Wound healing activities of ortosiphon stamineus leaves and protein extracts on human skin fibroblast in diabetic microenvironment

Abstract

Orthosiphon stamineus, also known as cat whiskers, is a well-known plant in herbal remedies and traditional medicine. The ethanolic leaves and transketolase protein extracts of the plant may be a possible diabetic wound healing. It has previously been shown that this plant extract has potential antihyperglycemic and wound healing effects in in vivo on diabetic induced rats under normal and hyperglycemic conditions. However, the effects of the O. stamineus leave extracts on human skin fibroblast (HSF 1184) cells under hyperglycemic microenvironment and protein extracts on HSF 1184 cells under normal and hyperglycaemic microenvironments are still unknown. The aim of this research is to study the cell viability and wound healing activity of the O. stamineus ethanolic and protein extract on HSF 1184 cells under both microenvironments. The cytotoxicity and migration assay were conducted in a various selected range of ethanolic and protein extract concentrations against HSF 1184 cells using MTT assay and wound healing assay. The cytotoxicity assay was evaluated after 72 hours of treatment under different concentrations of O. stamineus ethanolic (7.81, 15.63, 31.25, 62.5, 125, 250, 500 and 1000 ^g/mL) and protein extracts (1.25, 2.5, 5, 10 and 20 ng/mL) under both microenvironments. The results show that the highest cell proliferation activity at the concentration of O. stamineus ethanolic extract (7.81 ^g/mL) and protein extract (1.25 ng/mL), respectively. Conversely, the highest concentration of O. stamineus ethanolic (1000 ^g/mL) and protein extract (20 ng/mL) resulted in the lower cell proliferation activity under both microenvironments respectively. However, the O. stamineus ethanolic and protein extracts showed low cytotoxicity effects against the HSF 1184 cells under both microenvironments. The IC50 value obtained for the O. stamineus ethanolic extract under normal microenvironment was 151.36 ^g/mL and under hyperglycemic microenvironment was 141.26 ^g/mL respectively. Whereas, no IC50 values were obtained from the O. stamineus protein extracts. The wound healing assaywas determined based on the accelerated cell migration activity at 18, 20, 22 and 24 hours of incubation with various concentrations of O. stamineus ethanolic (40, 80 and 160 ^g/mL) and protein extracts (5, 10 and 20 ng/mL) under both microenvironments. Based on the results for the in vitro wound healing assay of O. stamineus ethanolic extracts, control has shown to exhibit higher cell migrating activity at 86.99% compared to the remaining concentration under normal microenvironment. On the other hand, O. stamineus ethanolic extracts at the concentration of 40 ^g/mL displayed rapid wound closure activity at 89.17% compared to control under the hyperglycemic microenvironment. Nevertheless, in vitro wound healing assay of O. stamineus protein extracts, treatment with 5 ng/mL has revealed a remarkable wound closure rate at 99.79% when compared to the control under normal microenvironment. While O. stamineus protein extracts at the concentration of 10 ng/mL showed enhanced wound closure effect at 99.83% when compared to control under hyperglycemic microenvironment. Collectively, these results suggested that O. stamineus ethanolic and protein extracts have potential in vitro in the acceleration of cell migration whichinfluences the traditional use of O. stamineus extracts in wound healing activity on HSF 1184 cells.