Oroxylin A induced apoptosis of human hepatocellular carcinoma cell line HepG2 was involved in its antitumor activity

We previously reported that wogonin, a flavonoid compound, was a potent apoptosis inducer of human hepatoma SMMC-7721 cells and murine sarcoma S180 cells. In the present study, the effect of oroxylin A, one wogonin structurally related flavonoid isolated from Scutellariae radix, on human hepatocellular carcinoma cell line HepG2 was examined and molecular mechanisms were also investigated. Oroxylin A inhibited HepG2 cell proliferation in a concentration- and time-dependent manner measured by MTT-assay. Treatment with an apoptosis-inducing concentration of oroxylin A caused typical morphological changes and apoptotic blebbing in HepG2 cells. DNA fragmentation assay was used to examine later apoptosis induced by oroxylin A. FACScan analysis revealed a dramatic increase in the number of apoptotic and G(2)/M phase arrest cells after oroxylin A treatment. The pro-apoptotic activity of oroxylin A was attributed to its ability to modulate the concerted expression of Bcl-2, Bax, and pro-caspase-3 proteins. The expression of Bcl-2 protein and pro-caspase-3 protein was dramatically decreased after treatment with oroxylin A. These results demonstrated that oroxylin A could effectively induce programmed cell death and suggested that it could be a promising antitumor drug.     

Role of TRPM2 in H(2)O(2)-Induced Cell Apoptosis in Endothelial Cells

Melastatin-like transient receptor potential channel 2 (TRPM2) is an oxidant-sensitive and cationic non-selective channel that is expressed in mammalian vascular endothelium. Here we investigated the functional role of TRPM2 channels in hydrogen peroxide (H(2)O(2))-induced cytosolic Ca(2+) ([Ca(2+)](i)) elavation, whole-cell current increase, and apoptotic cell death in murine heart microvessel endothelial cell line H5V. A TRPM2 blocking antibody (TM2E3), which targets the E3 region near the ion permeation pore of TRPM2, was developed. Treatment of H5V cells with TM2E3 reduced the [Ca(2+)](i) rise and whole-cell current change in response to H(2)O(2). Suppressing TRPM2 expression using TRPM2-specific short hairpin RNA (shRNA) had similar inhibitory effect. H(2)O(2)-induced apoptotic cell death in H5V cells was examined using MTT assay, DNA ladder formation analysis, and DAPI-based nuclear DNA condensation assay. Based on these assays, TM2E3 and TRPM2-specific shRNA both showed protective effect against H(2)O(2)-induced apoptotic cell death. TM2E3 and TRPM2-specific shRNA also protect the cells from tumor necrosis factor (TNF)-α-induced cell death in MTT assay. In contrast, overexpression of TRPM2 in H5V cells resulted in an increased response in [Ca(2+)](i) and whole-cell currents to H(2)O(2). TRPM2 overexpression also aggravated the H(2)O(2)-induced apoptotic cell death. Downstream pathways following TRPM2 activation was examined. Results showed that TRPM2 activity stimulated caspase-8, caspase-9 and caspase-3. These findings strongly suggest that TRPM2 channel mediates cellular Ca(2+) overload in response to H(2)O(2) and contribute to oxidant-induced apoptotic cell death in vascular endothelial cells. Down-regulating endogenous TRPM2 could be a means to protect the vascular endothelial cells from apoptotic cell death.     

