紫杉醇对肝癌HepG2细胞增殖及凋亡的影响

目的观察紫杉醇对肝癌HepG2细胞增殖及凋亡的影响。方法取对数生长期人肝癌HepG2细胞,分别置入5、10、20 nmol/L紫杉醇共同培养48 h（实验组）,另设阴性对照组。采用MTT法测定细胞增殖活性;Ho-echst33342荧光染色法观察紫杉醇处理后细胞形态的变化;DNA琼脂糖凝胶电泳检测DNA梯状条带;免疫组织化学法检测细胞中凋亡相关蛋白cFLIP、Survivin、Caspase-3的表达。结果紫杉醇浓度分别为5、10、20 nmol/L时,细胞增殖活性分别为0.51±0.02、0.49±0.05、0.38±0.09,与阴性对照组（0.61±0.07）比较P〈0.05或〈0.01;荧光显微镜观察可见典型的细胞凋亡形态学改变,以20 nmol/L紫杉醇作用最明显;DNA琼脂糖凝胶电泳显示,实验组细胞均可见典型的＂梯状＂条带;与阴性对照组比较,实验组能显著抑制cFLIP、Survivin蛋白的表达,促进Caspase-3蛋白的表达（P〈0.05或〈0.01）。结论紫杉醇可能通过下调cFLIP、Survivin蛋白的表达及上调Caspase-3蛋白的表达诱导人肝癌HepG2细胞凋亡。     

沙利度胺对人肝癌细胞株体外生长的影响

对人肝癌HepG2细胞株进行体外培养,分为空白对照组、阿霉素组、沙利度胺组、联合用药组,不同浓度的沙利度胺作用不同时间,应用MTT方法及流式细胞术分别观察沙利度胺单独及联合阿霉素对HepG2细胞增殖和凋亡影响。结果沙利度胺在200μg/ml对HepG2细胞有明显抑制作用,以48 h抑制率最高,联合用药组高于沙利度胺、阿霉素单独用药组;沙利度胺不同浓度组、阿霉素组、联合用药组对HepG2细胞凋亡指数有显著差异(P<0.05),联合用药组高于沙利度胺、阿霉素单独用药组。提示沙利度胺在一定浓度范围内能抑制HepG2细胞体外生长并与阿霉素有协同作用。     

原发性肝癌雌激素受体表达及内分泌治疗

目的研究原发性肝癌雌激素受体（ＥＲ）的表达及内分泌治疗的临床价值．方法应用Ｌｅｅ氏荧光配体细胞化学法检测４１例手术病理确诊的原发性肝癌ＥＲ的表达．本组男３８例，女３例，年龄２５岁～７２岁．同时检测肝癌的组织类型、分化程度、肿瘤大小、血清ＡＦＰ及ＣＥＡ．６例ＥＲ阳性者采用他莫昔芬治疗（每次２０ｍｇ，２次／ｄ，长期服用）并观察疗效．结果肝癌组织ＥＲ阳性２０／４１例，阳性率４８８％．ＥＲ阳性率与患者年龄、性别、ＡＦＰ、ＣＥＡ含量及组织类型（梁状型ＥＲ阳性４５０％，腺样型３３３％，实体型７１４％，硬化型６６７％，透明细胞型２５０％；Ｐ＞００５）无明显关系，与肿瘤体积（≥１０ｃｍ，ＥＲ阳性率７５０％；＜１０ｃｍ，ＥＲ阳性２３８％；Ｐ＜００１）和分化程度（分化好２６例，ＥＲ阳性３４６％；分化差１５例，ＥＲ阳性７３３％；Ｐ＜００５）有显著关系．本组ＥＲ阳性病例中６例经他莫昔芬内分泌治疗有效５／６（８３３％），其中４例ＡＦＰ下降．结论ＥＲ阳性肝细胞癌患者进行内分泌治疗有一定疗效     

