米非司酮诱导雄激素非依赖性前列腺癌细胞凋亡的实验研究

目的探讨米非司酮在体外诱导雄激素非依赖性前列腺癌DU-145、PC-3细胞凋亡的作用。方法采用四甲基偶氮唑蓝法检测1,10,50和100μmol／L米非司酮作用于前列腺癌DU-145、PC-3细胞24～120h的吸光度（A）值,用流式细胞仪检测10μmol／L米非司酮作用24和48h后DU．145、PC-3细胞凋亡率的变化;采用免疫组化法检测米非司酮作用DU-145、PC．3细胞后bax、bcl-2、血管内皮生长因子（VEGF）蛋白表达的变化。结果1μmol／L米非司酮组的A值与对照组相比,差异无统计学意义（P〉0．05）;10,50和100μmol／L米非司酮组的A值与对照组比较,差异有统计学意义（P〈0．01）;米非司酮对前列腺癌DU-145、PC-3细胞的抑制作用呈时间-剂量依赖性。10μmol／L米非司酮作用前列腺癌DU-145细胞24和48h的凋亡率分别为15．3％和30．4％,PC-3细胞的凋亡率分别为22．2％和32．0％。经10μmol／L米非司酮作用后,DU-145、PC-3细胞中VEGF和bcl-2蛋白的表达明显减少,而bax的表达显著增加（P〈0．05）。结论米非司酮以时间．剂量依赖性方式抑制激素非依赖性前列腺癌DU-145和Pc-3细胞的增殖,其作用可能是通过降低VEGF蛋白的表达。从而下调bcl-2、激活bax蛋白的表达来实现。     

ABL-N诱导前列腺癌细胞凋亡及其机制

目的 探讨旋覆花内酯(ABL)-N诱导前列腺癌细胞凋亡的作用及其机制.方法 0～40μmol/L ABL-N分别处理前列腺痛细胞后,利用噻唑蓝(MTF)检测对细胞增长的抑制作用;流式细胞学、TUNEL染色等方法检测其诱导凋亡的作用,并检测Capase活性,Western blot测定bax、bel-2水平变化.结果 ABL-N明显抑制前列腺癌细胞PC3、LNCaP及DU145的生长,并呈剂量依赖性.40 μmol/L ABL-N作用24 h后,(73.34±4.41)%的PrEC细胞存活,而3种前列腺癌细胞分别为(11.92±2.31)%、(12.55±1.94)%、(13.28±2.26)%.膜联蛋白/碘化丙锭(Annexin V/PI)及TUNEL染色表明ABL-N呈剂量依赖性诱导PC3凋亡.ABL-N可激活Caspase活性,尤其Caspase-3,20μmol/L ABL-N作用24 h后其活性是对照组的4.23倍;bax/bcl-2比率随浓度增加明显增高.结论 ABL-N可通过Caspase途径及bax/bcl-2蛋白途径,诱导PC3等前列腺癌细胞凋亡,抑制前列腺癌细胞增殖.

Abstract：

Objective To explore the ABL-N-induced apoptosis of human prostate cancer cells and the mechansim. Methods After administration of 0-40 μmol/L ABL-N for 24 h, the effects of ABL-N on the induction of apoptosis in human prostate cancer cells PC3 were measured by methyl thiazol tetrazolium ( MTT) colorimetry, Annexin V/propidium iodide staining and TUNEL staining. The levels of bax and bcl-2 were tested by Western blotting. Caspase activity was assayed. Results ABL-N treatment to PC3,LNCaP, and DU145 cells resulted in a dose-dependent inhibition of cell growth without any substantial effect on normal human prostate epithelial PrEc cells. About (73. 34 ±4. 41)% of PrEC cells were viable following a 24-h exposure to 40 μmol/L ABL-N, whereas only (11. 92 ± 2. 31) % of PC3, (12. 55 ±1. 94) % of LNcap, and (13. 28 ± 2. 26) % of DU145 cells survived under similar conditions of ABL-N treatment. ABL-N treatment resulted in a dose-dependent induction of apoptosis of PC3 cells. Furthermore,ABL-N induced the activation of Caspases, especially Caspase 3. The Caspase-3 activity of PC3 cells treated with ABL-N (20 μmol/L) (0. 95) was significantly increased by about 4. 2-fold of the untreated cells (0. 24) at 24 h. The ratio of bax/bcl-2 was also increased significantly. Conclusion ABL-N induces apoptosis though the activation of Caspase 3 and pro- and anti-apoptotic Bcl-2 family proteins in prostate cancer cells.     