L-茶氨酸对H2O2致L02细胞损伤的保护作用及其机制研究

研究L-茶氨酸对肝细胞损伤的保护作用及其机制。利用H2O2诱导的LO2肝细胞损伤模型,分别用MTT法检测细胞存活率、测定LDH、流式细胞术检测细胞凋亡率、Western blot法检测Caspase-3和PARP蛋白表达7LBax／Bcl-2比值的变化,评价L-茶氨酸是否能保护H2O2诱导的肝细胞损伤。结果表明,L-茶氨酸能提高H2O2损伤的L02细胞存活率,减少LDH的渗漏,降低肝细胞凋亡,且L-茶氨酸通过抑制caspase-3的激活和PARP的切割及Bax／Bcl-2比值的升高而发挥抗凋亡的作用。L-茶氨酸对肝细胞损伤有一定的治疗和保护作用。     

Roles of protein kinase C delta in the accumulation of P53 and the induction of apoptosis in H2O2-treated bovine endothelial cells

To clarify the signaling pathways of oxidative stress-induced apoptosis in bovine aortic endothelial cells (BAEC), we treated cells with 1 mM H 2 O 2 and investigated the roles of protein kinase C δ(PKC δ) and Ca 2+ in the accumulation of p53 associated with apoptosis. The treatment of cells with H 2 O 2 caused the accumulation of p53, which was inhibited by rottlerin (a PKC δinhibitor) but not by BAPTA-AM (an intracellular Ca 2+ chelator). PKC δitself was activated through the phosphorylation at tyrosine residues. H 2 O 2 induced the release of cytochrome c and the activation of caspases 3 and 9, and these apoptotic signals were inhibited by rottlerin and BAPTA-AM. These results suggest that PKC δcontributes to the accumulation of p53 and that Ca 2+ plays a role in downstream signals of p53 leading to apoptosis in H 2 O 2 -treated BAEC.     

Shikonin inhibits the proliferation and induces the apoptosis of human HepG2 cells

This study investigated the potential of shikonin as an anticancer agent against liver cancer and an in vitro human hepatoma cancer model system. The HepG2 cell line was the hepatoma cancer model in the present study. The inhibitory effect of shikonin on the growth of HepG2 cells was measured by MTT assay. To explore the underlying mechanism of cell growth inhibition of shikonin, the cell cycle distribution, DNA fragmentation, mitochondrial membrane potential (ΔΨm) disruption, and expression of Bax and Bcl-2 were measured in HepG2 cells. The activity of shikonin in inducing apoptosis was investigated through the detection of Annexin V signal and CD95 expression by flow cytometry and electron microscopy, respectively. Shikonin inhibited the growth of HepG2 cells in a dose-dependent manner. The IC50 value (inhibiting cell growth by 50%) was 4.30 mg/mL. Shikonin inhibited cell growth in a dose-dependent manner and blocked HepG2 cell cycle progression at the S phase. The changes in mitochondrial morphology, dose-dependently decreased in ΔΨm, were observed in different concentrations of the drug treatment group. Western blot analysis showed that cajanol inhibited Bcl-2 expression and induced Bax expression. Furthermore, we show that shikonin increases Annexin V signal and CD95 (Fas/APO) expression, resulting in apoptotic cell death of HepG2 cells. In addition, lump formation of intranuclear chromatin, pyknosis of cell nucleus, deletion of microvillus, vacuolar degeneration of mitochondria, reduction of rough endoplasmic reticulum, and resolution of free ribosome, etc., associated with apoptosis were discovered by electron microscopy in HepG2 cells after 48 h treatment. Shikonin inhibited HepG2 cells, possibly through the pathway of inducing early apoptosis, and was beneficial for restoring the apoptotic sensitivity of HepG2 cells by CD95, and should therefore be considered as a candidate agent for the prevention or treatment of human hepatoma.     

Activation of Na(+), K(+), Cl(-)-cotransport mediates intracellular Ca(2+) increase and apoptosis induced by Pinacidil in HepG2 human hepatoblastoma cells

