树莓体外抑制人肝癌细胞系HepG2生长的实验性研究

目的 研究树莓提取物体外抑制肝癌细胞系HepG2生长的情况;从mRNA水平检测树莓提取物对于肝癌细胞系VEGF表达的影响;探讨树莓提取物抑制肝癌细胞生长的机制.方法 MTT方法 检测树莓提取物对于肝癌细胞系HepG2生长的抑制活性;应用PCR(聚合酶链反应)方法 检测树莓提取物对于肝癌细胞VEGF表达的影响.结果 0.25 mg/mL至10 mg/mL的树莓提取物对肝癌细胞系HepG2的抑制率分别为7.17±1.39%、16.80±2.49%、28.00±1.80%、34.45±1.12%、46.01±1.63%、83.15±2.12%和86.16±1.36%(P<0.05);PCR实验结果 显示:树莓提取物在体外有较强的抑制VEGF表达的能力,随树莓提取物浓度升高,VEGF表达减弱.结论 树莓提取物可以抑制肝癌细胞系增殖、抑制肝癌细胞VEGF的表达.树莓中植物化学物质的上述活性可能是其抑制肝癌细胞生长的机制.     

树突状细胞体外诱导抗肝癌免疫抑制裸鼠人肝癌细胞系HepG2移植瘤发生与生长

为研究树突状细胞(DC)体外诱导抗肿瘤免疫能否预防裸鼠移植瘤发生及抑制裸鼠移植瘤生长,联合应用粒/巨噬细胞集落刺激因子(GM-CSF)及白介素4(IL-4)直接从肝癌患者外周血中培养出DC;以源于人肝癌细胞系HepG2的肿瘤抗原粗提物刺激DC,DC激活同源的T淋巴细胞;以被激活的T淋巴细胞预防性接种于裸鼠皮下,观察进一步接种的人肝癌细胞系HepG2移植瘤发生;观察被激活的T淋巴细胞抑制已接种于裸鼠的人肝癌细胞系HepG2移植瘤生长.发现,DC诱导的抗肿瘤免疫不仅能预防裸鼠人肝癌细胞系HepG2移植瘤发生,而且能抑制该移植瘤生长.提示经肿瘤抗原激活的DC作为一新概念上的抗肿瘤疫苗有可能在肝癌的预防及治疗中发挥重要作用.     

RNA干扰沉默ERK2基因体外诱导HepG2细胞凋亡的实验研究

目的：探讨RNA干扰技术对肝癌HepG2细胞ERK2基因表达的抑制作用及对HepG2细胞凋亡的作用.方法：实验分siRNA干扰质粒转染组、阴性质粒对照组和HepG2细胞空白对照组.应用DNA重组技术,制备针对ERK2基因的重组质粒pAAV-ERK2-siRNA-GFP,通过脂质体转染HepG2细胞.MTT细胞增殖实验,TUNEL法细胞凋亡染色及流式细胞仪凋亡细胞法检测体外诱导HepG2细胞凋亡的作用.结果：成功构建ERK2-siRNA重组质粒.MTT细胞增殖实验,TUNEL法细胞凋亡染色及流式细胞仪凋亡细胞检测法显示该重组质粒在体外能够有效抑制HepG2细胞生长,诱导HepG2细胞凋亡.结论：构建ERK2-siRNA重组质粒能抑制HepG2细胞增殖并诱导HepG2细胞凋亡,为肝癌治疗提供新思路.     

海胆肠提取物体外抗肿瘤活性的研究

目的 :研究海胆肠提取物对体外培养的SGC 790 1人胃腺癌细胞和Bel74 0 2人肝癌细胞系的作用 ,了解其体外抗肿瘤活性 ,并对其作用机制进行初步探讨。方法 :从海胆肠内提取水溶性多糖类成分。应用四氮唑蓝 (MTT)快速比色法测定肿瘤细胞生长抑制率。利用扫描电镜和透射电镜观察提取物对培养细胞超微结构的影响。结果 :MTT实验显示 ,海胆肠提取物对培养的SGC 790 1人胃腺癌细胞和Bel74 0 2人肝癌细胞的生长有明显的抑制作用 ,且抑制程度随浓度升高而增强 ;经海胆肠提取物作用后的细胞 ,透射电镜和扫描电镜观察均显示发生了特征性的形态学改变。结论 :海胆肠提取物在体外可显著抑制人癌细胞株生长 ,其作用机制主要是诱导肿瘤细胞凋亡     

