Establishment of a Chinese bladder cancer cell line (T921) with high metastatic activity

Muscle-invasive bladder cancer is prone to metastasis without a standard organ preference. The current cell lines used to study bladder cancer have primarily been derived from individuals in Western populations, and no human bladder cancer cell line has been established from the Chinese population. A bladder cancer cell line was derived from a female Chinese patient with muscle-invasive bladder cancer, and these cells were then xenografted into the bladders of three nude mice. Five weeks later, these mice were killed to observe local invasion and distant metastasis. The metastatic tumors were also removed and analyzed to assess the metastatic mechanism. This bladder cancer cell line, named T921, was successfully established, as evidenced by karyotype and immunohistochemistry analyses. Multi-organ metastases were observed in all three of the nude mice 5 wk after the orthotopic transfer of the cell line. In addition, epithelial–mesenchymal transition (EMT)-related genes were involved in the tumor metastases. The T921 bladder cancer cell line was successfully established, and EMT was observed to play a role in bladder cancer metastasis.     

Cervix tumour cell line established from tumour after long term passages as heterotransplant in nude mice.

One human cancer of the uterine cervix xenograft was established in tissue culture only after repeated passages in nude mice suggesting that with the repeated passages in nude mice, tumour cells acquire some properties which allow them to grow in vitro. Attempts to establish cell line tumours from earlier passages were not successful. The established cell line is tumourigenic. On inoculation of cultured cells in nude mice tumour take was found to be 100%. Karyotypic analysis revealed human origin.     

Human lung cancer cell lines in our laboratory: establishment of three large cell carcinoma cell lines and their biological characterization]

Three large cell carcinoma cell lines were established from tumors of lung cancer patients. The cell lines were named NUTLC-2, NUTLC-4 and NUTLC-5 and they were found to have the following biological characterization. 1) By chromosomal analysis, the tumor cells of two of the cell lines (NUTLC-2 and NUTLC-5) were human-origin cells. Numbers of chromosomes of these cells ranged from 52 to 59 in NUTLC-2 and from 68 to 75 in NUTLC-5, with the modal numbers of 56 and 71, respectively. 2) Primary tumor resected from the patient with lung cancer was heterotransplanted into the subcutis of a nude mouse. NUTLC-4 cell line was established in vitro from the tumor in nude mouse and the tumor cells were found to be mouse-origin cells by chromosomal analysis. Human DNA was not detected by Alu analysis. 3) The tumor cells of three cell lines could be heterotransplanted subcutaneously into nude mice. However, no natural distant metastasis in nude mouse was observed. 4) Drug sensitivity to NUTLC-2, NUTLC-4 and NUTLC-5 tumor cells differed individually according to MTT colorimetric assay, and characteristic drug sensitivity was not noted in histological types of lung cancer.     

人卵巢癌裸鼠移植实体瘤模型的建立

目的建立人卵巢癌裸鼠移植实体瘤模型。方法将1例人卵巢癌组织移植裸鼠,建立人卵巢癌裸鼠原代移植实体瘤模型的基础上,再将实体瘤皮下移植、实体瘤原位移植、实体瘤细胞移植裸鼠。观察裸鼠实体瘤生长和转移情况,称量其体重、子宫卵巢重、瘤重及瘤的大小,并作病理、电镜、染色体检查。结果成功地建立人卵巢癌裸鼠移植实体瘤模型,并已传至第18代,传代移植成功率100％,组织学和超微结构形态均证明该实体瘤保留了原人卵巢癌特征,有人卵巢癌染色体特征,并出现肝、脾转移。结论本研究建立人卵巢癌裸鼠移植实体瘤模型与人相似,通过18代传代和实验观察方法稳定可靠。为人卵巢癌的研究提供了理想的动物模型。     