The role of Na(+), K(+), Cl(-)-cotransport (NKCC) in apoptosis of HepG2 human hepatoblastoma cells was investigated. Pinacidil (Pin), an activator of ATP-sensitive K(+) (K(ATP)) channels, induced apoptosis in a dose- and time-dependent manner in HepG2 cells. Pin increased intracellular K(+) concentration ([K(+)](i)). Bumetanide and furosemide, NKCC inhibitors, significantly inhibited the Pin-induced increased [K(+)](i) and apoptosis, whereas K(ATP) inhibitors (glibenclamide and tolbutamide) had no effects. The Pin-induced [K(+)](i) increase was significantly prevented by reducing extracellular Cl(-) concentration, and Pin also increased intracellular Na(+) concentration ([Na(+)](i)), further indicating that these effects of Pin may be due to NKCC activation. In addition, Pin induced a rapid and sustained increase in intracellular Ca(2+) concentration ([Ca(2+)](i)), which was completely prevented by the NKCC inhibitors. Treatment with EGTA or BAPTA/AM markedly inhibited the Pin-induced apoptosis. Inhibitors of Na(+), Ca(2+)-exchanger, bepridil, and benzamil significantly prevented both [Ca(2+)](i) increase and apoptosis induced by Pin. Taken together, these results suggest that Pin increases [Na(+)](i) through NKCC activation, which leads to stimulation of reverse-mode of Na(+), Ca(2+) exchanger, resulting in [Ca(2+)](i) increase, and in turn, apoptosis. These results further suggest that NKCC may be a good target for induction of apoptosis in human hepatoma cells.     

Tumor necrosis factor induces apoptosis in hepatoma cells by increasing Ca(2+) release from the endoplasmic reticulum and suppressing Bcl-2 expression

Tumor necrosis factor (TNF) plays an import role in the control of apoptosis. The most well known apoptotic pathway regulated by TNF involves the TNFR1-associated death domain protein, Fas-associated death domain protein, and caspase-8. This study examines the mechanism of TNF-induced apoptosis in FaO rat hepatoma cells. TNF treatment significantly increased the percentage of apoptotic cells. TNF did not activate caspase-8 but activated caspase-3, -10, and -12. The effect of TNF on the expression of different members of the Bcl-2 family in these cells was studied. We observed no detectable changes in the steady-state levels of Bcl-X(L), Bax, and Bid, although TNF suppresses Bcl-2 expression. Dantrolene suppressed the inhibitory effect of TNF on Bcl-2 expression. TNF induced release of Ca(2+) from the endoplasmic reticulum (ER) that was blocked by dantrolene. Importantly, the expression of Bcl-2 blocked TNF-induced apoptosis and decreased TNF-induced Ca(2+) release. These results suggest that TNF induces apoptosis by a mechanism that involves increasing Ca(2+) release from the ER and suppression of Bcl-2 expression.     

Biological effects of 6 mT static magnetic fields: a comparative study in different cell types

The present work was a comparative study of the bio-effects induced by exposure to 6 mT static magnetic field (MF) on several primary cultures and cell lines. Particular attention was dedicated to apoptosis. Cell viability, proliferation, intracellular Ca(2+) concentration and morphology were also examined. Primary cultures of human lymphocytes, mice thymocytes and cultures of 3DO, U937, HeLa, HepG2 and FRTL-5 cells were grown in the presence of 6 mT static MF and different apoptosis-inducing agents (cycloheximide, H(2)O(2), puromycin, heat shock, etoposide). Biological effects of static MF exposure were found in all the different cells examined. They were cell type-dependent but apoptotic inducer-independent. A common effect of the exposure to static MF was the promotion of apoptosis and mitosis, but not of necrosis or modifications of the cell shape. Increase of the intracellular levels of Ca(2+) ions were also observed. When pro-apoptotic drugs were combined with static MF, the majority of cell types rescued from apoptosis. To the contrary, apoptosis of 3DO cells was significantly increased under simultaneous exposure to static MF and incubation with pro-apoptotic drugs. From these data we conclude that 6 mT static MF exposure interfered with apoptosis in a cell type- and exposure time-dependent manner, while the effects of static MF exposure on the apoptotic program were independent of the drugs used.     

