Induction of apoptosis by hydroxydibenzoylmethane through coordinative modulation of cyclin D3, Bcl-X(L), and Bax,release of cytochrome c,and sequential activation of caspases in human colorectal carcinoma cells

DBM (dibenzoylmethane) is a minor constituent of licorice that has antimutagenic activity. However, its other biological activities are not well-known. The structurally related beta-diketones hydroxydibenzoylmethane (HDB) and hydroxymethyldibenzoylmethane (HMDB) were able to induce apoptosis in colorectal carcinoma COLO 205 cells. Thus, the effect of structurally related beta-diketones on cell viability, DNA fragmentation, and caspase activity was assessed. The potency of these compounds on these features of apoptosis were in the order of HDB > HMDB > DBM in colorectal carcinoma COLO 205 cells. Here, we found that HDB-induced apoptotic cell death was accompanied by upregulation of cyclin D3, Bax, and p21 and down-regulation of Bcl-X(L), while HDB had no effect on the levels of Bcl-2 and Bad protein. These results indicate that HDB allows caspase-activated deoxyribonuclease to enter the nucleus and degrade chromosomal DNA and induces DFF-45 degradation. It is suggested that HDB-induced apoptosis is triggered by the release of cytochrome c into cytosol, procaspase-9 processing, activation of caspase-3 and caspase-2, degradation of PARP, and DNA fragmentation caused by the caspase-activated deoxyribonuclease through the digestion of DFF-45. The induction of apoptosis by HDB may provide a pivotal mechanism for its cancer chemopreventive action.     

Induction of apoptosis by shikonin through coordinative modulation of the Bcl-2 family, p27, and p53, release of cytochrome c, and sequential activation of caspases in human colorectal carcinoma cells

Shikonin is a main constituent of the roots of Lithospermum erythrorhizon that has antimutagenic activity. However, its other biological activities are not well-known. Shikonin displayed a strong inhibitory effect against human colorectal carcinoma COLO 205 cells and human leukemia HL-60 cells, with estimated IC(50) values of 3.12 and 5.5 microM, respectively, but were less effective against human colorectal carcinoma HT-29 cells, with an estimated IC(50) value of 14.8 microM. Induce apoptosis was confirmed in COLO 205 cells by DNA fragmentation and the appearance of a sub-G1 DNA peak, which were preceded by loss of mitochondrial membrane potential, reactive oxygen species (ROS) generation, cytochrome c release, and subsequent induction of pro-caspase-9 and -3 processing. Cleavages of poly(ADP-ribose) polymerase (PARP) and DNA fragmentation factor (DFF-45) were accompanied by activation of caspase-9 and -3 triggered by shikonin in COLO 205 cells. Here, we found that shikonin-induced apoptotic cell death was accompanied by upregulation of p27, p53, and Bad and down-regulation of Bcl-2 and Bcl-X(L), while shikonin had little effect on the levels of Bax protein. Taken together, we suggested that shikonin-induced apoptosis is triggered by the release of cytochrome c into cytosol, procaspase-9 processing, activation of caspase-3, degradation of PARP, and DNA fragmentation caused by the caspase-activated deoxyribonuclease through the digestion of DFF-45. The induction of apoptosis by shikonin may provide a pivotal mechanism for its cancer chemopreventive action.     

Induction of apoptosis by the oolong tea polyphenol theasinensin A through cytochrome c release and activation of caspase-9 and caspase-3 in human U937 cells

This study examined the growth inhibitory effects of theasinensin A (from oolong tea) and black tea polyphenols, including theaflavin (TF-1), a mixture (TF-2) of theaflavin-3-gallate (TF-2a) and theaflavin-3'-gallate (TF-2b), and theaflavin-3,3'-digallate (TF-3) in human cancer cells. Theasinensin A, TF-1, and TF-2 displayed strong growth inhibitory effects against human histolytic lymphoma U937, with estimated IC50 values of 12 microM, but were less effective against human acute T cell leukemia Jurkat, whereas TF-3 and (-)-epigallocatechin-3-gallate (EGCG) had lower activities. The molecular mechanisms of tea polyphenol-induced apoptosis as determined by annexin V apoptosis assay, DNA fragmentation, and caspase activation were further investigated. Loss of membrane potential and reactive oxygen species (ROS) generation were also detected by flow cytometry. Treatment with tea polyphenols caused rapid induction of caspase-3, but not caspase-1, activity and stimulated proteolytic cleavage of poly(ADP-ribose) polymerase (PARP). Pretreatment with a potent caspase-3 inhibitor, Z-Asp-Glu-Val-Asp-fluoromethyl ketone, inhibited theasinensin A induced DNA fragmentation. Furthermore, it was found that theasinensin A induced loss of mitochondrial transmembrane potential, elevation of ROS production, release of mitochondrial cytochrome c into the cytosol, and subsequent induction of caspase-9 activity. These results indicate that theasinensin A allows caspase-activated deoxyribonuclease to enter the nucleus and degrade chromosomal DNA and induces DFF-45 (DNA fragmentation factor) degradation. The results suggest that induction of apoptosis by theasinensin A may provide a pivotal mechanism for their cancer chemopreventive function.     