人乳腺癌MDA-MB-231细胞裸鼠移植瘤模型的建立及生物学特性研究

目的-建立人乳腺癌MDA-MB-231细胞株裸小鼠模型,研究其生物学特性,观察MDA-MB-231乳腺癌细胞在移植前后的形态学变化。方法-将人乳腺癌细胞MDA-MB-231接种于裸鼠腋窝处皮下,每3天测量肿瘤大小,第30天处死小鼠。肿瘤组织及相关脏器送病理切片。皮下肿瘤组织细胞及细胞株培养HE染色。结果-肿瘤生长较快,成功率为72%,病理检查符合人乳腺癌细胞特征。肿瘤组织细胞及培养细胞形态学未见显著差异。结论-人乳腺癌细胞株MDA-MB-231裸小鼠模型建立方法较简便,细胞形态无明显差异,且保持了人乳腺癌的生物学特性。     

新构建的含DF3启动子和白喉毒素A片段的重组表达载体对人乳腺癌细胞的特异性杀伤作用

目的研究新构建的含人乳腺癌DF3启动子和白喉毒素A片段的重组表达载体PGL3-DF3-DTA对人乳腺癌细胞的影响。方法构建重组表达载体PGL3-DF3-DTA,并用其转染DF3阳性和阴性的乳腺癌细胞MCF-7和MDA-MB-231。通过MTT法测定PGL3-DF3-DTA在体外对乳腺癌细胞生长的影响,建立裸鼠动物模型观察PGL3-DF3-DTA在体内对乳腺癌细胞的杀伤效应。结果成功构建出重组表达载体PGL3-DF3-DTA并建立了人乳腺癌裸鼠动物模型,经重组表达载体PGL3-DF3-DTA作用后的DF3阳性人乳腺癌细胞出现明显的凋亡现象,Ki-67、bcl-2基因表达水平降低,bax基因表达水平升高。结论重组表达载体PGL3-DF3-DTA能对DF3阳性的乳腺癌细胞产生特异性杀伤作用。     

Establishment and characterization of a human colonic cancer cell line expressing carbohydrate antigen 19-9 and carcinoembryonic antigen]

Colonic cancer cell strain KE43 was established from a human colonic cancer diagnosed histologically as a predominantly well differentiated adenocarcinoma with minute foci of poorly differentiated adenocarcinoma. The well differentiated adenocarcinoma cell line was identified as the major morphological picture in xenografts of KE43 and 58 in nude mice, but this changed to poorly differentiated adenocarcinoma in passage 105. Doubling time of this cancer cell line was 22.5 hours in passage 105. The modal numbers of chromosomes were 41 and 76. Cancer cells could be heterotransplanted in 100% of the nude mice. The tumor cells produced and secreted CA19-9, CEA and Laminin into the spent medium. This cell line appears to provide a useful system for studying colonic cancer in vivo and in vitro.     

A rat cell line derived from DMBA-induced mammary carcinoma

A new cell line, designated UHKBR-01, was successfully established from a 7,12-dimethylbenz[a]anthracene (DMBA)-induced rat mammary tumour. DMBA was administered orally at a dose of 4 mg/ml per rat on the first day of the experiment and thereafter at weekly intervals of same dosage, until the rats have reached a weight of around 150-200 g. The tumours grew rapidly after the injection, and were transplanted into nude mice one the harvest size (2.5 x 2 x 1 mm(3)) was reached, it was transplanted onto nude mice. We have developed a cell line from a portion of the DMBA-induced carcinoma of the nude mice. The UHKBR-01 cell exhibited a slow increase in growth rate during the time of culture and was highly tumourigenic in nude mice. The cells have been grown in culture for over 40 passages. Characterization of the cell line was performed. This included morphology by light and transmission electron microscopy, karyotype, growth rate, tumour antigen expression and xenograft implantation into nude mice. These cells exhibit ultrastructural and immunohistochemical features of epithelial cells of mammary origin. The above analyses also demonstrated that UHKBR-01 cells were oestrogen- and progesterone-receptor positive, in likeness to other established breast cancer cell lines such as MDA-MB-231 and MCF-7. The cell line grows as monolayers of oval-shaped cells with large folded nuclei accompanied by a rich supply of mitochondria. This report describes the first in vitro cell line from transplantable DMBA-induced mammary carcinoma of nude mice, which presents unique characteristics that may prove to be a good experimental model for investigating breast cancer biology.     