Modulation of prostate carcinoma cell growth and apoptosis by chromogranin A

PURPOSE: We elucidated the effect and possible pathways of chromogranin A in regulating prostatic cancer cell growth. MATERIALS AND METHODS: Chromogranin A expression in prostatic cancer cell lines were detected with immunofluorescence flow cytometry (FCM) and the inhibition of cell growth due to chromogranin A antibody was measured using microculture tetrazolium. Cell cycle and RNA changes were evaluated with acridine orange FCM. Intracellular chromogranin A variations were detected with FCM, as were apoptotic changes, p53, Fas and tumor necrosis factor-alpha. Apoptosis and caspase-3 expression of tumor cells was assessed with dual immunohistochemical staining. RESULTS: Increased chromogranin A expression was observed in PC-3, DU145 and LNCap cells independent of hormone dependence. Dose and time dependent growth inhibition occurred at 12 hours. Chromogranin A antibody arrested PC3 cells in the S-phase immediately after treatment. The number of G0/G1 and G2/M cells subsequently decreased. PC3 tumor cells had transiently increased RNA at 12 hours with a marked decrease at 48 hours. Decreasing chromogranin A expression started at 12 hours and was most prominent at 48 hours. Apoptotic cells markedly increased at 12 hours with an increase in p53, Fas and tumor necrosis factor-alpha (Fas more than the others). Increased apoptotic cell and caspase-3 expression was observed on immunohistochemical stains. CONCLUSIONS: Chromogranin A is an important neuropeptide regulating the growth of prostate cancer cells. Specific antibodies can suppress its function through apoptotic pathways (Fas and caspase-3), leading to programmed cell death. Chromogranin A antibody mediated apoptosis may be a specific alternative treatment for prostate cancer.     

Tetrandrine suppresses proliferation,induces apoptosis,and inhibits migration and invasion in human prostate cancer cells

Tetrandrine (TET), a traditional Chinese medicine, exerts remarkable anticancer activity on various cancer cells. However, little is known about the effect of TET on human prostate cancer cells, and the mechanism of function of TET on prostate cancer has not yet been elucidated. To investigate the effects of TET on the suppression of proliferation, induction of apoptosis, and inhibition of migration and invasion in human prostate cancer cell lines, DU145 and PC-3. Inhibition of growth was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and clone formation assay, and flow cytometry analysis was performed to detect the induction of apoptosis. Activation of poly (ADP-ribose) polymerase, caspase-3, Akt, phospho-Akt, Bcl-2, and Bax was analyzed by Western blotting. Wound healing assay and transwell migration assay were used to evaluate the effect of TET on migration and invasion of cancer cells. TET inhibited the growth of DU145 and PC–3 cells in a dose- and time-dependent manner. Cell cloning was inhibited in the presence of TET in DU145 and PC-3 cells. TET suppressed the migration of DU145 and PC-3 cells. Transwell invasion assay showed that TET significantly weakened invasion capacity of DU145 and PC-3 cells. TET exhibited strong inhibitory effect on proliferation, migration, and invasion of prostate cancer cells. In addition, TET induced apoptosis in a dose-dependent manner by activating the caspase cascade and inhibiting phosphoinositide 3-kinase-Akt signal pathway. The accumulating evidence suggests that TET could be a potential therapeutic candidate against prostate cancer in a clinical setting.     

