Suppression of the neoplastic phenotype by replacement of the RB gene in human cancer cells

Mutational inactivation of the retinoblastoma susceptibility (RB) gene has been proposed as a crucial step in the formation of retinoblastoma and other types of human cancer. This hypothesis was tested by introducing, via retroviral-mediated gene transfer, a cloned RB gene into retinoblastoma or osteosarcoma cells that had inactivated endogenous RB genes. Expression of the exogenous RB gene affected cell morphology, growth rate, soft agar colony formation, and tumorigenicity in nude mice. This demonstration of suppression of the neoplastic phenotype by a single gene provides direct evidence for an essential role of the RB gene in tumorigenesis.     

The human papilloma virus-16 E7 oncoprotein is able to bind to the retinoblastoma gene product

Deletions or mutations of the retinoblastoma gene, RB1, are common features of many tumors and tumor cell lines. Recently, the RB1 gene product, p105-RB, has been shown to form stable protein/protein complexes with the oncoproteins of two DNA tumor viruses, the adenovirus E1A proteins and the simian virus 40 (SV40) large T antigen. Neither of these viruses is thought to be associated with human cancer, but they can cause tumors in rodents. Binding between the RB anti-oncoprotein and the adenovirus or SV40 oncoprotein can be recapitulated in vitro with coimmunoprecipitation mixing assays. These assays have been used to demonstrate that the E7 oncoprotein of the human papilloma virus type-16 can form similar complexes with p105-RB. Human papilloma virus-16 is found associated with approximately 50 percent of cervical carcinomas. These results suggest that these three DNA viruses may utilize similar mechanisms in transformation and implicate RB binding as a possible step in human papilloma virus-associated carcinogenesis.     

茶多酚与银杏叶提取物联用对人乳腺癌细胞增殖的影响

[目的]研究茶多酚与银杏叶提取物联合作用对人乳腺癌细胞MDA-MB-231增殖的影响。[方法]采用MTT法测定茶多酚、银杏叶提取物单独及联合应用对人乳腺癌细胞系MDA-MB-231的抑制作用;利用光镜观察茶多酚、银杏叶提取物对人乳腺癌系MDA-MB-231形态的影响。[结果]MTT法显示银杏叶提取物与低浓度（10、20μg/ml）的茶多酚联合使用,对乳腺癌细胞抑制率明显高于单独用药组,但是联合用药组只有在一定的浓度范围内才会产生协同作用抑制乳腺癌细胞增殖;在细胞形态方面,与单独用药组、对照组相比,联合用药组乳腺癌细胞生长状况差,折光性弱,贴壁细胞皱缩,变圆。[结论]银杏叶提取物在低浓度下对人乳腺癌细胞系MDA-MB-231生长无明显抑制作用,茶多酚和茶多酚联合银杏叶提取物均可抑制MDA-MB-231细胞生长,联合用药组强于单独用药,在一定浓度范围内起协同作用。     

Transfection of v-rasH DNA into MCF-7 human breast cancer cells bypasses dependence on estrogen for tumorigenicity

The natural history of estrogen-responsive breast cancers often involves a phenotypic change to an estrogen-unresponsive, more aggressive tumor. The human breast cancer cell line, MCF-7, which requires estradiol for tumor formation in vivo and shows growth stimulation in response to estradiol in vitro, is a model for hormone-responsive tumors. The v-rasH onc gene was transfected into MCF-7 cells. The cloned MCF-7ras transfectants, which expressed the v-rasH messenger RNA and v-rasH p21 protein (21,000 daltons), were characterized. In contrast to the parental cell line, MCF-7ras cells no longer responded to exogenous estrogen in culture and their growth was minimally inhibited by exogenous antiestrogens. When tested in the nude mouse, the MCF-7ras cells were fully tumorigenic in the absence of estrogen supplementation. Thus, cells acquiring an activated onc gene can bypass the hormonal regulatory signals that trigger the neoplastic growth of a human breast cancer cell line.     

