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Background and objective Pulmonary emphysema is linked to T cell-mediated autoimmune inflammation, although the pathogenic role of specific pro-inflammatory cytokines remains unclear. The Th17 type response, characterized by the production of the cytokine interleukin ( IL)- 17A, is modulated in part by the IL-6/signal transducer and activator of transcription ( Stat)3 signalling axis and is associated with numerous autoimmune diseases. We therefore evaluated a causal role for IL- 17A in the IL-6-driven gp130F/ F mouse model for spontaneous pulmonary inflammation and emphysema. Methods The expression of Th17-related factors was quantified in the lungs of gp130F/ F mice and emphysematous patients, and the degree of pulmonary inflammation and emphysema was measured in gp130F/ F : Il17a−/− mice by immunohistochemistry, stereology and respiratory mechanics. Results In gp130F/ F mice, lung gene expression of Il17a and other Th17-related factors was augmented compared with gp130 +/+ (wild-type), gp130F/ F : Il6−/− and gp130F/ F : Stat3−/+ mice displaying normalized Stat3 activity and no lung inflammation. Importantly, genetic ablation of Il17a in gp130F/ F : Il17a−/− mice prevented lung inflammation; however, emphysema still developed. Additionally, messenger RNA expression of inflammatory genes Cxcl1, Cxcl2, Ccl2 and Tnfα; as well as Il6 and the Stat3-target gene, Socs3, were upregulated in the lungs of gp130F/ F mice compared with gp130F/ F : Il17a−/− and gp130+/+ mice. Consistent with these findings, augmented IL17A expression was observed in emphysema patients presenting with inflammation compared with inflammation-free individuals. Conclusions Collectively, our data suggest that the integration of IL- 17A into the IL-6/ Stat3 signalling axis mediates lung inflammation, but not emphysema, and that discrete targeting of IL- 17A may alleviate pulmonary inflammatory-related diseases. [ABSTRACT FROM AUTHOR]

Quantitative data on lung structure, such as volume, surface area and length, are used for assessment of the functional performance of the lung during normal development and inflammatory-related diseases such as chronic obstructive pulmonary disorder (COPD), and carcinogenesis, in animal models. Stereology is considered as the gold standard to obtain quantitative data on lung structure, with a key advantage being to quantify irregular three-dimensional structures on the basis of measurement made on two-dimensional sections. Therefore, preservation of original tissue dimensions without shrinkage is vital for stereology.Three steps, fixation, sampling and embedding, are essential requirements to minimise tissue shrinkage to obtain theoretically unbiased estimates of stereological parameters of lung structures. Perfusion fixation by intratracheal instillation with 1.5% glutaraldehyde/1.5% formaldehyde at a pressure of 25 cm fluid column is considered as one of the best methods. A systematic uniform random sampling scheme is then applied to the fixed lung to ensure each and every part of the lung is analysed, irrespective of homogeneity or heterogeneity of the structural distribution. The sampled tissue sections are then embedded in glycol methacrylate to minimise further tissue shrinkage. Here we describe the accurate fixation, sampling and embedding for stereological methods to quantify lung structures in mice.

Interleukin (IL)-6 is a potent immunomodulatory cytokine that is associated with emphysema, a major component of chronic obstructive pulmonary disease (COPD). IL-6 signaling via the gp130 coreceptor is coupled to multiple signaling pathways, especially the latent transcription factor signal transducer and activator of transcription (Stat)3. However, the pathological role of endogenous gp130-dependent Stat3 activation in emphysema is ill defined. To elucidate the role of the IL-6/gp130/Stat3 signaling axis in the cellular and molecular pathogenesis of emphysema, we employed a genetic complementation strategy using emphysematous gp130(F/F) mice displaying hyperactivation of endogenous Stat3 that were interbred with mice to impede Stat3 activity. Resected human lung tissue from patients with COPD and COPD-free individuals was also evaluated by immunohistochemistry. Genetic reduction of Stat3 hyperactivity in gp130(F/F):Stat3(-/+) mice prevented lung inflammation and excessive protease activity; however, emphysema still developed. In support of these findings, Stat3 activation levels in human lung tissue correlated with the extent of pulmonary inflammation but not airflow obstruction in COPD. Furthermore, COPD lung tissue displayed increased levels of IL-6 and apoptotic alveolar cells, supporting our previous observation that increased endogenous IL-6 expression in the lungs of gp130(F/F) mice contributes to emphysema by promoting alveolar cell apoptosis. Collectively, our data suggest that IL-6 promotes emphysema via upregulation of Stat3-independent apoptosis, whereas IL-6 induction of lung inflammation occurs via Stat3. We propose that while discrete targeting of Stat3 may alleviate pulmonary inflammation, global targeting of IL-6 potentially represents a therapeutically advantageous approach to combat COPD phenotypes where emphysema predominates.