茶多酚与茶色素对大鼠肝癌前病变组织细胞周期调节因子的影响

目的 利用大鼠肝癌前病变模型探讨茶多酚和茶色素对细胞周期因子的影响。方法 采用改良的Solt-Farber大鼠肝癌癌前病变模型,分别饮用0．1％和0．1％茶多酚,喂养56d。采用Westernblot方法观察细胞周期素D1、D21^WAF1/CIPI、GADD45和PCNA蛋白的表达;用RT－PCR在mRNA水平观察Ckd4的表达。结果 茶多酚和茶色素显著抑制细胞周期素D1、Ckd4和PCNA表达,诱导P21^WAF1／CIP1和GAD45蛋白表达。结论 茶多酚和茶色素通过调节细胞周期调节因子而诱导细胞周期阻滞,抑制细胞过度增值。     

曲古抑菌素A对鼻咽癌CNE3细胞增殖抑制作用

目的 探讨曲古抑菌素A(TSA)对鼻咽癌CNE3细胞周期阻滞与凋亡影响及作用机制.方法 以不同浓度TSA作用鼻咽癌CNE3细胞株,采用甲基噻唑基四唑(MTT)法检测细胞生长抑制率;流式细胞术测定TSA作用前后鼻咽癌细胞调亡情况及细胞周期变化;逆转录-聚合酶链反应(RT-PCR)分析鼻咽癌细胞中细胞周期素依赖性蛋白激酶相互作用蛋白1(p21)mRNA表达的变化.结果 100～500 nmol/L TSA对CNE3细胞生长有明显抑制作用,呈剂量依赖性,抑制率为17.74%～44.33%;400,500 nmol/L TSA可在DNA合成期(S)、DNA合成后期/有丝分裂期(G2/M)诱导CNE3细胞周期阻滞,并诱导细胞凋亡,24 h凋亡率为(14.67±0.21)%,(18.73±1.80)%,48 h凋亡率分别为(16.3±0.96)%,(39.17±1.27)%,与对照组(9.16±1.05)%比较差异均有统计学意义(P<0.05);TSA可诱导CNE3细胞内p21基因表达.结论 TSA可明显抑制CNE3细胞增殖,诱导细胞凋亡,其作用机制与p21基因异常表达有关.     

蛋白激酶B与卵巢癌化疗耐药性研究进展

蛋白激酶B(Akt/PKB)是卵巢癌化疗敏感性的关键调控因子,可促进细胞周期运行,血管形成及细胞的生长,抑制细胞凋亡。其过度表达和活化在卵巢癌化疗耐药性的产生中起着重要作用。Akt通过影响p53功能、线粒体凋亡及JNK/p38信号传导等多种途径抑制化疗药物诱导的卵巢癌细胞凋亡,同时还可通过调节抑癌基因PTEN及X-连锁的凋亡抑制蛋白(XIAP)导致化疗耐药的产生。因此,加深对Akt的研究有助于进一步明确卵巢癌化疗耐药产生的机制,并使之成为卵巢癌化疗的有效作用靶点。     

蛋白激酶B与卵巢癌化疗耐药性研究进展

蛋白激酶B(Akt/PKB)是卵巢癌化疗敏感性的关键调控因子,可促进细胞周期运行,血管形成及细胞的生长.抑制细胞凋亡.其过度表达和活化在卵巢癌化疗耐药性的产生中起着重要作用.Akt通过影响p53功能、线粒体凋亡及JNK/p38信号传导等多种途径抑制化疗药物诱导的卵巢癌细胞凋亡,同时还可通过调节抑癌基因PTEN及X-连锁的凋亡抑制蛋白(XIAP)导致化疗耐药的产生.因此,加深对Akt的研究有助于进一步明确卵巢癌化疗耐药产生的机制.并使之成为卵巢癌化疗的有效作用靶点.     