Human breast cancer: correlation of relapse and survival with amplification of the HER-2/neu oncogene

The HER-2/neu oncogene is a member of the erbB-like oncogene family, and is related to, but distinct from, the epidermal growth factor receptor. This gene has been shown to be amplified in human breast cancer cell lines. In the current study, alterations of the gene in 189 primary human breast cancers were investigated. HER-2/neu was found to be amplified from 2- to greater than 20-fold in 30% of the tumors. Correlation of gene amplification with several disease parameters was evaluated. Amplification of the HER-2/neu gene was a significant predictor of both overall survival and time to relapse in patients with breast cancer. It retained its significance even when adjustments were made for other known prognostic factors. Moreover, HER-2/neu amplification had greater prognostic value than most currently used prognostic factors, including hormonal-receptor status, in lymph node-positive disease. These data indicate that this gene may play a role in the biologic behavior and/or pathogenesis of human breast cancer.     

番木瓜种子中异硫氰酸苄酯（BITC）的抑癌试验

利用MTT法检测番木瓜种子中的异硫氰酸苄酯(BITC)在体外对肝癌细胞等6种常见癌细胞的生长抑制率。结果表明,番木瓜种子提取物中BITC的浓度达到5μmol.L-1(相当于0.745mg.L-1)时,对人肝癌HepG2细胞、人肺癌A549细胞、人乳腺癌MCF-7细胞、人大肠癌HCT-8细胞、人宫颈癌HeLa细胞、人前列腺癌DU-145细胞的生长抑制率均达到70%以上,其中,人肝癌细胞HepG2最敏感,cBITC=2.5μmol.L-1时抑制率可达87%,在cBITC=10μmol.L-1和5.0μmol.L-1时,人大肠癌HCT-8细胞的生长抑制率几乎相同。因此,BITC具有良好的抑癌效果。     

Cell cycle-dependent regulation of phosphorylation of the human retinoblastoma gene product

The human retinoblastoma gene (RB1) encodes a protein (Rb) of 105 kilodaltons that can be phosphorylated. Analysis of Rb metabolism has shown that the protein has a half-life of more than 10 hours and is synthesized at all phases of the cell cycle. Newly synthesized Rb is not extensively phosphorylated (it is "underphosphorylated") in cells in the G0 and G1 phases but is phosphorylated at multiple sites at the G1/S boundary and in S phase. HL-60 cells that were induced to terminally differentiate by various chemicals lost their ability to phosphorylate newly synthesized Rb at multiple sites when cell growth was arrested. These findings suggest that underphosphorylated Rb may restrict cell proliferation.     

Retinoblastoma: clues to human oncogenesis

The retinoblastoma gene can be considered a model for a class of recessive human cancer genes that have a "suppressor" or "regulatory" function. The loss or inactivation of both alleles of this gene appears to be a primary mechanism in the development of retinoblastoma. Such a mechanism is in direct contrast to that of putative human oncogenes which are thought to induce tumorigenesis following activation or alteration. The high incidence of second primary tumors among patients who inherit one inactive retinoblastoma allele also suggests that this cancer gene plays a key role in the etiology of several other primary malignancies. Finally, the observation that extra nonrandom copies of specific chromosomal regions occur in some of these tumors provides circumstantial evidence that an "expressor" gene (possibly an oncogene) may be involved in retinoblastoma development.     

转染抑癌基因PTEN对人乳腺癌ZR-75-1细胞的细胞周期和增殖能力的影响

目的：观察PTEN（phosphatase and tensin homology deleted on chromosome ten，第10号染色体上磷酸酶和张力蛋白同源缺失的基因）对人乳腺癌细胞ZR-75-1细胞增殖和细胞周期的影响。方法；利用脂质体介导法将携带有野生型和突变型PTEN cDNA的真核表达载体pBP—wt—PTEN和pBP—G129R—PTEN导八人乳腺癌ZR-75-1细胞（质粒转染成功后实验分为C—WT—PTEN组、CG129R—PTEN组和未转染质粒组即对照组）后，以RT—PCR、Western blot分析目的基因的表达，并采用MTT法和流式细胞术检测细胞增殖和细胞周期。结果：C—WT—PTEN组、C—G129R—PTEN组细胞PTEN mRNA及PTEN蛋白出现明显的高表达。C—WT—PTEN组细胞生长的抑制率可高达42．7％，与对照组比较．差异有显著性（P〈0．05）。但C—G129R—PTEN组细胞生长的抑制率与对照组比较，差异无显著性（P〉0．05）。流式细胞术显示C—WT—PTEN组细胞周期从G1期到S期已发生抑制。结论：野生型PTEN可依赖其磷酸酶活性抑制肿瘤细胞的增殖,并最终诱导细胞凋亡。     