Acacetin induces apoptosis in human gastric carcinoma cells accompanied by activation of caspase cascades and production of reactive oxygen species

Acacetin (5,7-dihydrocy-4'-methoxy flavone), which is a flavonoid compound, possesses anti-peroxidative and anti-inflammatory effects. The effects of acacetin on cell viability in human gastric carcinoma AGS cells were investigated. This study demonstrated that acacetin was able to inhibit cell proliferation and induce apoptosis in a concentration- and time-dependent manner. Acacetin-induced cell death was characterized with changes in nuclear morphology, DNA fragmentation, and cell morphology. The molecular mechanism of acacetin-induced apoptosis was also investigated. Treatment with acacetin caused induction of caspase-3 activity in a time-dependent manner, but not caspase-1 activity, and induced the degradation of DNA fragmentation factor (DFF-45) and poly(ADP-riobse) polymerase. Cell death was completely prevented by a pancaspase inhibitor, Z-Val-Ala-Asp-fluoromethyl ketone. Furthermore, treatment with acacetin caused a rapid loss of mitochondrial transmembrane potential, stimulation of reactive oxygen species (ROS), release of mitochondrial cytochrome c into cytosol, and subsequent induction of procaspase-9 processing. Antioxidants such as N-acetylcysteine and catalase, but not superoxide dismutase, allopurinol, or pyrrolidine dithiocarbamate, significantly inhibited acacetin-induced cell death. In addition, it was found that acacetin promoted the up-regulation of Fas and FasL prior to the processing and activation of pro-caspase-8 and cleavage of Bid, suggesting the involvement of a Fas-mediated pathway in acacetin-induced apoptosis. On the other hand, the results showed that acacetin-induced apoptosis was accompanied by up-regulation of Bax and p53, down-regulation of Bcl-2, and cleavage of Bad. Taken together, these results suggest that ROS production and a certain intimate link might exist between receptor- and mitochondria-mediated death signalings that committed to acacetin-induced apoptosis in AGS cells. The induction of apoptosis by acacetin may provide a pivotal mechanism for its cancer chemopreventive action.     

Induction of apoptosis by vitamin D2, ergocalciferol, via reactive oxygen species generation, glutathione depletion, and caspase activation in human leukemia Cells

This study demonstrated that ergocalciferol was able to inhibit leukemia cell growth in a concentration-dependent manner. Exploration of the acting mechanisms involved this event revealed that ergocalciferol induced DNA fragmentation and increased sub-G1 DNA contents in HL-60 cells, both of which are hallmarks of apoptosis. Analysis of the integrity of mitochondria demonstrated that ergocalciferol caused loss of mitochondrial membrane potential with release cytochrome c to cytosol, generation of reactive oxygen species (ROS), and depletion of glutathione (GSH), suggesting that ergocalciferol may induce apoptosis in HL-60 cells through a ROS-dependent pathway. Further results show that caspases-2, -3, -6, and -9 were all activated by ergocalciferol, together with cleavage of the downstream caspase-3 targets, DNA fragmentation factor (DFF-45), and poly(ADP-ribose) polymerase. In addition, ergocalciferol led to the increase in pro-apoptotic factor Bax accompanied with the decrease in anti-apoptotic member Mcl-1, and the reduced Mcl-1 to Bax ratio may be a critical event concerning mitochondrial decay by ergocalciferol. Furthermore, ergocalciferol also led to induction of Fas death receptor closely linked to caspase-2 activation, suggesting the involvement of a Fas-mediated pathway in ergocalciferol-induced apoptosis. Totally, these findings suggest that ergocalciferol causes HL-60 apoptosis via a modulation of mitochondria involving ROS production, GSH depletion, caspase activation, and Fas induction. On the basis of anticancer activity of ergocalciferol, it may be feasible to develop chemopreventive agents from edible mushrooms or hop.     

