title: The Wound Healing and Antibacterial Activity of Five Ethnomedical Calophyllum inophyllum Oils: An Alternative Therapeutic Strategy to Treat Infected Wounds.
abstract: BACKGROUND
  Calophyllum inophyllum L. (Calophyllaceae) is an evergreen tree ethno-medically used along the seashores and islands of the Indian and Pacific Oceans, especially in Polynesia. Oil extracted from the seeds is traditionally used topically to treat a wide range of skin injuries from burn, scar and infected wounds to skin diseases such as dermatosis, urticaria and eczema. However, very few scientific studies reported and quantified the therapeutic properties of Calophyllum inophyllum oil (CIO). In this work, five CIO from Indonesia (CIO1), Tahiti (CIO2, 3), Fiji islands (CIO4) and New Caledonia (CIO5) were studied and their cytotoxic, wound healing, and antibacterial properties were presented in order to provide a scientific support to their traditional use and verify their safety.
  
  
  METHODS
  The safety of the five CIO was ascertained using the Alamar blue assay on human keratinocyte cells. CIO wound healing properties were determined using the scratch test assay on human keratinocyte cells. CIO-stimulated antibacterial innate immune response was evaluated using ELISA by measuring β defensin-2 release in human derivative macrophage cells. CIO antibacterial activity was tested using oilogramme against twenty aerobic Gram- bacteria species, twenty aerobic Gram+ bacteria species, including a multi-drug resistant Staphylococcus aureus strain and two anaerobic Gram+ bacteria species e.g. Propionibacterium acnes and Propionibacterium granulosum. To detect polarity profile of the components responsible of the antibacterial activity, we performed bioautography against a Staphylococcus aureus strain.
  
  
  RESULTS
  Based on Alamar Blue assay, we showed that CIO can be safely used on keratinocyte cells between 2.7% and 11.2% depending on CIO origin. Concerning the healing activity, all the CIO tested accelerated in vitro wound closure, the healing factor being 1.3 to 2.1 higher compared to control when keratinocytes were incubated after scratch with CIO at 0.1%. Furthermore, our results showed that CIO exhibit two distinct antibacterial effects: one against Gram+ bacteria by direct inhibition of mitotic growth and another potent effect against Gram- bacteria due to increased release of β-defensin 2 peptide by macrophages. Interestingly, the needed concentrations of CIO to inhibit bacteria growth and to promote wound healing are lower than concentrations exhibiting cytotoxic effects on keratinocyte cells. Finally, we performed bioautography assay against Staphylococcus aureus to determine polarity profile of the components responsible for CIO antibacterial activity. Our results showed for the five tested CIO that components responsible of the bacterial growth inhibition are the more polar one on the TLC chromatographic profile and are contained in the resinous fraction of the oil.
  
  
  CONCLUSIONS
  This study was conducted to evaluate cytotoxicity, wound healing and antibacterial properties of five CIO traditionally used to treat infected wounds. Using cell and bacteria cultures, we confirmed the pharmacological effects of CIO as wound healing and antimicrobial agent. Moreover, we showed that concentration of CIO needed to exhibit therapeutic effects are lower than concentrations exhibiting cytotoxic effects in vitro. For the first time, this study provides support for traditional uses of CIO. These wound healing and antibiotic properties make CIO a valuable candidate to treat infected wounds especially in tropical areas.
authors.affiliation: Laboratoire Chimie-Toxicologie Analytique et Cellulaire-UMR CNRS COMETE 8638, Université Paris Descartes, Sorbonne Paris Cité, Faculté de Pharmacie, Paris, France.
authors.forename: Teddy
authors.initials: T
authors.lastname: Léguillier
authors.affiliation: Laboratoire Ecosystème Intestinal, Probiotiques, Antibiotiques-EA 4065, Université Paris Descartes, Sorbonne Paris Cité, Faculté de Pharmacie, Paris, France.
authors.forename: Marylin
authors.initials: M
authors.lastname: Lecsö-Bornet
authors.affiliation: Laboratoire de Pharmacognosie-UMR CNRS COMETE 8638, Université Paris Descartes, Sorbonne Paris Cité, Faculté de Pharmacie, Paris, France.
authors.forename: Christelle
authors.initials: C
authors.lastname: Lémus
authors.affiliation: INRA, Biologie du Développement et Reproduction-UMR 1198, Jouy-en-Josas, France.
authors.forename: Delphine
authors.initials: D
authors.lastname: Rousseau-Ralliard
authors.affiliation: Laboratoire Insulaire du Vivant et de l'Environnement-EA 4243, Université de la Nouvelle-Calédonie, Nouméa, Nouvelle Calédonie, France.
authors.forename: Nicolas
authors.initials: N
authors.lastname: Lebouvier
authors.affiliation: Laboratoire Insulaire du Vivant et de l'Environnement-EA 4243, Université de la Nouvelle-Calédonie, Nouméa, Nouvelle Calédonie, France.
authors.forename: Edouard
authors.initials: E
authors.lastname: Hnawia
authors.affiliation: Laboratoire Insulaire du Vivant et de l'Environnement-EA 4243, Université de la Nouvelle-Calédonie, Nouméa, Nouvelle Calédonie, France.
authors.forename: Mohammed
authors.initials: M
authors.lastname: Nour
authors.affiliation: Institute of Applied Sciences, University of the South Pacific, Laucala Campus, Suva, Fiji.
authors.forename: William
authors.initials: W
authors.lastname: Aalbersberg
authors.affiliation: Centre de recherche de BioMécanique et BioIngénierie-CNRS UMR 7338, Université de Technologie de Compiègne, Compiègne, France.
authors.forename: Kamelia
authors.initials: K
authors.lastname: Ghazi
authors.affiliation: Equipe Etude Intégrée des Métabolites Secondaires-UMR 241 EIO, Université de la Polynésie Française, Tahiti, FAA'A, Polynésie Française.
authors.forename: Phila
authors.initials: P
authors.lastname: Raharivelomanana
authors.affiliation: Laboratoire Chimie-Toxicologie Analytique et Cellulaire-UMR CNRS COMETE 8638, Université Paris Descartes, Sorbonne Paris Cité, Faculté de Pharmacie, Paris, France.
authors.forename: Patrice
authors.initials: P
authors.lastname: Rat
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D029759:Calophyllum / Q000737:chemistry*; D002478:Cells, Cultured; D020834:Ethnopharmacology; D005786:Gene Expression Regulation / Q000187:drug effects; D006090:Gram-Negative Bacteria / Q000187:drug effects; D006094:Gram-Positive Bacteria / Q000187:drug effects; D006801:Humans; D015603:Keratinocytes / Q000187:drug effects; D008264:Macrophages / Q000187:drug effects; D010139:Pacific Islands / Q000208:ethnology; D010938:Plant Oils / Q000494:pharmacology*; D012639:Seeds / Q000737:chemistry; D014945:Wound Healing / Q000187:drug effects*; D014946:Wound Infection / Q000188:drug therapy; D023083:beta-Defensins / Q000378:metabolism
chemical_list: D000900:Anti-Bacterial Agents; C106641:DEFB4A protein, human; D010938:Plant Oils; D023083:beta-Defensins