喜树碱诱导人膀胱癌细胞RT4和MGH凋亡的实验研究

目的 研究DNA拓扑异构酶Ⅰ抑制剂喜树碱(CPT)诱导人膀胱癌细胞凋亡的作用。方法 体外培养的膀胱癌RT4和MGH细胞用CFr处理，用FACScan流式细胞仪分析细胞凋亡的发生及变化，用Western blotting的方法检测了CFT处理不同时间段，bcl-2与bax表达的变化，以及天冬氨酸特异性半胱氨酸蛋白酶-3(Caspase-3)和多聚腺苷二磷酸核糖聚合物酶(PARP)的活性的改变。结果 CPT处理膀胱癌细胞24h后，细胞凋亡率明显上升，且CPT诱导的凋亡呈时间依赖性和剂量依赖性。CFT诱导膀胱癌细胞RT4和MGH后均未见bcl-2和bax表达水平的改变，但是可以观察到凋亡执行蛋白Caspase-3和PARP的激活。结论 CPT能够诱导膀胱癌细胞RT4和MGH发生凋亡，其抑制膀胱癌细胞的作用可能与肿瘤细胞发生凋亡有关。     

Resveratrol sensitizes androgen independent prostate cancer cells to death-receptor mediated apoptosis through multiple mechanisms

BACKGROUND: A critical factor in prostate cancer development and progression is the altered expression of apoptotic regulatory proteins which renders cells resistant to both hormone- and chemo-therapies. Resveratrol, a dietary component with chemopreventive properties has been reported to resensitize a variety of cancer cell types to apoptosis. In the current study, the ability of resveratrol pre-treatment to sensitize hormone refractory prostate cancer cell lines (PC-3 and DU145) to apoptosis and the mechanisms involved were investigated. METHODS: Apoptosis was assessed using several established parameters and protein expression was analyzed by Western blot and flow cytometry. IAP knockdown was achieved using RNAi while inhibition of Akt phosphorylation was achieved by pre-incubation with the PI3-kinase inhibitor LY294002. RESULTS: Pre-treatment with resveratrol sensitized PC-3 and DU145 cells to agents that specifically target death receptors (TRAIL, Fas, TNFalpha) but not agents that initiate apoptosis through other mechanisms (Etoposide, Paclitaxel, Tunicamycin, Thapsigargin). Resveratrol pre-treatment altered the expression of IAPs and Bax, and decreased Akt phosphorylation in PC-3 cells, leading to increased caspase activation and apoptosis. While knockdown of IAPs using siRNA did not mimic the effects of resveratrol, inhibition of Akt phosphorylation using LY294002 sensitized PC-3 cells to TRAIL induced apoptosis but not to etoposide or tunicamycin. CONCLUSION: Altering apoptotic susceptibility in advanced androgen independent disease requires manipulation of a broad signaling pathway. Use of resveratrol or inhibition of Akt phosphorylation may represent an important therapeutic approach in combination with conventional therapies for the treatment of prostate cancer.     

Prostate cancer cell proliferation is influenced by leptin

BACKGROUND: Obesity is considered a risk for many cancers. Serum leptin levels are often elevated in obese people. Leptin acts as a mitogenic agent in many tissues; therefore, it may act to promote cancer cell growth. We previously demonstrated that leptin acts as a growth factor for prostate cancer cells in vitro. The purpose of this study was to characterize leptin receptor isoform mRNA expression in leptin-treated DU145 and PC-3 prostate cancer cell lines. Expression levels of SOCS-3, a known leptin-inducible suppressor of leptin signaling, and known mitogenic signaling pathways of PI3K and ERK were also analyzed METHODS: DU145 and PC-3 cells were treated with 0, 4, 40, or 80 ng/ml leptin for 0, 0.5, 1, 2, 4, 24, or 48 h. Multiplex RT-PCR was performed to determine mRNA levels of the short (huOB-Ra) or the long (huOB-Rb) OB-R isoforms or SOCS-3. p-Akt and p-ERK were determined by Western blot. Cell viability and apoptosis were determined by MTT and nucleosomal fragmentation assay RESULTS: DU145 and PC-3 expressed huOB-Ra, huOB-Rb, and SOCS-3 mRNA. huOB-Ra mRNA levels increased in PC-3 at 48 h (P < 0.01); however, no significant changes were observed in DU145. huOB-Rb mRNA levels decreased at 48 h in DU145; however, a twofold increase at 48 h (P < 0.01) was observed with PC-3 and was dose-dependent (P < 0.05). Leptin increased SOCS-3 mRNA in DU145 at 24 and 48 h (P < 0.05) and in PC-3 at 1 h (2-fold) and 48 h (fivefold; P < 0.01). Leptin up-regulated p-Akt in a time- and dose-dependent manner in the DU145 prostate cancer cells via a suppression of apoptosis. Leptin up-regulated p-ERK in a time-dependent manner in PC-3 cells CONCLUSIONS: In prostate cancer cells, the mitogenic effects of leptin are not a consequence of altered receptor isoform mRNA expression. No defect in SOCS-3 signaling was observed, and proliferation appears to be working through the PI3K and MAPK leptin receptor-activated pathways, depending on cell type. Leptin stimulation may be selective for either pathway to suppress apoptosis, thereby enhancing prostate cancer growth.     

