大肠癌细胞DNA、AgNORs定量研究──两者相关性及预后意义的探讨

应用ＡｇＮＯＲｓ染色技术，对６３例大肠癌的ＡｇＮＯＲｓ颗粒进行了计数研究。其中３８例同时进行肿瘤细胞核ＤＮＡ含量静态测定。结果表明，大肠癌细胞ＡｇＮＯＲｓ数量与其ＤＮＡ含量之间有明显相关性（Ｐ＜０．０ｌ）。ＤＮＡ含量和ＡｇＮＯＲｓ计数愈低，肿瘤分化愈好，患者预后愈好；反之，ＤＮＡ含量和Ａｇ－ＮＯＲｓ计数愈高、肝瘤分化愈差，患者预后愈差。表明ＡｇＮＯＲｓ计数，ＤＮＡ含量检测均可作为大肠癌分级、判断预后的参考指标。     

肝针吸良、恶性细胞核内AgNORs图像分析的定量研究

应用真彩色图像分析技术对肝针吸细胞中的２０例肝癌及５例增生肝细胞核内ＡｇＮＯＲｓ进行了定量研究。结果显示，肝针吸肝癌细胞核内ＡｇＮＯＲｓ的平均面积、平均积分光密度、颗粒直径及平均颗粒数均明显高于增生肝细胞（Ｐ＜００１）；ＡｇＮＯＲｓ图像定量分析可为肝针吸癌细胞和增生肝细胞的鉴别诊断提供一种新的定量指标；ＡｇＮＯＲｓ在形态、数量及分布上的不同特征也可作为鉴别两者的重要依据。     

食管鳞癌组织AgNOR计数与DNA倍的关系

本文应用ＡｇＮＯＲ技术对２０例食管鳞癌手术切除标本中癌细胞核内ＡｇＮＯＲ进行计数，同时运用流式细胞分析技术测定其ＤＮＡ指数及Ｓ期细胞百分率，并与ＡｇＮＯＲ计数进行比较，结果显示：食管鳞癌细胞核内平均ＡｇＮＯＲ数目与Ｓ期细胞百分率呈明显正相关（ｒ＝０．７３）。而ＤＩ与ＡｇＮＯＲ计数无明显相关关系（ｒ＝０．３０）。     

宫颈病变细胞核AgNOR颗粒形态定量分析

为探讨不同程度宫颈病变的细胞核、ＡｇＮＯＲ的变化规律。利用自动图像分析技术分别对２８例宫颈癌，５５例宫颈非典型增生和１５例正常宫颈组织细胞核及ＡｇＮＯＲ颗粒形态定量参数进行测定。结果显示：除正常宫颈与轻度非典型增生宫颈组外，宫颈中度、重度非典型增生与宫颈鳞癌三组之间其平均细胞核面积、每个核ＡｇＮＯＲ颗粒数、平均颗粒面积、颗粒直径以及ＡｇＮＯＲ颗粒面积／细胞核面积之比值逐渐增加，而ＡｇＮＯＲ颗粒形状因子逐渐减小，各组之间存在着统计学差异。ＡｇＮＯＲ颗粒直径变化不显著。认为在宫颈组织发生癌变过程中，细胞核及核内ＡｇＮＯＲ颗粒大小、形态和数量均发生改变。提示细胞核及核内ＡｇＮＯＲ定量参数有可能作为估计肿瘤细胞恶性潜能的指标     

食管鳞癌组织AgNOR计数与DNA倍体的关系

本文应用ＡｇＮＯＲ技术对２０例食管鳞癌手术切除标本中癌细胞核内ＡｇＮＯＲ进行计数，同时运用流式细胞分析技术测定其ＤＮＡ指数（ＤＩ）及Ｓ期细胞百分率，并与ＡｇＮＯＲ计数进行比较。结果显示：食管鳞癌细胞核内平均ＡｇＮＯＲ数目与Ｓ期细胞百分率呈明显正相关（ｒ＝０．７３）。而ＤＩ与ＡｇＮＯＲ计数无明显相关关系（ｒ＝０．３０）。     

