CD3AK细胞在裸鼠体内抗转移作用的研究

目的研究肿瘤引流淋巴结的淋巴细胞在体外用ＣＤ３单克隆抗体激活（ＣＤ３ＡＫ）后在体内是否仍然具有抗肿瘤作用。方法取卵巢癌病人盆腔引流淋巴结淋巴细胞,在体外用ＣＤ３单抗激活后输注载高转移人卵巢癌裸鼠体内实验性治疗。实验分４组：顺铂组,ＣＤ３ＡＫ组,联合治疗组,对照组。治疗于接种移植瘤后１０天开始,历时８０天。结果观察到ＣＤ３ＡＫ组１只裸鼠移植瘤消退,２／７只出现转移灶,与对照组８／１０只裸鼠出现转移灶比较差异有显著性（Ｐ＜０．０５）。ＣＤ３ＡＫ组肿瘤体积（０．５７８８±０．２５４９）与对照组（１．５６８５±０．２８３０）比较差异有显著性（Ｐ＜０．０５）。ＣＤ３ＡＫ组抑瘤率达６３．１％。ＣＤ３ＡＫ组裸鼠的血清孕酮含量（３．３８４３±０．５３１４）明显低于对照组（６．３４８０±０．７６１５）,两组比较差异有显著性（Ｐ＜０．０５）。ＣＤ３ＡＫ组淋巴结窦性组织细胞增生（５９／６９枚）明显多于对照组（５５／９４枚）,两组比较差异有非常显著性（Ｐ＜０．０１）。结论ＣＤ３ＡＫ细胞在裸鼠体内具有抗肿瘤生长和抗转移作用,同时有提高宿主免疫功能的作用。     

鼻咽癌细胞裸鼠体内传代中转移及增殖动力学研究

目的：为研究鼻咽癌细胞在裸鼠体内演进中转移能力和增殖能力之间的关系。方法：将鼻咽癌单克隆细胞株（ＣＮＥ－２Ｚ－Ｈ５）移植于裸鼠皮下，待裸鼠发生转移时，取其淋巴结和（或）肺转移灶癌细胞再次移植裸鼠皮下传代，如此用转移灶癌细胞重复传２～４个代后，观察各代裸鼠的转移率和转移途径，同时观察各代癌细胞生长曲线、增殖指数及移植瘤瘤重和体积倍增时间的变化。结果：随着鼻咽癌转移灶癌细胞裸鼠体内的演进，各代裸鼠的转移率增高，且转移途径更为单一，各代癌细胞的增殖能力不断增强。结论：鼻咽癌细胞在裸鼠体内演进中转移能力和增殖能力均不断增强，且二者关系密切。     

应用连锁不平衡检验法进行单纯性先天性心脏病易感基因 …

将人类单纯性先天性心脏病（ｃｏｎｇｅｎｉｔａｌｈｅａｒｔｄｅｆｅｃｔ，ＣＨＤ）易感基因初步定位，为进一步对其克隆奠定基础，方法在胚胎心脏发育调控相关基因－ＨＯＸ基因Ａ簇、Ｂ簇所在在的染色体区域７ｐ１４－１５、１７ｑ２１内选择３个微卫星ＤＮＡ标记Ｄ７Ｓ１８０８、Ｄ７Ｓ６７３、Ｄ１７Ｓ７９１，应用荧光标记聚合酶边反应技术扩增微卫星征 段，对３９个单纯性ＣＨＤ核心家系的１１２名成员进行基因型，并进行遗传     

裸小鼠原位移植筛选高转移性人骨肉瘤细胞及其生物 …

目的 在裸小鼠体内筛选人骨肉瘤高肺转移性细胞亚群,为实验研究提供模型。方法 利用原位移植方法,将人骨肉瘤细胞系ＳＯＳＰ－９６０７皮下移植瘤接种于裸鼠胫骨骨髓腔。筛选到第２代时,有１例发生肺转移,以皮下扩增转移灶,再次原位移植,第３代肺转移率达８３％（５／６）。用组织块法培养收集。比较其与纱在形态结构、细胞核型,ｖｉｍｅｎｔｉｎ,ａｃｔｉｎ,ＮＳＥ抗原表达及体积倍时间待贩差别。结果 同母系相比较,该     