利用裸鼠建立人泌尿生殖系统肿瘤细胞系

目的建立人泌尿系肿瘤无限细胞系,为泌尿系肿瘤研究提供实验模型.方法无菌取下肿瘤标本后,将标本剪成大小约1.0mm3的组织块,在裸鼠右后肢皮下包埋,当皮下肿瘤块发生明显增殖并长到一定程度后,再行裸鼠体内传代两次,最后取下组织块进行原代培养.培养细胞传代超过20代后按建系标准[2]进行检测.结果共取40例标本,裸鼠体内传代F1代成功6例,F3代成功3例,该3例标本行原代培养后建成3个无限细胞系人肾透明细胞癌RCC-9863,人膀胱癌BC-6,人前列腺癌PC-98106,全部细胞传代1年以上,生长稳定,传代周期固定,其形态结构,分化程度与原发瘤保持一致,染色体形态仍为人类核型.结论裸鼠肿瘤皮下种植法是泌尿系肿瘤建系的一个较好方法.     

An ascites form of human mammary carcinoma transplantable in nude mice

Mammary carcinoma cells from pleural effusion of a patient were inoculated into the peritoneal cavity of nude mice, and a large amount of ascites was produced about 120 days later. From the ascites, serial passages in the same form were successful in nude mice by intraperitoneal injection of 10(7) or more cells. The ascites cells retained the morphology almost similar to that of the patient tumor cells, whereas specific estrogen-binding proteins in the cytoplasm disappeared after growing in male nude mice. The results were compared with those of other established human cancer cell lines in nude mice.     

Regulation of zinc transporters by dietary zinc supplement in breast cancer

Zinc is essential for cell growth and is a co-factor for more than 300 enzymes, representing over 50 different enzyme classes. Two gene families have been identified involved in zinc homeostasis. ZnT transporters reduce intracellular zinc while ZIP transporters increase intracellular zinc. Previous studies have shown that zinc concentration in breast cancer tissues is higher than that in normal breast tissues. However, the mechanisms involved and the relations to zinc transporters are still unknown. A series of zinc transporters are characterized in this article and several of that are emphasized in view of their unique tissue-specific expressions. Established human breast cancer in a nude mice model is used. With a dietary zinc supplement treatment, ZnT-1 mRNA expression in established human breast cancer is raised by 24%, and is nearly 2 times of that in basal diet. ZIP1, ZIP2 and LIV-1 mRNA are the same between two treatment groups. Moreover, no significant changes of these zinc transporters expressions are found between differential breast cancer cell lines in the nude mice model. This is the first report, which detects the zinc transporters expressions in established human breast cancer in nude mice model. These results lead to the constitutive expression and response to zinc in different tissues. In addition to that, ZnT-1 seems to have played an important role in zinc homeostasis, even in breast cancer.     

人OC-3-VGH卵巢癌细胞裸小鼠肿瘤模型的建立

目的建立人OC-3-VGH细胞株卵巢癌裸小鼠模型并观察该肿瘤生物学生长特性。方法OC-3-VGH细胞株复苏后加入10 mL RPMI-1640培养液,放入培养箱,传2～3代后,取细胞悬液,均以4×106个细胞,每只0.2 mL分别接种至BALB/c雌性裸小鼠皮下,2月后处死取材,观察肿瘤生长特性和转移情况。结果皮下接种一周后,裸鼠长出肿瘤,并随时间而增大,体积呈指数增长,第42天始,明显增大（P〈0.05）。组织学检查发现裸鼠皮下肿瘤细胞均细胞体积较大,细胞核大而染色深,核分裂相较多、异型性明显,接种2个月时,未发生其他组织转移。结论建立了新的卵巢癌动物模型,并初步研究了其生物学特性,为卵巢癌治疗方法的研究拓宽了道路。     

Establishment and characterization of a CA19-9 producing human gastric cancer cell line, STKM-1

A human gastric cancer cell line, STKM-1, was established from the malignant cells in pleural effusion of a 41-year-old female patient. The primary gastric cancer revealed histologically a poorly differentiated adenocarcinoma. The cells have been cultured with RPMI-1640 medium supplemented with 10% fetal bovine serum and grew as monolayers following a doubling time of 31.4 hour at passage 30. The mode of chromosome number was 52. The STKM-1 cell was tumorigenic in nude mice. The STKM-1 cell cultured in vitro secreted CA19-9, into the medium as a tumor marker. Cells in tumors grown in nude mice were immunohistochemically recognized positively by anti-CA19-9 antibody. The STKM-1 will provide a useful information to clarify the mechanism of CA19-9 secretion.     