生长抑素对人肝癌HepG2细胞增殖及cyclinD1、cyclinE蛋白表达的影响

目的探讨生长抑素治疗肝癌的作用机制。方法取对数生长期HepG2细胞以不含血清的培养液使其周期同步化,以含血清培养液继续培养24h后随机分为观察Ⅰ、Ⅱ、Ⅲ组及对照组,前三组分别加入质量浓度为100、200、400μg／（kg·d）的生长抑素,对照组加入等体积RPMll640培养液24—72h。MTT比色法观察各组细胞增殖抑制率,流式细胞仪检测细胞周期分布,免疫组化法检测周期素D1（cyclinD1）、周期素E（cyclinE）蛋白表达。结果观察Ⅰ、Ⅱ、Ⅲ组不同时间点细胞增殖抑制率均显著高于对照组,且同一时间点观察Ⅲ组〉观察Ⅱ组〉观察I组,同组中72h〉48h〉24h（P〈0．05、0．01）;观察Ⅰ、Ⅱ、Ⅲ组G,期细胞比例均显著高于对照组,观察Ⅱ、Ⅲ组S期细胞比例均显著低于对照组（P〈0．05、0．01）;观察Ⅰ、Ⅱ、Ⅲ组cyclinD1、cyclinE蛋白水平均显著低于对照组,且观察Ⅲ组〈观察Ⅱ组〈观察Ⅰ组（P〈0．05、0．01）。结论生长抑素可通过下调cyclinD1、cyclinE表达抑制HepG2细胞增殖,此可能为其治疗肝癌的作用机制之一。     

原发性肝癌雌二醇及其受体的表达及意义

目的通过检测肝癌组织雌二醇（E2）及其受体（ER）表达，探讨雌二醇及其受体与肝癌的关系和意义。方法应用组织芯片和免疫组化S-P法检测48例肝癌、30例乙型肝炎肝硬化组织E2和ER表达．以正常肝组织为对照组，分析E2和ER与肝癌的关系和意义。结果肝癌组、肝硬化组及对照组E2阳性率分别为62．5％、56．7％、10．0％，肝癌组和肝硬化组阳性率均较对照组明显增加（P〈0．01）．肝癌组与肝硬化组两者无显著性差异；肝癌组ER阳性率（43．8％）与肝硬化组ER阳性率（63．3％）相比无显著性差异，但均显著高于对照组（15．0％，P〈0．05）。结论肝癌及肝硬化存在E2、ER高表达。在肝硬化期就已存在E2、ER含量增多，在肝硬化向肝癌的转变过程中，E2、ER发挥重要的促进作用。E2、ER与肝癌的发生发展有密切关系。     

RNA干扰对肝癌细胞增殖及其Bcl-2表达的影响

目的观察靶向Bcl-2基因的shRNA对肝癌HepG-2细胞增殖及其Bcl-2表达的影响。方法构建靶向Bcl-2基因的shRNA,转染对数生长期HepG-2细胞,设空白组、空载体组、pGenesil—B1组、pGenesil-B2组、pGenesil-B3组、pGenesil-S组;MTT法检测HepG-2细胞增殖抑制率,流式细胞仪检测HepG-2细胞凋亡率,RT-PCR法检测Bcl-2 mRNA,免疫细胞化学SP法检测Bcl-2蛋白。结果空白组、空载体组、pGenesil—B1组、pGenesil-B2组、pGenesil-B3组、pGenesil—S组细胞抑制率分别为0、4．6％、3．8％、46．4％、8．3％、0．13％,细胞凋亡率分别为33．95％、48．80％、69．14％、90．50％、58．14％、50．96％,Bcl-2 mRNA的表达抑制率分别为0、3％、8．3％、49．1％、4．8％、1．8％,Bel-2蛋白阳性率分别为38．15％±3．46％、35．25％±2．67％、15．21％±2．32％、9．51％±2．51％、14．15％±2．67％、36．83％±3．35％,pGenesil-B2组与其他各组相比,P均〈0．01。结论shRNA可致HepG-2细胞凋亡,并抑制Bcl-2的表达。     

羟基喜树碱对人肝癌细胞增殖影响及凋亡诱导

目的 探讨羟基喜树碱对人肝癌细胞株SMMC-7721增殖的影响及其诱导凋亡的作用。方法 以不同浓度的羟基喜树碱在体外作用于人肝癌细胞株SMMC-7721,采用MTT法检测细胞增殖;化学荧光法检测Bcl-2的细胞表达;电镜观察、流式细胞仪与原位末端标记方法检测细胞凋亡情况。结果 与空白对照组比较,羟基喜树碱浓度在3.125μg/mL～100μg/mL组,对肿瘤细胞的增殖抑制率显著增高(P<0.01);在0.05mg/mL组出现细胞早期凋亡现象,Bcl-2表达明显减少;在0.1mg/mL组形成凋亡小体;细胞凋亡比例随羟基喜树碱浓度增加而增高。结论 羟基喜树碱在体外可抑制人肝癌细胞株SMMC-7721增殖,其作用机制可能系通过抑制Bcl-2表达诱导细胞凋亡。     

综合疗法治疗原发性肝癌24例

综合疗法治疗原发性肝癌２４例王在国王影张爱玲丁福全刘光中四川省肿瘤医院成都市６１００４１主题词受体，雌激素／分析他莫昔芬／治疗应用灌注，局部肝肿瘤／药物疗法Ｓｕｂｊｅｃｔｈｅａｄｉｎｇｓｒｅｃｅｐｔｏｒｓ，ｅｓｔｒｏｇｅｎ／ａｎａｌｙｓｉｓｔａｍｏｘ...     

