二烯丙基三硫通过Caspase-3途径诱导人胃癌MGC803细胞凋亡

背景与目的:大蒜及其烯丙基硫化物具有抗肿瘤作用,但其具体的作用机制尚不十分清楚。本研究探讨二烯丙基三硫(diallyltrisulfide,DATS)诱导人胃癌MGC803细胞凋亡及凋亡过程中Caspase-3的变化及其意义。方法:采用荧光显微镜和电子显微镜观察以及流式细胞术和Westernblot检测等方法,观察DATS诱导MGC803细胞凋亡的作用及对活化的Caspase-3的影响。结果:在荧光显微镜及电子显微镜下,DATS作用的胃癌细胞出现了典型的凋亡形态学改变;流式细胞术检测出现明显的亚G1峰;12mg/L的DATS处理MGC803细胞0、4、8、12、24h后,活化的Caspase-3的表达率分别为1.9%、3.0%、7.3%、14.4%、27.6%,提示DATS呈时间依赖性促进活化的Caspase-3的表达;用特异的Caspase-3抑制剂预处理细胞后,再用12mg/LDATS处理细胞24h,与单用DATS组相比,凋亡率从23.0%降低到5.1%(P<0.01),提示Caspase-3的抑制剂能抑制DATS诱导的胃癌MGC803细胞凋亡。Westernblot显示酶原形式的Caspase-3被水解活化。结论:DATS诱导人胃癌MGC803细胞凋亡可能与活化Caspase-3有关。     

Finasteride induces apoptosis via Bcl-2, Bcl-xL, Bax and caspase-3 proteins in LNCaP human prostate cancer cell line

We investigated the effects of finasteride, a 5alpha-reductase inhibitor, on cell death machinery through the induction of apoptosis in an in vitro model for prostate cancer. Finasteride treatment of the LNCaP hormone-dependent human prostate cancer cell line caused the loss of cell viability and accelerated apoptosis in a concentration-dependent manner. The contents of immunoreactive procaspase-3 were examined by immunoblot analysis and the results suggest that the apoptosis induced by finasteride involves the increase of caspase-3 activity. Early cell changes that occur during apoptosis are associated with mitochondrial changes mediated by members of the Bcl-2 family of proteins. Therefore, Bcl-2, Bcl-xL and Bax were evaluated by the Western blot analysis. The immunoreactivity for pro-apoptotic Bax was markedly increased whereas antiapoptotic Bcl-2 and Bcl-xL expression was significantly reduced after incubation of cells with finasteride. These findings suggest that finasteride induces apoptosis in LNCaP cells via proteins of the Bcl-2 and caspase family.     

Dihydroartemisinin induces apoptosis of cervical cancer cells via upregulation of RKIP and downregulation of bcl-2

Treatment of recurrent and metastatic cervical cancer remains a challenge, especially in developing countries, which lack efficient screening programs. In recent years, artemisinin and its derivatives, such as dihydroartemisinin (DHA), which were traditionally used as anti-malarial agent, have been shown to inhibit tumor growth with low toxicity to normal cells. In this study, we investigated mechanisms underlying the anti-tumor effect of DHA in cervical cancer. We evaluated the role of DHA on the expression of bcl-2 and Raf kinase inhibitor protein (RKIP), which is a suppressor of metastasis. The MTT assay was used to compare the proliferation of untreated and DHA-treated Hela and Caski cervical cancer cells. Flow cytometry was used to determine the percentage of cells at each stage of the cell cycle in untreated and DHA-treated cells. We used RT-PCR and western blots to determine the expression of bcl-2 and RKIP mRNA and proteins. We evaluated the effect of DHA treatment in nude mice bearing Hela or Caski tumors. DHA-treated cells showed a time- and dose-dependent inhibition of proliferation and a significant increase in apoptosis. The expression of RKIP was significantly upregulated and the expression of bcl-2 was significantly downregulated in DHA-treated cells compared with control cells. DHA treatment caused (1) a significant inhibition of tumor growth and (2) a significant increase in the apoptotic index in nude mice bearing Hela or Caski tumors. Our data suggest that DHA inhibits cervical cancer growth via upregulation of RKIP and downregulation of bcl-2.     