人巨细胞肺癌裸鼠移植瘤中mts1和nm23-H1表达及其与肺癌淋巴结转移关系

目的 探讨肿瘤转移相关基因mts1和nm23-H1在人巨细胞肺癌裸鼠移植瘤中的表达及其与啼癌淋巴结转移的关系。方法 用高转移的人巨细胞肺癌瘤株PLA-801—D和低转移的人巨细胞肺癌瘤株PLA-801—C在裸鼠中建立转移模型,观察瘤细胞在裸鼠淋巴结及肺中的转移情况;原位杂交检测mts1和nm23-H1mRNA在移植瘤中的表达。结果 PLA-801—D在12只移植裸鼠中均有淋巴结转移,转移率100%,2只裸鼠中有肺转移,转移率17％;PLA-801—C在12只移植的裸鼠中均无淋巴结及肺转移。mts1mRNA在PLA-801—D、PLA-801-C裸鼠移植瘤中阳性表达分别为12例、2例;nm23-H1mRNA在PLA-801—D、PLA-801—C裸鼠移植瘤中阳性表达分别为12例、11例。结论 mts1 mRNA表达与人巨细胞啼癌裸鼠移植瘤淋巴结转移呈正相关,nm23-H1 mRNA表达与人巨细胞肺癌裸鼠移植瘤淋巴结转移不相关。     

中国人胃癌组织微卫星DNA的不稳定性研究

目的为探明中国人胃癌的微卫星ＤＮＡ的不稳定性（ｍｉｃｒｏｓａｔｅｌｉｔｅｉｎｓｔａｂｉｌｉｔｙ，ＭＳＩ）频率。方法选择了２９个多态微卫星标记，采用ＰＣＲ－聚丙烯酰胺凝胶电泳及银染技术对４２例胃癌组织进行了ＭＳＩ分析。结果中国人胃癌，在２９个位点上平均ＭＳＩ频率为３３．９％。在Ｄ３Ｓ１０６７、Ｄ３Ｓ１５７７、Ｄ８Ｓ２７９、Ｄ９Ｓ２５７、Ｄ１Ｓ２４８、Ｄ７Ｓ５２０及Ｄ２Ｓ１４７等位点上存在高频率的ＭＳＩ，其中Ｄ３Ｓ１５７７和Ｄ３Ｓ１０６７（５１．３５％）ＭＳＩ发生频率最高。胃癌的不同病理类型表现出不同的ＭＳＩ。低分化胃癌和印戒细胞癌与高分化癌比，ＭＳＩ频率明显增高（Ｐ＜０．０１）；而低分化胃癌和印戒细胞癌之间ＭＳＩ无显著差异（Ｐ＞０．０５）。结论ＭＳＩ在中国人胃癌中起着重要作用，尤其在低分化胃癌中。这些资料进一步证明，高低分化的胃癌可能具有不同的发病机理     

高转移人卵巢癌细胞系HO-8910PM的建立及其生物学特性

我们在建立高转移人卵巢癌裸鼠皮下移植瘤模型的基础上［１］，又建立高转移人卵巢癌体外细胞系ＨＯ８９１０ＰＭ。现将建系过程及其生物学特性介绍如下。一、材料和方法１材料来源：取高转移人卵巢癌皮下移植瘤模型第７代３７０号裸鼠皮下肿块，在无菌条件下剪碎至１...     