恩度与阿霉素联用对人肝癌HepG2细胞生长的抑制作用

目的:体外观察恩度与阿霉素联用对人肝癌细胞系HepG2细胞增殖的影响.方法:采用MTT 法观察不同浓度阿霉素、恩度及联合应用对HepG2细胞生长的抑制作用,采用流式细胞仪检测上述药物单独或联合应用对HepG2细胞的凋亡及周期影响.结果:化疗药物单独应用时,对HepG2细胞的抑制作用呈明显的剂量效应依赖关系.阿霉素与恩度均可诱导细胞凋亡,阻滞细胞周期,且联用时有协同作用.结论:恩度与化疗药物联合应用时的具有协同效应.     

树突状细胞诱导抗裸鼠人肝癌移植瘤免疫

[目的]讨究树突状细胞（DC）体外诱导的抗肝癌细胞免疫能否预防裸鼠人肝癌移植癌发生及抑制课鼠人肝癌移植瘤生长。[方法]联合应用粒／巨噬细胞集落刺激因子（GM－CSF）及白介素－4（IL－4）直接从肝癌患者外周血中培养DC：以人肝癌细胞系HepG2肿瘤细胞的肿瘤抗原粗提物刺激DC；DC激活同源的T淋巴细胞产生细胞毒性T淋巴细胞（CYL）；以DC激活的CIL预防性接种于探鼠皮下观察其后接种的人肝癌细胞系HepG2移植瘤发生率；并观察该CTL对已接种裸鼠的人肝癌细胞系HepG2移植瘤生长的影响。[结果]DC诱导的工体外对HepG2肿瘤细胞具有高效而特异杀伤作用，在裸鼠体内不能预防探鼠人肝癌细胞系HepG2移植瘤发生，而且能抑制该移植瘤生长。[结论]经肝癌肿瘤相关抗原激活的肝癌患者外用血DC体外诱导的抗肿瘤细胞免疫在体内外均具有良好的抗肿瘤作用。提示DC作为一新概念上的抗肿瘤疫苗有可能在人肝癌的预防及治疗中发挥重要作用。     

钩藤酸E对人肝癌HepG2细胞的抑制作用及其机制研究

目的:通过体外实验研究钩藤酸E(UAE)抑制人肝癌细胞HepG2的细胞增殖作用,并进一步研究其作用机制.方法:MTT法考察钩藤酸E对人肝癌HepG2细胞的生长抑制作用,通过电镜观察细胞微细结构变化,流式细胞术进一步验证细胞凋亡,并检测细胞周期变化.结果:研究发现UAE能抑制HepG2细胞增殖.电镜分析证实UAE引起细胞凋亡.流式细胞术检测细胞凋亡主要发生于G0/G1期.结论:钩藤酸E诱导人肝癌细胞HepG2的细胞增殖,诱导其发生凋亡.     

D-氨基葡萄糖衍生物抑制HepG2细胞生长及诱导凋亡的初步研究

目的研究D-葡萄糖衍生物对人肝癌HepG2细胞生长增殖的影响,进而探讨D-葡萄糖衍生物诱导HepG2细胞凋亡的作用.方法用不同浓度的D-葡萄糖衍生物处理HepG2,采用MTT法测定其对HepG2细胞生长增殖的抑制作用,通过荧光显微镜、透射电镜、流式细胞术和DNA琼脂糖凝胶电泳观察凋亡细胞的形态结构及其对HepG2细胞诱导凋亡的效应.结果D-氨基葡萄糖衍生物能抑制HepG2细胞的生长增殖,并呈一定的浓度、时间依赖性;D-氨基葡萄糖衍生物能诱导肝癌细胞凋亡;荧光显微镜、透射电镜可见肝癌细胞出现典型的凋亡细胞形态学改变;DNA琼脂糖凝胶电泳呈现典型的DNA梯形条带图谱;流式细胞仪检测出现典型的亚二倍体凋亡峰.结论D-氨基葡萄糖衍生物在体外可显著抑制人肝癌细胞株生长,其作用机制主要是诱导肿瘤细胞凋亡.     

