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Published ahead of print on April 29, 2004, doi:10.1164/rccm.200308-1071OC
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American Journal of Respiratory and Critical Care Medicine Vol 170. pp. 154-161, (2004)
© 2004 American Thoracic Society

Original Article

Differential Cytokine Gene Expression in the Diaphragm in Response to Strenuous Resistive Breathing

Theodoros Vassilakopoulos, Maziar Divangahi, George Rallis, Osama Kishta, Basil Petrof, Alain Comtois and Sabah N. A. Hussain

Critical Care and Respiratory Divisions, Department of Medicine, McGill University Hospital Center; and Meakins-Christie Laboratories, McGill University, Montreal, Quebec, Canada

Correspondence and requests for reprints should be addressed to Theodoros Vassilakopoulos, M.D., Department of Critical Care and Pulmonary Services, University of Athens Medical School, 45–47 Ipsilandou Street 10675, Athens, Greece. E-mail:tvassil{at}med.uoa.gr

Strenuous resistive breathing induces plasma cytokines that do not originate from circulating monocytes. We hypothesized that cytokine production is induced inside the diaphragm in response to resistive loading. Anesthetized, tracheostomized, spontaneously breathing Sprague-Dawley rats were subjected to 1, 3, or 6 hours of inspiratory resistive loading, corresponding to 45–50% of the maximum inspiratory pressure. Unloaded sham-operated rats breathing spontaneously served as control animals. The diaphragm and the gastrocnemius muscles were excised at the end of the loading period, and messenger ribonucleic acid expression of tumor necrosis factor-{alpha}, tumor necrosis factor-ß, interleukin (IL)-1{alpha}, IL-1ß, IL-2, IL-3, IL-4, IL-5, IL-6, IL-10, IFN-{gamma}, and two housekeeping genes was analyzed using multiprobe RNase protection assay. IL-6, IL-1ß, and, to lesser extents, tumor necrosis factor-{alpha}, IL-10, IFN-{gamma}, and IL-4 were significantly increased in a time-dependent fashion in the diaphragms but not the gastrocnemius of loaded animals or in the diaphragm of control animals. Elevation of protein levels of IL-6 and IL-1ß in the diaphragm of loaded animals was confirmed with immunoblotting. Immunostaining revealed IL-6 protein localization inside diaphragmatic muscle fibers. We conclude that increased ventilatory muscle activity during resistive loading induces differential elevation of proinflammatory and antiinflammatory cytokine gene expression in the ventilatory muscles.

Key Words: interleukin • loaded breathing • respiratory muscles • ribonuclease protection assay




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