人食管癌细胞系的建立及其转移相关基因PCNA的表达

目的:建立人食管癌细胞系,并对其肿瘤标志物(caycinoembryanic antigen,CEA)和转移相关基因增殖细胞核抗原(proliferating cell nuclear antigen,PCNA)的表达进行研究。方法:将人食管癌手术标本移植至裸鼠皮下,成瘤后取组织块体外原代培养,光镜和电镜观察细胞形态,测定细胞周期和染色体众数。取部分裸鼠皮下移植瘤接种至裸鼠食管,观察裸鼠体内肿瘤转移特征,采用免疫组化法测定肝转移灶PCNA基因的表达。结果:裸鼠皮下移植瘤组织16d,可形成明显的瘤结节,体外培养获得大量的肿瘤细胞,命名为YXA-1,其形态和超微结构符合鳞状细胞癌的形态学特征,染色体保持人类肿瘤染色体的特点,众数为70～76,细胞中肿瘤标志物CEA。食管癌标本原位移植裸鼠21d时,发生了广泛的肝内侵袭和转移,转移灶中PCNA表达为阳性。结论:人食管癌细胞系YXA-1维持了原发肿瘤的特征,而裸鼠其原位移植模型体现了肿瘤的转移特性,可用于肿瘤的转移研究。     

一个新的微卫星多态标记(D14S1435)的FISH定位

目的将从人染色体１４ｑ２４．３显微切割制备的ＤＮＡ文库中分离得到的一个新的微卫星多态标记（Ｄ１４Ｓ１４３５，ＣＡ重复序列，人群中存在１１种等位形式，ＰＩＣ值为０．８５）进行染色体荧光原位杂交（ＦＩＳＨ）返回定位。方法以此多态标记筛选人Ｌａｍｂｄａ／ＤＡＳＨ基因组文库，得到的阳性重组噬菌体ＤＮＡ，通过ＢａｍＨⅠ完全酶切，低熔点胶电泳回收插入片段，再经Ｓａｕ３ＡⅠ酶切，接头捕获ＰＣＲ（ｌｉｎｋｅｒ－ｃａｔｃｈＰＣＲ）法标记探针进行ＦＩＳＨ。结果将这一新的ＳＴＲ精确地定位于人染色体１４ｑ２４．３，证实了从染色体显微切割探针池分离染色体区带特异性遗传标记的可靠性。结论经染色体显微切割、ＰＣＲ及微克隆方法所得到的遗传标记，通过染色体荧光原位杂交证实其确实来自所切割区域，增加了这一区域可用于连锁分析的遗传标记，为定位在１４ｑ２４．３区带内若干遗传病的基因诊断和基因克隆，提供一个新的有价值的遗传标记。     

介导RA538与反义c—myc腺病毒对肿瘤细胞的作用及其分 …

目的 比较全反式维甲酸诱导基因ＲＡ５３８及反义ｃ－ｍｙｃ对人胃癌细胞的生物学特性，并探讨其作用的分子机理。方法 采用细胞生长曲线、ＤＮＡ梯度降解试验、原位末端标记、流式细胞仪、逆转录－聚合酶链反应、蛋白质印迹分析、裸鼠致瘤性、裸鼠皮下移植瘤模型实验等方法，对ＲＡ５３８、反义ｃ－ｍｙｃ重组腺病毒在人胃癌细胞系（ＳＧＣ７９０１）中的生物学作用及其分子机理进行体外研究。结果 ＲＡ５３８及反义ｃ－ｍｙｃ重     

Microsatellite instability in early-onset breast cancer

Breast cancer in a young person is considered a rare and very aggressive disease. The theories addressing the underlying genetic mechanisms of this disease are controversial. Therefore, additional genetic concepts playing a possible role in its pathogenesis and prognosis must be investigated. Microsatellite instability (MSI) characterized by a mutational process of insertions or deletions in microsatellite repeats might constitute a sensitive indicator for genomic instability in cancer. MSI has been described in a wide variety of tumors, particularly in hereditary non-polyposis colorectal cancer. The reports regarding its occurrence and prognostic significance in breast cancer are in conflict with each other. The purpose of this study was to investigate MSI in early-onset breast cancer and to correlate its occurrence with clinicopathological prognisticators. In this study, 16 female patients with primary breast cancer under 35 years of age (range 29-34) were investigated for the incidence of MSI in five microsatellite loci (D2S123, D3S1611, D17S807, D17S796 and Xq11-12) by comparing paired normal and tumor tissue DNA after PCR amplification from paraffin-embedded tissues. No instability was found in any of these five microsatellite loci. Although care must be taken not to overstate the importance of this result due to the inadequate number of microsatellite markers and DNA samples studied, this preliminary report indicates that MSI phenotype is uncommon in human early-onset breast cancer. Therefore, it does not appear to be related to the prognosis of disease.     

