Effect of two homeopathic remedies at different degrees of dilutions on the wound closure of 3T3 fibroblasts in in vitro scratch assay

Background: Since ancient times, preparations from traditional medicinal plants e.g. Arnica montana, Calendula officinalis or Hypericum perforatum have been used for different wound healing purposes. The aim of this study was to investigate the efficacy of the commercial low dilution homeopathic remedy Similasan® Arnica plus Spray, a preparation of Arnica montana 4x, Calendula officinalis 4x, Hypericum perforatum 4x and Symphytum officinale 6x (0712-2) and medium diluted SIM WuS (Petroleum 15x, Arnica montana 15x, Calcium fluoratum 12x, Calendula officinalis 12x, Hepar sulfuris 12x and Mercurius solubilis 15x; 1101-4), on the wound healing in cultured NIH 3T3 fibroblasts. Both remedies were from Similasan AG (Jonen, Switzerland) and prepared according the German Homoeopathic Pharmacopoeia (GHP) following descriptions 4a for arnica, 3a for marigold and St. John’s wort, 2a for comfrey, 5a for petroleum, and 6 for calcium fluoride, hepar sulfuris and mercurius solubilis. Materials and Methods: Cell proliferation, migration and wound closure promoting effect of the preparations (0712-2, 1101- 4) and their succussed solvents (0712-1, 1101-3) were investigated on mouse NIH 3T3 fibroblasts. Cell viability was determined by WST-1 assay, cell growth using BrdU uptake, cell migration by chemotaxis assay and wound closure by CytoSelect ™Wound Healing Assay Kit which generated a defined wound area. All assays were performed in three independent controlled experiments. In some experiments diluted unsuccussed alcohol (0712-3) was also investigated. Results: Preparations (0712-1), (0712-2), (0712-3), (1101-3) and (1101-4) were investigated at decimal dilution steps from 1x to 4x. Cell viabilty was not affected by any of the substances and (0712-1) and (0712-2) showed no stimulating effect on cell proliferation. Preparation (0712-2) exerted a stimulating effect on fibroblast migration (31.7%) vs 15% with succussed solvent (0712-1) at 1:100 dilutions (p0.05). Positive control 2 ng/ml EGF increased migratory activity of cells by 49.8%. Preparation (0712-2) at a dilution of 1:100 promoted in vitro wound closure by 59.5% and differed significantly (p