Sekretomodulatorische Effekte des proinflammatorischen Zytokins GM-CSF auf die Muzinsekretion an der isolierten Rattentrachea nativer und NO2-exponierter Tiere

Eine Facette des klinischen Erscheinungsbildes der „chronic obstructive pulmonary disease“ (COPD) ist die chronische Bronchitis mit einem überwiegend inflammatorischen Korrelat. Der Pathogenese der chronischen Bronchitis liegen komplexe pathophysiologische Vorgänge zugrunde, welche bislang nur unv...

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Bibliographic Details
Main Author: Krohm, Inga
Contributors: Wagner, Ulrich (Prof. Dr. med.) (Thesis advisor)
Format: Doctoral Thesis
Published: Philipps-Universität Marburg 2019
Online Access:PDF Full Text
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One facet of the clinical manifestation of chronic obstructive pulmonary disease (COPD) is chronic bronchitis, with inflammation being the predominant correlating factor. The pathogenesis of chronic bronchitis is based on complex pathophysiological processes, which are currently not fully known and have not yet been fully clarified, in detail. The definition from the "global initiative on obstructive lung disease" (GOLD) mentions an abnormal inflammatory reaction of the lung to harmful particles and gases, thus putting inflammation at the centre of the pathogenesis. Cigarette smoke and important environmental noxae such as nitrogen dioxide (NO2) cause changes in the respiratory epithelium of humans and other species through eliciting inflammation and stimulate the production of pro-inflammatory cytokines, such as granulocyte-macrophage colony stimulating factor (GM-CSF). These structural changes also involve the mucin-producing epithelial components. One relevant clinical symptom of chronic bronchitis is hypersecretion. The regulation of mucin synthesis and secretion is subject to complex neural, neuro-endocrine and paracrine mechanisms. With the aim of characterising the underlying regulatory mechanisms of mucus secretion in the chronic inflammatory disease COPD, the focus of this work is on the significance of the pro-inflammatory cytokine GM-CSF. The following questions that are of importance for the regulation of mucus secretion will be investigated in this paper: What influence do long- and short-term exposure of NO2 have on tracheobronchial mucin secretion? What influence does the peptidergic substance substance P (SP) have on the mucin secretion of exposed animals, and how does this compare by species? Does the pro-inflammatory cytokine GM-CSF act as a secretagogue? Are there interactive processes between the cytokine GM-CSF and the tachykinin SP with regard to secretion modulation? An established animal model was chosen for the experimental clarification of these questions. To study the mode of response of respiratory tracts altered through inflammation, inhalative exposure to NO2 was implemented in rats. Mucin secretion was investigated in tracheal explants of Fischer rats (F344 rats) and Sprague-Dawley rats (SD rats) using the established Ussing chamber method. In order to quantify the secretion of tracheal submucosal glands, the mucin underwent radioactive labelling with 35S. In order to study tracheal mucin secretion, inhalative exposure was implemented, with damage induced and acute and subacute to chronic tracheobronchitis brought about. Using standardised methods, exposure times of 1, 3 and 20 days were selected for male rats of the species in question, as well as of 2 and 28 days, in some cases, to NO2 at a concentration of 10 ppm. Control animals were subject to an atmosphere without NO2. In addition to basal secretion, initially, the maximum stimulated secretion due to acetylcholine (ACh) was recorded on the tracheal implants in stages of artificial inflammation and in the natural state. With an understanding of these foundations, the secreto-modulatory effect of the peptide SP and the dose–response relationship of the pro-inflammatory cytokine GM-CSF can then be investigated over time under exposure to NO2. In addition, the question of any additive effect of GM-CSF and SP in terms of secretion result was investigated in tracheas exposed long-term. The secretion analysis showed that NO2 exposure of 10 ppm does not significantly influence tracheal mucin secretion in rats of either species in a time-dependent manner. However, similar trends were demonstrated that were of no significance over the course of secretion. In the species comparison, SD rats exhibited a greater secretory potential compared to F344 rats, in line with an almost parallel shift in secretion level. What is more, in both species, the natural secretion rate and that up until an exposure duration of 20 days was significantly increased by acetylcholine compared to basal secretion. Here, again, in a direct comparison with the native Fischer rats, significant differences were noted after 3 and 20 days. Significant secretagogic effects were able to be demonstrated for substance P in Fischer rats and SD rats, both in the control animals and in almost all the exposure groups investigated. GM-CSF was shown to stimulate tracheal mucin secretion in our COPD rat model in a manner that was both dose- and exposure-time-dependent. A significant secretory response was able to be demonstrated in the 3-day exposure group for 0.1 ng/ml GM-CSF and for 0.4 ng/ml GM-CSF. In addition, significance was noted in the 20-day exposure group for 0.1 and 0.15 as well as 0.2 ng/ml GM-CSF. A highly significant result was obtained for the dose of 0.2 ng/ml GM-CSF. Simultaneous administration of GM-CSF and SP did not indicate any measurable additive secretagogic effect over long-term exposure and resulted in the secretion level of SP. This was investigated after 20 days of exposure, since for this group, a significant secretory response was seen for both individual substances. The supposedly different secretagogic mechanisms of SP and GM-CSF are compatible with the observation that SP is in any case effective regardless of the presence of any respiratory inflammation. These results contribute to the understanding of the complexity of the tracheobronchial epithelium pathomechanisms involved in the regulation of mucin secretion in chronic bronchitis. The secretagogic effect of GM-CSF in the presence of chronic inflammation was demonstrated. Here, a targeted therapeutic approach is conceivable, with the aim of influencing hypersecretion in cases of COPD. The use of neutralising antibodies against GM-CSF or its receptor is plausible as a specific therapy concept. This should be viewed in addition to the established anti-inflammatory therapeutic options for COPD and, by contrast, enables targeted intervention in the inflammatory cascade. Further research is needed to substantiate the role of such an approach.