Identification of a I-cM region of common deletion on 13q14 associated with human prostate cancer.

Frequent allelic losses on chromosome arm 13q are observed in carcinomas of the head and neck, breast, ovary, and pituitary gland. We analyzed 59 primary prostate tumors (stage B, 18 patients; C, 12 patients; D1, 4 patients; and endocrine therapy-resistant cancer death, 25 patients), as well as 18 metastatic tissues from 14 of the 25 cancer death patients for loss of heterozygosity (LOH) using 35 microsatellite markers on chromosome arm 13q. Of the 59 primary tumors, 31 (53%) showed LOH involving at least one locus. Detailed deletion mapping identified a distinct commonly deleted region in the I-cM interval flanked by D13S153 and D13S273 on 13q14 and this region overlapped a part of the RB1 gene. Paired DNAs were available from both primary and metastatic tumors in the 14 cases of cancer death; among those pairs, we detected LOH on 13q in seven (50%) primary tumors, and in all metastatic foci (P = 0.0029). Moreover, the regions lost in metastatic tissues were more extensive than those seen in the corresponding primary tumors. These results suggest that inactivation of a putative tumor suppressor gene(s) including the RB1 gene on 13q14 plays an important role in human prostate cancer.     

Allelic loss of chromosomal arm 8p in breast cancer progression.

Loss of heterozygosity (LOH) of chromosomal arm 8p has been reported to occur at high frequency for a number of common forms of human cancer, including breast cancer. The objectives of this study were to define the regions on this chromosomal arm that are likely to contain breast cancer tumor suppressor genes and to determine when loss of chromosomal arm 8p occurs during breast cancer progression. For mapping the tumor suppressor gene loci, we evaluated 60 cases of infiltrating ductal cancer for allelic loss using 14 microsatellite markers mapped to this chromosomal arm and found LOH of 8p in 36 (60%) of the tumors. Whereas most of these tumors had allelic loss at all informative markers, five tumors had partial loss of 8p affecting two nonoverlapping regions. LOH for all but one of the tumors with 8p loss involved the region between markers D8S560 and D8S518 at 8p21.3-p23.3, suggesting that this is the locus of a breast cancer tumor suppressor gene. We then studied LOH of 8p in 38 cases of ductal carcinoma in situ (DCIS) with multiple individually microdissected tumor foci evaluated for each case. LOH of 8p was found in 14 of the DCIS cases (36%), including 6 of 16 cases of low histological grade and 8 of 22 cases of intermediate or high histological grade. In four of these DCIS cases, 8p LOH was seen in some but not all of the multiple tumor foci examined. These data suggest that during the evolution of these tumors, LOH of 8p occurred after loss of other chromosomal arms that were lost in all tumor foci. Thus, LOH of 8p, particularly 8p21.3-p23, is a common genetic alteration in infiltrating and in situ breast cancer. Although 8p LOH is common even in low histological grade DCIS, this allelic loss often appears to be preceded by loss of other alleles in the evolution of breast cancer.     

Rapid detection of xenotransplanted human tissues using in situ hybridization.

A rapid and sensitive method for the identification of human tissues xenotransplanted in nude mice was developed. An in situ hybridization technique made it possible to distinguish between cells of human origin and cells of murine origin in formalin-fixed paraffin sections. High-molecular-weight DNAs extracted from human or mouse tissues were sonicated, nick-translated with 32P-dCTP, and used as hybridization probes. Dot blot hybridization of 32P-labeled probes revealed clear species-specific signals. Formalin-fixed paraffin-embedded tissue samples from repopulated tracheal transplants, containing either human tracheal epithelial cells or human renal tubular cells, were used. Cells of human and murine origin were distinguishable by in situ hybridization with sonicated DNA probes. This method has several advantages; simple preparation of probes, high sensitivity, and applicability to formalin-fixed paraffin-embedded tissue sections. In situ hybridization with sonicated DNA probes should provide a powerful tool for verifying the human origin of xenotransplanted tissues in nude mice.     