AgNORs检验对胃粘膜良,恶性病变的诊断价值

为探讨ＡｇＮＯＲｓ变化对胃粘膜良，恶性病变的诊断价值，应用银染技术对５０例胃粘膜良，恶性病变进行ＡｇＮＯＲｓ计数，其中正常胃粘膜４例，萎缩性胃炎５例，异型增生９例，胃癌３２例结果。正常胃粘膜，萎缩性胃炎和异型增生细胞核内ＡｇＮＯＲｓ颗粒均数，分别与胃癌各型癌细胞核内均数比较均有显著性差异     

形态计量学在辐射及化学致癌物诱发体外细胞转化研究中的应用

本研究应用细胞化学和形态计量学的实验手段，对６０Ｃｏγ射线及苯并芘在体外诱发人胚肺细胞转化过程中，细胞核内ＡｇＮＯＲｓ颗粒数量、大小等形态计量学参数的改变进行了研究，并对其生物学意义进行了分析。结果表明，γ射线和苯并芘均可在体外诱发人胚肺细胞发生转化。γ射线和苯并芘作用后早期，当人胚肺细胞尚未发生转化时，细胞核内ＡｇＮＯＲｓ颗粒计数及部分形态计量学参数已发生明显改变，并具有一定的剂量效应关系。这些规律性的变化，可用于定量地研究辐射或化学致癌因子体外诱发细胞转化的剂量效应关系，亦可望成为早期观察细胞转化程度的指标。物理化学因素复合效应研究表明，γ射线和苯并芘复合作用，对人胚肺细胞转化有复合加强作用，同时对细胞内ＡｇＮＯＲｓ颗粒数及形态计量学参数的改变亦表现出复合加强的使用。     

胃癌前病变AgNOR面积及形态与其DNA含量的对照研究

本文应用计算机图象分析系统对１０例肠化、３０例不同程度异型增生的胃粘膜，１０例正常胃粘膜和１０例高分化腺癌细胞核内ＡｇＮＯＲ蛋白的面积及其颗粒的形状进行了定量，并测定了其ＤＮＡ含量，结果显示，随胃粘膜病变程度的加重，其核内ＡｇＮＯＲ蛋白与ＤＮＡ含量依次增加，而ＡｇＮＯＲ颗粒的形状因子呈递减趋势。ＡｇＮＯＲ蛋白面积／核和其颗粒的形状因子与ＤＮＡ含量均有良好的线性相关关系，前者为正相关（ｒ＝０．９９Ｐ＜０．００１），后者为负相关（ｒ＝－０．９５２Ｐ＜０．００１）。我们认为，ＡｇＮＯＲ面积及其颗粒的形状可以反映出胃粘膜的病变程度。可作为胃癌前病变诊断及病情监测的有用指标。     

甲状腺肿瘤细胞核DNA含量及AgNORs的图像定量分析

甲状腺肿瘤细胞核ＤＮＡ含量及ＡｇＮＯＲｓ的图像定量分析余力，区士欢，肖莎本文作者通过图像分析技术对３８例甲状腺肿瘤患者的肿瘤细胞核面积、ＤＮＡ含量及ＡｇＮＯＲｓ进行定量分析，现将结果报告于下。１材料与方法甲状腺腺癌３０例，其中乳头状腺癌２２例，滤泡状...     

AgNOR的图像分析在判断子宫内膜癌预后方面的研究

应用图像分析技术对１０３例子宫内膜癌癌细胞中ＡｇＮＯＲ进行了定量研究。结果表明，低分化、肌层浸润＞１／２者的癌细胞核面积、每核ＡｇＮＯＲ颗粒总面积、颗粒数及颗粒平均面积明显大于高分化及无肌层浸润者。Ⅱ、Ⅳ期、复发、死亡患者的癌细胞的核面积及ＡｇＮＯＲ颗粒平均面积较Ⅰ期、无复发、存活者明显增高。ＡｇＮＯＲ的颗粒平均面积多在１．２０μｍ×１．２０μｍ以上，并且ＡｇＮＯＲ颗粒分布多呈聚集型；而预后较好的患者，ＡｇＮＯＲ颗粒平均面积多在亚．１５μｍ×１．１５μｍ左右，颗粒多呈弥散分布。结果提示，细胞内ＡｇＮＯＲ的含量与子宫内膜癌的生物学行为及预后有关，图像分析进行ＡｇＮＯＲ的定量研究较传统的光镜下计数ＡｇＮＯＲ颗粒数目更为客观、准确。     