Anticancer effect of tert-butyl-2(4,5-dihydrogen-4,4,5,5-tetramethyl-3-O-1H-imidazole-3-cationic-1-oxyl-2)-pyrrolidine-1-carboxylic ester on human hepatoma HepG2 cell line

Tert-butyl-2(4,5-dihydrogen-4,4,5,5-tetramethyl-3-O-1H-imidazole-3-cationic-1-oxyl-2)-pyrrolidine-1-carboxylic ester (L-NNP) is a stable nitroxyl nitroxide radical, which have displayed cytotoxicity on human breast cancer MCF-7 and MDA-MB-231 cell lines. In the present study, we investigated the selective cytotoxicity of L-NNP on isogenetic human hepatoma HepG2 and normal L-02 cell lines. Cell growth inhibition, intracellular reactive oxygen species production, the mitochondrial membrane potential loss, malondialdehyde generation and glutathione levels were analyzed. The expression of Bax, Bcl-2 and NF-κBp65 proteins was also examined. The anticancer activity was evaluated in a HepG2 cell xenograft nude mice model. The results showed that 10, 20, 40 μg/ml L-NNP exposure for 48 h caused 52%, 82% and 91% cell growth inhibition of HepG2 cells, compared with 5%, 10% and 15% that of L-02 cells (p < 0.01). Concentrations of 10, 20, 40 μg/ml L-NNP induced cell death by increasing the generation of intracellular reactive oxygen species and MDA, by depolarizing the mitochondrial membrane potential, and by decreasing intracellular GSH levels in HepG2 cells. Western blot assay showed that Bax, Bcl-2 and NF-κBp65 might be implicated in L-NNP-induced selective HepG2 cell death. L-NNP was also found to inhibit HepG2 hepatoma growth and extend the life span of nude mice model (p < 0.01). The pretreatment and co-treatment of 10 mM N-acetyl-cysteine alleviated L-NNP exposure induced intracellular reactive oxygen species increase and cell growth inhibition demonstrated that L-NNP exhibited neoplasm-selective cytotoxicity and pro-apoptotic activities via reactive oxygen species mediated oxidative damage in HepG2 cells. It might be promising for developing a new class of anticancer agent for liver cancer.     

H(2)O(2): a mediator of esophagitis-induced damage to calcium-release mechanisms in cat lower esophageal sphincter

We previously reported that induction of acute experimental esophagitis by repeated perfusion of HCl may affect release of intracellular Ca(2+) stores. We therefore measured cytosolic Ca(2+) in response to a maximally effective dose of ACh in fura 2-AM-loaded lower esophageal sphincter (LES) circular muscle cells and examined the contribution of H(2)O(2) to the reduction in Ca(2+) signal. In normal cells, the ACh-induced Ca(2+) increase was the same in normal-Ca(2+) and Ca(2+)-free medium and was abolished by the phosphatidylinositol 4,5-bisphosphate-specific phospholipase C inhibitor U-73122, confirming that the initial ACh-induced contraction depends on Ca(2+) release from intracellular stores through production of inositol trisphosphate. In LES cells, the ACh-induced Ca(2+) increase in normal-Ca(2+) medium was significantly lower in esophagitis than in normal cells and was further reduced ( approximately 70%) when the cells were incubated in Ca(2+)-free medium. This reduction was partially reversed by the H(2)O(2) scavenger catalase. H(2)O(2) measurements in LES circular muscle showed significantly higher levels in esophagitis than in normal cells. When normal LES cells were incubated with H(2)O(2), the ACh-induced Ca(2+) increase was significantly reduced in normal-Ca(2+) and Ca(2+)-free medium and was similar to that observed in animals with esophagitis. The initial ACh-induced contraction was also reduced in normal cells incubated with H(2)O(2). H(2)O(2), when applied to cells at sufficiently high concentration, produced a visible and prolonged Ca(2+) signal in normal cells. H(2)O(2)-induced cell contraction was also sensitive to depletion of stores by thapsigargin (TG); conversely, H(2)O(2) reduced TG-induced contraction, suggesting that TG and H(2)O(2) may operate through similar mechanisms. Ca(2+)-ATPase activity measurement indicates that H(2)O(2) and TG reduced Ca(2+)-ATPase activity, confirming similarity of mechanism of action. We conclude that H(2)O(2) may be at least partly responsible for impairment of Ca(2+) release in acute experimental esophagitis by inhibiting Ca(2+) uptake and refilling Ca(2+) stores.     

