Wound healing from the outback: novel wound healing therapeutics from native Australian plants

Introduction Chronic wounds are an area of major concern. The on-going and in-direct costs are substantial, reaching far beyond the costs of the hospitalization and associated care. As a result, pharmacological therapies have been developed to address treatment insufficiencies, however, the availability of drugs capable of promoting the wound repair process still remain limited. The wound healing properties of various herbal plants is well recognised amongst indigenous Australians. Hence, based on traditional accounts, we evaluated the wound healing potential of two Australian native plants. Methods Bioactive compounds were methanol extracted from dried plant leaves that were commercially sourced. Primary keratinocyte (Kc) and fibroblast (Fib) cells (denoted as Kc269, Kc274, Kc275, Kc276 and Fib274) obtained from surgical discarded tissue were cultured in 48-well plates and incubated (37⁰C, 5% CO2) overnight. The growth media was discarded and replaced with fresh growth media plus various concentrations (15.12 µg/mL, 31.25 µg/mL, 62.5 µg/mL, 125 µg/mL, 250 µg/mL and 500 µg/mL) of the plant extracts. Cellular responses were measured using the alamarBlue® assay and the CyQUANT® assay. Plant extracts in the aqueous phase were prepared by boiling whole leaves in water and taking aqueous phase samples at various (1, 2 , 5 minutes boiling) time points. Plant leaves were either added before the water was boiled (cold boiled) or after the water was boiled (hot boiled). The final concentrations of the aqueous plant extracts were 3.3 ng/mL (± 0.3 ng/mL) per sample. The antimicrobial properties of the plant extracts were tested using the well diffusion assay method against Staphylococcus aureus, Klebsiella pnuemoniae and methicillin resistant S. aureus and Bacillus cereus. Results Assay results from the almarBlue® and CYQUANT® assays indicated that extracts from both native plants at various time points (0, 24 and 48 hours) and concentrations (31.25 mg/mL, 62.5 mg/mL, and 125 mg/mL) were significantly higher (n=3, p=0.03 for Kc269, p=0.04 for Kc274, p=0.02 for Fib274, p=0.04 for Kc275 and p=0.001 for Kc276) compared with the untreated controls. Neither plant extract demonstrated cytotoxic effects. Significant antimicrobial activity against methicillin resistant Staphylococcus aureus (p=0.0009 for hot boiled plant A, n=2, p=0.034 for cold boiled plant A, n=2) K. pnuemoniae (p=0.0009 for hot boiled plant A, n=2, p=0.002 for cold boiled plant A, n=2) and B. cereus (p=0.0009 for hot boiled plant A, n=2, p=0.003 for cold boiled plant A, n=2) was observed at concentrations of 3.2 ng/mL for plant A and 3.4 ng/mL for plant B. Conclusion Both native plants contain bioactive compounds that increase cellular metabolic rates and total nucleic acid content. Neither plant was shown to be cytotoxic. Furthermore, both exhibited significant antimicrobial activity.