Melatonin and prostate cancer cell proliferation: interplay with castration,epidermal growth factor,and androgen sensitivity

BACKGROUND: Potential modulatory effects of melatonin on the proliferation of androgen-sensitive LNCaP and androgen-insensitive PC-3 and DU 145 prostate cancer cells were reported recently. In this study, we investigated the effects of combined melatonin and castration on LNCaP tumor growth in vivo, the interactions between melatonin and epidermal growth factor (EGF) on LNCaP cell proliferation, and melatonin actions on the proliferation of PC-3 and DU 145 cells. METHODS: Tumor development and growth in castrated nude mice inoculated with LNCaP cells or in intact animals inoculated with DU 145 cells, with or without daily melatonin treatment, were monitored by observation and caliper measurement. MT(1) receptor expression in native or transfected prostate cancer cell lines was examined by immunocytochemistry or 2-[(125)I]iodomelatonin binding. Cyclin D1 expression in LNCaP cells was assessed by Western blotting, and cell proliferation was measured by thymidine incorporation and/or cell count. RESULTS: Melatonin treatment was associated with further decreases in LNCaP tumor incidence and growth rate in castrated nude mice. Melatonin and 2-iodomelatonin (a melatonin receptor agonist) attenuated EGF-stimulated increases in LNCaP cell proliferation and cyclin D1 levels. Melatonin had no effect on the proliferation or growth of MT(1) receptor-expressing DU 145 cells, and of PC-3 cells in which MT(1) receptor protein was undetectable. The proliferation of transfected PC-3 cells expressing MT(1) receptor was unaffected by 2-iodomelatonin. CONCLUSION: Together with previous data, the present results indicate synergistic action of melatonin and castration in inhibiting the growth of androgen-sensitive LNCaP tumor. Androgen-sensitive prostate cancer cell proliferation may be modulated by opposite changes in cyclin D1 levels induced by activated MT(1) and EGF receptors. In androgen-insensitive prostate cancer cells, MT(1) receptor-mediated signal transduction may become defective not only through changes in membrane receptor protein expression and/or functions, but also by means of alterations in downstream postreceptor signaling events.     

FOXN2在前列腺癌组织的表达及对前列腺癌细胞生物学的影响

探讨双头框蛋白N2（FOXN2）在前列腺癌（PCa）组织中的表达以及对PCa细胞生物学的影响。方法 选取宜宾市第二人民医院50例接受手术治疗的PCa患者的标本及癌旁组织,通过RT-qPCR实验和Western blot实验分别检测PCa组织和癌旁组织中的FOXN2 mRNA及蛋白的表达水平,并分析PCa组织中FOXN2与患者临床病理特征的相关性。通过RT-qPCR实验检测正常前列腺细胞RWPE-1、PCa细胞PC-3、DU145、LNCaP中FOXN2 mRNA的表达水平。选取PC-3细胞为研究对象,分为FOXN2过表达组、空载体对照组以及对照组,分别通过MTT实验、流式细胞实验、Transwell实验以及Western blot实验检测各组细胞增殖、凋亡、迁移和侵袭情况以及相关蛋白Bax、CyclinD1、MMP-2的表达量。结果 与癌旁组织相比,FOXN2的mRNA和蛋白表达水平显著降低（P<0.05）,与TCGA数据库中结果一致。FOXN2低表达与PCa患者淋巴转移、TNM分期以及Gleason评分有关（P=0.003、0.005、0.002）。与人正常前列腺细胞RWPE-1相比,FOXN2在PCa细胞PC-3、DU145、LNCaP中均呈现低表达（P<0.05）,其中PC-3细胞中表达量最低。与对照组和空载体对照组相比,FOXN2过表达组的PC-3细胞在作用24 、48 、72 h后增殖能力显著下降（F=290.400、57.735、113.014,P<0.05）,CyclinD1蛋白表达水平显著下降（P<0.05）;PC-3细胞的凋亡率显著升高（P<0.05）,Bax蛋白表达水平显著升高（P<0.05）;PC-3细胞的迁移和侵袭能力显著下降（P<0.05）,MMP-2蛋白表达水平显著下降（P<0.05）。结论 FOXN2在PCa组织及细胞中低表达,过表达FOXN2可抑制PCa细胞的增殖、迁移和侵袭,并促进细胞凋亡。     