Polymethoxylated flavones induce Ca(2+)-mediated apoptosis in breast cancer cells

Flavonoids, polyphenolic phytochemicals which include flavones and isoflavones, are present in the common human diet. It has been suggested that these compounds may exert anticancer activity; however, the mechanisms involved remain unknown. We have recently shown (Sergeev, 2004, Biochem Biophys Res Commun 321: 462-467) that isoflavones can activate the novel apoptotic pathway mediated by cellular Ca(2+). Here, we report that polymethoxyflavones (PMFs) derived from sweet orange (Citrus sinensis L.) inhibit growth of human breast cancer cells via Ca(2+)-dependent apoptotic mechanism. The treatment of MCF-7 breast cancer cells with 5-hydroxy-3,6,7,8,3',4'-hexamethoxyflavone (5-OH-HxMF) and 3'-hydroxy-5,6,7,4'-tetramethoxyflavone (3'-OH-TtMF) induced a sustained increase in concentration of intracellular Ca(2+) ([Ca(2+)](i)) resulting from both depletion of the endoplasmic reticulum Ca(2+) stores and Ca(2+) influx from the extracellular space. This increase in [Ca(2+)](i) was associated with the activation of the Ca(2+)-dependent apoptotic proteases, mu-calpain and caspase-12, as evaluated with the calpain and caspase-12 peptide substrates and antibodies to active (cleaved) forms of the enzymes. Corresponding non-hydroxylated PMFs, 3,5,6,7,8,3',4'-heptamethoxyflavone (HpMF) and 5,6,7,3',4'-pentamethoxyflavone (PtMF), were dramatically less active in inducing Ca(2+)-mediated apoptosis. Our results strongly suggest that the cellular Ca(2+) modulating activity of flavonoids underlies their apoptotic mechanism and that hydroxylation of PMFs is critical for their ability to induce an increase in [Ca(2+)](i) and, thus, activate Ca(2+)-dependent apoptotic proteases.     

Redox status of thioredoxin-1 （TRX1） determines the sensitivity of human liver carcinoma cells （HepG2） to arsenic trioxide-induced cell death

lntracellular redox homeostasis plays a critical role in determining tumor cells＇ sensitivity to drug-induced apoptosis. Here we investigated the role of thioredoxin-1 （TRX1）, a key component of redox regulation, in arsenic trioxide （AS2O3）-induced apoptosis. Over-expression of wild-type TRX1 in HepG2 cells led to the inhibition of As2O3-induced cytochrome c （cyto c） release, caspase activation and apoptosis, and down-regulation of TRX1 expression by RNAi sensitized HepG2 cells to As2O3-induced apoptosis. Interestingly, mutation of the active site of TRX1 from Cys^32/35 to Ser^32/35 converted this molecule from an apoptotic protector to an apoptotic promoter. In an effort to understand the mechanisms of this conversion, we used isolated mitochondria from mouse liver and found that recombinant wild-type TRX1 could protect mitochondria from the apoptotic changes. In contrast, the mutant form of TRX1 alone elicited mitochondria-related apoptotic changes, including the mitochondrial permeability transition pore （mPTP） opening, loss of mitochondrial membrane potential, and cyto c release from mitochondria. These apoptotic effects were inhibited by cyclosporine A （CsA）, indicating that mutant TRX1 targeted to mPTP. Alteration of TRX1 from its reduced form to oxidized form in vivo by 2,4-dinitrochlorobenzene （DNCB）, a specific inhibitor ofTRX reductase, also sensitized HepG2 cells to As203-induced apoptosis. These data suggest that TRX1 plays a central role in regulating apoptosis by blocking cyto c release, and inactivation of TRX1 by either mutation or oxidization of the active site cysteines may sensitize tumor cells to As2O3-induced apoptosis.     