毛蚶中天然牛磺酸的提取和活性研究

[目的]提取和纯化毛蚶(Scapharca subcrenata)中的天然牛磺酸,并对其活性进行研究。[方法]采用薄层色谱法(TLC)对提取的牛磺酸纯度进行分析,并对其结构用红外光谱(IR)和核磁共振波谱(NMR)进行鉴定,然后对其细胞毒活性进行检测。[结果]毛蚶富含牛磺酸,对人白血病细胞HL-60、肝癌细胞SMMC-7721、肺癌细胞A-549、乳腺癌细胞MCF-7和结肠癌细胞SW480没有明显的细胞毒活性。[结论]该研究为天然牛磺酸的开发和利用提供了依据。     

Growth arrest and morphological change of human breast cancer cells by dibutyryl cyclic AMP and L-arginine

The growth in vitro of human breast cancer cells, line MCF-7, was inhibited by a daily supplement of L-arginine (1 milligram per milliliter). Arginine acted synergistically with dibutyryl adenosine 3',5'-monophosphate (cyclic AMP) (10(-6) molar) to enhance the growth inhibitory effect: the cell replication ceased completely within 2 days after treatment. The growth arrest accompanied a change in cell morphology and was preceded by increases in the cellular concentration of cyclic AMP, adenylate cyclase, and type II cyclic AMP-dependent protein kinase activities as well as a decrease of estrogen binding activity. The results suggest that growth of human breast cancer cells is subject to cyclic AMP-mediated regulation and that arginine may play a specific role in this process.     

视网膜母细胞瘤109例分析

目的：分析视网膜母细胞瘤的发病特点和诊治规律。方法；收集本院近２３ａ来的１０９例ＲＢ病毒，对其临床特点，病理表现，治疗，预后及家系调查作一回顾性分析。结果：７７．１％的病例就诊年龄小于５岁。２２例为双眼发病，９例具有肿瘤家族史。１０９例中以晚期虱阳多见，病理检查大多可见有视神经和巩膜浸润。     

漆姑草石油醚提取物化学成分分析及抗肿瘤活性筛选

[目的]对漆姑草[Sagina japonica（Sw.）Ohwi]石油醚提取物的化学成分进行分析鉴定,并对其抗肿瘤活性进行研究。[方法]采用GC-MS联用技术分析漆姑草石油醚提取物的化学成分;选用3种人癌细胞株（人白血病K562细胞株、人宫颈癌Hela细胞株和人乳腺癌MCF-7细胞株）,采用MTT法和SRB法,对其抗肿瘤活性进行研究。[结果]从漆姑草石油醚提取物中分离鉴定出25种成分,其中脂肪酸含量最多,亚油酸含量达49.318%;漆姑草石油醚提取物对人白血病K562细胞株的生长具有微弱的抑制作用,对人宫颈癌Hela细胞株和人乳腺癌MCF-7细胞株均无抑制作用。[结论]该研究可为漆姑草资源在临床上的广泛应用奠定基础。     