Mechanisms of cancer chemoprevention by hop bitter acids (beer aroma) through induction of apoptosis mediated by Fas and caspase cascades

The bitter acids of hops (Humulus lupulus L.) mainly consist of alpha-acids, beta-acids, and their oxidation products that contribute the unique aroma of the beer beverage. Hop bitter acids displayed a strong growth inhibitory effect against human leukemia HL-60 cells, with an estimated IC(50) value of 8.67 microg/mL, but were less effective against human histolytic lymphoma U937 cells. Induction of apoptosis was confirmed in HL-60 cells by DNA fragmentation and the appearance of a sub-G1 DNA peak, which were preceded by dissipation of mitochondrial membrane potential, cytochrome c release, and subsequent induction of pro-caspase-9 and -3 processing. Cleavages of PARP and DFF-45 were accompanied with activation of caspase-9 and -3 triggered by hop bitter acids in HL-60 cells. The change in the expression of Bcl-2, Bcl-X(L), and Bax in response to hop bitter acids was studied, and the Bcl-2 protein level slightly decreased; however, the Bcl-X(L) protein level was obviously decreased, whereas the Bax protein level was dramatically increased, indicating that the control of Bcl-2 family proteins by hop bitter acids might participate in the disruption of mitochondrial integrity. In addition, the results showed that hop bitter acids promoted the up-regulation of Fas and FasL prior to the processing and activation of pro-caspase-8 and cleavage of Bid, suggesting the involvement of a Fas-mediated pathway in hop bitter acids-induced cells. Taken together, these findings suggest that a certain intimate link might exist between receptor- and mitochondria-mediated death signalings that committed to cell death induced by hop bitter acids. The induction of apoptosis by hop bitter acids may offer a pivotal mechanism for their chemopreventive action.     

Effect of caffeic acid phenethyl ester, an antioxidant from propolis, on inducing apoptosis in human leukemic HL-60 cells.

Caffeic acid phenethyl ester (CAPE) is an active component isolated from propolis. The aim of this study was to investigate the mechanism of CAPE-induced apoptosis in human leukemic HL-60 cells. It was found that CAPE entered HL-60 cells very quickly and then inhibited their survival in a concentration- and time-dependent manner. CAPE induced characteristic DNA fragmentation and morphological changes typical of apoptosis in these cells. Estimation of the apoptotic percentage showed a time-dependent increase after CAPE (6 microg/mL) treatment (up to 66.7 +/- 2.0% at 72 h). Treatment with CAPE caused rapid activation of caspase-3 after 4 h, down-regulation of Bcl-2 expression after 6 h, and up-regulation of Bax expression after 16 h. These results suggest that CAPE is a potent apoptosis-inducing agent; its action is accompanied by activation of caspase-3, down-regulation of Bcl-2, and up-regulation of Bax in human leukemic HL-60 cells.     

Induction of apoptosis by the Anthocyanidins through regulation of Bcl-2 gene and activation of c-Jun N-terminal kinase cascade in hepatoma cells

Anthocyanidins that are reddish pigments widely distributed in fruit and vegetables have been reported to possess antioxidant and anticancer activities. To understand the molecular basis of the putative anticancer activity of anthocyanidins, we investigated the antiproliferation effects of anthocyanidins in human hepatoma cell lines. Delphinidin, cyanidin, and malvidin exhibited strong growth inhibitory effects against human hepatoma HepG(2), but were less effective against Hep3B. According to the appearance of the caspase-3 fragments and stimulated proteolytic cleavage of poly (ADP-ribose) polymerase (PARP) in time-dependent studies, delphinidin induced apoptotic cell death characterized by internucleosomal DNA fragmentation and caused a rapid induction of caspase-3 activity. RT-PCR and Western blot data revealed that delphinidin stimulated an increase in the c-Jun and JNK phosphorylation expression at mRNA and protein levels, respectively. Moreover, delphinidin-induced apoptotic cell death was accompanied by up-regulation of Bax and down-regulation of Bcl-2 protein. Dephinidin-induced DNA fragmentation was blocked by N-acetyl-l-cysteine and catalase, suggesting that the death signaling was triggered by oxidative stress. Our experiments provide evidence that delphinidin is an effective apoptosis inducer in HepG(2) cells through regulation of Bcl-2 family moleculars and activation of c-Jun N-terminal kinase cascade. The results suggest that induction of apoptosis by anthocyanidins is a pivotal mechanism of their cancer chemopreventive functions.     

