Effect of Hedyotis diffusa water extract on protecting human hepatocyte cells (LO₂) from H₂O₂-induced cytotoxicity

Context: Natural products are good sources of natural dietary antioxidants that are believed to protect the body against hepatotoxic effect induced by oxidative stress. Hedyotis diffusa Willd (Rubiaceae) (HDW) is a traditional Chinese medicinal herb that has been shown to possess a variety of antioxidant properties. Objective: The present study examines and explains the cell protective property of HDW water extract (WEHDW). Materials and methods: 2,2-Diphenyl-1-(2,4,6-trinitrophenyl) hydrazyl (DPPH) assay was used to measure the free radical scavenging property of WEHDW (0.001–10 mg/mL). The protective effect of WEHDW (0.3–10 mg/mL 2 h pretreatment) against hydrogen peroxide (H₂O_2, 200 μM for 6 h) induced cytotoxicity in human hepatic cells, LO₂, was evaluated using cell viability assay and nuclear staining. The molecular pathway of WEHDW's effect was investigated by using Western blot assay. Results: WEHDW had a 50% scavenging concentration (SC₅₀) at 0.153 mg/mL in the DPPH assay. Exposure of LO₂ cells to H₂O₂ resulted in apoptosis which could be markedly attenuated by pre-treating WEHDW in a concentration-dependent manner (0.5, 1, 3, 5, or 10 mg/mL) (all with p < 0.001, versus control). Moreover, Hoechst (nuclear) staining showed that 1 mg/mL WEHDW could protect LO₂ cells by attenuating apoptotic cell death mediated by H₂O₂. It was found that WEHDW reversed H₂O₂-induced activation of MEK/ERK pathway and H₂O₂-induced inhibition of P13-K/AKT/GSK3β pathway in LO₂ cells. Discussion and conclusion: WEHDW may help to improve the antioxidant defense system, resulting in prevention of oxidative stress-related fatty liver diseases.