Scratch wound assay

For experiments, fibroblasts were harvested through trypsin-EDTA (Gibco) treatment, seeded into 4-well culture slides (BD Biosciences, Germany) at a density of 40,000 cells/cm², and cultured for 48 h to confluence. Afterwards, fibroblasts monolayers were scratched with a sterile pipette tip and washed with medium to remove any loose cells. Then, wound dressing samples were placed directly on the scratch and weighed down with a small polypropylene (PP) weight (sterilized Nunc™ Cap Mats, ThermoScientific, Germany). Test specimens were incubated with the cells for 4, 24, 48, and 144 hours in complete DMEM (contains 1% antibiotic-antimycotic solution and 10% FCS). Cells cultured in complete DMEM alone served as untreated controls. After the respective incubation periods, dressing samples were removed and cells were stained with hematoxylin and eosin for evaluation. Microscopic evaluation was carried out using the Axio Scope A.1 microscope (Carl Zeiss GmbH, Germany) and images were obtained with the digital camera AxioCam MRc (Carl Zeiss GmbH, Germany). Scratch width was determined using the ScratchCalculator (Mario Kanka, Jena). The progression of ‘healing’ of the scratch wound, designated as wound area in [%], was calculated in percent comparing scratch width at each time point to the initial scratch width at 4 hours. Furthermore, the slope was calculated by linear regression (Microsoft ® Excel 2000) for comparison of the healing progression over time.