Knockdown of zinc finger protein X-linked inhibits prostate cancer cell proliferation and induces apoptosis by activating caspase-3 and caspase-9

Zinc finger protein X-linked (ZFX) is a highly conservative mammalian gene with related functions in cell survival and proliferation. However, there are limited reports on regulation of ZFX as a therapeutic target in cancer treatment. In this study, the expression of ZFX in prostate cancer with matched normal adjacent tissues (n=45) and benign prostatic hyperplasia (BPH) tissues (n=16) were observed by immunohistochemical analysis. The effect of lentiviral siRNA (small interference RNA)-mediated dysfunction of ZFX on the proliferation of prostate cancer cells was studied. ZFX mRNA and protein expression levels in prostate cancer cells (PC-3 and DU145) were analyzed by western blotting and real-time polymerase chain reaction (RT-PCR). The effects of siRNA targeting ZFX on growth, cell cycle and apoptosis of PC-3 cells were evaluated by MTT assay and flow cytometry. We also investigated the effect of ZFX deletion on the activation of caspase-1, -3 and -9 by western blotting and colorimetric assay. Prostate cancer specimens appeared significantly higher (42.2% of cases being positive) than that observed in normal adjacent tissues (11.8% of cases being positive) and BPH tissues (12.5% of cases being positive). Repression of ZFX in the prostate cancer cells effectively suppressed the cellular proliferation and colony-formation ability, and led to G1 phase cell cycle arrest. Moreover, inhibition of ZFX-induced cell apoptosis by activating caspase-1, -3 and -9. In conclusion, ZFX represents the prominent role in the progression of prostate cancer and may be a promising therapy target for prostate cancer.     

Kinetin riboside preferentially induces apoptosis by modulating Bcl-2 family proteins and caspase-3 in cancer cells

Here, we demonstrate that kinetin riboside (KR), a cytokinin analog, induces apoptosis in HeLa and mouse melanoma B16F-10 cells. KR disrupted the mitochondrial membrane potential and induced the release of cytochrome c and activation of caspase-3. Bad were upregulated while Bcl-2 was down-regulated under KR exposure. A tumor growth in mice was dramatically suppressed by KR. In contrast, human skin fibroblast CCL-116 and bovine primary fibroblast cells show resistances to KR and no significant changes in Bad, Bcl-X(L,) and cleaved PARP were observed. Our data suggest that KR selectively induces apoptosis in cancer cells through the classical mitochondria dependent apoptosis pathway.     

Diallyl sulfide induces cell cycle arrest and apoptosis in HeLa human cervical cancer cells through the p53, caspase- and mitochondria-dependent pathways

Diallyl sulfide (DAS), one of the main active constituents of garlic, causes growth inhibition of cancer cells in vitro and promotes immune responses in vivo in experimental settings. However, its effects on the induction of cell cycle and apoptosis in human cervical cancer cells are still unclear. The aims of this study were to explore the anti-cancer effects of DAS in HeLa human cervical cancer cells and to investigate the underlying mechanisms in vitro. Cytotoxicity and apoptosis in HeLa human cervical cancer cells were examined by the morphological changes, viability assay, 4',6-Diamidino-2-phenylindole dihydrochloride (DAPI) staining, comet assay, Western blotting and confocal microscopy examination. The results showed that DAS treatment for 24-72 h resulted in a marked decrease in cell viability time- and dose-dependently. Flow cytometric analysis showed that a 48-h treatment of 75 μM DAS induced G0/G1 cell cycle arrest and sub-G1 phase (apoptosis) in HeLa cells. Typical apoptotic nucleus alterations were observed by fluorescence microscopy in HeLa cells after exposure to DAS using DAPI staining. Cells treated with different concentrations of DAS also showed changes typical of apoptosis such as morphological changes, DNA damage and fragmentation, dysfunction of mitochondria, cytochrome c release and increased expression of pro-caspase-3 and -9. DAS also promoted the release of AIF and Endo G from mitochondria in HeLa cells. In conclusion, DAS induced G0/G1 cell cycle arrest and apoptosis in HeLa cells through caspase- and mitochondria and p53 pathways providing further understanding of the molecular mechanisms of DAS action in cervical cancer. This study, therefore, revealed that DAS significantly inhibits the growth and induces apoptosis of human cervical cancer HeLa cells in vitro.     

