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.     

Characterization of a new human cell line derived from a xenografted embryonal carcinoma

Summary A human cell line has been established from a transplantable xenografted human testicular tumor, which, both in the original tumor and in the xenograft, exhibited the histological characteristics of an undifferentiated malignant teratoma (embryonal cell carcinoma). The cells in culture were undifferentiated by biochemical, morphological, and ultrastructural criteria, growing as small islands of cells that tended to form aggregates at high density. The cells showed some variation in chromosome number with 30 to 40% of the cells having a normal human karyotype. The cells expressed high levels of alkaline phosphatase, which by heat inactivation and inhibition studies was 40 to 50% placental type alkaline phosphatase. None of the cultures produced human chorionic gonadotrophin, alphafetoprotein, carcinoembryonic antigen, or fibronectin, although at high cell densities plasminogen activator could be detected at low levels. Cell surface studies showed that the cells shared antigens with the murine embryonal carcinoma cell line F9, expressedβ ₂-microglobulin at very low and variable levels, and bound the lectin peanut agglutinin. These studies suggest that this cell line has some of the characteristics described for murine embryonal carcinoma cell lines.     

Calcium-dependent morphogenesis in a cell line derived from human uterine cervical carcinoma

Cells of the cervical carcinoma line C-4I replicate in culture media with 0.8–1.8 mM calcium and concommitantly form multilayered colonies with tonofilament bundles, desmosomes and an intercellular organization resembling stratified squamous epithelium. In medium with 0.02 mM calcium, the cells grow as monolayers of loosely cohesive cells, without desmosomes and with an altered tonofilament distribution. Upon return to 1.8 mM calcium, the morphological characteristics of stratified squamous epithelium are restored. The proliferative and morphogenetic response of this cervical cell line to extracellular calcium levels closely resembles the response of epidermal carcinoma cells and suggests that calcium may play a similar role in the regulation of differentiation in these two stratified squamous epithelia.     

Supernatant derived from a human hepatocellular carcinoma cell line (PLC/PRF/5) activates a population of T-suppressor cells

Harvest fluid derived from a primary hepatocellular carcinoma cell line (PLC/PRF/5) inhibited the incorporation of 3H-thymidine into PHA-activated human lymphocytes. A similar effect was observed when lymphocytes were pre-incubated with the tumour supernatant and washed prior to mitogen activation. Not only did the tumour supernatant inhibit 3H-thymidine incorporation by mitogen-activated lymphocytes, but it also inhibited production of the lymphokine leucocyte inhibitory factor (LIF). In experiments designed to establish whether a component of the tumour harvest fluid was activating a population of suppressor cells, normal mononuclear (MN) cells were treated with the PLC/PRF/5 or embryonic fibroblast supernatant for 48 h, after which they were washed and added to normal mitogen-activated lymphocyte cultures. Only cells pretreated with the PLC/PRF/5 supernatant suppressed mitogenesis. The cell responsible for the suppressor effect was a T cell, which after a further 24 h in culture liberated a suppressor factor responsible for inhibiting lymphocyte function. Although the nature of the factor/s in the PLC/PRF/5 supernatant responsible for activation of the T-suppressor cell population is unknown, it is suggested that this mechanism may be important in protecting the tumour from the immune response.     

Comparative characterization of a human large cell lung carcinoma cell line and the xenograft derived cell line.

The HK-1 cell line established from a human large cell lung carcinoma shows a high transformed phenotype and undifferentiated characteristics. This cell line grows as an adherent monolayer in fetal calf serum-supplemented medium, shows a high proliferation index, is able to grow in semi-solid agar and is tumorigenic in athymic nude rats. The cell line HK-2 derived from the HK-1 xenograft in athymic nude rats shows basically the same features found in the original HK-1 cell line, which include aneuploid nuclear DNA content, abnormal chromosomal number. rearranged marker chromosomes and abnormally localised nucleolar organizer regions. Cytokeratin and vimentin intermediary-sized filaments were found in both cell lines as well as in the original and induced tumour, while neither oncofetal antigens (alphafeto-protein, carcinoembryonic antigen, chorionic gonatropin and human placental lactogen) nor neural differentiation markers (neurofilament and neural specific enolase) were expressed. Analysis of sulphated glycosaminoglycans in the cell cultures and in the nude rat induced tumour showed high expression of chondroitin sulphate.     