Bcl-x(L)-mediated changes in metabolic pathways of breast cancer cells: from survival in the blood stream to organ-specific metastasis

Bcl-x(L) protein plays a role in breast cancer dormancy, promoting survival of cells in metastatic foci by counteracting the proapoptotic signals in the microenvironment. The aim of this study was to identify phenotypes mediated by Bcl-x(L) in breast cancer cells that enhance in vivo survival of clinical metastases. 435/Bcl-x(L) or 435/Neo human breast cancer cells were injected into the inguinal mammary gland of nude mice, and tumors, metastases in lymph node, lung, and bone, and bloodstream surviving cells were examined. Proteomic analysis identified 17 proteins that were overexpressed (more than twofold) or underexpressed (less than twofold) in metastases. A protein interaction program allowed us to functionally associate peroxiredoxin 3, peroxiredoxin 2, carbonyl reductase 3, and enolase 1, suggesting a role for cellular responses to oxidative stress in metastasis organ selection. The prediction included proteins involved in redox systems, kinase pathways, and the ATP synthase complex. Furthermore, the interaction of redox proteins with enolase 1 suggests a connection between glycolysis and antioxidant pathways, enabling achievement of a high metastatic activity. In conclusion, Bcl-x(L) mediates a phenotype in which redox pathways and glycolysis are coupled to protect breast cancer metastatic cells during transit from the primary tumor to the metastatic state.     

Spontaneous length variation in microsatellite dna from human T-cell clones

Recently, much interest has been focused on instability of microsatellite DNA sequences such as di- and tri-nucleotide repeats in human cancers. Certain tumors show an increased frequency of mutation leading to repeat length variation at microsatellite loci, and it is thought that such instability may be a marker for the transformed phenotype. However, the spontaneous frequency by which repetitive DNA such as CA-repeats undergoes size changes in normal human somatic cells is not known. Therefore, it is not possible to decide if there is an increase in the frequency of microsatellite mutation in specific tumors or if the change observed simply reflects the frequency of microsatellite mutation in the cell population from which the tumor originates. To investigate this we have established panels of T-tymphocyte clones from 28 healthy males and determined the spontaneous length variations at three CA-repeat markers that are often used to investigate satellite instability: D2S123, D9S180, and D10S197. We found 3 T-cell clones with altered microsatellite size in a total of 178. This corresponds to a background frequency of 3 somatic microsatellite mutations in 1,028 alleles studied, i.e., 2.9 × 10^?3. This frequency is comparable to that found in many tumors of the breast, brain, ovary, and skin but is considerably lower than the frequency of microsatellite mutation in tumors related to hereditary non-polyposis colorectal cancer.     

Sub-cellular accumulation of magnetic nanoparticles in breast tumors and metastases

In this study, the sub-cellular accumulation of superparamagnetic iron oxide nanoparticles (SPIONs) in breast tumors and peripheral organs were investigated. MNPs were conjugated with luteinizing hormone releasing hormone (LHRH), whose receptors are expressed by most types of breast cancer cells. After the nanoparticles were injected into female nude mice bearing MDA-MB-435S.luc tumors, the mice were sacrificed to collect tumors and peripheral organs for biological and TEM analyses. LHRH conjugated SPIONs (LHRH- SPIONs) were found to accumulate in cancer cells, mainly in the primary tumors and the metastatic lungs, where they aggregated to form clusters. In contrast, most of the unconjugated SPIONs were collected in the liver cells. The results suggest that LHRH- SPIONs can be used to target cancer cells in the primary breast tumors and the lung metastases. TEM is also shown to be a useful tool for the studies of sub-cellular distributions of SPIONs in tumors and tissues.     