Blazeispirol A from Agaricus blazei fermentation product induces cell death in human hepatoma Hep 3B cells through caspase-dependent and caspase-independent pathways

Currently, liver cancer is a leading cause of cancer-related death in the world. Hepatocellular carcinoma is the most common type of liver cancer. Previously, it was reported that blazeispirol A (BA) is the most active antihepatoma compound in an ethanolic extract of Agaricus blazei fermentation product. The aim of this study was to understand the antihepatoma mechanism of BA in human liver cancer Hep 3B cells. The results showed that BA inhibited the growth of Hep 3B cells and increased the percentage of cells in sub-G1 phase in a concentration- and time-dependent manner. In addition, BA treatment resulted in DNA fragmentation, caspase-9 and caspase-3 activations, poly(ADP-ribose)polymerase (PARP) degradation, down-regulation of Bcl-2 and Bcl-xL expressions, up-regulation of Bax expression, and disruption of the mitochondrial membrane potential (MMP) in Hep 3B cells. Furthermore, z-VAD-fmk, a caspase inhibitor, did not enhance the viability of BA-treated Hep 3B cells, and BA induced the release of HtrA2/Omi and apoptosis-inducing factor (AIF) from mitochondria into the cytosol. These findings suggested that BA with novel chemopreventive and chemotherapeutic potentials causes both caspase-dependent and caspase-independent cell death in Hep 3B cells.     

苏云金芽胞杆菌Bt9875杀虫晶体蛋白可诱导Caspase和Bcl-2家族参与调控HL-60细胞凋亡

研究Caspase家族与Bcl-2家族参与调控苏云金芽胞杆菌(Bacillus thuringiensis,Bt)Bt9875杀虫晶体蛋白对人急性髓细胞性白血病细胞HL-60的影响.本实验采用MTT法检测了杀虫晶体蛋白诱导HL-60细胞凋亡后的Caspase家族的活性和Caspase凋亡酶抑制剂对杀虫晶体蛋白诱导HL-60细胞凋亡的影响;采用Western blot检测了杀虫晶体蛋白诱导多聚ADP-核糖聚合酶(PARP)降解、Bcl-2/Bax调控和细胞色素C的释放.研究结果表明,杀虫晶体蛋白作用HL-60细胞后,激活了Caspase-3、Caspase-8和Caspase-9,在48 h内Caspase家族抑制剂(Z-VAD-FMK)、Caspase-3抑制剂(z-DEVD-FMK)和Caspase-9抑制剂(Z-LEHD-FMK)均可显著抑制杀虫晶体蛋白诱导的细胞凋亡;杀虫晶体蛋白可明显上调促凋亡蛋白Bax的表达,同时下调抗凋亡蛋白BCl-2的表达,并观察到胞浆中细胞色素C的释放.初步证明了Bt9875杀虫晶体蛋白诱导的HL-60细胞凋亡是由Caspase家族和Bcl-2家族共同调控的,线粒体途径在诱导细胞凋亡过程中起着重要作用.     

Induction of apoptosis by 1-(2-hydroxy-5-methylphenyl)-3-phenyl-1,3-propanedione through reactive oxygen species production, GADD153 expression, and caspases activation in human epidermoid carcinoma cells

This study examined the growth inhibitory effects of the structurally related beta-diketones compounds in human cancer cells. Here, we report that 1-(2-hydroxy-5-methylphenyl)-3-phenyl-1,3-propanedione (HMDB) induces growth inhibition of human cancer cells and induction of apoptosis in A431 cells through modulation of mitochondrial functions regulated by reactive oxygen species (ROS). ROS generation occurs in the early stages of HMDB-induced apoptosis, preceding cytochrome c release, caspase activation, and DNA fragmentation. The changes occurred after single breaks in DNA were detected, suggesting that HMDB induced irreparable DNA damage, which in turn triggered the process of apoptosis. Up-regulation of Bad and p21; down-regulation of Bcl-2, Bcl-XL, Bid, p53, and fatty acid synthase; and cleavage of Bax were found in HMDB-treated A431 cells. Glutathione and N-acetylcysteine (NAC) suppress HMDB-induced apoptosis. HMDB markedly enhanced growth arrest DNA damage inducible gene 153 (GADD153) mRNA and protein in a time- and concentration-dependent manner. NAC prevented up-regulation of GADD153 mRNA expression caused by HMDB. These findings suggest that HMDB creates an oxidative cellular environment that induces DNA damage and GADD153 gene activation, which in turn helps trigger apoptosis in A431 cells.     

