title: Comparison of In-Vitro and Ex-Vivo Wound Healing Assays for the Investigation of Diabetic Wound Healing and Demonstration of a Beneficial Effect of a Triterpene Extract.
abstract: Diabetes mellitus is a frequent cause for chronic, difficult-to-treat wounds. New therapies for diabetic wounds are urgently needed and in-vitro or ex-vivo test systems are essential for the initial identification of new active molecules. The aim of this study is to compare in-vitro and ex-vivo test systems for their usability for early drug screening and to investigate the efficacy of a birch bark triterpene extract (TE) that has been proven ex-vivo and clinically to accelerate non-diabetic wound healing (WH), in a diabetic context. We investigated in-vitro models for diabetic WH, i.e. scratch assays with human keratinocytes from diabetic donors or cultured under hyperglycaemic conditions and a newly developed porcine ex-vivo hyperglycaemic WH model for their potential to mimic delayed diabetic WH and for the influence of TE in these test systems. We show that keratinocytes from diabetic donors often fail to exhibit significantly delayed WH. For cells under hyperglycaemic conditions significant decrease is observed but is influenced by choice of medium and presence of supplements. Also, donor age plays a role. Interestingly, hyperglycaemic effects are mainly hyperosmolaric effects in scratch assays. Ex-vivo models under hyperglycaemic conditions show a clear and substantial decrease of WH, and here both glucose and hyperosmolarity effects are involved. Finally, we provide evidence that TE is also beneficial for ex-vivo hyperglycaemic WH, resulting in significantly increased length of regenerated epidermis to 188±16% and 183±11% (SEM; p<0.05) compared to controls when using two different TE formulations. In conclusion, our results suggest that microenvironmental influences are important in WH test systems and that therefore the more complex hyperglycaemic ex-vivo model is more suitable for early drug screening. Limitations of the in-vitro and ex-vivo models are discussed. Furthermore our data recommend TE as a promising candidate for in-vivo testings in diabetic wounds.
authors.affiliation: University Medical Center Hamburg-Eppendorf, Department of Dermatology and Venerology, Hamburg, Germany.
authors.forename: Christopher
authors.initials: C
authors.lastname: Ueck
authors.affiliation: University Medical Center Hamburg-Eppendorf, Department of Dermatology and Venerology, Hamburg, Germany.
authors.forename: Thomas
authors.initials: T
authors.lastname: Volksdorf
authors.affiliation: University Medical Center Hamburg-Eppendorf, Department of Dermatology and Venerology, Hamburg, Germany.
authors.forename: Pia
authors.initials: P
authors.lastname: Houdek
authors.affiliation: University Medical Center Hamburg-Eppendorf, Department of Dermatology and Venerology, Hamburg, Germany.
authors.forename: Sabine
authors.initials: S
authors.lastname: Vidal-Y-Sy
authors.affiliation: University Medical Center Hamburg-Eppendorf, Institute of Medical Biometry and Epidemiology, Hamburg, Germany.
authors.forename: Susanne
authors.initials: S
authors.lastname: Sehner
authors.affiliation: Fraunhofer Institute for Molecular Biology and Applied Ecology IME, ScreeningPort, Hamburg, Germany.
authors.forename: Bernhard
authors.initials: B
authors.lastname: Ellinger
authors.affiliation: Klinikum Stuttgart, Department for Endocrinology, Diabetology and Geriatrics, Stuttgart, Germany.
authors.forename: Ralf
authors.initials: R
authors.lastname: Lobmann
authors.affiliation: University Medical Center Hamburg-Eppendorf, Department of Vascular Medicine, Hamburg, Germany.
authors.forename: Axel
authors.initials: A
authors.lastname: Larena-Avellaneda
authors.affiliation: University Medical Center Hamburg-Eppendorf, Department and Clinic of Pediatric Surgery, Hamburg, Germany.
authors.forename: Konrad
authors.initials: K
authors.lastname: Reinshagen
authors.affiliation: University Medical Center Hamburg-Eppendorf, Department and Clinic of Pediatric Surgery, Hamburg, Germany.
authors.forename: Ina
authors.initials: I
authors.lastname: Ridderbusch
authors.affiliation: University Medical Center Hamburg-Eppendorf, Department of Dermatology and Venerology, Hamburg, Germany.
authors.forename: Klaas
authors.initials: K
authors.lastname: Kohrmeyer
authors.affiliation: University Medical Center Hamburg-Eppendorf, Department of Dermatology and Venerology, Hamburg, Germany.
authors.forename: Ingrid
authors.initials: I
authors.lastname: Moll
authors.affiliation: Eberhard Karls University Tübingen, Department of Pharmaceutical Technology, Tübingen, Germany.
authors.forename: Rolf
authors.initials: R
authors.lastname: Daniels
authors.affiliation: Albert-Ludwigs-University Freiburg, Department of Pharmaceutical Biology and Biotechnology, Freiburg, Germany.
authors.forename: Philipp
authors.initials: P
authors.lastname: Werner
authors.affiliation: Albert-Ludwigs-University Freiburg, Department of Pharmaceutical Biology and Biotechnology, Freiburg, Germany.
authors.forename: Irmgard
authors.initials: I
authors.lastname: Merfort
authors.affiliation: University Medical Center Hamburg-Eppendorf, Department of Dermatology and Venerology, Hamburg, Germany.
authors.forename: Johanna M
authors.initials: JM
authors.lastname: Brandner
mesh_terms: D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000818:Animals; D002675:Child, Preschool; D016207:Cytokines / Q000378:metabolism; D003920:Diabetes Mellitus / Q000188:drug therapy* / Q000473:pathology*; D004848:Epithelium / Q000187:drug effects / Q000473:pathology; D005260:Female; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism; D005947:Glucose / Q000494:pharmacology; D006801:Humans; D006943:Hyperglycemia / Q000150:complications / Q000188:drug therapy; D015603:Keratinocytes / Q000187:drug effects / Q000473:pathology; D008297:Male; D034421:Sus scrofa; D014315:Triterpenes / Q000494:pharmacology* / Q000627:therapeutic use*; D014945:Wound Healing / Q000187:drug effects*
chemical_list: D016207:Cytokines; D014315:Triterpenes; D005947:Glucose