海胆肠提取物体外抗肿瘤活性的研究

目的：研究海胆肠提取物对体外培养的SGC-7901人胃腺癌细胞和Bel7402人肝癌细胞系的作用，了解其体外抗肿瘤活性，并对其作用机制进行初步探讨。方法：从海胆肠内提取水溶性多糖类成分。应用四氮唑蓝(MTT)快速比色法测定肿瘤细胞生长抑制率。利用扫描电镜和透射电镜观察提取物对培养细胞超微结构的影响。结果：MTT实验显示，海胆肠提取物对培养的SGC-7901人胃腺癌细胞和Bel7402人肝癌细胞的生长有明显的抑制作用，且抑制程度随浓度升高而增强；经海胆肠提取物作用后的细胞，透射电镜和扫描电镜观察均显示发生了特征性的形态学改变。结论：海胆肠提取物在体外可显著抑制人癌细胞株生长，其作用机制主要是诱导肿瘤细胞凋亡。     

D-氨基葡萄糖衍生物抑制HepG2细胞生长及诱导凋亡的初步研究

目的：研究D-葡萄糖衍生物对人肝癌HepG2细胞生长增殖的影响,进而探讨D-葡萄糖衍生物诱导HepG2细胞凋亡的作用.方法：用不同浓度的D-葡萄糖衍生物处理HepG2,采用MTT法测定其对HepG2细胞生长增殖的抑制作用,通过荧光显微镜、透射电镜、流式细胞术和DNA琼脂糖凝胶电泳观察凋亡细胞的形态结构及其对HepG2细胞诱导凋亡的效应.结果：D-氨基葡萄糖衍生物能抑制HepG2细胞的生长增殖,并呈一定的浓度、时间依赖性;D-氨基葡萄糖衍生物能诱导肝癌细胞凋亡;荧光显微镜、透射电镜可见肝癌细胞出现典型的凋亡细胞形态学改变;DNA琼脂糖凝胶电泳呈现典型的DNA梯形条带图谱;流式细胞仪检测出现典型的亚二倍体凋亡峰.结论：D-氨基葡萄糖衍生物在体外可显著抑制人肝癌细胞株生长,其作用机制主要是诱导肿瘤细胞凋亡.     

Antitumor Activities of Apple Extracts

OBJECTIVE To examine the antitumor activities of fresh apple extracts. METHODS Fuji apple extracts were tested for their anti-LS~174T-pro-liferative activities, for their effect on expression of PCNA, and ability to induce apoptosis in a LS-174T cell line. RESULTS Apple extracts inhibited LS-174T cellular proliferation in a concentration and time dependent manner. The apple extracts equivalent to a concentration of 50 mg/ml inhibited the proliferation of the LS-174T cells by 34.5±1.2% after 48 h and 47.5±1.8% after 72 h respectively. Apple extracts inhibited PCNA expression and induced apoptosis of the LS-174T cells at concentrations above 12.5mg/ml. CONCLUSION Apple extracts can inhibit PCNA expression and induction of apoptosis in LS-174T cells which may contribute to their inhibitio-ry effect on cellular proliferation.     

Inhibition of human tumor xenograft growth in nude mice by a novel monoclonal anti-HSPG isolated from human liver