BLV-miRNA跨界调控人类靶基因预测及生物信息学分析

【目的】评估牛白血病病毒（BLV）来源的miRNAs跨界调控人源基因的风险。对BLV-miRNA可能带来的食品安全问题及对人体健康可能造成何种影响进行前瞻性研究,为未来实际生产中地方流行性白血病防控措施执行的必要性研究奠定基础,对BLV与人类疾病间关联性的研究提供理论指导。【方法】首先使用mirbase网站对BLV miRNA的成熟序列进行查询,通过miRanda软件对BLV编码的10种miRNA（BLV-miR-B1-3P,5P、BLV-miR-B2-3P,5P、BLV-miR-B3-3P,5P、BLV-miR-B4-3P,5P、BLV-miR-B5-3P,5P）进行靶基因预测,并选取每个BLV-miRNA评分前10的候选靶基因（去除重复基因后共88个）进行功能分析,对受到多个BLV miRNA共同调控的候选靶基因使用RNAhybrid软件进行二次预测验证,并对其功能进行分析。【结果】 BLV编码的10种miRNA经预测后分别获得1 630—16 383个靶基因不等。对评分前十的共计88个候选靶基因进行功能分析后发现,其中18个基因无相关功能报道;36个候选靶基因与肿瘤性疾病的发生发展存在相关性。2个候选靶基因可以对细胞周期起调控作用;16个候选靶基因参与细胞信号转导的调控;14个候选靶基因在细胞结构/骨架蛋白的形成中发挥作用;细胞的增殖与凋亡的功能表现成拮抗关系,往往促进增殖的基因同时也可以抑制细胞凋亡,共有13个基因对细胞的增殖和凋亡起调节作用,有趣的是,这13的候选靶基因对细胞增殖凋亡功能的调节是双向性的,但不能明确BLV miRNA对细胞的调节到底是更趋向于增殖还是凋亡,因此仍需要后续研究深入探讨;2个候选靶基因对细胞分化起调节作用;16个候选靶基因对细胞的迁移/侵袭功能起调节作用,再次提示BLV miRNA与肿瘤性疾病可能存在更重要的关联性。7个候选靶基因可能在乳腺细胞的分化、迁移、侵袭过程中发挥重要作用,提示BLV与人乳腺癌相关性的研究中,可以从BLV miRNA的角度深入探讨;BLV-B4-3P的2个候选靶基因Ⅰ型胶原α1链基因（COL1A1）、断裂点簇集区（BCR）对人急性淋巴细胞白血病（ALL）具有调节作用。此外,可以被多个BLV miRNA共同靶向的候选靶基因均属于黏蛋白家族（MUC5B、MUC12和 MUC16）,且均可以在结肠中表达,对结肠黏膜的形成产生影响。【结论】外源性BLV miRNA可能跨界调控细胞周期、信号转导、结构/骨架、增殖、凋亡、分化、迁移/侵袭相关等细胞功能相关基因,破坏细胞结构;BLV miRNA与人乳腺癌的相关性可能表现在人乳腺癌细胞的分化、迁移、和侵袭过程中;而BLV-miR-B4-3p本身与白血病相关miR 29a共享同一种子区域,可能对人急性淋巴细胞白血病的发生发展造成影响;外源性BLV miRNA具有靶向抑制黏蛋白基因（MUC5B、MUC12、MUC16）表达,通过破坏肠黏膜形成这一途径,跨界调控人源基因的风险。     

黄芩苷通过上调miR-126诱导乳腺癌细胞凋亡的机制研究

目的 用黄芩苷干预人乳腺癌细胞株MDA-MB-231,观察黄芩苷对乳腺癌细胞增殖、凋亡的影响及作用机制。方法 用qRT-PCR检测miR-126的表达变化,Western-blot检测Bcl-2、Caspase-9、Caspase-3、p-p38和p53的表达,MTT法检测细胞增殖,流式细胞术检测细胞凋亡。结果 miR-126在乳腺癌MDA-MB-231细胞中的表达比正常乳腺细胞低,黄芩苷干预乳腺癌细胞后miR-126上调最为明显（P<0.05）。用miR-126 mimics、miR-126 inhibitors转染乳腺癌细胞,Western-blot显示黄芩苷及miR-126 mimics作用于人乳腺癌细胞后Bcl-2表达水平下降,Caspase-9和Caspase-3的裂解产物、p-p38、p53表达增加,差异有统计学意义（P<0.05）。MTT法显示黄芩苷和miR-126 均可抑制乳腺癌细胞的增殖,流式细胞术显示黄芩苷与miR-126 mimics促进癌细胞凋亡。结论 黄芩苷可以抑制乳腺癌细胞的增殖,促进其凋亡,其机制可能与通过上调miR-126调节凋亡相关基因有关。