Our results showed that compared to the control group, treatment of cells with 80 M tricetin dramatically increased dichlorofluorescein (DCF) fluorescence at 6 and 8 h after treatment (Physique 5A). apoptosis. Dichlorofluorescein (DCF) staining showed that intracellular reactive oxygen species (ROS) levels were higher in tricetin-treated HL-60 cells compared to the control group. Moreover, an ROS scavenger, honey, appears to have potent anti-inflammatory properties which may be responsible for its beneficial effects [6]. Recently, tricetin has garnered much attention in relation to its anticancer activities such as antiproliferative and antimetastatic activities in many solid tumor cell models including breast [7], liver [8], lung [9], bone [10], and brain [11] tumors. Although it is quite obvious that tricetin can inhibit the growth or metastasis of various solid tumor cells, the precise impact of tricetin on nonsolid tumors is still unclear. Apoptosis is an active process of endogenous programmed cell death. The recognized characteristics of apoptosis include morphologic changes such as condensation and fragmentation of nuclei, cell membrane shrinkage, and loosening of organelle positions in the cytoplasm. In addition to morphological changes, sophisticated molecular procedures and mechanisms are also involved. Apoptosis can be initiated either through a death receptor followed by caspase-8 and -10 activation or the mitochondrial pathway including caspase-9 [12]. One of the hallmarks of malignancy is the deregulation of apoptosis; thus increasing apoptosis in tumors is one of the best ways for anticancer brokers to treat all types of malignancy. Actually, there are several plant-derived anticancer brokers such as alkaloids, taxines, and podophyllotoxin already in clinical use [13]. The mitogen-activated protein kinase (MAPK) pathway is an important route that communicates extracellular signals in intracellular responses and was correlated with many physiological processes such as cell growth, differentiation, and apoptosis. In mammalian cells, you will find three well-characterized subfamilies of MAPKs: extracellular signal-regulated kinases (ERKs), c-Jun N-terminal kinases (JNKs), and p38 MAPKs [14]. JNK was reported to be phosphorylated/activated after exposure of cells to nerve-racking stimuli, such as irradiation and malignancy chemotherapeutics, and it plays an important role in chemotherapeutic drug-mediated apoptosis [15]. Recently, it was reported that a JNK-activation defect confers chemoresistance in solid tumors such as ovarian and liver cancers [16,17]. Notably, involvement of the JNK-activation defect in anthracycline-containing chemotherapy resistance was also characterized in AML, and JNK targeting might be a new therapeutic approach for AML [18]. Although it is usually entirely obvious about the anti-metastatic and anti-tumor growth effects of tricetin in Ethylmalonic acid various solid tumor cells, the exact impact of tricetin on nonsolid tumors is SOCS2 still unknown. This is the first study to determine the cell growth-inhibitory activity and molecular mechanisms of tricetin in different French-American-British (FAB) types of AML cells (THP-1, U937, HL-60, and MV4-11). Our results exhibited that tricetin suppressed proliferation of these four AML cell lines. We found that superoxide was overproduced in HL-60 AML cells during tricetin treatment, which initiated a signal leading to activation of JNK-mediated apoptosis. Moreover, a combination of tricetin and an ERK inhibitor may be a better strategy than tricetin alone for treating AML. This study should provide a scientific basis for the clinical use of tricetin to effectively inhibit AML. 2. Results 2.1. Tricetin Inhibited Proliferation of Human Acute Myeloid Leukemia (AML) Cells The chemical structure of tricetin is usually shown in Physique 1A. In this study, we first examined the effect of tricetin around the growth of human AML cell lines Ethylmalonic acid using the cell counting kit-8 (CCK-8) assay. After treating cells with tricetin for 24 h, the tricetin concentration dependently inhibited the proliferation of four AML cell lines which represent different FAB types (M2: HL-60 and M5: MV4-11, U937, and THP-1) (Physique 1B,C). Among these four AML cell lines, HL-60 cells were the most sensitive to tricetin treatment. Therefore, we selected HL-60 cells for subsequent experiments. We further analyzed the long-term antiproliferative potential of tricetin against HL-60 cells by trypan blue exclusion assay. As illustrated in Physique 1D, tricetin time- and concentration-dependently