Isoliquiritigenin induces G2 and M phase arrest by inducing DNA damage and by inhibiting the metaphase/anaphase transition

Isoliquiritigenin, a natural flavonoid found in licorice, shallots, and bean sprouts, has been demonstrated to inhibit proliferation and to induce apoptosis in a variety of human cancer cells. We attempted to ascertain the underlying mechanism by which isoliquiritigenin induced cell cycle arrest and cytotoxicity in HeLa human cervical cancer cells. Isoliquiritigenin treatment arrested cells in both G2 and M phase. The cells arrested in interphase (G2) showed markers for DNA damage including the formation of γ-H2AX foci and the phosphorylation of ATM and Chk2, whereas the cells arrested in M phase evidenced separate poles and mitotic metaphase-like spindles with partially unaligned chromosomes. The induction of DNA damage and blockade at the metaphase/anaphase transition implied that isoliquiritigenin might function as a topoisomerase II poison, which was further demonstrated via an in vitro topoisomerase II inhibition assay. These results show that isoliquiritigenin inhibits topoiosmerase II activity, and the resultant DNA damage and arrest in mitotic metaphase-like stage contributes to the antiproliferative effects of isoliquiritigenin.     

Targeted cancer therapy with ribosome biogenesis inhibitors: a real possibility?

The effects of many chemotherapeutic drugs on ribosome biogenesis have been underestimated for a long time. Indeed, many drugs currently used for cancer treatment – and which are known to either damage DNA or hinder DNA synthesis – have been shown to exert their toxic action mainly by inhibiting rRNA synthesis or maturation. Moreover, there are new drugs that have been proposed recently for cancer chemotherapy, which only hinder ribosome biogenesis without any genotoxic activity. Even though ribosome biogenesis occurs in both normal and cancer cells, whether resting or proliferating, there is evidence that the selective inhibition of ribosome biogenesis may, in some instances, result in a selective damage to neoplastic cells. The higher sensitivity of cancer cells to inhibitors of rRNA synthesis appears to be the consequence of either the loss of the mechanisms controlling the cell cycle progression or the acquisition of activating oncogene and inactivating tumor suppressor gene mutations that up-regulate the ribosome biogenesis rate. This article reviews those cancer cell characteristics on which the selective cancer cell cytotoxicity induced by the inhibitors of ribosome biogenesis is based.     

核仁组成区测定对胃癌和癌前病变诊断上的意义

应用核仁组成区嗜银蛋白染色法(AgNoRs),对130例胃粘膜病变进行定量测定。结果表明单纯肠上皮化生、异型增生和分化型腺癌的每核平均AgNoRs颗粒量,随病变程度加重而递增。除重度异型增生与腺癌明显交叉重叠外,各组之间皆有非常显著性差异(P<0.01)。因此,我们认为AgNoRs测定,可作为胃癌和癌前病变诊断、分级上较客观的辅助性定量指标。高活性rRNA的出现可能是重要癌变标志。     

Nucleolar organiser regions (AgNORs) as predictors in transitional cell bladder cancer.

The predictive value of silver stained nucleolar organiser regions (AgNORs) was assessed in 229 patients with transitional cell bladder cancer followed up for over 10 years. The AgNORs were enumerated in pretreatment biopsy specimens. The AgNORs were related to clinical stage (T) (P = 0.0111), papillarity (P less than 0.0001), WHO grade (P less than 0.0001), DNA ploidy (P = 0.0010) and S-phase fraction (P less than 0.0001). Tumours presenting with pelvic lymph node involvement (P = 0.0085) or metastasis (P = 0.0780) at the time of diagnosis had more AgNORs than tumours confined to the bladder wall. Progression in T-, N- and M-categories (P = 0.0010-0.0030) was related to AgNORs and consequently they predicted bladder cancer related survival (P = 0.0005). The diploid tumours could be regrouped according to survival by AgNORs (P = 0.0001). In papillary tumours AgNORs predicted progression (P = 0.0110) and survival (P = 0.0038). In Ta-T1 tumours AgNORs predicted progression (P = 0.11) and survival (P = 0.0751) and also in T2-T3 tumours AgNORs contributed to survival significantly (P = 0.0039). The AgNORs subdivided WHO grade III tumours according to their ability to progress during the follow-up time (P = 0.0711). In a multivariate analysis AgNORs predicted progression independently in Ta-T1 category (P = 0.0165). AgNORs predicted recurrence free period like SPF (P = 0.0010). In conclusion, AgNORs are inferior to classic prognostic factors or DNA flow cytometric variables in muscle invasive bladder cancers whereas they have independent predictive value in superficial cancers.     