Heparan sulfate proteoglycans (HSPGs) were isolated from normal human liver and α monoclonal antibody (MAb) was raised against them. Preliminary studies showed that MAb clone 1E4-1C2 was able to react with many cell lines tested, including hematopoietic cells and solid tumors. MAb1E4-1C2 was used to study whether HSPG was involved in growth and proliferation of human liver cancer using hepatocellular carcinoma (HCC) cell line (HepG2) as a model. Inhibition by MAb1E4-1C2 of HepG2 cell proliferation was studied in vitro by MTT assay. For in vivo assay, xenograft induction in athymic mice was performed. The results showed that MAb1E4-1C2 inhibited proliferation of HepG2 cells significantly, compared to isotype and medium control. MAb1E4-1C2 also suppressed the growth of tumor, resulting in smaller tumor size and weight. The investigation also showed that MAb1E4-1C2 inhibited proliferation and restricted tumor growth through the induction of apoptosis. The results suggest that HSPG might be involved in liver cancer cell proliferation. Therefore, a specific MAb that was raised against liver HSPG might be an alternative therapeutic agent for the treatment of human liver cancer.     

Clinacanthus Nutans Hexane Extracts Induce Apoptosis Through a Caspase-Dependent Pathway in Human Cancer Cell Lines

Background: Clinacanthus nutans (C.nutans) is a plant consumed as a cancer treatment in tropical Asia. Despite the availability of numerous anecdotal reports, evaluation of active anticancer effects has remained elusive. Therefore we here examined antiproliferative, reactive oxygen species (ROS)-inducing and apoptosis mechanisms of whole plant extracts in different cancer cell lines. Methods: Antiproliferative actions of five solvent extracts (hexane, chloroform, ethyl acetate, methanol and water) of C.nutans were tested on non-small cell lung cancer (A549), nasopharygeal cancer (CNE1) and liver cancer (HepG2) cells using MTT assay. The most potent anticancer extract was then assessed by flow cytometry to study cell cycle changes . Intracellular levels of ROS were quantified by DCFH-DA assay. Involvement of the caspase pathway in induction of apoptosis was assessed using caspase assay kits. GC-MS analysis was performed to identify phytoconstituents in the extracts. Results: Hexane and chloroform extracts were antiproliferative against all three cell lines, while the ethyl acetate extract, at 300 μg/mL, was antiproliferative in the CNE1 but not A549 and HepG2 cases. Methanol and water extracts did not inhibit cancer cell proliferation. The most potent anticancer hexane extract was selected for further testing. It induced apoptosis in all three cell lines as shown by an increase in the percentage of cell in sub-G1 phase. Dose-dependent increase in ROS levels in all three cell lines indicated apoptosis to be possibly modulated by oxidative stress. At high concentrations (>100 μg/mL), hexane extracts upregulated caspases 8, 9 and 3/7 across all three cell lines. GC-MS analysis of the hexane extract revealed abundance of 31 compounds. Conclusion : Among the five extracts of C.nutans, that with hexane extract demonstrated the highest antiproliferative activity against all three cancer cell lines tested. Action appeared to be via ion of intracellular ROS, and induction of apoptosis via intrinsic and extrinsic caspase pathways.     

miR-526b-3p通过靶基因TNKS2调控肝癌细胞增殖、侵袭、迁移的分子机制探讨

目的:探讨miR-526b-3p对肝癌细胞增殖、迁移及侵袭的影响及其潜在的作用机制。方法:运用qRT-PCR检测人肝癌细胞HepG2、SMMC-7721、BEL-7402和正常肝细胞L02中miR-526b-3p和TNKS2的mRNA表达水平。建立miR-526b-3p过表达和TNKS2抑制表达的HepG2细胞株,采用MTT法检测细胞增殖活力,Transwell小室检测细胞的迁移及侵袭能力,Western blot检测TNKS2蛋白的表达水平;双荧光素酶报告基因分析法验证miR-526b-3p可能的靶基因。结果:与正常肝细胞L02相比,肝癌细胞HepG2、SMMC-7721及BEL-7402中miR-526b-3p的表达显著降低（P<0.05）,TNKS2的表达显著升高（P<0.05）。过表达miR-526b-3p或抑制表达TNKS2均可抑制HepG2细胞的增殖、迁移及侵袭（P<0.05）。TNKS2是miR-526b-3p的靶基因。过表达TNKS2可部分逆转过表达miR-526b-3p对HepG2细胞增殖、迁移及侵袭的抑制作用。结论:miR-526b-3p可通过下调TNKS2的表达,从而抑制肝癌细胞的增殖、迁移及侵袭能力。     

