Numerous studies have shown that Hedyotis diffusa WILLD (H. diffusa) could promote apoptosis in several cancer cell lines. However, the cellular and molecular mechanisms responsible for this phenomenon are still poorly understood. The current study determines the role of Fas/FasL in THP-1 cell apoptosis induced by 2-hydroxy-3-methylanthraquinone from H. diffusa.THP-1 cells were treated with 2-hydroxy-3-methylanthraquinone from H. diffusa. The effect on the cell viability of THP-1 cells was evaluated using the trypan blue assay, and cell apoptosis was measured by Giemsa staining and flow cytometry. Protein expression was assayed using the Western blot method.Our results showed that 2-hydroxy-3-methylanthraquinone from H. diffusa induced THP-1 cell apoptosis in a time- and dose-dependent manner. Apoptosis was associated with a more prominent induction expression of Fas/FasL, DR4 and TRAIL in a time-dependent manner. Moreover, treatment of THP-1 cells with 2-hydroxy-3-methylanthraquinone from H. diffusa resulted in activation of caspase-8.For the first time, our study suggests that 2-hydroxy-3-methylanthraquinone from H. diffusa is able to enhance apoptosis of THP-1 cells, at least in part, through activation of Fas/FasL, DR4 and TRAIL. Moreover, triggering of caspase-8 activation mediated apoptotic induction.