Originally discovered in bacteria, subtilisin-like proteases are also abundant described in plants such as Oryza sativa L., Arabidopsis thaliana (L.) Heynh. and Solanum lycopersicum L. Proteases have the potential to induce inflammation through activation of protease-activated receptors (PAR) [1]. It is known, that plant latices induce strong skin inflammation. Therefore, we investigate the plant latex from plants of Euphorbia mauritanica L. (Euphorbiaceae Juss.) which contains amongst to other secondary plant ingredients diterpenes [2] and proteins [3]. We focused on the inflammatory potential of the combination with diterpenes and proteases. Mauritanicain, a Subtilisin-like protease from E. mauritanica, was isolated with anion exchange chromatography, followed by size exclusion chromatography. Fibroblasts were isolated from juvenile human foreskin. Various concentrations were tested on HaCaT keratinocytes and the isolated fibroblasts. PAR2 selective agonist peptide 2-Fluoryl-LIGRO-amide, diterpene Phorbol-12-myristat-13-acetate (PMA) and Subtilisin were used as single treatments. PAR positive studies were made with PCR. IL-8 was detected with ELISA. The results show, that the combination of Mauritanicain and the diterpene PMA increased the release of IL-8 both, in HaCaT cell line (1.5-fold after 48h compared with Mauritanicain) and in primary fibroblasts (4-fold after 24h – compared with PMA), compared to single treatments with Mauritanicain, Subtilisin, or PMA.
[1] Guenther F, Melzig MF. Protease-activated receptors and their biological role - focused on skin inflammation.; J Pharm Pharmacol 2015; 67: 1623-1633
[2] Vasas A, Hohmann J. Euphorbia diterpenes: isolation, structure, biological activity, and synthesis (2008-2012). Chem Rev 2014; 114: 8579-8612
[3] Sytwala S, Guenther F, Melzig MF. Lysozyme- and chitinase activity in latex bearing plants of genus Euphorbia – A contribution to plant defense mechanism. Plant Physiology and Biochemistry 2015; 95: 35-40