干扰PRMT2基因对肝癌细胞系生物学行为的影响

目的:研究抑制蛋白质精氨酸甲基转移酶2(protein arginine methyltransferase 2,PRMT2)基因的表达对肝癌细胞系生物学行为的影响。方法:qRT-PCR检测人肝癌细胞系(HepG2和SMMC-7721)和人正常肝细胞(LO2)中PRMT2的表达情况;设计并合成针对PRMT2基因的3对小干扰RNA,体外通过脂质体Lipofectamine 3000转染到相对高表达PRMT2的人肝癌细胞系HepG2和SMMC-7721中抑制PRMT2基因的表达,并设置阴性对照组（NC组）和空白对照组。qRT-PCR、Western blot检测转染后细胞PRMT2 mRNA水平和蛋白水平的变化。CCK-8法检测转染后细胞增殖能力,Transwell小室检测细胞体外迁移及侵袭能力。结果:与人正常肝细胞LO2相比,PRMT2基因在人肝癌细胞系HepG2和SMMC-7721中相对高表达。将siPRMT2转染HepG2和SMMC-7721细胞后,与NC组相比,siPRMT2-1和siPRMT2-2组的PRMT2 mRNA和蛋白表达均明显降低（P均<0.05）,NC组与空白对照组无明显差异。与NC组相比,将siPRMT2-1和siPRMT2-2转染进HepG2和SMMC-7721细胞后,细胞增殖速度减慢、迁移和侵袭能力显著减弱（P均<0.05）,NC组与空白对照组无明显差异。结论:RNA干扰抑制PRMT2表达后,抑制肝癌细胞的增殖、迁移和侵袭,PRMT2可能影响肝癌的预后。     

Antitumour effects of a protease inhibitor,nelfinavir, in hepatocellular carcinoma cancer cells

Abstract

Nelfinavir is a protease inhibitor with potential antitumour activity against certain cancer types. The objective of this study was to evaluate the antitumour effects of nelfinavir in hepatocellular carcinoma cell lines, HepG2 and WCH-17. Results indicate that nelfinavir inhibited the proliferation of Hep G2 and WCH-17 cell lines, with IC50 of 5·1 and 62·0 μmol/l, respectively. Nelfinavir induced apoptosis in both cell lines, although the extent as indicated by Annexin V staining varied. The concentration of nelfinavir needed to induce apoptosis in liver cancer cells were 10 and 100 μmol/l for HepG2 and WCH-17, respectively. At the same concentrations, nelfinavir induced cell cycle arrest at G0/G1 phase in HepG2 and WCH-17 cell lines. Our results suggest that nelfinavir inhibit hepatoma cell growth, through the induction of apoptosis and cell cycle arrest. However, the clinical relevance of these findings warrants further investigation.     

Effects of Genistein and Synergistic Action in Combination with Tamoxifen on the HepG2 Human Hepatocellular Carcinoma Cell Line

Introduction: The flavonoids comprise a diverse group of polyphenolic compounds with antioxidant activity that is present in edible plants like soybeans and soy products. In vivo studies have concentrated on the effects of flavonoids on cancer and genistein (GE), a soy-derived isoflavone, has been reported to reduce prostate, colon, hepatic and breast adenocarcinoma risk. Tamoxifen (TAM) is an important drug for cancer treatment worldwide, which can induce apoptosis in various cancers, including examples in the liver, breast and ovaries. The aim of the present study was to evaluate the effects of GE and TAM, alone and in combination, on proliferation and apoptosis in the human hepatocellular carcinoma (HCC) HepG2 cell line. Materials and Methods: HepG 2 cells were treated with GE, TAM and GE/TAM and then MTT and flow cytometry assays were conducted to determine effects on viability and apoptosis, respectively. Results: GE and TAM inhibited cell proliferation and induced apoptosis in the HepG 2 cell lines. Discussion: Our findings clearly indicated that GE and TAM may exert inhibitory and apoptotic effects in liver cancer cells. Conclusion: GE and TAM can significantly inhibit growth of HCC cells and play a significant role in apoptosis.