去甲二氢愈创木酸对恶性胶质瘤细胞周期的影响

 目的　观察去甲二氢愈创木酸(NDGA)对人恶性胶质瘤细胞系SHG-44细胞周期的影响。方法　采用细胞培养、免疫组化、细胞计数及流式细胞仪检测等技术方法。结果　①在200μM浓度范围内,NDGA对SHG-44细胞增殖有抑制作用,使细胞增殖受阻于G1→S期,且这种作用与浓度呈正相关;②在NDGA处理96小时内,NDGA对SHG-44细胞的增殖抑制作用以处理后24小时最明显;③NDGA处理后,SHG-44细胞cyclin D1和CDK4蛋白表达减弱,p16蛋白表达增强。结论　NDGA对SHG-44细胞周期有明显的增殖抑制作用,且这种作用可能与CDK4蛋白激酶活性降低有关。     

去甲二氢愈创木酸诱导体外恶性胶质瘤细胞凋亡的研究

目的：观察去甲二氢愈创木酸（nordihydroguaiaretic acid,NDGA)对人恶性胶质瘤细胞系SHG-44细胞凋亡的影响,并探讨其发生的可能机理。方法：采用MTT（methy thiazolyl tetrazolium)法检测细胞增殖抑制的情况。利用光镜和电镜观察及TUNEL法检测体外培养细胞的凋亡情况,用免疫组织化学,原位杂交和图像分析等方法检测bcl-2基因mRNA和蛋白的表达水平。结果：（1）一定浓度（100μmol/L）的NDGA处理SHG-44细胞不同时间后,在NDGA抑制细胞增殖的同时,也可诱导此细胞发生凋亡,且随着作用时间的延长,凋亡细胞数增加越明显;（2）免疫组化染色结果表明,一定浓度的NDGA处理细胞不同时间后,出现SHG-44细胞bcl-2蛋白表达水平降低,且随着作用时间的延长,这种趋势更加明显,与细胞凋亡的发生呈负相关;（3）原位杂交结果显示;NDGA处理细胞不同时间后,出现SHG-44细胞bcl-2基因mRNA的表达水平降低,结果与免疫组化检测一致。结论：NDGA可诱导SHG-44细胞的凋亡,这种作用可能与其下调bcl-2基因的表达有关,确切机制有待进一步深入研究。     

5-氟尿嘧啶、长春新碱与去甲二氢愈创木酸配伍对人恶性胶质瘤细胞株的作用

 目的 观察体外应用去甲二氢愈创木酸（ＮＤＧＡ）、予氟尿嘧啶（５－Ｆｕ）、长春新碱（ＶＣＲ）三种药物单用或合用对人恶性胶质瘤细胞系 ＳＨＧ－４４细胞的作用，并探讨其作用的可能机制。方法用 ＭＴＴ法检测药物作用效应，用免疫组化染色法检测细胞 ｃｙｃｌｉｎ Ｄ１基因的蛋白表达情况。结果 （１）三种药物单用及两药合用时随着药物浓度的增加其抗肿瘤效应也增加，且两药合用时药物的效应增强；（２）先给ＮＤＧＡ ２４ｈ后再给 ５－Ｆｕ或 ＶＣＲ，与同时给药对该细胞的抗肿瘤效应无显著差异（Ｐ＞０．０５），而先给 ５－Ｆｕ或 ＶＣＲ ２４ｈ后再给 ＮＤＧＡ，与同时给药对细胞的抗肿瘤效应有显著差异（Ｐ＜０．０５）；（３）免疫组化染色结果表明，与对照组相比，ＮＤＧＡ处理后该细胞 ｃｙｃｌｉｎ Ｄ１基因的蛋白表达明显降低。结论 ＮＤＧＡ与 ５Ｆｕ或 ＶＣＲ间有协同作用，且这种作用可能与 ＮＤＧＡ降低细胞 ｃｙｃｌｉｎ Ｄ１基因的蛋白表达有关。     