姜黄素活化caspase-3诱导食管癌Ec-109细胞凋亡的研究

目的探讨姜黄素活化caspase-3诱导食管癌Ec-109细胞凋亡的作用机制。方法20、40、80mmol/L姜黄素作用于食管癌Ec-109细胞,流式细胞仪检测细胞亚二倍体凋亡峰变化及线粒体膜电位变化;Western-blot检测细胞色素C释放及caspase-3表达。结果姜黄素呈剂量依赖性诱导食管癌Ec-109细胞凋亡,20、40、80mmol/L姜黄素作用后,细胞亚二倍体凋亡峰分别为（16.07±1.71）%、（25.54±1.25）%、（37.67±1.09）%,对照组为（3.65%±0.56）%,比较有统计学意义（F=6.14,P〈0.01）。上述浓度姜黄素作用6小时后线粒体膜电位出现降低,比较均有统计学意义（F=5.37,P〈0.05）;12小时出现细胞色素C表达,24小时检测到caspase-3表达,且姜黄素浓度越高,细胞色素C及caspase-3表达越明显。结论姜黄素通过损伤细胞线粒体,使之释放出细胞色素C,激活caspase-3,而且其呈剂量依赖性诱导食管癌细胞凋亡。     

2'-Nitroflavone induces cell cycle arrest and apoptosis in HeLa human cervical carcinoma cells

The mechanism of antitumor action of a synthetic nitroflavone derivative, 2'-nitroflavone, was evaluated in vitro in HeLa human cervix adenocarcinoma cells. We showed that the nitroflavone derivative slowed down the cell cycle at the S phase and increase the population of cells at the G(2)/M phase after 24h of incubation. The treatment with 2'-nitroflavone also induced an apoptotic response, characterized by an increase of the sub-G1 fraction of cells, by cells with chromatin condensation and membrane blebbing, by a typical ladder of DNA fragmentation and by detection of apoptotic cells stained with Annexin V. The observed apoptosis was regulated by caspase-8 and -9, both contributing to the activation of the effector caspase-3. In addition, inhibitors of caspase-8 or -9 partially protected HeLa cells from 2'-nitroflavone-induced cell death. We also found that 2'-nitroflavone did not affect the total amount of Bax and Bcl-2 proteins, although a translocation of Bax from cytosol to mitochondria was evident after 6h of exposure. Furthermore, 2'-nitroflavone decreased the expression of the anti-apoptotic Bcl-X(L) protein, induced the release of cytochrome C to cytosol and increased the levels of Fas and Fas-L. Our results indicated that both death receptor and mitochondria-dependent pathways are involved in the apoptotic cell death triggered by 2'-nitroflavone and suggest that this derivative could be a potentially useful agent for the treatment of certain malignancies.     

Canthaxanthin induces apoptosis in human cancer cell lines

To investigate the possibility that canthaxanthin inhibits cancer cell growth by inducing apoptosis, human WiDr colon adenocarcinoma and SK- MEL-2 melanoma cells were treated with two different doses of the carotenoid for 48 h. Canthaxanthin was incorporated and/or associated to cells. The treatment with the carotenoid caused growth inhibition in both cell types. Concomitantly, apoptosis was induced. Increasing time of exposure and carotenoid concentration, this effect was more pronounced. At 48 h, the percentages of apoptotic cells were 13 and 15, using 1 microM canthaxanthin, and 18 and 20, using 10 microM canthaxanthin in WiDr and SK-MEL-2 cells, respectively. This study represents the first demonstration that canthaxanthin is able to induce apoptosis in tumour cells.      

