCTORAL FELLOWSSudakshina Ghosh, Ph.D.Shuo Zheng, Ph.D.SENIOR TECHNOLOGISTTannishia Goggans, B.S.RESEARCH COORDINATORJacqueline Pyle, R.N.TECHNOLOGISTAllison Janocha, B.S.TECHNICAL ASSOCIATEDaniel LaskowskiGRADUATE STUDENTFares Masri54Inducible Nitric Oxide Synthase (NOS2)Expression Key to Airway InflammationOur laboratory studies the molecularmechanisms that initiate and maintainairway inflammation. In this context, wehave focused on the role of antioxidants andreactive oxygen (ROS) and nitrogen species(RNS) in the pathogenesis of lung diseases.Recently, the lungs have been shown toproduce significant amounts of RNS, i.e., nitricoxide (NO). NO is present in exhaled air ofnormal individuals and increased in that of asthmaticindividuals. Although excessive NO may bean injurious oxidantspecies, NO is endogenouslyproduced inhealthy human lungs,suggesting it has aphysiologic role.Three isoforms ofnitric oxide synthase(NOS), the enzymesresponsible for endogenousNO production, are foundin cells: inducible NOS(NOS2) and two isoformsof constitutive NOS(NOS1 and 3). Expressionof NOS2 has traditionallybeen found in cellsstimulated by inflammatorycytokines such as interleukin-1β (IL-1β), tumornecrosis factor-α (TNF-α), and interferon-γ(IFN-γ). However, having identified and clonedthe NOS isoform from freshly obtained healthyhuman airway epithelium, we found that thiscontinuously expressed airway NOS is NOS2.Ours was the first conclusive demonstration ofcontinuous expression of the NOS2 gene innormal, non-inflamed tissues and suggests uniqueKaneko, F.T., Arroliga, A.C., Dweik, R.A., Comhair, S.A., Laskowski, D., Oppedisano, R.,Thomassen, M.J., and S.C. Erzurum (1998) Biochemical reaction products of nitric oxide asquantitative markers of primary pulmonary hypertension. Am. J. Respir. Crit. Care Med.158:917-923.Comhair, S.A., Bhathena, P.R., Dweik, R.A., Kavuru, M., and S.C. Erzurum (2000) Rapid lossof superoxide dismutase activity during antigen-induced asthmatic response. Lancet 355:624.Beall, C.M., Laskowski, D., Strohl, K.P., Soria, R., Villena, M., Vargas, E., Alarcon, A.M.,Gonzales, C., and S.C. Erzurum (2001) Pulmonary nitric oxide in mountain dwellers. Nature414:411-412.Chung-man Ho, J. [Ho, J.C.] Zheng, S., Comhair, S.A., Farver, C., and S.C. Erzurum (2001)Differential expression of manganese superoxide dismutase and catalase in lung cancer.Cancer Res. 61:8578-8585.Zheng, S., De, B.P., Choudhary, S., Comhair, S.A., Goggans, T., Slee, R., Williams, B.R.,Pilewski, J., Haque, S.J., and S.C. Erzurum (2003) Impaired innate host defense causessusceptibility to respiratory virus infections in cystic fibrosis. Immunity 18:619-630.Xu, W., Comhair, S.A., Zheng, S., Chu, S.C., Marks-Konczalik, J., Moss, J., Haque, S.J., andS.C. Erzurum (2003) STAT-1 and c-Fos interaction in nitric oxide synthase-2 gene activation. Am.J. Physiol. Lung Cell. Mol. Physiol. 285:L137-L148.The Department of Cancer Biologymechanisms for regulation of NOS2 in airwayepithelial cells.However, increased expression of theNOS2 gene may contribute to the pathogenesis ofinflammatory airway diseases, i.e., asthma andbronchitis. We found that asthmatic epitheliumexpresses 15-fold higher levels of NOS2 thannormal airway epithelium. Interestingly, the upregulationof NOS2 may be rel-ated to the level ofROS present in the airway. For example, uponexposure of cells to increasing O 2, increasedamounts of NO are produceddue to increased amounts ofNOS2 expression. In contrast,decreased NO may also lead tolung disease. We recently demonstratedthat individuals withprimary pulmonary hypertensionhave decreased NO levelscompared with healthy controls.Thus, airway epithelialNOS2 may mediate pulmonaryvascular response by generatingNO, pharmacologically definedas a potent vasodilator. Furthermore,we recently identifiedthat loss of NO synthesisSerpil C. Erzurum, M.D. in cystic fibrosis (CF) contributesto the susceptibility of CFneonates to viral infection. Weare now studying the source, regulation and role ofNO in the lung; our long-term goal is, by determininghow NO is mechanistically involved in lungdiseases, to design effective therapies.Evidence has emerged in recent years suggestinga role for increased ROS and RNS in lungdiseases, specifically in the airway inflammationcharacteristic of chronic bronchitis and asthma.However, the lungs are well equipped with antioxidantdefenses, and in some experimental systems,antioxidants increase in response to ROS tominimize inflammation and injury. We have shownthat augmentation of antioxidant defenses throughgene therapy prevents cell injury. We investigatedthe response of antioxidants to increased ROS inlung. Our work has identified alterations in theantioxidants glutathione (GSH), Cu-Zn superoxidedismutase (SOD) and glutathione peroxidase(GPx) in cigarette-smoke-exposed lungs, lung cancer,asthma and pulmonary hypertension. SOD degradessuperoxide and exists in three forms, includingintracellular manganese SOD and Cu-Zn SODand an extracellular SOD. GPx removes hydrogenperoxide and organic hydroperoxides by oxidizingGSH, a water-soluble, low-molecular-weight tripeptide(L-γ-glutamyl-L-cysteinyl glycine) that isabundant in lung epithelial lining fluid. Currentwork is focused on identifying the consequences ofloss of antioxidant defenses and increase of ROSandRNS-mediated modifications of proteins inlung diseases.