去甲二氢愈创木酸抗肿瘤作用的研究

去甲二氢愈创木酸(NDGA)是一种从植物中提取的天然药物成分,是脂氧化酶的强抑制剂,一度作为抗氧化剂,以防止食物腐败。近来发现去甲二氢愈创木酸有抗肿瘤的作用,能抑制肿瘤细胞的增殖并诱导凋亡,本文就去甲二氢愈创木酸抗肿瘤的作用及机制做一综述。     

去甲二氢愈创木酸抑制宫颈癌SiHa细胞的生长与上调p21基因组蛋白乙酰化相关性的研究

目的:研究去甲二氢愈创木酸(nordihydroguaiaretic acid, NDGA)对宫颈癌SiHa细胞增殖的影响及作用机制.方法:MTT法检测NDGA对SiHa细胞生长的影响,FCM法检测NDGA对SiHa细胞周期及凋亡的影响,RT-PCR检测NDGA对p21基因转录的影响,Western印迹法检测NDGA对组蛋白H3总乙酰化水平的影响,染色质免疫沉淀(chromatin immunoprecipitation,ChIP)-PCR法检测NDGA对p21基因近启动子区域组蛋白H3乙酰化的影响.结果:NDGA能明显抑制SiHa细胞的生长,且呈时间和剂量依赖性;可使SiHa细胞的细胞周期阻滞于G1期,而凋亡率却下降;能明显提高p21基因的转录水平和组蛋白H3的总乙酰化水平,ChIP-PCR检测结果表明NDGA可明显促进p21基因近启动子区域组蛋白H3的乙酰化水平.结论:NDGA促进p21基因近启动子区域及细胞内总体组蛋白H3的乙酰化可能是其抑制宫颈癌SiHa细胞生长的潜在机制.     

PTEN在人脑胶质瘤细胞中的表达及其对细胞增殖的影响

背景与目的:PTEN基因的缺失及突变与多种肿瘤有关,而脑胶质瘤是与PTEN基因变异关系最为密切的恶性肿瘤之一.本研究旨在观察PTEN基因对人脑胶质瘤细胞生物学特性的影响,为PTEN用于胶质瘤基因治疗提供科学的实验依据.方法:(1)应用RT-PCR方法扩增人脑胶质瘤细胞U251、SHG-44中PTEN基因,序列测定分析.(2)采用阳离子聚合物转染试剂,将携带野生型PTEN基因的重组真核表达载体质粒转染至胶质瘤细胞,G418筛选出稳定转染的细胞并扩增培养.通过细胞形态学、细胞生长曲线观察PTEN基因表达对细胞形态和增殖的影响,应用Western blot、免疫细胞化学法检测相关蛋白的表达.结果:(1)胶质瘤细胞U251和SHG-44的PTEN mRNA分别存在着突变型、缺失型两种表达方式.(2)稳定转染的U251和SHG-44细胞株的生长曲线显示细胞增殖明显受抑制,第7天细胞计数分别为未转染对照组细胞数的39.1%、27.8%;Westem blot显示稳定转染细胞株有外源性PTEN蛋白的表达;细胞免疫化学法显示胶质纤维酸性蛋白(glial fibrillary acidic protein,GFAP)表达量增加,从而表现出对细胞形态影响的差异,稳定转染的U251细胞出现向星形细胞分化的特征,SHG-44细胞形态转染前后无明显改变.结论:恢复野生型PTEN基因表达对胶质瘤细胞存在差异性的诱导分化作用.     

四氧化三铁纳米粒子增强二氢卟吩e6对胶质瘤U251细胞的声动力治疗效果

目的:探讨四氧化三铁(Fe3O4)纳米粒子(PION)作为药物载体增强二氢卟吩e6(chlorin e6,Ce6)在胶质瘤中的增效作用.方法:采用高温降解法和相转移法制备PEG-Fe3O4@Ce6复合纳米粒子(PION@E6),用水合粒径分析、透射电镜、胶体稳定性分析、紫外可见光吸收光谱等方法对PION@E6进行鉴定....     