Moving irradiation in the combined radiation treatment of cervical cancer]

Application of moving telegamma therapy as an adjunct to the associated radiation treatment in 399 patients allowed an increase of the dosage up to 6500 rad in the parametrium area. A 5-year survival in stage I was 91.6 +/- 10.5%, in stage II--63.3 +/- 4.7% and in stage III--48 +/- 5.6%. Late postirradiation rectitis was noted in 24 patients, cystitis--in 8 and rectal fistula--in 7.     

survivin和caspase-3在非小细胞肺癌中的表达及其与肿瘤细胞凋亡的关系

背景与目的survivin是新近发现的凋亡抑制因子,在肿瘤的发生中起重要作用。survivin可直接抑制caspase3和caspase7的活性。本研究旨在探讨survivin和caspase3在非小细胞肺癌(NSCLC)中的表达情况及其与肿瘤细胞凋亡的关系。方法采用免疫组织化学SP法检测88例NSCLC患者手术标本和29例正常肺组织标本中survivin和caspase3的表达,分析其与NSCLC临床病理参数之间的关系,同时用原位末端标记法(TUNEL)标记肺癌组织中的凋亡细胞。结果88例NSCLC患者标本中survivin的阳性表达率为61.4%(54/88),明显高于正常肺组织(13.8%,4/29)(P<0.01);survivin在NSCLC中的表达与肿瘤的组织学分型、病理分级和淋巴结转移均无明显相关性(P>0.05),但与TNM分期有相关性(P<0.05)。caspase3在正常肺组织和NSCLC中阳性表达率分别为89.7%(26/29)和73.9%(65/88),但二者差异无显著性(P>0.05);caspase3在NSCLC中的阳性表达率随肿瘤的病理分级增高而降低,差异有显著性(P<0.05)。survivin蛋白阳性表达的肺癌组织中平均细胞凋亡指数(1.63±0.58)明显低于阴性表达组(3.29±0.76)(P<0.05),caspase3阳性组平均细胞凋亡指数(2.42±0.59)明显高于阴性组(1.28±0.65)(P<0.05);survivin和caspase3呈负相关(P<0.05)。结论凋亡抑制蛋白survivin在NSCLC组织中高表达,caspase3与survivin的表达呈负相关,survivin可能通过抑制caspase3的激活来发挥其抑制细胞凋亡的作用。     

The roles of caspase-3 and bcl-2 in chemically-induced apoptosis but not necrosis of renal epithelial cells

The kidney is a target for toxicants including cisplatin and S-(1,2-dichlorovinyl)-L-cysteine (DCVC), a metabolite of the environmental contaminant, trichloroethylene. Necrosis is well characterized in kidney cells, but pathways leading to apoptosis are less clear. Cysteine conjugates are useful toxicants because they induce either necrosis or apoptosis depending on chemical structure or antioxidant status. Herein, we show that in the renal epithelial cell line LLC-PK1, activation of caspase-3 (CPP32/Yama/apopain) is crucial for apoptosis, but not necrosis. Apoptosis was blocked by zVAD.fmk, and partially by a cathepsin inhibitor. Caspase-3 activity and cleavage of poly(ADP-ribose) polymerase (PARP) was detected only during apoptosis. S-(1,1,2,2-Tetrafluoroethyl)-L-cysteine (TFEC), a metabolite of tetrafluoroethylene, kills cells only by necrosis, and did not activate caspases under any conditions. Apoptosis and activation of caspase-3 by cisplatin, but not DCVC, was prevented by bcl-2. Thus, caspase-3 activation by bcl-2-dependent and -independent mechanisms is a terminal event in chemical-apoptosis of renal epithelial cells.     