SHP-2酪氨酸磷酸酶对肝癌细胞增殖的影响

目的 建立SHP-2野生过表达型(WT)及激活突变型(MT)稳定转染的肝癌HepG2细胞系,研究SHP-2激活突变及野生过表达对肝癌细胞增殖能力的影响.方法 将已经构建成功的WT SHP-2(WT组)和MT SHP-2(MT组)及空载体pcDNA3.1(空载组)分别转染到人肝癌HepG2细胞中,同时设未转染细胞作对照组;Westernblot法检测细胞中SHP-2蛋白的表达水平;MTT法检测细胞增殖情况;平板克隆和软琼脂集落形成实验检测细胞集落、克隆形成能力.结果 成功的将SHP-2突变型和野生型真核表达载体转染到人肝癌HepG2细胞中;MTT结果显示,对照组和空载组细胞增殖速度接近且增殖慢,而MT组和WT组细胞增殖速度明显加快,WT、MT组与空载组、对照组比较,P均＜0.01;细胞克隆形成的数量多于空载组和对照组,P均＜0.01.结论 成功构建了WT及MT稳定表达SHP-2的HepG2细胞株;SHP-2激活突变及野生过表达促进肝癌细胞增殖能力.     

雌二醇影响MCA-38细胞增殖及雌激素受体表达的实验研究

目的研究雌二醇（E2）影响小鼠结肠腺癌细胞系MCA-38细胞增殖与凋亡及雌激素受体（ER）表达变化的特点与机制。方法将MCA-38细胞分为5个实验组（分别含E2浓度为0.01 nmol/L、0.1 nmol/L、1 nmol/L、10 nmol/L、100 nmol/L的培养液培养）和空白对照组,体外培养24 h。检测各组肿瘤细胞E2处理前后ERα和ERβ表达（Western blot法）、细胞增殖（MTT法）以及caspase-3表达（Western blot法）变化。结果 MCA-38细胞表达ERα、ERβ,后者是其主要ER。不同剂量的E2均影响ERα和ERβ表达,E2上调ERα蛋白表达,0.01 nmol/L、0.1 nmol/L、1 nmol/L浓度范围的E2作用最明显,与对照组相比分别使ERα蛋白表达量增加了4.7%、5.5%和5.9%（P〈0.05）。E2则使ERβ表达下降,0.1 nmol/L、1 nmol/L组最明显,分别使ERβ蛋白表达量下降了10.2%和3.9%（P〈0.05）;与对照组相比,0.01 nmol/L、0.1 nmol/L、1 nmol/L、10 nmol/L浓度E2使MCA-38细胞数目分别增加了20.47%、25.29%、37.59%和30.95%（P〈0.05）;经E2处理,MCA-38细胞的caspase-3蛋白表达量明显下降,其中以0.1 nmol/L和1 nmol/L组最为明显,分别降低20.2%和32.9%（P〈0.05）。结论 E2对MCA-38细胞的ERα和ERβ表达、细胞增殖及凋亡均产生影响,MCA-38细胞增殖和凋亡分别与ERα和ERβ表达变化密切相关。     

Use of tamoxifen in hepatocellular carcinoma: a review and paradigm shift

Hepatocellular carcinoma is often diagnosed at a late, inoperable stage for which there are no uniformly efficacious treatment available presently. The oral anti-oestrogen drug, tamoxifen, has been used in such patients, based on the belief that the growth of hepatocellular carcinoma is promoted by endogenous oestrogen via a receptor-mediated process. In this review, we examine the trials reported in the literature using tamoxifen in hepatocellular carcinoma. Randomized controlled trials with tamoxifen have so far revealed mixed results. We propose that this may be due to the fact that the mechanism of action of tamoxifen in hepatocellular carcinoma is via an oestrogen-receptor independent pathway that requires much higher doses of tamoxifen for activation than those used in the trials so far. Thus there must be a paradigm shift to dissociate the action of tamoxifen from oestrogen receptors in hepatocellular carcinoma. This means that future trials with tamoxifen in hepatocellular carcinoma should use higher doses of tamoxifen, at least four to eight-fold that of the dose that is efficacious in an oestrogen-receptor dependent mechanism.     