二甲亚砜和维甲酸对人卵巢癌细胞COC1的生长抑制和超微结构的影响

本实验以人卵巢癌细胞株COC1为模型，观察了诱导分化剂二甲亚砜（DMS0）与维甲酸（RA）对该细胞生长增殖及超微结构的影响。结果表明，DMSO和RA可明显抑制COC1细胞的生长，并改变其恶性细胞的结构特征，使之向正常细胞的方向逆转，从而提示DMSO和RA对人卵巢癌细胞有一定的诱导分化作用。     

二氢卟吩e6光动力对人结肠癌细胞SW620生物学行为的影响

目的探讨二氢卟吩e6(Ce6)光动力学疗法(PDT)对人结肠癌SW620细胞增殖、迁移及克隆形成能力等生物学行为的影响。方法Ce6光动力处理SW620细胞,采用MTT法检测细胞增殖,划痕实验检测细胞迁移能力,平板克隆形成实验检测细胞克隆形成能力。结果Ce6浓度和光照剂量增加时,Ce6-PDT对SW620细胞生长抑制作用逐渐增加,呈现浓度依赖、光照剂量依赖关系。划痕实验结果显示Ce6-PDT能够抑制细胞的迁移。Ce6-PDT能够减弱SW620细胞克隆形成能力。结论Ce6-PDT能够抑制SW620细胞增殖、迁移及克隆形成能力。     

诺帝对HPV16亚基因永生化人宫颈上皮细胞增殖、凋亡和分化的影响

目的：研究诺帝（Nordy）对HPV 16型亚基因（E6、E7）永生化人宫颈上皮细胞（H8细胞）增殖、分化和凋亡的影响。方法：MTT法检测诺帝对H8细胞增殖的抑制作用，SP法检测增殖细胞核抗原mcm 5的表达，FCM分析细胞周期和凋亡，光学显微镜、电子显微镜观察细胞形态变化，端粒重复序列扩增-酶联免疫吸附法（telomerase repeat amplification protocol-enzyme linked immunosorbent assay，TRAP—ELISA）法检测端粒酶活性变化。结果：10～100μmol/L浓度的诺帝能够不同程度地抑制H8细胞的增殖，细胞核内mcm 5的表达明显下降，可阻滞细胞周期于G0／G1期、S期细胞减少，细胞凋亡率增多，细胞形态上出现向成熟分化的趋势，端粒酶活性明显降低。结论：诺帝具有抑制H8细胞增殖，促进凋亡，诱导其分化的作用，这种作用可能是通过阻滞细胞周期，降低端粒酶活性来实现的。     

Identification of Genetic Alterations in Thai Breast Cancer Patients by Arbitrarily Primed Polymerase Chain Reaction

Breast cancer is the most common cancer among women worldwide. Genetic alterations prevalent in breastcancer are still being elucidated. In this report, changes in 30 breast cancer tissues, in compariosn with normaltissues from Thai patients, were analyzed by arbitrarily primed polymerase chain reaction (AP-PCR). Geneticinstability was detected by DNA fingerprinting obtained with 13 of 60 random primers. Of these, at least oneamplification band, the incidence ranging from 27 to 80%, was observed in DNA amplified with 8 primers,whereas a band loss was exhibited with from 6 primers, the incidences ranging from 23 to 40%. Likewise, anamplification band amplified from primer D15 was observed in 80% of this patient group and a band lossproduced from primer B12 presented in 40% of all cases. These results showed that AP-PCR is effective for thedetection of genetic alterations in breast cancer tissues.     