Arsenic trioxide induces apoptosis in human gastric cancer cells through up-regulation of p53 and activation of caspase-3

Arsenic trioxide (As(2)O(3)) can induce clinical remission in patients suffering from acute promyelocytic leukemia, through induction of apoptosis and activation of caspases. We investigated the potential use of As(2)O(3) in human gastric cancer and its possible mechanisms. Human gastric cancer cell lines AGS and MKN-28 were treated with various concentrations (0.1 to 100 microM) of As(2)O(3) for 24 to 72 hr. Apoptosis was determined by acridine orange staining, flow cytometry and DNA fragmentation. Protein levels of p53, p21(waf1/cip1), c-myc, bcl-2 and bax were detected by Western blotting. Effects of As(2)O(3) on caspase-3 protease activity, its protein concentration and cleavage of poly(ADP)-ribose polymerase (PARP) were also studied. As(2)O(3) inhibited cell growth and induced apoptosis in both cell lines, though AGS cells were more sensitive. As(2)O(3) induced apoptosis in AGS cells in a concentration- and time-dependent manner. Treatment resulted in a marked increase in p53 protein levels as early as 4 hr. Co-incubation with p53 anti-sense oligo-nucleotide suppressed As(2)O(3)-induced intracellular p53 over-expression and apoptosis. As(2)O(3) increased the activity of caspase-3, with appearance of its 17 kDa peptide fragment, and cleavage of PARP, with appearance of the 85 kDa cleavage product, both in parallel with the induction of apoptosis. Both the tripeptide caspase inhibitor zVAD-fmk and the specific caspase-3 inhibitor DEVD-fmk partially suppressed As(2)O(3)-induced caspase-3 activation and apoptosis. As(2)O(3) inhibits cell growth and induces apoptosis in gastric cancer cells, involving p53 over-expression and activation of caspase-3. The potential use of this compound in the treatment of gastric cancer is worth further investigation.     

The synergistic anticancer effect of troglitazone combined with aspirin causes cell cycle arrest and apoptosis in human lung cancer cells

Troglitazone (TGZ) is a synthetic thiazolidinedione drug belonging to a group of potent peroxisome proliferator‐activated receptor γ (PPARγ) agonists known to inhibit proliferation, alter cell cycle regulation, and induce apoptosis in various cancer cell types. TGZ is an oral anti‐type II diabetes drug that can reverse insulin resistance. For more then 100 yr, aspirin, a nonselective cyclooxygenase (COX) inhibitor, has been successfully used as an anti‐inflammatory drug. Recently, Aspirin (ASA) and some other nonsteroidal anti‐inflammatory drugs (NSAIDs) have drawn much attention for their protective effects against colon cancer and cardiovascular disease; it has been observed that ASA's anti‐tumor effect can be attributed to inhibition of cell cycle progression, induction of apoptosis, and inhibition of angiogenesis. In this report we demonstrate for the first time that, when administered in combination, TGZ and ASA can produce a strong synergistic effect in growth inhibition and G₁ arrest in lung cancer CL1‐0 and A549 cells. Examination by colony formation assay revealed an even more profound synergy. In Western blot, combined TGZ and ASA also could downregulate Cdk2, E2F‐1, cyclin B1, cyclin D3 protein, and the ratio of phospho‐Rb/Rb. Importantly, apoptosis was synergistically induced by the combination treatment, as evidenced by caspase‐3 activation and PARP cleavage. The involvement of PI3K/Akt inhibition and p27 upregulation, as well as hypophosphorylation of Rac1 at ser71, were demonstrated. Taken together, these results suggest that clinically achievable concentrations of TGZ and ASA used in combination may produce a strong anticancer synergy that warrants further investigation for its clinical applications. © 2009 Wiley‐Liss, Inc.     