Role of free radicals in an adriamycin-resistant human small cell lung cancer cell line

In two Adriamycin (Adr) resistant sublines (GLC4-Adr1 and GLC4-Adr2) of a human small cell lung carcinoma cell line, GLC4, cross-resistance for radiation was found. GLC4-Adr1 has an acquired Adr resistance factor of 44 after culturing without Adr for 20 days and GLC4-Adr2, the same subline cultured without Adr for 3 months, has a decreased but stable resistance factor of 8. One of the assumed mechanisms of Adr is that the effect is mediated through the formation of free radicals. Therefore free radical scavenging might play a role in these Adr resistant cell lines. Adr, H2O2, and X-ray induced cytotoxicity were evaluated. Glutathione (GSH) levels and activities of associated enzymes were determined as well as Adr, H2O2, and X-ray induced DNA breaks and repair. GSH level was decreased in GLC4-Adr1, but restored to the normal level in GLC4-Adr2. Superoxide dismutase, catalase, glutathione-peroxidase, and glutathione S-transferase were not elevated in the resistant sublines. Adr induced a decreased amount of DNA breaks in GLC4-Adr1 compared to GLC4. For X-ray and H2O2 a comparable amount of DNA damage was found. GLC4-Adr1 was able to repair DNA breaks induced by Adr, X-ray, and H2O2 better than GLC4. In conclusion, no increased enzyme capacity for detoxification of free radicals could be detected in the cytosol of the resistant cells. The resistance against free radicals in the GLC4-Adr1 line may at least in part be a result of increased DNA repair.     

Apoptosis of human melanoma cells induced by the novel compounds propolin A and propolin B from Taiwenese propolis

We recently demonstrated that two new prenylflavanones, propolin A and propolin B, isolated and characterized from Taiwanese propolis, induced cytotoxicity effect in human melanoma A2058 cells and shows a strong capability to scavenge free radicals. In this study, propolin A effectively induced a cytotoxic effect on five different cancer cell lines. Similar results were obtained for propolin B. DNA flow cytometric analysis and DNA fragmentation ladder indicated that propolin A and propolin B actively induced apoptosis in A2058 cells. To address the mechanism of the apoptosis effect of propolin A and propolin B, we evaluated the apoptosis-related proteins in A2058 cells. The levels of procaspase-8, Bid, procaspase-3, DFF45, and PARP were decreased in dose- and time course-dependent manners. Furthermore, also found propolin A and propolin B was capable of releasing cytochrome c from mitochondria to cytosol. The findings suggest that propolin A and propolin B may activate a mitochondria-mediated apoptosis pathway. On the other hand, our data show that propolin B inhibitied xanthine oxidase activity more efficiently than propolin A or CAPE. However, CAPE suppressed ROS-induced DNA strand breakage more efficiently than propolin A or propolin B. All these results indicated that propolin A and propolin B may trigger apoptosis of A2058 cells through mitochondria-dependent pathways and also shown that propolin A and propolin B were strong antioxidants.     

