title: Nitric oxide and apoptosis during human head and neck squamous cell carcinoma development.
abstract: OBJECTIVE
  Apoptosis index (AI), Bcl-2, and Bax have shown prognostic significance in head and neck squamous cell carcinoma (HNSCCa). Other areas of research have implicated nitric oxide (NO) or its various intermediate species in both proapoptotic and antiapoptotic processes. We have previously shown that NO-generating enzymes are significantly increased during the stepwise progression to HNSCCa. The aim of this study was to explore the interrelationship of NO and a known consequence of NO-related oxidative stress, apoptosis, during this step-wise process.
  
  
  METHODS
  Formalin fixed-paraffin embedded tissue samples of 10 normal oral mucosa, 15 reactive/dysplastic lesions, and 17 HNSCCa lesions studied previously were subjected to the terminal deoxynucleotidyl transferase (TdT)-mediated dUTP labeling (TUNEL) assay as well as immunohistochemical staining against Bcl-2, Bax, and p53. Patient charts were reviewed and clinical data were compared. The study pathologist (G.K.H) reviewed these slides blinded to patient identifiers or clinical data. The number of immunopositive cell nuclei or staining intensity was graded, noting the pattern of immunostaining. These staining characteristics were compared with the results of immunostaining previously obtained for endothelial constitutive NO synthase (ecNOS) and nitrotyrosine.
  
  
  RESULTS
  Compared with normal oral mucosa, the AI, Bcl-2, Bax, Bcl-2/Bax intensity and frequency ratios, and mutant p53 intensity significantly changed in reactive/dysplastic and HNSCCa lesions (P <.001 for all). Correlations between the staining characteristics of the antigens studied are presented. Furthermore, perilesional inflammatory cells showed staining in the TUNEL assay.
  
  
  CONCLUSIONS
  In a set of tissue samples previously well characterized, these new findings implicate a link between NO and the induction of apoptotic cell death in HNSCCa development.
authors.affiliation: Department of Otolaryngology-Head & Neck Surgery, Northwestern University Medical Center, 303 East Chicago Ave., Chicago, IL 60611-3008, USA.
authors.forename: Brandon G
authors.initials: BG
authors.lastname: Bentz
authors.forename: Rakesh
authors.initials: R
authors.lastname: Chandra
authors.forename: G Kenneth
authors.initials: GK
authors.lastname: Haines
authors.forename: Alan M
authors.initials: AM
authors.lastname: Robinson
authors.forename: Pinky
authors.initials: P
authors.lastname: Shah
authors.forename: James A
authors.initials: JA
authors.lastname: Radosevich
mesh_terms: D000911:Antibodies, Monoclonal; D017209:Apoptosis / Q000502:physiology*; D002294:Carcinoma, Squamous Cell / Q000235:genetics / Q000378:metabolism* / Q000473:pathology*; D019254:Genes, bcl-2 / Q000235:genetics; D006258:Head and Neck Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000473:pathology*; D006801:Humans; D007150:Immunohistochemistry; D020287:In Situ Nick-End Labeling / Q000379:methods; D008875:Middle Aged; D009367:Neoplasm Staging; D009569:Nitric Oxide / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000235:genetics; D019253:Proto-Oncogene Proteins c-bcl-2; D051028:bcl-2-Associated X Protein
chemical_list: D000911:Antibodies, Monoclonal; C494826:BAX protein, human; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D051028:bcl-2-Associated X Protein; D009569:Nitric Oxide