过氧化氢预处理对H9c2心肌细胞氧化应激损伤的保护作用

目的：活性氧介导的氧化损伤是缺血再灌注损伤的重要机制,本研究通过观察H2O2预处理对氧化损伤的H9c2心肌细胞存活率和细胞凋亡的影响,探讨其保护H9c2心肌细胞的作用机制。方法：体外培养H9c2心肌细胞,取对数生长期细胞用于实验研究。建立H2O2预处理抵抗高浓度H：O：诱导的细胞氧化损伤模型,实验分组如下：（1）正常对照组（CTL）;（2）损伤组（INJURY）;（3）预处理组十损伤组（PC）。应用CCK8法检测细胞存活率;试剂盒检测胞内MDA水平和T．sOD活性;Hoechst33258染色观察凋亡形态;Annexin-V／PI双染与流式细胞术检测细胞凋亡率。结果：25vLmol／L的H202预处理90rain能明显地保护H9c2心肌细胞抵抗400μmol／LH2O2诱导的氧化损伤,提高细胞存活率,下调MDA水平,上调SOD活性,抑制细胞凋亡,降低细胞凋亡率。结论：低浓度H2O2预处理能减轻H9c2心肌细胞的氧化损伤,抑制氧化损伤诱导的心肌细胞凋亡,具有很好的抗氧化损伤和抗心肌细胞凋亡的保护作用,其作用机制可能与细胞SOD活性上调有关。H2O2预处理为临床治疗心肌缺血／再灌注损伤提供了一项新策略。     

Glibenclamide induces apoptosis through inhibition of cystic fibrosis transmembrane conductance regulator (CFTR) Cl(-) channels and intracellular Ca(2+) release in HepG2 human hepatoblastoma cells.

Glibenclamide, an inhibitor of cystic fibrosis transmembrane conductance regulator (CFTR) Cl(-) channels, induced apoptosis in a dose- and time-dependent manner in HepG2 human hepatoblastoma cells. Glibenclamide increased intracellular Ca(2+) concentration, which was significantly inhibited by Ca(2+) release blockers dantrolene and TMB-8. BAPTA/AM, an intracellular Ca(2+) chelator, and the Ca(2+) release blockers significantly inhibited glibenclamide-induced apoptosis. Glibanclamide also increased intracellular Cl(-) concentration, which was significantly blocked by CFTR Cl(-) channel activators levamisole and bromotetramisole. These activators also significantly inhibited both intracellular Ca(2+) release and apoptosis induced by glibenclamide. The expression of CFTR protein in the cells was confirmed by Western blot analysis. These results suggest that glibenclamide induced apoptosis through inhibition of CFTR Cl(-) channels and intracellular Ca(2+) release and that this protein may be a good target for treatment of human hepatomas.     

Role of ca(2+) in the control of h(2)o(2)-modulated phosphorylation pathways leading to eNOS activation in cardiac myocytes