多肽K237对PC-3M细胞增殖及bax、bcl-2 mRNA表达的影响

目的:研究多肽K237对体外培养的雄激素非依赖性前列腺癌PC-3M细胞的抑制作用,及其可能的作用机制。方法:将培养的PC-3M细胞分为4组:实验组(分别以50、100、200μmol/L的多肽K237处理48h)和对照组(K237浓度为0μmol/L),采用MTT法观察多肽K237对前列腺癌PC-3M细胞增殖的影响,用RT-PCR法检测bax、bcl-2mRNA表达的变化。结果:不同浓度的多肽K237处理48h后,PC-3M细胞形状变圆,体积变小,胞质透亮度下降,部分细胞脱落悬浮于培养液中。在50、100、200μmol/L的多肽K237作用48h后,MTT法检测的细胞生长抑制率分别为(12.6±0.95)%、(17.8±0.99)%、(27.2±1.12)%。RT-PCR结果显示:50、100、200μmol/L实验组和对照组的bax/β-actin值分别为0.919±0.071、0.971±0.083、0.992±0.102,(0.889±0.06),bcl-2/β-actin值分别为0.896±0.085、0.791±0.084、0.764±0.702,0.922±0.097,3组中上述两项指标均较对照组有明显变化(P均<0.01),其中,baxmRNA表达水平上调,而bcl-2mRNA表达水平下调,上述作用呈现剂量效应关系。结论:多肽K237可能通过影响bax、bcl-2mRNA的表达来诱导PC-3M细胞凋亡,从而抑制前列腺癌细胞的增殖。     

Prostate stromal cell-derived hepatocyte growth factor induces invasion of prostate cancer cell line DU145 through tumor-stromal interaction.

BACKGROUND: In prostate cancer, several growth factors derived from stromal cells regulate tumor cell growth. Hepatocyte growth factor (HGF) possesses biological activities that promote cancer proliferation and invasion through tumor-stromal interaction. We examined how prostate stromal cell-derived HGF affects invasion of prostate cancer cells through this interaction. METHODS: The effects of HGF, various growth factors (transforming growth factor (TGF)-alpha, TGF-beta1, basic fibroblast growth factor, keratinocyte growth factor, and platelet-derived growth factor), and conditioned medium (CM) from prostate stromal cells (PrSC) on prostate cancer cells (LNCaP, PC-3, and DU145) were determined by collagen gel invasion assay. DU145 cells and PrSC were cocultured for Matrigel invasion chamber assay. Induction activity of CM from cancer cells to stimulate HGF production by PrSC was studied by the ELISA method and Western blotting. RESULTS: LNCaP and PC-3 cells did not respond to any of the factors examined. Invasion of DU145 cells into the collagen gel matrix was induced by HGF and TGF-beta1, but not by any of the other factors tested. When DU145 cells were cultured in CM from PrSC or cocultured with PrSC, the cells acquired invasive potential, and this invasion was inhibited by an antibody against HGF, but not against TGF-beta1. Native-type HGF production in PrSC was enhanced by some unknown inducer(s) produced by cancer cells. CONCLUSIONS: PrSC-derived HGF enhanced invasive activity of the prostate cancer cell line DU145 through tumor-stromal interaction, wherein DU145 cells secreted some HGF-inducer(s) for PrSC.     