肝细胞肝癌微血管密度与雌激素受体表达的临床病理意义

研究肝细胞肝癌(Hepatocellular Carcinoma,HCC)微血管密度(Microvessel Density,MVD)与雌激素受体(Estrogen Receptor,ER)表达之间的关系,以及此二者与临床病理学特征之间的关系.36例HCC的患者分为ER(+)组和ER(-)组.按MVD值分为MVD＜中位值组和MVD＞中位值组,比较各组有预后意义的临床病理学参数之间的差别.ER(+)14例,ER(-)22例.全组MVD自3至121(44.67±32.15中位值为36).ER(+)的HCC为直径较小、多有完整的包膜和较低的血清AFP浓度.MVD较低的HCC多为单结节、直径较小、细胞分化较好、血清AFP浓度较低.ER含量与MVD呈负相关.由此可见(1)ER(+)或MVD较低的HCC分别比ER(-)或MVD较高的HCC恶性度较低.(2)ER与MVD均是有价值的预后指标.     

没药甾酮对人肝癌细胞HepG2增殖和凋亡的影响

目的研究没药甾酮对人肝癌细胞HepG2增殖和凋亡的影响。方法以正常人肝细胞L-02作为对照,采用3-（4,5-二甲基噻唑-2）-2,5-二苯基四氮唑溴盐法观察不同浓度没药甾酮（5～100μmol/L）对人肝癌细胞HepG2和L-02细胞增殖的影响并观察细胞形态的变化;应用流式细胞术检测细胞周期变化和凋亡发生。结果不同浓度没药甾酮均可显著抑制人肝癌细胞HepG2生长,并呈时间、剂量依赖性,最大抑制率可达81.9%±1.92%（100μmol/L）;没药甾酮可使G0/G1期细胞比例增多,G2/M期细胞比例下降,可将细胞阻滞于G0/G1期;没药甾酮诱导人肝癌细胞HepG2发生凋亡,50μmol/L和75μmol/L没药甾酮早期细胞凋亡率分别为24.91%±2.41%、53.03%±2.28%,与对照组相比,差异均具有统计学意义（P〈0.05）。结论没药甾酮可抑制人肝癌细胞HepG2增殖并诱导凋亡,其作用可能与干扰细胞周期有关。     

奥曲肽对人肝癌细胞生长增殖、甲胎蛋白合成及细胞凋亡的作用

目的　研究奥曲肽对肝癌细胞生长增殖及细胞凋亡的作用 ,探讨其对肝癌的作用机制 ,为临床应用提供实验依据。方法　采用MTT法、生长曲线观察奥曲肽对肝癌细胞HepG2生长增殖的影响 ,电化学发光法测定培养上清液中甲胎蛋白 (AFP)含量 ,并用荧光染色、透射电镜和流式细胞仪检测细胞凋亡。结果　奥曲肽在 0 .0 0 5～ 80 μg/ml浓度范围内呈剂量依赖性方式抑制肝癌细胞HepG2的生长增殖 ,并能显著减少肝癌细胞AFP合成。奥曲肽 (0 .2 5～ 4 .0 μg/ml)作用 4 8h后 ,荧光染色与透射电镜可见部分HepG2细胞呈典型的凋亡形态学改变。流式细胞仪检测 ,出现凋亡峰 ,与对照组相比 ,细胞的凋亡率显著升高 (P <0 .0 5 )。结论　奥曲肽能够显著抑制肝癌细胞生长 ,并诱导肝癌细胞凋亡 ,有望成为治疗肝细胞癌的一个有效药物     