suppressed the growth of cultured HL-60 cells. Open in a separate window Physique 1 Tricetin treatment results in Ethylmalonic acid reduced cell viability of human acute myeloid leukemia (AML) cell lines. (A) The chemical structure of tricetin; (B,C) Four human AML cell lines (HL-60, U937, THP-1, and MV4-11) were treated with the vehicle (DMSO) or tricetin (0~160 M) in serum-containing medium for 24 h; (D) HL-60 cells were treated with different concentrations.Consistent results from previous studies also indicated that an ERK inhibitor enhances docetaxel- and DAPT-induced apoptosis in androgen-independent prostate malignancy and gastric malignancy, respectively [41,42]. and JNK by their specific inhibitors respectively promoted and abolished tricetin-induced cell apoptosis. Dichlorofluorescein (DCF) staining showed that intracellular reactive oxygen species (ROS) levels were higher in tricetin-treated HL-60 cells compared to the control group. Moreover, an ROS scavenger, honey, appears to have potent anti-inflammatory properties which may be responsible for its beneficial effects [6]. Recently, tricetin has garnered much attention in relation to its anticancer activities such as antiproliferative and antimetastatic activities in many solid tumor cell models including breast [7], liver [8], lung [9], bone [10], and brain [11] tumors. Although it is quite obvious that tricetin can inhibit the growth or metastasis of various solid tumor cells, the precise impact of tricetin on nonsolid tumors is still unclear. Apoptosis is an active process of endogenous programmed cell death. The identified characteristics of apoptosis include morphologic changes such as condensation and fragmentation of nuclei, cell membrane shrinkage, and loosening of organelle positions in the cytoplasm. In addition to morphological changes, sophisticated molecular procedures and mechanisms are also involved. Apoptosis can be initiated either through a death receptor followed by caspase-8 and -10 activation or the mitochondrial pathway including caspase-9 [12]. One of the hallmarks of malignancy is the deregulation of apoptosis; thus increasing apoptosis in tumors is one of the best ways for anticancer brokers to treat all types of malignancy. Actually, there are several plant-derived anticancer brokers such as alkaloids, taxines, and podophyllotoxin already in clinical use [13]. The mitogen-activated protein kinase (MAPK) pathway is an important route that communicates extracellular signals in intracellular responses and was correlated with many physiological processes such as cell growth, differentiation, and apoptosis. In mammalian cells, you will find three well-characterized subfamilies of MAPKs: extracellular signal-regulated kinases (ERKs), c-Jun N-terminal kinases (JNKs), and p38 MAPKs [14]. JNK was reported to be phosphorylated/activated after exposure of cells to nerve-racking stimuli, such as irradiation and malignancy chemotherapeutics, and it plays an important role in chemotherapeutic drug-mediated apoptosis [15]. Recently, it was reported that a JNK-activation defect confers chemoresistance in solid tumors such as ovarian and liver cancers [16,17]. Notably, involvement of the JNK-activation defect in anthracycline-containing chemotherapy resistance was also characterized in AML, and JNK targeting might be a new therapeutic approach for AML [18]. Although it is usually entirely obvious about the anti-metastatic and anti-tumor growth effects of tricetin in various Ethylmalonic acid solid tumor cells, the exact impact of tricetin on nonsolid tumors is still unknown. This is the first study to determine the cell growth-inhibitory activity and molecular mechanisms of tricetin in different French-American-British (FAB) types of AML cells (THP-1, U937, HL-60, and MV4-11). Our results exhibited that tricetin suppressed proliferation of these four AML cell lines. We found that superoxide was overproduced in HL-60 AML cells during tricetin treatment, which initiated a signal leading to activation of JNK-mediated apoptosis. Moreover, a combination of tricetin and an ERK inhibitor may be a better strategy than tricetin alone for treating AML. This study should provide a scientific basis for the clinical use of tricetin to effectively inhibit AML. 2. Results 2.1. Tricetin Inhibited Proliferation of Human Acute Myeloid Leukemia (AML) Cells The chemical structure of tricetin is usually shown in Physique 1A. In this study, we first examined the effect of tricetin around the growth of human AML cell lines using the cell counting kit-8 (CCK-8) assay. After treating.