硫利达嗪对胃癌细胞生长的抑制作用与机制研究

目的:探讨硫利达嗪对胃癌细胞体外生长的抑制作用及其机制。方法:不同浓度的硫利达嗪作用于胃癌SGC-7901细胞24 h后,分别用MTT法与流式细胞仪检测胃癌细胞的增殖与凋亡情况,以及用Western blot检测凋亡相关蛋白bax、bcl-2、caspase-3的表达情况。结果:硫利达嗪作用后,SGC-7901细胞的增殖明显抑制,凋亡率明显增加,抗凋亡蛋白bcl-2表达下调、促凋亡蛋白bax表达上调、caspcase-3蛋白表达上调,且均呈浓度依赖性(均P0.05)。结论:硫利达嗪对人胃癌细胞体外的生长有明显抑制作用,该作用可能与其并活化caspase-3依赖的凋亡途径有关。     

Leptin induces cell migration and the expression of growth factors in human prostate cancer cells

BACKGROUND: The adipocyte hormone leptin has been shown to increase migration and angiogenesis in epithelial cells. Therefore, we hypothesized that leptin would induce prostate cancer cell migration and growth factor expression in vitro. METHODS: Prostate cancer cell lines DU145 and PC-3 (androgen-resistant) were treated with leptin over time. Supernatants were assayed for growth factor expression via enzyme-linked immunosorbent assay (ELISA). Becton Dickinson-Falcon Transwell systems were used to assay leptin-induced migration. RESULTS: Leptin significantly induced expression of vascular endothelial growth factor (VEGF), transforming growth factor-beta1 (TGF-beta1), and basic fibroblast growth factor (bFGF) in DU145 and PC-3 cells. Prostate cancer cell migration was enhanced by leptin and inhibited 50% to 70% with the addition of mitogen-activated protein kinase (MAPK) and phosphatidylinositol 3-kinase (PI3K) inhibitors. CONCLUSIONS: The mitogenic effects of leptin on cancer cells, in combination with the increased migration and expression of growth factors, overall likely contributes to the progression of prostate cancer. Therefore, obesity associated with high leptin levels should be considered a risk factor in prostate cancer patients.     

雷帕霉素抑制人前列腺癌细胞增殖及其作用机制

目的 观察雷帕霉素(Rapamycin)对体外培养的人前列腺癌PC-3M-2B4细胞增殖及凋亡的影响,探讨其机制.方法 分别用不同浓度的雷帕霉素(100、200、400、800μg/L)对细胞进行干预后,采用噻唑蓝(MTT)比色法检测细胞增殖变化,流式细胞术检测细胞凋亡变化,Western blot 法检测凋亡相关蛋白bcl-2及bax表达的变化.结果 雷帕霉素能明显抑制PC-3M-2B4细胞的增殖活性,此作用呈现量-效、时-效关系.雷帕霉素呈浓度依赖性诱导细胞凋亡.雷帕霉素作用PC-3M-2B4细胞后,细胞内凋亡抑制蛋白bcl-2的表达明显降低,bax蛋白的表达明显增加.结论 雷帕霉素能够通过调节凋亡相关蛋白bcl-2和bax的表达比例,诱导前列腺癌细胞凋亡,从而抑制肿瘤生长.

Abstract：

Objective To investigate the effects of Rapamycin on the growth and apoptosis of human prostate carcinoma cell line PC-3M-2B4. Methods The inhibitory effect of Rapamycin was observed at 100,200,400,800μg/L on the growth of human prostate carcinoma cell line PC-3M-2B4 in serum-free medium for different concentrations by methyl thiazol tetrazolium (MTF) assays. Flow cytometry (FCM)analysis was used to study the changes of cell apoptosis. The expression level of bcl-2 and bax was determined by Western blotting. Results Rapamycin caused dose-dependent inhibition on the growth of human prostate carcinoma cell line PC-3M-2B4 in a concentration-and time dependent manner. Rapamycin induced the apoptosis of PC-3M-2B4 cells in a concentration-dependent manner. The levels of bcl-2 protein were reduced gradually with the increase of concentration or action time. Conclusion Rapamycin, a mTOR inhibitor, inhibits the growth of human prostate cancer cell and induces apoptosis of human prostate cancer cell. mTOR might be a potential target for anti-prostate cancer.     