A mixture of trans, trans conjugated linoleic acid induces apoptosis in MCF-7 human breast cancer cells with reciprocal expression of Bax and Bcl-2

The growth inhibitory effect of a mixture of trans, trans conjugated linoleic acid isomers (t, t CLA) was investigated in a human breast cancer cell line, MCF-7, with references to c9, t11 CLA, t10, c12 CLA, and linoleic acid. The t, t CLA treatment effectively induced a cytotoxic effect in a time-dependent (0-6 days) and concentration-dependent (0-40 microM) manner, as compared to the reference and control treatments. The apoptotic parameters were measured on cells treated with 40 microM t, t CLA for 4 days. The occurrence of the characteristic morphological changes and DNA fragmentation confirmed apoptosis. The t, t CLA treatment led to an increase in the level of p53 tumor suppressor protein and Bax protein, but suppressed the expression of Bcl-2 protein. In addition, cytochrome c was released from the mitochondria into the cytosol, and the activation of caspase-3 led to the cleavage of poly(ADP-ribose) polymerase (PARP). Moreover, the composition of the linoleic and arachidonic acids was decreased in cellular membranes. These findings suggest that incorporation of t, t CLA in the membrane induces a mitochondria-mediated apoptosis that can enhance the antiproliferative effect of t, t CLA in MCF-7 cells.     

Growth inhibition and induction of apoptosis in NB4 promyelocytic leukemia cells by trypsin inhibitor from sweet potato storage roots

The objective of this study was to investigate the antiproliferative effect and the mechanism of trypsin inhibitor (TI) from sweet potato [Ipomoea batatas (L.) Lam. 'Tainong 57'] storage roots on NB4 promyelocytic leukemia cells. The results showed that TI inhibited cellular growth of NB4 promyelocytic leukemia cells in a time-dependent and dose-dependent manner, and treatment for 72 h induced a marked inhibition of cellular growth, showing an IC50 of 57.1 +/- 8.26 microg/mL. TI caused cell cycle arrest at the G1 phase as determined by flow cytometric analysis and apoptosis as shown by DNA laddering. TI-induced cell apoptosis involved p53, Bcl-2, Bax, and cytochrome c protein in NB4 cells. P53 and Bax proteins were accumulated, and antiapoptotic molecule Bcl-2 was decreased in the tested cells in a time-dependent manner during TI treatment. TI also induced a substantial release of cytochrome c from the mitochondria into the cytosol. Hence, TI induced apoptosis in NB4 cells through a mitochondria-dependent pathway, which was associated with the activation of caspase-3 and -8. These results demonstrated that TI induces NB4 cell apoptosis through the inhibition of cell growth and the activation of the pathway of caspase-3 and -8 cascades.     

Lucidenic acid B induces apoptosis in human leukemia cells via a mitochondria-mediated pathway

Ganoderma lucidum is known as a medicinal mushroom used in traditional Chinese medicine. In the present study, the effect of lucidenic acids (A, B, C, and N) isolated from a new G. lucidum (YK-02) on induction of cell apoptosis and the apoptotic pathway in HL-60 cells were investigated. The results demonstrated that lucidenic acids decreased cell population growth of HL-60 cells, assessed with the MTT assay. The cell cycle assay indicated that treatment of HL-60 cells with lucidenic acid A, C, and N caused cell cycle arrest in the G 1 phase. Lucidenic acid B (LAB) did not affect the cell cycle profile; however, it increased the number of early and late apoptotic cells but not necrotic cells. Treatment of HL-60 cells with LAB caused loss of mitochondria membrane potential. Moreover, the ratio of expression levels of pro- and antiapoptotic Bcl-2 family members was changed by LAB treatment. LAB-induced apoptosis involved release of mitochondria cytochrome c and subsequently induced the activation of caspase-9 and caspase-3, which were followed by cleavage of poly(ADP-ribose) polymerase (PARP). Pretreatment with a general caspase-9 inhibitor (Z-LEHD-FMK) and caspase-3 inhibitor (Z-DEVD-FMK) prevented LAB from inhibiting cell viability in HL-60 cells. Our finding may be critical to the chemopreventive potential of lucidenic acid B.     

