Investigations into the bioactive entities of hawthorn extract WS® 1442 responsible for its endothelial barrier protecting activity

Our previous studies indicate that hawthorn (Crataegus spp.) extract WS® 1442 effectively protects against endothelial barrier dysfunction and subsequent edema formation in vitro and in vivo by influencing key regulation systems of endothelial permeability. Aim of the present study was to gain a first insight into the bioactive principles of this multi-component system. We used 4 different fractions (A-D) of WS® 1442 (Sephadex LH-20 column chromatography, kindly provided by Dr. Willmar Schwabe GmbH & Co. KG, Karlsruhe, Germany) and examined their impact on key parameters of endothelial barrier function in human endothelial cells. Fractions B (small phenolic compounds, flavonoids), C (oligomeric proanthocyanidins), and D (polymeric proanthocyanidins) inhibited the thrombin-induced endothelial hyperpermeability (Transwell® assay). Interestingly, these fractions differenzially affected the signaling pathways triggered by WS® 1442: Fractions C and D induced a clear augmentation of cAMP concentrations (ELISA), leading to an increase in VE-cadherin stability and an enhancement of cortical F-actin bundles (confocal microscopy). In contrast, the thrombin-induced rise of intracellular calcium was primarily attenuated by fraction B (ratiometric imaging). Fractions B and C were further sub-fractionated (preparative RP-HPLC, Schwabe). Concerning fraction B, two sub-fractions out of ten clearly affected the calcium signaling. Four out of six sub-fractions of fraction C induced cortactin phosphorylation and cortical F-actin redistribution. Currently, these sub-fractions undergo further investigations focusing on their impact on calcium signaling. In summary, we showed for the first time that the different signaling mechanisms triggered by the extract can clearly be assigned to distinct fractions, i.e. phytochemical groups of WS® 1442.