Meta-analysis of vascular and neoplastic events associated with tamoxifen

OBJECTIVE: Tamoxifen reduces the risk of developing breast cancer but also affects the risks of certain vascular and neoplastic events. Our purpose was to estimate the effects of tamoxifen on potentially life-threatening vascular and neoplastic outcomes. DESIGN: Random effects meta-analysis of published randomized controlled trials. PATIENTS: Participants in all trials in which a treatment arm that included tamoxifen was compared to a similar control arm. Breast cancer risk reduction and treatment trials were included. INTERVENTIONS: Tamoxifen at variable dose and duration. MEASUREMENTS AND MAIN RESULTS: Thirty-two trials (52,929 patients) reported one or more outcomes of interest. Tamoxifen was associated with significantly increased risks of endometrial cancer (relative risk [RR] 2.70; 95% CI, 1.94 to 3.75), gastrointestinal cancers (RR 1.31; 95% CI, 1.01 to 1.69), strokes (RR 1.49; 95% CI, 1.16 to 1.90), and pulmonary emboli (RR 1.88; 95% CI, 1.77 to 3.01). Tamoxifen had no effect on secondary malignancies other than endometrial and gastrointestinal cancers (RR 0.96; 95% CI, 0.81 to 1.13). In contrast, tamoxifen significantly decreased myocardial infarction deaths (RR 0.62; 95% CI, 0.41 to 0.93) and was associated with a statistically insignificant decrease in myocardial infarction incidence (RR 0.90; 95% CI, 0.66 to 1.23). Postmenopausal women had greater risk increases for neoplastic outcomes. CONCLUSIONS: This meta-analysis of randomized trials found tamoxifen use to be significantly associated with several neoplastic and vascular outcomes. Consideration of tamoxifen use requires balance of potential benefits and risks.     

Apoptosis and proliferation of human hepatocellular carcinoma

Abstract: To investigate the contribution of apoptosis, a major mechanism of cell death, in the growth of hepatocellular carcinoma, we analyzed both apoptosis and cell proliferation in human hepatocellular carcinoma. We used the terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling method and proliferative cell nuclear antigen staining, respectively. Among 21 hepatocellular carcinoma specimens examined, four were well, ten were moderately, and seven were poorly differentiated hepatocellular carcinoma. Terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling-positive cells in hepatocellular carcinoma were scattered individually or were sometimes clustered in the tumors. The terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling indices were 0.35±0.09, 0.81±0.29, and 1.9±0.94 in well, moderately, and poorly differentiated hepatocellular carcinoma, respectively. The proliferative cell nuclear antigen labeling indices were 6.6±0.9, 13.1±3.5, and 26.7±6.3 in hepatocellular carcinoma in the same respective order of differentiation. The differences in both terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labelling indices and proliferative cell nuclear antigen labeling indices (p>0.05) were significant between well, moderately and poorly differentiated hepatocellular carcinoma. There was a positive correlation between the terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling and proliferative cell nuclear antigen labeling indices in hepatocellular carcinoma (r=0.84, p>0.001). This study showed that the proliferation rate and the incidence of apoptosis increased as the differentiation grade of hepatocellular carcinoma was lowered, suggesting a rapid turnover of cancer cells in the lower differentiation grades. Apoptosis may thus play an important role in the growth of hepatocellular carcinoma.     

Distribution of asialoglycoprotein receptor in human hepatocellular carcinoma

ABSTRACT— Altered expression of asialoglycoprotein (ASGP) receptors on hepatocytes has been reported during hepatic neoplasia mostly in animal models. In this study, we examined immunohistochemically the distribution of the ASGP receptor in humans with various liver diseases, including ten cases of hepatocellular carcinoma (HCC). In livers of acute hepatitis, chronic hepatitis, cirrhosis and the non-cancerous tissues (mostly cirrhosis) adjacent to HCC, the receptor was present in its normal distribution, i.e. mostly along the sinusoidal margin and partly on the lateral surface of hepatocytes. In four of six well-differentiated HCCs, the receptor was also normally distributed on the plasma membrane; by immunoelectron microscopy, it was seen in the endoplasmic reticulum and in pits in the plasma membrane but not on bile canaliculus-like structures, suggesting that it was synthesized, transported, and integrated into the plasma membrane in a polar manner. In contrast, there was no surface expression of the ASGP receptor in the remaining six HCCs (two well-differentiated and four poorly differentiated). In two of the poorly differentiated HCCs, the receptor, although absent from the cell surface, was prominent in the endoplasmic reticulum, suggesting disturbed transport of the ASGP receptor to the cell surface. When we examined proliferative activity of HCCs by immunohistochemical labeling of DNA polymerase a, HCCs with high percentages (above 30%) of DNA polymerase α-positive cells had lost the cell-surface expression of the receptor. Thus, the expression of the ASGP receptor in human HCC appears to be closely related to differentiation and proliferative activity of the tumor cells.