石英致MAPK/cyclinD1-CDK4信号转导通路的活化

目的探讨石英诱导的人胚肺成纤维细胞(HELF)中丝裂素活化蛋白激酶(MAPK)/细胞周期蛋白和细胞周期蛋白依赖激酶(cyclin D1-CDK4)信号转导通路的活化。方法两种处理方式:(1)石英刺激细胞2h后,收获细胞;(2)石英长时间(2个月)作用于细胞,使细胞具有部分转化细胞的特征(S-HELF),检测信号蛋白因子和细胞周期的变化。分别采用Western blot、免疫细胞化学和流式细胞术方法检测细胞内信号蛋白和细胞周期的改变。选用特异化学抑制剂或分子抑制剂抑制上游激酶,检测下游激酶的变化。结果HELF暴露于石英粉尘2 h后,可以导致MAPK家族中ERK1/2、p38和JNK1/2 3个亚家族的磷酸化水平升高。在S-HELF中,只有细胞外调节蛋白激酶(ERK)和C-Jun氨基末端激酶[JNK1(p46)]较未处理的HELF磷酸化水平增高,而JNK2(p54)的磷酸化水平没有变化,p38的磷酸化水平反而下降。cyclin D1和CDK4蛋白在S-HELF中较HELF中表达增多。抑制ERK和JNK活化或者抑制核转录因子活化蛋白1(AP- 1)的活化后,S-HELF中cyclin D1和CDK4蛋白过表达得到控制。而抑制p38的活性不能改变cyclin D1和CDK4蛋白过表达。结论石英刺激HELF2h后可诱导ERK、JNK和p38的活化,而S-HELF中ERK、JNK活化,p38没有被活化。S-HELF中,cyclin D1和CDK4蛋白质过表达与ERK、JNK和AP-1活化有关。     

茶多酚对人肝癌细胞激活蛋白1及细胞周期影响

目的 观察茶多酚对人肝癌细胞激活蛋白1（AP—1）及细胞周期的影响。方法 构建AP-1报告质粒测量AP-1活性，免疫组化测量细胞周期素D1和细胞周期素依赖激酶4表达变化，流式细胞仪分析细胞周期。结果 茶多酚抑制AP-1活性，细胞周期索Dl和细胞周期素依赖激酶4表达降低，细胞阻滞在Gl期。结论 茶多酚可能通过下调AP-1活性使细胞周期索Dl表达降低；而茶多酚使细胞周期素依赖激酶4表达降低，可能与AP-1的活性无关。     

汉黄芩素增敏TNF促肿瘤细胞凋亡的实验研究

摘要:目的　研究JNK通路在汉黄芩素增敏TNF促进肺癌干细胞凋亡的作用机制。方法　采用汉黄芩素和TNF单独或联合用药,LDH检测细胞凋亡,免疫蛋白印记（Western Bloting,WB）检测JNK磷酸化水平和凋亡蛋白的表达。结果　汉黄芩素显著增强TNF诱导的抗肿瘤作用,并且这个作用通过促进TNF诱导的JNK 通路的磷酸化来实现。同时伴随潜在的TNF诱导的caspase 8活化,引起凋亡的启动。凋亡抑制因子zVAD-fmk和JNK通路抑制因子SP600125均可以扭转这种凋亡。更为重要的是,汉黄芩素对于正常的支气管上皮细胞则不会出现增敏TNF诱导细胞凋亡的作用。结论　汉黄芩素可能通过促进JNK通路磷酸化增敏TNF抗肺癌作用。     

TNF-α诱导人蜕膜细胞凋亡基因表达谱的分析

目的:研究TNF-α诱导人蜕膜细胞异常凋亡后基因表达谱的变化,为了解病理妊娠蜕膜细胞基因表达及药物治疗研究提供基础.方法:体外常规进行人蜕膜细胞培养,选用适宜浓度的TNF-α诱导建立人蜕膜细胞异常凋亡模型,采用人全基因组17KcDNA基因芯片分析TNF-α作用后的基因表达谱变化情况,分析基因的变化趋势.结果:TNF-α诱导蜕膜细胞后检出基因9 320条,上调基因36条,下调基因48条.表达上调的已知基因主要为细胞周期依赖性蛋白激酶和抑制剂基因、DNA损伤及修复基因,另外还有氧化压力和炎症损伤基因等.表达下调的基因与细胞营养、细胞生长、细胞周期及信号转导相关.结论:TNF-α诱导蜕膜细胞异常凋亡.     