多囊蛋白1氨基端肽对常染色体显性多囊肾囊肿衬里上皮细胞增殖和凋亡的影响

目的 探讨多囊蛋白1氨基端肽（PC-1NTP）对常染色体显性多囊肾病（ADPKD）肾囊肿衬里上皮细胞增殖、细胞周期和凋亡的影响.方法 用噻唑蓝（MTT）法检测PC-1NTP对ADPKD囊肿衬里上皮细胞增殖的影响.用流式细胞仪分析细胞周期与凋亡状况.用荧光定量PCR检测细胞中细胞周期素cyclinD1、p21WAF1、bax、bcl-2和小染色体维护蛋白2（MCM-2）的mRNA表达.结果 PC-1NTP使ADPKD囊肿衬里上皮细胞G0/G1期细胞百分比显著增多,S期细胞百分比显著减少,并明显抑制其增殖和凋亡;同时细胞中p21和bcl-2 mRNA表达明显增高（P＜0.01）,cyclinD1、bax和MCM-2的mRNA表达明显减弱（P＜0.01或P＜0.05）.结论 PC-1NTP能明显抑制ADPKD囊肿衬里上皮细胞的增殖和凋亡,减慢细胞周期进程,其作用机制可能是通过调节G1/S关卡调节因子cyclinD1/p21WAF1和凋亡调节蛋白bcl-2/bax的表达实现的.PC-1NTP有希望成为治疗ADPKD的可选药物.     

Vitamin D3 analogue inhibits keratinocyte growth factor signaling and induces apoptosis in human prostate cancer cells.

BACKGROUND: Prostate cancer is a worldwide significant health care problem, due to its high incidence and mortality. In particular, androgen-independent tumors have the worst prognosis, because they are refractory to almost all kinds of available therapy. Hence, there is the need of new treatment opportunities targeting androgen-independent, growth factor-mediated, tumor signaling. One of these new promising opportunities is vitamin D3 and its related analogues. METHODS: We investigated the effect of a vitamin D3 analogue, analogue (V), on proliferation of several human prostate cancer cells in basal condition and after treatment with KGF, one of the intraprostatic growth factors that might participate in the progression of prostate cancer. In addition, in the androgen-independent cell line DU 145, we also studied the effect of analogue (V), KGF, and their mutual interaction on protein tyrosine phosphorylation, bcl-2 expression and apoptosis. RESULTS: Overall, we found that analogue (V) dose-dependently decreased basal and KGF-induced prostate cancer cell growth, although to a different extent. Maximal effect was obtained in DU 145 cells. In these cells, KGF stimulated tyrosine phosphorylation of a protein corresponding to its receptor, induced bcl-2 expression, and prolonged cell survival. Analogue (V) not only counteracted all these KGF-mediated events, but also decreased basal bcl-2 expression, therefore, allowing DU 145 cells to undergo an apoptotic program. CONCLUSIONS: Our results indicated that in prostate cancer cells analogue (V) decreased basal and KGF-induced cell proliferation. This effect, at least in DU 145 cells, is in part mediated by negative interactions with cell survival and KGF signaling.     

bcl-2/bax ratio as a predictive marker for therapeutic response to radiotherapy in patients with prostate cancer