Establishment of a cell line derived from a canine prostate carcinoma with a highly rearranged karyotype

Akin to the situation in humans, dogs are frequently affected by tumors of the prostate. The malignancies share many similarities between both species, for example, median age at the onset of the disease and metastatic behavior. In human prostatic tumor samples, investigations of prepared metaphase spreads showed a variety of chromosomal aberrations, with trisomies of chromosomes 7, 8, and 17 as the leading cytogenetic abnormalities. In this article we present one case of a canine adenocarcinoma of the prostate, including clinical examination and establishment of a cell line from a tumor sample obtained from the affected 10-year-old male Briard. Searching for similarities between both species in respect to chromosomal changes within the tumor samples, we investigated prepared metaphases of the canine cell line cytogenetically. These investigations presented a highly rearranged karyotype showing a large biarmed marker consisting of material from chromosomes 1 and 2 in addition to centromeric fusions between dog chromosomes 1 and 5 that both could be identified in every metaphase investigated, while centric fusions of chromosomes 4 and 5 occurred in up to 50% of the metaphases. The cell line grew very well and showed evidence of being spontaneously immortalized when it crossed the 20th passage.     

New cell line derived from a human chondrosarcoma

Summary The morphological, chromosomal, and biological characteristics of a cell line derived from a human chondrosarcoma have been described. At the time of this report, the cell line has undergone 102 passages and continues to exhibit an epithelioid appearance, a modal number of 69 chromosomes, and has retained malignant properties. This cell line is easily cultivated in vitro or in vivo and may prove useful in human cancer research.     

Establishment and characterization of a new human hepatocellular carcinoma cell line

Summary A human hepatocellular carcinoma cell line (FOCUS—Friendship of China and United States) was derived from a patient with primary hepatocellular carcinoma. This cell line has been in continuous culture over an 18-mo period. The morphological and ultrastructural features of FOCUS are consistent with its neoplastic hepatocellular orgin. FOCUS cells contain aspartate aminotransferase and glucose-6-phosphatase activity. In addition, α₁-antitrypsin, fibrinogen, alpha fetoprotein, and carcinoembryonic antigens were detectble in the cytoplasm of the cultured cells by immunochemical staining techniques. The karyotype of the FOCUS cell is human in origin and it contains human DNA sequences as detected by molecular hybridization analysis. The FOCUS cells do not show evidence of density-dependent inhibition of growth under confluent conditions. Repeated growth curves over an 18-mo period were identical, revealing a doubling time of 42 to 48 h. The malignant potential of FOCUS cells was further demonstrated by their ability to lead to gross tumor formation after subcutaneous infection into nude mice. From one of the solid tumors grown in nude mice, recultured cell lines have been established and found to have properties identical to the original FOCUS cell line. This FOCUS cell line represents an additional model for further investigation of tumor specific antigens and the relationship between hepatitis B virus (HBV) and hepatocellular carcinoma. Preliminary molecular characterization has indicated the existence of integrated HBV sequences within the FOCUS genome.     

Establishment and characterization of a new human hepatocellular carcinoma cell line

A human hepatocellular carcinoma cell line (FOCUS--Friendship of China and United States) was derived from a patient with primary hepatocellular carcinoma. This cell line has been in continuous culture over an 18-mo period. The morphological and ultrastructural features of FOCUS are consistent with its neoplastic hepatocellular origin. FOCUS cells contain aspartate aminotransferase and glucose-6-phosphatase activity. In addition, alpha 1-antitrypsin, fibrinogen, alpha fetoprotein, and carcinoembryonic antigens were detectable in the cytoplasm of the cultured cells by immunochemical staining techniques. The karyotype of the FOCUS cell is human in origin and its contains human DNA sequences as detected by molecular hybridization analysis. The FOCUS cells do not show evidence of density-dependent inhibition of growth under confluent conditions. Repeated growth curves over an 18-mo period were identical, revealing a doubling time of 42 to 48 h. The malignant potential of FOCUS cells was further demonstrated by their ability to lead to gross tumor formation after subcutaneous injection into nude mice. From one of the solid tumors grown in nude mice, recultured cell lines have been established and found to have properties identical to the original FOCUS cell line. This FOCUS cell line represents an additional model for further investigation of tumor specific antigens and the relationship between hepatitis B virus (HBV) and hepatocellular carcinoma. Preliminary molecular characterization has indicated the existence of integrated HBV sequences within the FOCUS genome.     

Establishment and characterization of a cell line derived from a human serous surface papillary carcinoma of the ovary

A novel serous surface papillary carcinoma of the ovary (SSPC) cell line, HYKSSPC, was established successfully. Carcinoma cells were obtained from ascitic fluid of a 60-year-old Japanese woman. The population doubling time was 51.4 h. A phase contrast micrograph showed a pavement stone-like arrangement without contact inhibition. The chromosome number showed a wide distribution of aneuploidy, and the mode was in 46-47. An immunocytochemical study showed that CA125, BerER4 and cytokeratin were positive and that CEA, calretinin and thrombomodulin were negative. This cell line preserved some characters of the adenocarcinoma while growing in vitro. A chemosensitivity test revealed that HYKSSPC cells were sensitive to CDDP (cis-platinum), 5-fluorouracil, mitomycin C, paclitaxel and irinotecan. To our knowledge, HYKSSPC is the first established cell line derived from SSPC, and it may offer some useful information for investigating this disease.     