用于活体成像的小鼠肺癌移植瘤模型的建立

本研究旨在建立可用于活体成像的小鼠肺癌移植瘤模型。利用脂质体将荧光素酶表达载体pGL4.17(luc2/neo)转染至人非小细胞肺癌细胞株A549,经G418筛选获得稳定表达荧光素酶的细胞克隆。根据体外生物发光情况及细胞的生长特性,从中挑选合适克隆,进行裸鼠皮下接种,SCID鼠尾静脉接种,建立肺癌移植瘤模型。利用活体成像系统监测肿瘤的生长转移情况,并用切片HE染色进一步验证小鼠模型移植瘤的原位成瘤和转移能力。实验结果表明:本研究成功地构建了可用于活体成像的小鼠肺癌移植瘤模型,模型稳定可靠、直观、灵敏,为肿瘤生长转移机制的研究及抗肿瘤药物的研发提供了重要工具。     

Adeno-associated virus sensitizes HeLa cell tumors to gamma rays.

Infection with the helper virus-dependent human parvovirus adeno-associated virus (AAV) is known to interfere with cellular transformation in vitro and oncogenesis in vivo. Here we report on sensitization to gamma irradiation by AAV infection of cells in culture and of tumors established from HeLa cells grafted into immunodeficient (nude) mice: infection of HeLa cells with AAV type 2 enhanced cell killing and reduced plating efficiency after irradiation compared with uninfected cells. Similarly, HeLa cell tumors in nude mice displayed a reduced growth rate and were more sensitive to gamma irradiation when the animals were infected with AAV type 2 prior to or after tumor cell inoculation. Since no pathogenicity is known for AAV, the ability of this virus to render radiotherapy of human tumor cells more efficient may up open novel approaches in cancer treatment.     

Polyamine depletion by ODC-AdoMetDC antisense adenovirus impairs human colorectal cancer growth and invasion in vitro and in vivo

BACKGROUND: Polyamine biosynthesis is controlled primarily by ornithine decarboxylase (ODC) and S-adenosylmethionine decarboxylase (AdoMetDC). Polyamine concentrations are elevated in colorectal cancer. Depletion of polyamine content in colorectal cancer by chemotherapy is related to tumor regression and impaired tumorigenicity. The current study evaluates the therapeutic effects of antisense ODC and AdoMetDC sequences on colorectal cancer in vitro and in vivo. METHODS: Antisense ODC and AdoMetDC sequences were cloned into an adenoviral vector (Ad-ODC-AdoMetDCas). The human colon cancer cell lines, HT-29 and Caco-2, were infected with Ad-ODC-AdoMetDCas as well as with control vector. Viable cell counting, determination of polyamine concentrations, cell cycle analysis, and Matrigel invasion assays were performed in order to assess properties of tumor growth and invasiveness. Furthermore, the antitumor effects of Ad-ODC-AdoMetDCas were also evaluated in vivo in a nude mouse tumor model. RESULTS: Our study demonstrated that adenovirus-mediated ODC and AdoMetDC antisense expression inhibits tumor cell growth through a blockade of the polyamine synthesis pathway. This inhibitory effect cannot be reversed by the administration of putrescine. Tumor cells were arrested at the G1 phase of the cell cycle after gene transfer and had reduced invasiveness. The adenovirus also induced tumor regression in established tumors in nude mice. CONCLUSIONS: Our study suggests that Ad-ODC-AdoMetDCas has antitumor activity and therapeutic potential for the treatment of colorectal cancer.     