Objectives. Markers predictive of therapeutic response of prostatic tumors to radiotherapy may have major significance in optimizing effective treatment of prostate cancer. Because inherent cellular radioresistance plays a critical role in the failure of radiotherapy, in this study, we investigated whether there is a correlation between the ratio of two apoptosis regulators, bcl-2 (apoptosis suppressor) and bax (apoptosis inducer) in prostatic tumors and the clinical response to radiotherapy in patients with localized prostate cancer.Methods. A retrospective review of records of 41 patients who underwent external beam radiotherapy for prostate cancer was conducted. On the basis of post-treatment prostate biopsy and prostate-specific antigen (PSA) criteria, the cancers of 20 patients were classified as radiation nonresponders and 21 as radiation responders. Immunohistochemical analysis was performed on paraffin-embedded prostate sections to determine the level of expression of the two apoptotic proteins, bcl-2 and bax, in tumor cells.Results. bcl-2 immunoreactivity was significantly higher in prostatic tumors not responsive to radiotherapy (38.6 ± 4.1), compared with the radiation responders (24.1 ± 4.6) (P <0.001). Expression of bax protein was lower in nonresponders, but values were not significantly different from the responders. The resulting significantly higher bcl-2/bax ratio (P <0.01) correlated with poor therapeutic responsiveness of prostate cancer to radiotherapy (1.12 ± 0.12 and 0.56 ± 0.13, for nonresponders and responders, respectively). This correlation (r = 0.67) was independent of age, PSA, and Gleason score.Conclusions. These findings suggest that patients with an elevated bcl-2/bax ratio are at increased risk of their cancer failing to respond to radiotherapy. This study suggests a predictive value for the bcl-2/bax ratio as a potential molecular marker for predicting radioresistance of prostatic tumors.     

Apigenin drives the production of reactive oxygen species and initiates a mitochondrial mediated cell death pathway in prostate epithelial cells

BACKGROUND: Phytoestrogens may reduce tumorigenesis in prostate cancer. We screened five phytoestrogens for their effect on cell growth and apoptosis in PWR-1E, LNCaP, PC-3, and DU145 prostate epithelial cells in vitro. METHODS: We assessed cell number, proliferation, and apoptosis using crystal violet assays, flow cytometric analysis, and TUNEL. Focusing specifically on apigenin we assessed the ability of calpain, serine protease, caspase, estrogen receptor, and ceramide synthase inhibitors to block apigenin induced apoptosis. We also analyzed caspase 3, 7, 8, 9, Bcl-2, Bax, Bid, and cytochrome C by Western analysis, and mitochondrial permeability and reactive oxygen species production by flow cytometry using mitosensor(TM) and DCFH-DA, respectively. RESULTS: Apigenin and silybinin significantly reduced cell number, with apigenin inducing apoptosis in PWR-1E, LNCaP, PC-3, and DU145 cells. The PC-3 and DU145 cells were less susceptible to apigenin induced apoptosis then LNCaP and PWR-1E cells. The induction of apoptosis by apigenin was caspase dependent. Apigenin generated reactive oxygen species, a loss of mitochondrial Bcl-2 expression, mitochondrial permeability, cytochrome C release, and the cleavage of caspase 3, 7, 8, and 9 and the concomitant cleavage of the inhibitor of apoptosis protein, cIAP-2. The overexpression of Bcl-2 in LNCaP B10 cells reduced the apoptotic effects of apigenin. CONCLUSIONS: Apigenin induces cell death in prostate epithelial cells using a mitochondrial mediated cell death pathway. Bcl-2 has a role in inhibiting apigenin induced cell death in prostate epithelial cells.     

Inhibitors of apoptosis proteins in prostate cancer cell lines

BACKGROUND: The caspases are the central executioners of apoptosis. The inhibitors of apoptosis proteins (IAPs) are a family of recently described caspase inhibitors. We hypothesised that tumor resistance to apoptosis could be due in part to IAP expression. METHODS: The expression of NAIP, cIAP-1, cIAP-2, XIAP, and survivin was investigated in the prostate cancer cell lines LNCaP, PC3, and DU145. RNase protection assays and Western blotting were used to assess RNA and protein expression. Apoptotic susceptibility was determined using etoposide and assessed by propidium iodide (PI) DNA incorporation using flow cytometry. RESULTS: DU145 and PC3 cells were more resistant to apoptosis than LNCaP cells. All the IAPs were identified in the cell lines with variation in IAP expression between different cell types. Immunohistochemistry demonstrated cIAP-1 expression in PC3 cells was nuclear, while the expression of cIAP-2 and XIAP was perinuclear. Growing LNCaP cells in charcoal-stripped or androgen-supplemented medium resulted in no alteration in IAP expression. CONCLUSIONS: This study characterises the expression of IAP in three of the most commonly used prostate cancer cells. IAP may make an important contribution to apoptotic resistance in patients with prostate cancer.