title: Activity of Thymus capitellatus volatile extract, 1,8-cineole and borneol against Leishmania species.
abstract: In the search for new leishmanicidal agents, Thymus capitellatus Hoffmanns. & Link (family Lamiaceae) volatile extract and its major compounds, 1,8-cineole and borneol, were tested against Leishmania infantum, Leishmania tropica and Leishmania major. Plant volatile extract (essential oil) was analysed by GC and GC-MS and the activity of essential oil on Leishmania promastigotes viability was assessed using tetrazolium-dye colorimetric method (MTT). The MTT test was also used to assess the cytotoxicity of essential oil on macrophages and bovine aortic endothelial cells. Effects on parasites were also analyzed by flow cytometry in order to assess mitochondrial transmembrane electrochemical gradient (JC-1), analyze phosphatidylserine externalization (annexin V-FITC, propidium iodide) and evaluate cell cycle (DNase-free, RNase, PI). Morphological and ultrastructural studies were performed by light, scanning and transmission electron microscopy. T. capitellatus volatile extract exhibited anti-parasite activity on Leishmania species, with IC50 values ranging from 35 to 62 μg/ml. However, major compounds 1,8-cineole and borneol did not showed biological activity suggesting that these monoterpenes are not responsible for the antileishmanial activity of T. capitellatus essential oil. Appearance of aberrant-shaped cells, mitochondrial swelling and autophagosomal structures were some of the ultrastructural alterations exhibited among treated promastigote cells. T. capitellatus promoted leishmanicidal effect by triggering a programmed cell death as evidenced by externalization of phosphatidylserine, loss of mitochondrial membrane potential, and cell-cycle arrest at the G(0)/G(1) phase. The volatile extract did not induced cytotoxic effects on mammalian cells. Taken together, these results suggest that T. capitellatus may represent a valuable source for therapeutic control of leishmaniasis in humans and animals.
authors.affiliation: Faculty of Pharmacy/CEF, University of Coimbra, Azinhaga de Santa Comba, 3030-548 Coimbra, Portugal; Pharmacy Department, School of Health Vale do Ave/Centre for Research in Health Technologies CESPU-IPSN, Gandra, Portugal.
authors.forename: M
authors.initials: M
authors.lastname: Machado
authors.affiliation: Department of Life Sciences, Faculty of Science and Technology, University of Coimbra, Coimbra, Portugal.
authors.forename: A M
authors.initials: AM
authors.lastname: Dinis
authors.affiliation: Institute of Immunology, Faculty of Medicine, University of Coimbra, Coimbra, Portugal.
authors.forename: M
authors.initials: M
authors.lastname: Santos-Rosa
authors.affiliation: Institute of Immunology, Faculty of Medicine, University of Coimbra, Coimbra, Portugal.
authors.forename: V
authors.initials: V
authors.lastname: Alves
authors.affiliation: Faculty of Pharmacy/CEF, University of Coimbra, Azinhaga de Santa Comba, 3030-548 Coimbra, Portugal.
authors.forename: L
authors.initials: L
authors.lastname: Salgueiro
authors.affiliation: Faculty of Pharmacy/CEF, University of Coimbra, Azinhaga de Santa Comba, 3030-548 Coimbra, Portugal.
authors.forename: C
authors.initials: C
authors.lastname: Cavaleiro
authors.affiliation: Faculty of Pharmacy/CEF, University of Coimbra, Azinhaga de Santa Comba, 3030-548 Coimbra, Portugal. Electronic address: mcsousa@ci.uc.pt.
authors.forename: M C
authors.initials: MC
authors.lastname: Sousa
mesh_terms: D000818:Animals; D000981:Antiprotozoal Agents / Q000494:pharmacology / Q000633:toxicity; D001892:Camphanes / Q000494:pharmacology*; D002417:Cattle; D002453:Cell Cycle / Q000187:drug effects; D002460:Cell Line; D002478:Cells, Cultured; D003511:Cyclohexanols / Q000494:pharmacology*; D042783:Endothelial Cells / Q000187:drug effects; D000077591:Eucalyptol; D020128:Inhibitory Concentration 50; D007891:Leishmania / Q000187:drug effects*; D008264:Macrophages / Q000187:drug effects; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D008855:Microscopy, Electron, Scanning; D046529:Microscopy, Electron, Transmission; D039821:Monoterpenes / Q000494:pharmacology*; D009822:Oils, Volatile / Q000737:chemistry / Q000494:pharmacology* / Q000633:toxicity; D010718:Phosphatidylserines / Q000378:metabolism; D010938:Plant Oils / Q000737:chemistry / Q000494:pharmacology / Q000633:toxicity; D046930:Thymus Plant / Q000737:chemistry*
chemical_list: D000981:Antiprotozoal Agents; D001892:Camphanes; D003511:Cyclohexanols; D039821:Monoterpenes; D009822:Oils, Volatile; D010718:Phosphatidylserines; D010938:Plant Oils; C022871:isoborneol; D000077591:Eucalyptol
keywords: Apoptosis; Drug discovery; Essential oil; Leishmaniasis; Monoterpenes; Protozoa