Nitric oxide (NO) and hydrogen peroxide (H(2)O(2)) play key roles in physiological and pathological responses in cardiac myocytes. The mechanisms whereby H(2)O(2)-modulated phosphorylation pathways regulate the endothelial isoform of nitric oxide synthase (eNOS) in these cells are incompletely understood. We show here that H(2)O(2) treatment of adult mouse cardiac myocytes leads to increases in intracellular Ca(2+) ([Ca(2+)](i)), and document that activity of the L-type Ca(2+) channel is necessary for the H(2)O(2)-promoted increase in sarcomere shortening and of [Ca(2+)](i). Using the chemical NO sensor Cu(2)(FL2E), we discovered that the H(2)O(2)-promoted increase in cardiac myocyte NO synthesis requires activation of the L-type Ca(2+) channel, as well as phosphorylation of the AMP-activated protein kinase (AMPK), and mitogen-activated protein kinase kinase 1/2 (MEK1/2). Moreover, H(2)O(2)-stimulated phosphorylations of eNOS, AMPK, MEK1/2, and ERK1/2 all depend on both an increase in [Ca(2+)](i) as well as the activation of protein kinase C (PKC). We also found that H(2)O(2)-promoted cardiac myocyte eNOS translocation from peripheral membranes to internal sites is abrogated by the L-type Ca(2+) channel blocker nifedipine. We have previously shown that kinase Akt is also involved in H(2)O(2)-promoted eNOS phosphorylation. Here we present evidence documenting that H(2)O(2)-promoted Akt phosphorylation is dependent on activation of the L-type Ca(2+) channel, but is independent of PKC. These studies establish key roles for Ca(2+)- and PKC-dependent signaling pathways in the modulation of cardiac myocyte eNOS activation by H(2)O(2).     

Bcl-x(L) blocks a mitochondrial inner membrane channel and prevents Ca2+ overload-mediated cell death

Apoptosis is an active process that plays a key role in many physiological and pathological conditions. One of the most important organelles involved in apoptosis regulation is the mitochondrion. An increase in intracellular Ca(2+) is a general mechanism of toxicity in neurons which occurs in response to different noxious stimuli like excitotoxicity and ischemia producing apoptotic and necrotic cell death through mitochondria-dependent mechanisms. The Bcl-2 family of proteins modulate the release of pro-apoptotic factors from the mitochondrial intermembrane space during cell death induction by different stimuli. In this work, we have studied, using single-cell imaging and patch-clamp single channel recording, the mitochondrial mechanisms involved in the neuroprotective effect of Bcl-x(L) on Ca(2+) overload-mediated cell death in human neuroblastoma SH-SY5Y cells. We have found that Bcl-x(L) neuroprotective actions take place at mitochondria where this antiapoptotic protein delays both mitochondrial potential collapse and opening of the permeability transition pore by preventing Ca(2+)-mediated mitochondrial multiple conductance channel opening. Bcl-x(L) neuroprotective actions were antagonized by the Bcl-x(L) inhibitor ABT-737 and potentiated by the Ca(2+) chelator BAPTA-AM. As a consequence, this would prevent free radical production, mitochondrial membrane permeabilization, release from mitochondria of pro-apoptotic molecules, caspase activation and cellular death.     

速生桉叶水溶提取物对HepG2细胞增殖及凋亡的影响

目的:探讨速生桉叶水溶提取物对HepG2细胞的抑制增殖作用及凋亡的影响。方法:采用MTT法检测速生桉叶水溶提取物对HepG2细胞增殖抑制作用;采用流式细胞仪Annexin V-FITC/PI检测速生桉叶水溶提取物处理后HepG2细胞凋亡的情况。结果:MTT分析显示,当细胞培养24 h时,稀释5、10倍HepG2细胞抑制率分别为(47.32±1.11)%、(15.76±3.50)%;当细胞培养48 h时,稀释5、10倍HepG2细胞抑制率分别为(44.13±10.93)%、(25.93±8.37)%;当细胞培养72 h时,稀释5、10倍HepG2细胞抑制率分别为(59.47±6.90)%、(41.02±4.27)%。流式细胞术结果显示,速生桉叶水溶提取物稀释30倍时可诱导31.03%的HepG2细胞进入早期凋亡阶段,随着稀释倍数的减少被诱导进入凋亡阶段的细胞数目逐渐升高。另外,随着水溶提取物作用时间的延长,进入凋亡阶段的细胞数目也逐渐升高。结论:速生桉叶水溶提取物对HepG2细胞增殖与凋亡具有量-效、时-效关系。