新型脑组织特异性基因LRRC4的功能研究

目的研究LRRC4基因的抑瘤作用,探讨LRRC4基因对U251细胞的生物学功能影响。方法将转染LRRC4基因的U251细胞、转染空白载体PcDNA3.1(+)的U251细胞和阴性对照的U251细胞分别进行HE染色、DNA染色和核仁组成区嗜银蛋白(AgNORs)染色,并采用图像分析系统分别进行平面形态参数、DNA含量、DNA倍体、细胞周期和AgNORs定量分析,流式细胞仪检测各组细胞的细胞周期分布,MTT检测各组细胞的增殖活性。结果转染LRRC4基因U251细胞的面积为(100.6±7.6)μm2,周长为(42.4±2.0)μm,细胞直径为(9.0±1.2)μm,平均DNA含量为(46.8±8.7)pg,AgNORs平均颗粒数为(1.2±0.4)个,AgNORs平均面积为(16.9±2.0)μm2,均显著低于转染空白载体和阴性对照细胞,DNA异倍体也显著低于转染空白载体和阴性对照细胞。转染LRRC4基因的U251细胞的G0/G1期百分比明显增高,而S期和G2/M期细胞减少。转染LRRC4基因的U251细胞增殖活性显著低于转染空白载体和阴性对照细胞(P<0.01)。结论LRRC4基因通过抑制细胞的DNA复制,减少细胞核仁组成区相关蛋白质的合成,使细胞停滞于G0/G1期,抑制细胞的增殖活性,进而发挥抑瘤功能。形态定量检测技术结合流式细胞检测、MTT检测等实验,能更加全面地阐明新基因的生物学功能。     

Protection against Adriamycin cytotoxicity and inhibition of DNA topoisomerase II activity by 3,4-dihydroxybenzoic acid

The mechanism of Adriamycin (ADR) induced cytotoxicity is not completely understood. While a variety of mechanisms have been proposed, the production of free radicals by redox cycling of the semiquinone has been implicated in cytotoxicity, specifically for cardiotoxicity. To determine whether a scavenger of free radicals would modify the cytotoxicity of ADR, the benzoic acid derivative 3,4-dihydroxybenzoic acid (DHB) was investigated for its ability to protect against ADR-induced cytotoxicity and DNA double strand breaks in Chinese hamster V79 cells. V79 cells were treated with ADR, or its non-redox cycling analog iminodaunomycin, in the presence or absence of DHB. DHB provided significant protection (dose-modifying factor greater than 2.5 for ADR, and nearly 2 for iminodaunomycin) and also caused a dose-dependent decrease in DNA double strand breaks as measured by pulsed field gel electrophoresis. Assays of topoisomerase II activity showed that DHB inhibited topoisomerase II in a concentration-dependent manner, but did not inhibit topoisomerase I. Another non-toxic topoisomerase II inhibitor, the radioprotector WR-1065, also protected against ADR-induced cytotoxicity. These data identify DHB as a non-toxic inhibitor of DNA topoisomerase II and suggest that much of the cytotoxicity of ADR in actively growing V79 cells is due to mechanisms other than redox cycling by the semiquinone.     

PIG3: A novel link between oxidative stress and DNA damage response in cancer

Reactive oxygen species (ROS), the most prominent free radicals produced in cells, can have both beneficial and detrimental effects on them. Many genes are known to be involved in ROS regulation. P53 inducible gene 3 (PIG3 or TP53I3) was identified in an analysis of genes induced by p53 before the onset of apoptosis. It is a widely conserved gene between many species. Until now it has been shown to exert two disparate cellular roles. The first is that of ROS producer linked to p53 induced apoptosis. In this context, it exhibits a NADPH dependent reductase activity with orthoquinones. The second is that of a component of the DNA damage response pathway. While it is considered as a p53 dependent pro-apoptotic gene, it is rarely affected in cancer. This data does not support an anti-tumor activity. In the present review we present and discuss aspects on the regulation and function of this factor and how it is implicated in cancer. We conclude by proposing that PIG3 may possibly have a role in cancer cell survival.     

Comparison of nucleolar organizer regions (NORs) on chromosomes of cultured lymphocytes from cancer patients, high risk cancer family members and normal subjects

Using the technique of AG-NORs, the number of silver-stained NORs on chromosomes of the peripheral lymphocytes from 10 cancer patients, 10 high risk cancer family members and 10 normal individuals was compared. It was found that the total mean value of AG-NORs was 5.46 +/- 0.00/per cell in cancer patients, 5.62 +/- 0.01/per cell in high risk cancer family members and 7.03 +/- 0.07/per cell in normal subjects. The total mean value of AG-NORs was markedly lower in the first two groups than in the third group (P less than 0.001). It seems to be due to the decrease of the transcribed activity of rRNA gene on chromosomes of the cultured lymphocytes in the first two groups. A probable relation between the carcinogenesis and decrease of NORs is shown. The transcribed activity of rRNA gene gives the expression to AG-NORs numbers with which the relation to carcinogenesis is briefly discussed with a review of the literature.