Apoptosis Induced by Prednisolone Occurs without Altering the Promoter Methylation of BAX and BCL-2 Genes in Acute Lymphoblastic Leukemia Cells CCRF-CEM

Objective: one of the main mechanisms in which cancer cells are resistant to chemotherapy drugs and therapeutic strategies is resistance to apoptosis due to these anticancer factors. Regulating the expression of genes through epigenetics, especially regulation through methylation, is one of the key aspects of regulating gene expression and the function of genes, which is also regulated by the pathways regulating the pathway of apoptosis. The epigenetic regulatory phenomenon in cancer cells can undergo a change in regulation and induces resistance to apoptosis against chemotherapy and anticancer factors. The purpose of the present scrutiny was defined to probe the effect of subtoxic prednisolone dose on the level of promoter methylation and gene expression of BAX and BCL2 in the CCRF-CEM cells. Methods: The treated cells by prednisolone, cultured in RPMI 1640 medium in standard condition. Alteration in promoter DNA methylation was analyzed by use of methylation specific-PCR (MSP) technique after the defined intervened time of Prednisolone treatment with a subtoxic dose. Results: Prednisolone can induce apoptosis via alteration in BAX and BCL2 genes, based on our previous scrutiny. This essay shows no varies in the Pattern of DNA methylation of examined genes; however, prednisolone changes the expression of examined genes. Conclusion: Lack of alteration through prednisolone treatment in DNA methylation template of BAX and BCL2 genes make this possible that Prednisolone affects apoptotic gene expression via different pathways, which need more research to be done about it.     

An <Emphasis Type="Italic">in vitro</Emphasis> Study on the Suppressive Effect of Glioma Cell Growth Induced by Plasmid-Based Small Interference RNA (siRNA) Targeting Human Epidermal Growth Factor Receptor

Summary Objectives:To study the inhibitory effects of plasmid-based siRNA targeting human epidermal growth factor receptor (EGFR) on tumor proliferation and invasion of TJ905 glioblastoma cells. Methods: Two siRNA expression constructs targeting human EGFR extracellular domain (516–536) and catalytic domain (2400–2420) were transfected into TJ905 cell as mediated by Lipofectamine. Immunofluorescence assay and Western blotting were used to detect EGFR expression. Cell cycle was analyzed by flow cytometry, cell proliferative activity was measured by MTT assay. The expression and enzymatic activity of MMP9 were measured by Western blotting and gelatin zymography. Cell invasive capability was evaluated by Transwell method. Results: The expression of EGFR was knocked-down by 90 and 92, respectively in siRNA constructs transfected groups as indicated by immunofluorescence assay and Western blotting. The flow cytometric analysis showed that the S phase fraction (SPF) was lowered in both siRNAs transfected cells than that in parental cells and the cells transfected with empty vector. Compared to parental cells and the cells transfected with empty vector, the survival rates of glioma cells transfected with the siRNAs dramatically dropped down from the first day after implantation (P<0.05) as indicated by MTT assay. Meanwhile, the expression and enzymatic activity of MMP9 decreased significantly in siRNAs transfected in TJ905 cells, and cell invasive potential was also greatly inhibited in the Transwell study. Conclusion: The siRNA expression constructs targeting EGFR could specifically suppress EGFR expression, inhibit cell growth, induce cell cycle arrest and suppress invasion. The plasmid-based siRNA targeting human EGFR approach should be a new strategy for gene therapy of malignant gliomas.     

中药加化疗治疗局部晚期胰腺癌的疗效分析

观察中医药加由健择和顺铂组成的化疗方案时局部晚期胰腺癌的疗效。遵照患者意愿,有选择地将37例局部晚期胰腺癌患者分为治疗组(中西医结合组)和对照组(单纯化疗组)。治疗组化疗期间和化疗后继续服中药巩固疗效。结果两组近期疗效差异无统计学意义,临床获益率和远期疗效治疗组明显优于对照组,P<0．01。初步研究结果提示,中医药配合化疗可提高局部晚期胰腺癌的临床获益率,延长生存期。