金雀异黄素对胶质瘤细胞株细胞生长周期影响

目的 研究金雀异黄素(genistein)对人胶质瘤U251MG细胞生长和细胞周期的影响.方法采用噻唑蓝(MTT)比色法观察不同浓度genistein在不同作用时间下对U251MG细胞生长的影响,用流式细胞仪分析细胞周期分布,蛋白免疫印迹( western blotting)技术检测细胞周期素Bl(cyclinBl)和细胞周期素依赖性蛋白激酶1(CDK1)蛋白表达.结果Genistein对胶质瘤细胞生长有明显抑制作用,使细胞生长停滞于G2/M期,并使细胞cyclin B1、CDK1蛋白表达下降(均P＜0.01),且呈剂量依赖性.结论Genistein对U251 MG细胞生长有明显抑制作用,诱导G2/M期阻滞可能与下调cyclinB1和CDK1蛋白表达有关.     

姜黄素抗肿瘤作用的研究进展

姜黄素是一类无毒副作用、安全多效的天然活性物质,具有抑制肿瘤细胞增殖、诱导肿瘤细胞凋亡的作用。其抗肿瘤机制主要有：在基因水平上抑制真核转录因子核因子-κB活化、调控癌基因蛋白和凋亡调控蛋白;在细胞水平上诱导细胞周期停滞、诱导细胞凋亡;在组织水平上抑制肿瘤血管生成等。目前的研究已经发现姜黄素可预防和拮抗消化道肿瘤,抑制乳腺癌MCF-7细胞增殖,此外,姜黄素对鼻咽癌、肺腺癌、前列腺癌等多种肿瘤的治疗作用也有报道。本文就姜黄素在抗肿瘤作用的研究进展进行综述。     

番茄红素对前列腺癌细胞PC-3增殖和细胞周期的影响

目的研究番茄红素对体外培养的雄激素非依赖性前列腺癌细胞PC-3的抑制作用及其可能的作用机制。方法采用MTT法和H3-TdR掺入法观察番茄红素对癌细胞增殖的影响,流式细胞仪观察同步化的细胞经番茄红素作用后细胞周期及凋亡的变化,RT-PCR检测cyclin D1、bcl-2、bax的mRNA的表达的变化。结果番茄红素抑制PC-3细胞的增殖和DNA合成、诱导其凋亡、改变细胞周期分布(使G0/G1期细胞增多、而S期和G2/M期细胞减少)。RT-PCR结果显示,cyclin D1和bcl-2的mRNA表达水平下调,而bax的mRNA表达水平上调。上述作用呈剂量效应关系。结论番茄红素可诱导PC-3细胞凋亡、改变细胞周期分布、影响cyclin D1、bcl-2、bax的mRNA的表达,从而抑制肿瘤细胞增殖。     

活化蛋白-1和细胞周期蛋白在石英诱导的细胞周期改变中的作用

目的 探讨在石英刺激的人胚肺成纤维细胞(HELF)中AP-1/细胞周期蛋白D1 (cyclin D1)通路在细胞周期改变中的作用.方法 建立稳定转染空载体PXJ的HELF系(HELF-PXJ)及空载体PXJ与反义cyclin D1质粒或反义细胞周期蛋白依赖激酶(CDK4)质粒分别共转染的HELF系(简称anti-D1和anti-K4),将HELF-PXJ、anti-D1和anti-K4细胞分为对照组和石英刺激组(共6组),各对照组不加任何处理,石英刺激组用200 μg/ml石英处理细胞24h.检测AP-1对石英诱导的cyclin D1、CDK和E2F-4表达改变的影响.AP-1的化学抑制剂姜黄素(curcumin) 20μmol/L预处理细胞1h后,200 μg/ml石英刺激24 h,将HELF分为4组,分别为HELF对照组、HELF+石英组、HELF+curcumin对照组、HELF+curcumin+石英组.用免疫印迹(Western blot)法和免疫荧光法检测cyclin D1、CDK4和E2F-1/4蛋白表达;运用反义RNA技术证明通路的上下游关系;用流式细胞术检测细胞周期变化.结果 200 μg/ml石英处理HELF细胞,cyclin D1和CDK4蛋白表达升高,E2F-4蛋白表达明显降低,而E2F-1蛋白的表达没有明显改变.HELF-PXJ+石英组G1期细胞所占比例下降,S期细胞所占比例升高,与HELF-PXJ对照组的差异有统计学意义(P＜0.05).抑制cyclin D1或CDK4表达后,与对照组比较,anti-D1+石英组和anti-K4+石英组G1期细胞和S期细胞百分比无明显变化.抑制AP-1活性,与HELF+石英组比较,HELF+curcumin+石英组cyclin D1和CDK表达均减低,E2F-4表达增多.结论200μg/ml石英可诱导cyclin D1和CDK4蛋白表达增高,E2F-4蛋白表达减少,并通过AP-1/cyclin D1通路诱导细胞周期改变.     