Patterns of allelic loss at the BRCA1 locus in Arabic women with breast cancer

Loss of heterozygosity (LOH) of BRCA1, a tumor suppressor gene, is one mechanism of genetic inactivation in both sporadic and familial forms of breast cancer. Studies reported in breast cancers from women of Northern European descent have shown LOH in 30-50% of sporadic tumors. Microsatellite instability (MSI) has served as evidence for involvement of DNA repair genes. This study investigates the extent of allelic imbalance at the BRCA1 region in Arabic women with breast cancer. Paired normal and tumor tissue were available for DNA analysis in 13 cases. Results using fluorescent tagged primers to microsatellite markers D17S1323, D17S1325 and D17S855 intragenic to BRCA1 were analyzed using an ABI 310 DNA sequencer. As compared to normal DNA, MSI and LOH were recognized as a gain and a loss, respectively, of one signal in one allele in the tumor DNA. Microsatellite analyses showed 12 of 13 (92%) cases with LOH or MSI or both. Three cases demonstrated LOH alone, 3 cases with MSI alone. Six cases indicated both LOH and MSI; 2 cases with either LOH or MSI in separate markers. The combined finding of LOH and MSI in the same marker was detected only with D17S1325 in 4/6 cases. The proportion of aberrant findings of the BRCA1 locus in breast cancer appears to be higher in Arabic women than in other populations studied to date.     

Microsatellite instability at a single locus (D11S988) on chromosome 11p15.5 as a late event in mammary tumorigenesis

Replication errors at microsatellite repeats are markers forgenomic instability in hereditary nonpolyposis colon carcinomaand in some sporadic cancers. Microsatellite sequences may showalterations in one or both alleles in some tumors, suggestingan error in the DNA replication of dinucleotide repeats. Wehave investigated microsatellite instability (MSI) in sporadicbreast tumors at several loci on the short arm of chromosome11. Among microsateliltes studied we found a high frequencyof MSI at one specific locus, D11S988 on chromosome 11p15.5.Most colorectal tumors that exhibit MSI display abnormalitiesof at least one other locus and usually more. By contrast wehave detected only one abnormal microsatellite in all the tumorsexamined. This marker lies between the TH and HBB genes, a subregionpreviously suggested to harbor a putative tumor suppressor genefor breast cancer. Loss of heterozygosity for chromosome markersat 11 p15 has earlier been correlated with poor prognosis. Inan unselected panel of primary breast tumors, we observed that20 of 69 showed mobility shifts of D11S988 in tumor comparedwith corresponding normal DNA samples. Tumors with instabilityat D11S988 were rapidly proliferating compared with tumors withoutMSI. DNA aneuploidy, estrogen receptor positivity and moderateto poorly differentiated tumor phenotype were also characteristicsof tumors with MSI at this locus and the majority also exhibitedloss of heterozygosity at one or more of the six 11p loci analyzed.Taken together these data suggest that MSI at the D11S988 locusis a late event in mammary tumorigenesis and may be associatedwith progression of breast carcinomas.     

Dietary stearate reduces human breast cancer metastasis burden in athymic nude mice

Stearate is an 18-carbon saturated fatty acid found in many foods in the western diet, including beef and chocolate. Stearate has been shown to have anti-cancer properties during early stages of neoplastic progression. However, previous studies have not investigated the effect of dietary stearate on breast cancer metastasis. In this study, we present evidence that exogenously supplied dietary stearate dramatically reduces the size of tumors that formed from injected human breast cancer cells within the mammary fat pads of athymic nude mice by approximately 50% and partially inhibits breast cancer cell metastasis burden in the lungs in this mouse model system. This metastatic inhibition appears to be independent of primary tumor size, as stearate fed animals that had primary tumors comparable in size to littermates fed either a safflower oil enriched diet or a low fat diet had reduced lung metastasis. Also stearate fed mice sub-groups had different primary tumor sizes but no difference in metastasis. This anti-metastasis effect may be due, at least in part, to the ability of stearate to induce apoptosis in these human breast cancer cells. Overall, this study suggests the possibility of dietary manipulation with selected long-chain saturated fatty acids such as stearate as a potential adjuvant therapeutic strategy for breast cancer patients wishing to maximize the suppression of metastatic disease.