去甲斑蝥素对人乳腺癌细胞系的凋亡诱导作用及bcl-2基因的表达

目的：探讨去甲斑蝥素抗肿瘤作用的分子机制。方法：用１０μｇ／ｍｌ去甲斑蝥素处理体外培养的人乳腺癌细胞素ＭＣＦ－７。处理后,在不同时间点,采用普通光镜、电子显微观察去甲斑蝥素对乳腺癌细胞的诱导凋亡现象。利用流式细胞仪分析凋亡细胞百分比,用蛋白印迹杂交方法对凋亡抑制基因ｂｃｌ－２的表达情况进行检测。结果：经１０μｇ／ｍｌ去甲斑蝥素处理１２ｈ后,可观察到ＭＣＦ－７细胞变形、出泡,从培养瓶底脱离。细胞染色和电子显微镜可观察到染色质浓聚、边集,且随着药物作用时间的延长,凋亡细胞百分比逐渐增加。与对照组相比,凋亡抑制基因ｂｃｌ－２的表达降低。结论：诱导肿瘤细胞凋亡可能是去甲斑蝥素抗肿瘤作用的分子机制之一。     

Ursolic acid induces Bax-dependent apoptosis through the caspase-3 pathway in endometrial cancer SNG-II cells

The goal of this study was to examine the effect of ursolic acid, a pentacyclic triterpenoid compound, on growth of the endometrial cancer cell line SNG-II. We found that ursolic acid strongly inhibited the growth of SNG-II cells in a dose- and time-dependent manner. Morpholgical changes characteristic of apoptosis were observed in treated cells, such as the presence of apoptotic bodies and fragmentation of DNA into oligonucleosomal-sized fragments. We also investigated the active forms of caspase-3, -8 and -9 in ursolic acid-treated SNG-II cells. At 25 and 50 microM strength, ursolic acid induced marked increases in caspase-3 activity to approximately 5-fold that of control cells. Levels of cleaved caspase-3 increased in a time- and dose-dependent manner. Activation of caspases also led to the cleavage of target proteins, such as PARP. Ursolic acid treatment also resulted in a cleavage of poly (ADP-ribose) polymerase in a dose-dependent manner. Testing whether caspase-3 activation and DNA polymerase activity were inhibited by addition of Ac-DEDV-HCO during ursolic acid treatment showed that 50 microM Ac-DEDV-HCO inhibited caspase-3 activity in treated cells. Although DNA fragmentation was observed after ursolic acid treatment, DNA fragmentation did not occur in SNG II cells treated with both Ac-DEDV-HCO and ursolic acid. Because some researchers have suggested that mitochondrial pathways are involved in ursolic acid-induced apoptosis secondary to induction of mitochondrial cytochrome c release, we studied mitochondrial events in ursolic acid-induced apoptosis in these cell lines. After ursolic acid treatment, the anti-apoptotic Bcl-2 protein decreased and Bax expression was enhanced. Our results indicated that ursolic acid induced apoptotic processes in the endometrial cancer SNG-II cell line through mechanisms involving mitochondrial pathways and Bcl-2 family proteins.     

Induction of apoptosis by isothiocyanate sulforaphane in human cervical carcinoma HeLa and hepatocarcinoma HepG2 cells through activation of caspase-3

Sulforaphane (SFN) is an isothiocyanate that is found in abundant quantities in many cruciferous vegetables including broccoli and cauliflower. Its inhibitory effects on tumor cell growth in vitro and in vivo, which is dependent on the direct effect on cancer cells, has attracted considerable attention. This study examined the effects of SFN on the growth of human cervical carcinoma HeLa and hepatocarcinoma HepG2 cells. The results showed that SFN inhibits the viability of both HeLa and HepG2 cells by inducing apoptosis, as evidenced by the formation of apoptotic bodies and the accumulation of the sub-G1 phase. RT-PCR and immunoblotting showed that treating the cells with SFN caused the down-regulation of anti-apoptotic Bcl-2 and Bcl-XL, and the up-regulation of pro-apoptotic Bax expression. SFN-induced apoptosis was associated with the proteolytic activation of caspase-3, and the degradation/cleavage of poly (ADP-ribose) polymerase and the beta-catenin protein. z-DEVD-fmk, a caspase-3 specific inhibitor, blocked the activation of caspase-3 and increased the survival of the SFN-treated HeLa and HepG3 cells, suggesting that caspase-3 activation is essential for SFN-induced apoptosis.     