细胞周期蛋白D1在石英致人细胞恶性转化中的作用

目的探讨细胞周期蛋白D1在石英致人胚肺成纤维细胞(HELF)恶性转化过程中所起的作用。方法采用基因重组与基因导入的方法将表达正义和反义细胞周期蛋白D1 RNA的pXJ41-细胞周期蛋白D1导入石英恶性转化HELF细胞中。采用原位杂交和免疫组化的方法检测细胞中细胞周期蛋白D1基因表达情况,分析细胞周期蛋白D1导入前后细胞生长速度、倍增时间、细胞周期分布、软琼脂克隆形成能力的改变。结果石英诱导HELF细胞恶性转化过程中细胞周期蛋白D1基因过表达,反义细胞周期蛋白D1 RNA可抑制石英恶性转化细胞的生长增殖。与石英恶性转化细胞相比,反义pXJ41-细胞周期蛋白D1转染细胞培养至第8天时,生长速率下降58．69％,倍增时间从21．0h延长到31．4h,G1期细胞比例从45．1％增加到52．7％,S期细胞比例由40．3％下降到33．1％,克隆形成率显著下降,克隆明显变小。结论细胞周期蛋白D1的异常表达与石英恶性转化细胞有密切的关系,高水平表达的细胞周期蛋白D1对维持恶性转化细胞的恶性性状起重要的作用。     

Immunomodulatory Effects of Dietary Polyphenols

Functional and nutraceutical foods provide an alternative way to improve immune function to aid in the management of various diseases. Traditionally, many medicinal products have been derived from natural compounds with healing properties. With the development of research into nutraceuticals, it is becoming apparent that many of the beneficial properties of these compounds are at least partly due to the presence of polyphenols. There is evidence that dietary polyphenols can influence dendritic cells, have an immunomodulatory effect on macrophages, increase proliferation of B cells, T cells and suppress Type 1 T helper (Th1), Th2, Th17 and Th9 cells. Polyphenols reduce inflammation by suppressing the pro-inflammatory cytokines in inflammatory bowel disease by inducing Treg cells in the intestine, inhibition of tumor necrosis factor-alpha (TNF-α) and induction of apoptosis, decreasing DNA damage. Polyphenols have a potential role in prevention/treatment of auto-immune diseases like type 1 diabetes, rheumatoid arthritis and multiple sclerosis by regulating signaling pathways, suppressing inflammation and limiting demyelination. In addition, polyphenols cause immunomodulatory effects against allergic reaction and autoimmune disease by inhibition of autoimmune T cell proliferation and downregulation of pro-inflammatory cytokines (interleukin-6 (IL-6), IL-1, interferon-γ (IFN-γ)). Herein, we summarize the immunomodulatory effects of polyphenols and the underlying mechanisms involved in the stimulation of immune responses.     

茶多酚和茶色素对肝癌细胞株HepG2细胞周期的影响

为探讨茶对肝癌细胞周期的影响 ,体外培养人肝癌细胞株HepG2 ,加入终浓度为 50mg L和 1 0 0mg L茶多酚和茶色素作用 48h后 ,流式细胞仪分析DNA含量 ,Westernblot观察对细胞周期蛋白P2 1 WAF1 CIP1 和细胞周期素D1 (cyclinD1 ) ,RT PCR检测细胞周期素依赖激酶 4 (Cdk4)在mRNA水平上的表达情况。结果表明 ,茶多酚和茶色素引起了细胞周期G1 期阻滞 (G1 arrest) ,抑制了cyclinD1蛋白的表达 ,诱导了P2 1 WAF1 CIP1 蛋白的表达增加 ,并且显著抑制了Cdk4在mRNA水平上的表达。因此 ,诱导细胞周期阻滞可能是茶预防肿瘤的一个重要机制     

