The effect of carbonyl-sulphide containing medicinal water bath treatment on LPS-induced airway inflammation in mice

A Markovics, A Kemeny, A Perkecz, ZS Helyes, E Pinter. University of Pecs, Department of Pharmacology and Pharmacotherapy 7624, Hungary

Balneotherapy has been long used in the treatment of immune-mediated skin and arthritic diseases. Our research group showed that the bath treatment with the Harkany medicinal water containing carbonyl-sulphide exerts significant anti-inflammatory effects in the oxazolon-induced mouse model of allergic contact dermatitis. Carbonyl-sulphide is well-absorbed and converted to hydrogen-sulphide (H2S) in the body. It stimulates capsaicin-sensitive sensory nerves and induces the release of sensory neuropeptides, therefore it is likely to modulate the inflammatory processes and neuro-immune interactions. Therefore, the aim of the present study was to investigate the effect of this medicinal water bath on a pneumonitis model of the mouse.

Male CD1 mice (25-30 g, n=15/group) were bathed in 37 oC medicinal water or the same temperature distilled water for 20 min/day, during a 14-day experimental period. Airway inflammation was induced with intranasal administration of E. coli (serotype 083) endotoxin (lipopolysaccharide: LPS, dissolved in saline, 167 µg/ml, 60 µl) under short-lasting aether anaesthesia. Airway hyper-reactivity to inhaled carbachol (50 µl/mouse carbachol, 11 and 22 mM) was determined in conscious mice with the Buxco whole body plethysmograph before and 24 h after LPS administration. Saline (PBS)-treated mice served as controls. Mice were sacrificed at the end of the study under deep ketamin-xylazine anaesthesia. Bronchoalveolar lavage (BAL) was performed with sterile ice-cold PBS to determine the inflammatory cell profile of the airways with flow-cytometry. The right lung of each animal was fixed in formaldehyde, embedded in paraffin and stained with hematoxylin and eosin for histopathological investigation and semiquantitative scoring. The left lung was frozen in liquid nitrogen for measuring the inflammatory cytokine interleukin 1 beta (IL-1β) with ELISA from the homogenates. Statistical evaluation was performed with one-way ANOVA and in case of the histopathological scores with Kruskall-Wallis test.

Bronchial responsiveness to carbachol inhalation remarkably increased 24 h after LPS instillation compared to PBS administration in distilled water-treated mice. Medicinal water bath pretreatment did not influence this inflammatory airway hyper-responsiveness. The BAL fluid samples of distilled water-treated control mice receiving LPS 24 h earlier contained 2307.976±617.18 lymphocytes, 823.0317±236.18 granulocytes and 1951.05±418.31 macrophages. Medicinal water pretreatment significantly reduced the number of these inflammatory cells; there were 956.2232±353.408 lymphocytes (p=0.0391), 251.1658±76.305 granulocytes (p=0.0154) and 989.2377±255.84 macrophages (p=0.0145). The characteristic histopathological parameters, such as peribronchial oedema formation, neutrophil accumulation, macrophage infiltration, and goblet cell hyperplasia, as well as IL-1β concentration were also significantly reduced in response to medicinal water therapy.

We provided evidence that the carbonyl-sulphide-containing Harkany medicinal water attenuates inflammatory processes in the mouse model of endotoxin-evoked subacute airway inflammation. Therefore, besides skin and joint diseases, it migh also provide a beneficial effect for the treatment of airway inflammation.