Antcin B and its ester derivative from Antrodia camphorata induce apoptosis in hepatocellular carcinoma cells involves enhancing oxidative stress coincident with activation of intrinsic and extrinsic apoptotic pathway

The triterpenoids methylantcinate B (MAB) and antcin B (AB), isolated from the medicinal mushroom Antrodia camphorata , have been identified as strong cytotoxic agents against various type of cancer cells; however, the mechanisms of MAB- and AB-induced cytotoxicity have not been adequately explored. This study investigated the roles of caspase cascades, reactive oxygen species (ROS), DNA damage, mitochondrial disruption, and Bax and Bcl-2 proteins in MAB- and AB-induced apoptosis of hepatocellular carcinoma (HCC) HepG2 cells. Here, we showed that MAB and AB induced apoptosis in HepG2 cells, as characterized by increased DNA fragmentation, cleavage of PARP, sub-G1 population, chromatin condensation, loss of mitochondrial membrane potential, and release of cytochrome c. Increasing the levels of caspase-2, -3, -8, and -9 activities was involved in MAB- and AB-induced apoptosis, and they could be attenuated by inhibitors of specific caspases, indicating that MAB and AB triggered the caspase-dependent apoptotic pathway. Additionally, the enhanced apoptotic effect correlates with high expression of Fas, Fas ligand, as well as Bax and decreased protein levels of Bcl-(XL) and Bcl-2, suggesting that both the extrinsic and intrinsic apoptosis pathways were involved in the apoptotic processes. Incubation of HepG2 cells with antioxidant enzymes superoxide dismutase and catalase and antioxidants N-acetylcysteine and ascorbic acid attenuated the ROS generation and apoptosis induced by MAB and AB, which indicate that ROS plays a pivotal role in cell death. NADPH oxidase activation was observed in MAB- and AB-stimulated HepG2 cells; however, inhibition of such activation by diphenylamine significantly blocked MAB- and AB-induced ROS production and increased cell viability. Taken together, our results provide the first evidence that triterpenoids MAB and AB induced a NADPH oxidase-provoked oxidative stress and extrinsic and intrinsic apoptosis as a critical mechanism of cause cell death in HCC cells.     

Pyrrolidine dithiocarbamate inhibition of luteolin-induced apoptosis through up-regulated phosphorylation of Akt and caspase-9 in human leukemia HL-60 cells

Previously, we observed that luteolin effectively inhibited cell growth and induced apoptosis in HL-60 cells. In that study, we also explored the modulatory effects and molecular mechanisms of pyrrolidine dithiocarbamate (PDTC) on the cytotoxicity of luteolin to HL-60 cells. In this study, we found that PDTC was able to inhibit luteolin-induced cell apoptosis in a dose-dependent manner. When HL-60 cells were treated with PDTC for 0.5 h before 60 microM luteolin treatment, the DNA ladder disappeared. Moreover, flow cytometry showed that PDTC had dose dependently decreased the percentage of apoptotic HL-60 cells and had not interfered with luteolin's ability to change the mitochondrial membrane potential or its ability to trigger the release of cytochrome c to cytosol. Detection by Western blotting, however, did show that PDTC had interfered with luteolin's ability to cleave poly(ADP-ribose)polymerase and DNA fragmentation of factor-45. Three hours after the PDTC-pretreated HL-60 cells were treated with 60 microM luteolin, the product cleaved from Akt started to appear. Therefore, not only was PDTC able to stop the apoptosis of HL-60 cells treated with luteolin, it was also found to increase phosphorylation of Akt and caspase-9. These results suggest that in the luteolin-induced apoptotic pathway, phosphorylation of procaspase-9 by survival signals might play an important role in the ultimate fate of HL-60 cells.     