The Effect of Different Treatments of (–)-Epigallocatechin-3-Gallate on Colorectal Carcinoma Cell Lines

Backgroud: (-)-Epigallocatechin-3-gallate (EGCG), the major component of green tea, is well documented to induce apoptosis and cell cycle arrest in cancer by targeting multiple signal transduction pathways. However, EGCG is extremely unstable in general culture conditions and rapidly degraded. So, to what extent EGCG or which degradation products of EGCG play a role in anti-tumor is still unknown. In this study, we evaluated the effect of different treatments of EGCG on HCT116 cells.

Design: MTT assay was applied to evaluated the inhibitory effect of different treatments of EGCG on HCT116 cells. Cell cycle and apoptosis were performed by flow cytometry. Finally, western blot analysis was used to elucidate the molecular mechanism associated with cell cycle arrest and apoptosis.

Results: Compared with control, both EGCG and O-EGCG (i.e., EGCG being pre-incubated at 37°C for 3 h) significantly inhibited HCT116 cells growth. Surprisingly, we found that the inhibitory effect of O-EGCG was stronger than that of EGCG. The IC₅₀ values of EGCG and O-EGCG were 8.75 and 5.40 μM, respectively. Cell cycle analysis showed that 20 μM of EGCG simultaneously caused cell cycle arrest at G1 and G2 phase in HCT116 cells, differing from O-EGCG which exclusively caused cell cycle arrest at G2. This result suggested that parent EGCG at the early treatment might cause cell cycle arrest at G1. As time went on, EGCG disappeared and degraded products of EGCG were formed which might cause cell cycle arrest at G2. Further studies revealed that EGCG induced cell cycle arrest at G1 by downregulation of cyclin E and cyclin D1 and upregulation of p21. On the other hand, O-EGCG induced HCT116 cells apoptosis mainly by increasing the expression of p53 and cleaved caspase-3, which might be the underlying reason why O-EGCG had stronger inhibitory effect on HCT116 cells line than EGCG.

Conclusions: The pretreatment of EGCG may be an effective way to enhance its antitumor effect.     

Lycopene differentially induces quiescence and apoptosis in androgen-responsive and -independent prostate cancer cell lines

BACKGROUND & AIMS: Lycopene has been credited with a number of health benefits including a decrease in prostate cancer risk. Our study investigates the molecular mechanism underlying anti-cancer activity of lycopene-based products in androgen-responsive (LNCaP) and androgen-independent (PC3) cells. METHODS: The effect of lycopene-based agents on prostate cancer growth and survival were examined using proliferation assays, bromodeoxyuridine incorporation and flow cytometric analysis of cellular DNA content. Biochemical effects of lycopene treatment were investigated by immunoblotting for changes in the absolute levels and phosphorylation states of cell cycle regulatory and signalling proteins. RESULTS: LNCaP and PC3 cells treated with the lycopene-based agents undergo mitotic arrest, accumulating in G0/G1 phase. Immunoblot screening indicated that lycopene's antiproliferative effects are likely achieved through a block in G1/S transition mediated by decreased levels of cyclins D1 and E and cyclin dependent kinase 4 and suppressed Retinoblastoma phosphorylation. These responses correlated with decreased insulin-like growth factor-I receptor expression and activation, increased insulin-like growth factor binding protein 2 expression and decreased AKT activation. Exposure to lycopene at doses as low as 10 nM for 48 h induced a profound apoptotic response in LNCaP cells. In contrast PC3 cells were resistant to apoptosis at doses up to 1 microM. CONCLUSIONS: Lycopene exposure can suppress phosphatidylinositol 3-kinase-dependent proliferative and survival signalling in androgen-responsive LNCaP and androgen-independent PC3 cells suggesting that the molecular mechanisms for the cytostatic and cytotoxic actions of lycopene involve induction of G0/G1 cell cycle arrest. This study supports further examination of lycopene as a potential agent for both the prevention and treatment of prostate cancer.