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ISSN: 1505-8409
Przewodnik Lekarza/Guide for GPs
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1/2008
vol. 11
 
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abstract:

Hedera helix – mode of action evidenced by cell biological and biophysical investigations

Hanns Häberlein

Przew Lek 2009; 1: 255-256
Online publish date: 2008/03/03
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Extracts obtained from the leaves of Hedera helix (Ivy) are often used in the treatment of upper respiratory tract conditions characterised by hypersecretion of a viscous mucus (i.e. catarrh) and coughing. These extracts have also been applied as adjuvants for the treatment of inflammatory bronchial disease. Moreover, ivy leaf extracts are spasmolytic, reducing smooth muscle spasm, as well as bronchodilatory and antibacterial, which is mainly due to their triterpene saponin content. Hederagenin, a-hederin, and hederacoside C are the major saponins. Internalization of b2-adrenergic receptors (b2-AR) after stimulation with 1 µM terbutaline and 10 nM Alexa-NA (fluorescent b2-AR agonist) was inhibited by pre-incubation of A549 cells (alveolar type II cell line) with 1 µM a-hederin for 24 h. Compared with the positive control only 31±4% of internalised receptors were found, which was determined by laser cell scanning and quantification of intracellular fluorescence. This finding was confirmed by fluorescent microscopy imaging after immunocytochemical detection of b2-AR on A549 cells. After incubation of A549 cells with 5 nM Alexa-NA, two different diffusion time constants were found for b2-AR-Alexa-NA complexes by fluorescence correlation spectroscopy (FCS). Evaluation of the autocorrelation curve revealed an average slow diffusion time constant tbound2 of 95.2±21.8 ms (n=20) for inactive receptor-ligand complexes with hindered mobility (localized e.g. in coated pits or early endosomes) and a faster diffusion time constant tbound1 of 3.3±0.6 ms (n=20) found for active receptor-ligand complexes with free lateral mobility. Free Alexa-NA showed a tfree of 45.0±0.4 µs (n=20). Distribution of diffusion time constants in control experiments was 46±4% (n=20) for tfree, 30±3% (n=20) for tbound1, and 24±3% (n=20) for tbound2. A549 cells pre-treated with 1 µM a-hederin for 24 h showed alterations in this distribution with 37±3% (n=6) for tfree, 52±5% (n=6) for tbound1, and 11±2% (n=6) for tbound2. Because of the increased number of active receptor-ligand complexes with tbound1 and the decreased number of inactive receptor-ligand complexes with tbound2, one can conclude that the number of activated b2-AR of a-hederin treated cells is still high even under stimulating conditions. This was confirmed by determination of intracellular cAMP levels of a-hederin pre-treated HASM (human airway smooth muscle) cells after stimulation with terbutaline/forskolin, which was increased by approx. 30% compared to control cells. For alveolar type II cells an increased intracellular cAMP level could cause increased secretion of surfactant, which could thus explain the secretolytic effect of ivy extracts. In this manner one could also expect both a decrease in the intracellular Ca2+ concentration and an increase in phosphorylation of the Ca2+/calmodulin-dependent myosin light-chain kinase (MLCK) in bronchial smooth muscle cells, which could itself explain the bronchospasmolytic effect of ivy extracts.
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