Extracts of Australian native stingless bee (Tetragonula carbonaria) cerumen modulate dermal fibroblast proliferation and myofibroblast differentiation in vitro

Introduction. Cerumen is a resinous, plant-derived product of stingless bees. We previously identified extracts and fractions of Australian native stingless bee (Tetragonula carbonaria) cerumen with anti-oxidant and anti-inflammatory activities in vitro; however, their wound-healing potential are currently unknown. Aims. To examine the effects of T. carbonaria cerumen extracts and active fractions on the proliferation of human fibroblasts obtained from normal dermis (NF) and chronic wounds (CWF), and on transforming growth factor (TGF)-1-mediated myofibroblast differentiation. Methods. T. carbonaria cerumen collected from hives in South-East Queensland was partitioned into methanol and methanol-water extracts. Two active fractions (Fractions 1 and 9) were obtained from the methanol-water extract by preparative reversed-phase HPLC fractionation. Cultured NFs and CWFs were incubated in the absence or presence of cerumen extracts and fractions (0.3-20 μg/mL). MTT dye-reduction assays measured cellular proliferation over 120 h. The effects of extracts and fractions on myofibroblast differentiation were also investigated in TGF-1-stimulated NFs (10 ng/mL; 72 h), by examining -smooth muscle actin (SMA) immunoreactivity. Results. Increased proliferative activity was observed in NFs exposed to 5 μg/mL methanol extract (129.9±11.2%; 120 h), 5 μg/mL methanol-water extract (124.0±15.8%; 24 h), 1 μg/mL Fraction 1 (131.1±17.7%; 120 h) and 3 μg/mL Fraction 9 (214.6±26.4%; 120 h) (P7<0.05). Proliferation was also enhanced in CWFs treated with 5 μg/mL methanol extract (134.6±10.0%; 120 h) and 3 μg/mL Fraction 9 (134.8±5.7%; 120 h) (P