Feasibility and limits of an orthotopic human colon cancer model in nude mice

We sought to develop an accurate colorectal cancer model in nude mice with stable local growth, tumor cell dissemination, and reproducible metastatic capacity. To this end, we orthotopically transplanted histologically intact human colorectal cancer tissue from 10 human patients into nude mice. After successful local tumor growth, tumor tissues were retransplanted as many as 9 times in serial passage. All specimens were transplanted using microsurgical techniques. Histologic, immunohistochemical, and polymerase chain reaction techniques were used to determine tumor growth rates and kinetics, development of regional lymph node and distant hepatic metastases, and the induction of minimal residual disease (MRD). Stable local tumor growth rates with variable growth kinetics were detected in 73.4% of all mice. The lymph node and hepatic metastasis rates were low, at 18.4% and 4.9%, respectively. MRD, as reflected by CK20 positivity of the bone marrow in animals with lymph node and hepatic metastases, was present in 22.2%. The orthotopic colorectal cancer model described here is feasible for the induction of reproducible local tumor growth but is limited by variable growth kinetics and the low rate of lymph node and hepatic metastases. Cytokeratin-positive cells indicative of MRD could be detected in the bone marrow of approximately 25% of the nude mice with metastases. The observed induction of MRD after orthotopic implantation of intact human colon cancer in animals with lymph node and hepatic metastases might be improved if established colon cancer cell lines were used.     

胰腺癌原位种植瘤中VEGF-C的器官差异表达和作用

目的 研究胰腺癌裸鼠原位种植瘤自发性淋巴结转移模型中VEGF-C表达的器官差异,以及VEGF-C反义寡核苷酸对不同生长部位胰腺癌细胞生长、凋亡能力的影响。方法 建立人胰腺癌细胞株PANC-1裸鼠原位种植瘤自发性淋巴结转移模型,分离、原代培养原发灶和自发性淋巴结转移灶中的胰腺癌细胞,并应用荧光定量PCR、MTT、流式细胞术检测VEGF-C反义寡核苷酸转染对原发胰腺癌细胞和淋巴结转移的胰腺癌细胞各自生长特性、凋亡能力的影响。结果 淋巴结转移胰腺癌细胞VEGF-C的mRNA表达水平显著高于原发灶胰腺癌细胞（P〈0.05）。VEGF-C反义核苷酸抑制胰腺癌细胞VEGF-C的表达后,淋巴结转移灶中胰腺癌细胞生长抑制率、凋亡率均显著提高（P〈0.01）,而原发灶中胰腺癌细胞无明显影响（P〉0.05）。结论 VEGF-C反义寡核苷酸能显著抑制淋巴结转移灶中胰腺癌细胞生长、促进凋亡,但对原发灶胰腺癌细胞无影响;VEGF-C基因的表达和作用存在器官差异性。     

Targeting breast and prostate cancers through their hormone receptors

A targeted treatment that effectively destroys human breast, prostate, ovarian, and testicular cancer cells that express luteinizing hormone/chorionic gonadotropin (LH/CG) receptors has been developed. The treatment consists of a conjugate of a membrane-disrupting lytic peptide (Hecate, Phor14, or Phor21) and a 15-amino acid segment of the beta chain of CG. Because these conjugates act primarily by destroying cell membranes, their effects are independent of cell proliferation. The conjugates are relatively small molecules, are rapidly metabolized, and are not antigenic. In a series of independent experiments conducted in three different laboratories, the validity of the concept has been established, and it has been shown that the LH/CG receptor capacity of the cancer cells is directly related to the sensitivity of the lytic peptide conjugates. Sensitivity to the drugs can be increased by pretreating prostate or breast cancer cells with FSH or estradiol to up-regulate LH/CG receptors. A series of 23 in vivo experiments involving a total of 1630 nude mice bearing xenografts of human prostate or breast cancer cells showed convincingly that all three lytic peptide-betaCG compounds were highly effective in destroying tumors and reducing tumor burden. Hecate-betaCG was less effective in mice bearing ovarian epithelial cancer cell xenografts, but was highly effective in treating granulosa cell tumors in transgenic mice. In addition, Hecate-betaCG and Phor14-betaCG were highly effective in targeting and destroying prostate and breast cancer cell metastases in the presence or absence of the primary tumors. Although effective in vitro, neither Hecate nor Phor14 alone were effective in reducing primary tumor volume or burden in nude mice bearing prostate or breast cancer xenografts.