Free radical scavenging activity and antiulcer activity of garcinol from Garcinia indica fruit rind

Garcinol, a polyisoprenylated benzophenone derivative, was purified from Garcinia indica fruit rind, and its free radical scavenging activity was studied using electron spin resonance (ESR) spectrometry. In the hypoxanthine/xanthine oxidase system, emulsified garcinol suppressed superoxide anion to almost the same extent as DL-alpha-tocopherol by weight. In the Fenton reaction system, garcinol also suppressed hydroxyl radical more strongly than DL-alpha-tocopherol. In the H(2)O(2)/NaOH/DMSO system, garcinol suppressed superoxide anion, hydroxyl radical, and methyl radical. It was thus confirmed that this derivative is a potent free radical scavenger and able to scavenge both hydrophilic and hydrophobic ones including reactive oxygen species. Orally administered garcinol prevented acute ulceration in rats induced by indomethacin and water immersion stress caused by radical formation. These results suggested garcinol might have potential as a free radical scavenger and clinical application as an antiulcer drug.     

The chalcone flavokawain B induces G2/M cell-cycle arrest and apoptosis in human oral carcinoma HSC-3 cells through the intracellular ROS generation and downregulation of the Akt/p38 MAPK signaling pathway

Chalcones have been described to represent cancer chemopreventive food components that are rich in fruits and vegetables. In this study, we examined the anti-oral cancer effect of flavokawain B (FKB), a naturally occurring chalcone isolated from Alpinia pricei (shell gingers), and revealed its molecular mechanism of action. Treatment of human oral carcinoma (HSC-3) cells with FKB (1.25-10 μg/mL; 4.4-35.2 μM) inhibited cell viability and caused G(2)/M arrest through reductions in cyclin A/B1, Cdc2, and Cdc25C levels. Moreover, FKB treatment resulted in the induction of apoptosis, which was associated with DNA fragmentation, mitochondria dysfunction, cytochrome c and AIF release, caspase-3 and caspase-9 activation, and Bcl-2/Bax dysregulation. Furthermore, increased Fas activity and procaspase-8, procaspase-4, and procaspase-12 cleavages were accompanied by death receptor and ER-stress, indicating the involvement of mitochondria, death-receptor, and ER-stress signaling pathways. FKB induces apoptosis through ROS generation as evidenced by the upregulation of oxidative-stress markers HO-1/Nrf2. This mechanism was further confirmed by the finding that the antioxidant N-acetylcysteine (NAC) significantly blocked ROS generation and consequently inhibited FKB-induced apoptosis. Moreover, FKB downregulated the phosphorylation of Akt and p38 MAPK, while their inhibitors LY294002 and SB203580, respectively, induced G(2)/M arrest and apoptosis. The profound reduction in cell number was observed in combination treatment with FKB and Akt/p38 MAPK inhibitors, indicating that the disruption of Akt and p38 MAPK cascades plays a functional role in FKB-induced G(2)/M arrest and apoptosis in HSC-3 cells.     

Effects of capsaicin on induction of apoptosis and inhibition of adipogenesis in 3T3-L1 cells

Currently, at the beginning of the 21st century, obesity has become the leading metabolic disease in the world. It is a serious health problem in industrialized countries. Previous research has suggested that decreased preadipocyte differentiation and proliferation and decreased lipogenesis are mechanisms to reduce obesity. In the present study, the effects of capsaicin on the induction of apoptosis and inhibition of lipid accumulation in 3T3-L1 preadipocytes and adipocytes were investigated. The results demonstrated that capsaicin decreased cell population growth of 3T3-L1 preadipocytes, assessed with the MTT assay. Flow cytometric analysis of 3T3-L1 preadipocytes exposed to capsaicin showed that apoptotic cells increased in a time- and dose-dependent manner. Treatment with capsaicin decreased the number of normal cells and increased the number of early apoptotic and late apoptotic cells in a dose-dependent manner. The treatment of cells with capsaicin caused the loss of mitochondria membrane potential (delta psi m). The induction of apoptosis in 3T3-L1 preadipocytes by capsaicin was mediated through the activation of caspase-3, Bax, and Bak, and then through the cleavage of PARP and the down-regulation of Bcl-2. Moreover, capsaicin significantly decreased the amount of intracellular triglycerides and glycerol-3-phosphate dehydrogenase (GPDH) activity in 3T3-L1 adipocytes. Capsaicin also inhibited the expression of PPARgamma, C/EBPalpha, and leptin, but induced up-regulation of adiponectin at the protein level. These results demonstrate that capsaicin efficiently induces apoptosis and inhibits adipogenesis in 3T3-L1 preadipocytes and adipocytes.