Cisplatin-induced apoptosis proceeds by caspase-3-dependent and -independent pathways in cisplatin-resistant and -sensitive human ovarian cancer cell lines.

We have assessed in detail the effect of cisplatin-activated programmed cell death in the cisplatin-sensitive human ovarian cancer cell line A2780 and two drug-resistant subclones, CP70 and C30. To determine whether the differential extent of apoptosis observed between the sensitive and resistant ovarian cancer cell lines was the result of dissimilar upstream signaling events, we assessed the execution of apoptotic events that precede target protein proteolysis and subsequent chromosomal DNA degradation. Proteolytic degradation of procaspase-3 was observed in both the CP70 and C30 cells following IC50 cisplatin treatment, whereas no proteolyzed caspase-3 subunits were detected in the A2780 cells. However, using a direct enzymatic assay measuring cleavage of the synthetic peptide substrate (N-acetyl-Asp-Glu-Val-Asp-p-nitroanilide), activity was detected in extracts prepared from A2780 cells treated at the IC90 level of cisplatin and was 2-3-fold less than that of extracts prepared from CP70 and C30 cells. Because the activation of procaspase-3 by caspase-9 requires the release of cytochrome c into the cytoplasm, we determined the level of cytoplasmic cytochrome c in each cell line in response to cisplatin treatment. Consistent with the caspase-3 activation data, a very small increase in cytoplasmic cytochrome c was observed in A2780 cells following cisplatin treatment, whereas dramatic increases were evident in both the CP70 and C30 cell lines. The expression of the mitochondrial factors Bcl-2, Bcl-x, and Bax was determined because each has been implicated in the regulation or release of cytochrome c at the level of the mitochondria. Bcl-2 and Bcl-xL proteins remained relatively unchanged in expression for over 48 h after exposure to cisplatin in the A2780 cell lines. However, within the same time period, expression of Bcl-2 decreased in the CP70- and C30-resistant cell lines, whereas an increase in Bcl-xL expression was observed. Expression of the proapoptotic Bcl-xS protein was observed in only the resistant CP70 and C30 cell lines independent of cisplatin treatment. A change in the expression of Mr 24,000 Bax to a Mr 21,000 isoform was evidenced in the A2780 cells within 48 h of cisplatin treatment and, to a greater extent, in the CP70 and C30 cells, which also expressed a Mr 16,000 Bax variant. Evidence for an alternative apoptotic pathway in A2780 cells was obtained by demonstrating increased FADD expression in response to cisplatin treatment. These results support a model in which cisplatin-induced programmed cell death in the cisplatin-sensitive A2780 and -resistant CP70 and C30 cells proceeds via caspase-3-independent and -dependent pathways, respectively.     

Ursolic acid induces apoptosis through caspase-3 activation and cell cycle arrest in HaCat cells

Ursolic acid (UA) is a pentacyclic triterpene compound isolated from many kinds of medicinal plants and present in human diet. In this study, we investigated the pro-apoptotic effect of UA on HaCat derived keratinocyte cell line. Treatment with UA decreased the viability of HaCat cells in a concentration- and time-dependent manner. In addition, cell cycle analysis revealed that UA treated HaCat cells were blocked predominantly in G1 phase. Moreover, expression of p21WAF1, a cell cycle regulator, was increased by UA, indicating that UA-induced cell cycle arrest could be mediated through p21WAF1. During UA treatment, we also demonstrated that p53 was phosphorylated at serine 392 and translocated to the nucleus. It is well established that p53 achieves its tumor suppressor activity by inducing apoptosis on cells. To define the apoptotic process in our system, we examined effect of UA on caspase activities, and demonstrated caspase-3 activation. In conclusion, our results suggest that UA induces: i) cell cycle arrest concomitantly with the apparition of the apoptotic sub group G1 peak, and ii) cell death through apoptosis, which is mediated by caspase-3.