Oxidant/antioxidant imbalance is implicated in the pathogenesis of chronic obstructive pulmonary disease (COPD).The current study examined the expression of antioxidant and pro-oxidant enzymes, haem oxygenases (HO) and inducible nitric oxide synthase (iNOS) respectively, in patients with severe COPD and control smokers without lung function impairment. Immunoreactivity for HO-1, HO-2, iNOS and nitric oxide-derived oxidants expressed as nitrotyrosine (N-Tyr) was quantified in peripheral lung.HO-1+ alveolar macrophages were decreased in severe COPD compared to control smokers, whereas no difference was observed in iNOS+ macrophages. In contrast, severe patients had significantly higher numbers of iNOS+ cells in alveolar walls. These iNOS+ cells were identified as type 2 pneumocytes and their number was inversely related to HO-1+ macrophages. There were no significant differences in N-Tyr immunostaining between the two groups. However, the rate of protein nitration in lungtissue was directly related to iNOS expression and associated with lower valuesofforced expiratory volume in one second/forced vital capacity. HO-2 was constitutively expressed by type 2 pneumocytes and these cells were increased in severe COPD.In conclusion, the results suggest that the enzymes involved in the oxidative stressresponse may have a different role in the lung defence and that imbalance betweenhaemoxygenase-1 and inducible nitric oxide synthase may be associated withthe development of severe impairment in chronic obstructive pulmonary disease patients.

RNA recovered from paraffin-embedded tissue has been reported to be a suitable substrate for polymerase chain reaction. During tissue fixation and paraffin embedding, RNA undergoes degradation, but with certain restrictions, it can be used for gene expression studies. At the same time, formalin-fixed, paraffin embedded histopathology archives contain an unestimable collection, which could be analyzed to investigate changes in mRNA expression in pathologic processes. To decide for future tissue conservation of pathology samples, it would be reasonable to satisfy both histologic and molecular biologic needs. The effect of three different fixation methods, RNAlater (SIGMA R 0901, St Louis, MO), acetone, and formalin, were compared by histology, immunohistochemistry, and real-time PCR. To assess tissue structure preservation and antigenicity, hematoxylin-eosin staining and immunohistochemistry were performed; to assess RNA quality, RNA was extracted and the transcription of different amplicon sizes (121, 225, 406 bp for GAPDH; 166, 310, 536 bp for β globin) were examined on human endometrium samples. The most adequate tissue preservation was found in case of formalin fixation, while there were no significant differences in the three fixatives’ yields for various size real-time PCR amplicons. Longer amplicons (above ∼225 bp) have limited use for gene expression studies, while shorter amplicons could give more reliable results.