Biological characterization of a cell line derived from the pig oviduct

Summary A cell line has been derived from the inner lining of the pig oviduct. During 33 months in continous culture, the cells have been subcutured 145 times. Early passage cells had an epithelial-like morphology with clear and abundant cytoplasm. A spontaneous morphological alteration occurred at the 50th subculture by emergence of fibroblast-like cells. Evolution towards establishment of a cell line was suggested by the spontaneous morphological alteration, increase of the maximum population density without a significant change in the diploid state (92nd subculture), increase, in size and number of nucleoli, and increase in the nuclear-cytoplasmic ratio and of cytoplasmic basophilia. Evidence of pig origins is subtantiated by the karyotype (38, XX), the persistence of Barr bodies indicating female origin (140th subculture) and the persistence of species specificity by fluorescence (141th subculture). Preliminary studies indicate that nine viruses will replicate in both epithelial-like and fibroblast-like cells.     

Establishment and characterization of a human hepatocellular carcinoma cell line JHH-4

A human hepatocellular carcinoma cell line, JHH-4 was established from resected liver tumor. Morphological diagnosis of the original tumor was hepatocellular carcinoma, Edmondson type III. This cell line was composed of polygonal shaped cells. Subcellular organelle were observed in cytoplasm. Furthermore, bile canaliculi adhering junction was also remained at the cell surface. The growth rate of JHH-4 cell is slow, peaks of the chromosome number was 75 and 79, and plating efficiency was 3.0%. JHH-4 cell is transplantable to nude mouse. Furthermore, this cell line functionally synthesized and secreted human albumin, AFP and other proteins in vitro.     

Establishment and characterization of a clear cell carcinoma cell line,designated NOCC,derived from human ovary

A cell line, designated NOCC, was established from the ascites of a patient with clear cell adenocarcinoma of the ovary. The cell line has been grown without interruption and continuously propagated by serial passaging (more than 76 times) over 7 years. The cells are spherical to polygonal-shaped, display neoplastic, and pleomorphic features, and grow in a jigsaw puzzle-like pattern while forming monolayers without contact inhibition. The cells proliferate rapidly, but are easily floated as a cell sheet. The population doubling time is about 29 h. The number of chromosomes ranges from 60 to 83. The modal number of chromosomes is 70–74 at the 30th passage. NOCC cells secreted 750.5 ng/ml of VEGF over 3 days of culture. Hypoxia inducible factor-1α (HIF-1α) is a primary regulator of VEGF under hypoxic conditions. NOCC cells were not sensitive to the anticancer drugs BEV, DOX, GEM, ETP, CDDP, or TXT. The graft of NOCC cells to a scid mouse displayed similar histological aspects to the original tumor. Both the NOCC cells and the graft of the NOCC cells gave a positive PAS reaction.     

Establishment and characterization of a cell line derived from a spontaneous murinelung carcinoma (M109)

Summary The Madison lung (M109) tumor cell line, initiated from a “spontaneous”, anaplastic murine lung carcinoma, has been propagated continuously in vitro for more than 300 cell generations. Cytogenetic analysis revealed a mouse karyotype with a mode of 78 chromosomes (2n=40). Three distinct marker chromosomes were identified by trypsin-giemsa banding. The cells piled up in culture and had a short generation time and high plating efficiency. Electron microscopy revealed highly undifferentiated cells with little rough endoplasmic reticulum, an abundance of free polysomes, the presence of few and often oddshaped mitochondria, lipid bodies and phagocytic vacuoles. Virus particles of the C-type were found frequently. The subcutaneous transplantation of M109 cultured cells at a relatively low cell inoculum produced highly metastatic tumors in syngeneic BALB/c mice.     

A new human hepatocellular carcinoma cell line (KYN-1) with a transformation to adenocarcinoma

Summary A human hepatocellular carcinoma (HCC) cell line (KYN-1) has been established from a resected HCC of a 58-yr-old Japanese, male patient with HCC. Original resected HCC was moderately differentiated and proliferated in a solid pattern with vague trabecular structure in part. This cell line has been maintained for 10 mo. through 50 passages. Morphological features of KYN-1 cells demonstrated one or more large, round-to-oval nuceli with prominent nucleoli and eosinophilic polygonal-to-spindle abundant cytoplasm. In addition, some of these cells contained mucicarmin-positive materials in the cytoplasm. The cells exhibited a typical epithelial feature with pavementlike cell arrangement, and lacked contact inhibition. The doubling times of the cells grown in a serum-containing and a serum-fre, medium were about 31 h and 10 to 11 d, respectively. Functonally, KYN-1 cells produced albumin, α-fetoprotein (AFP), carcinoembryonic antigen (CEA), ferritin, β₂-microglobulin (BMG), and α₁-anti-trypsin (AAT). Positive reactions for albumin, AFP, CEA, and ferritin were identified in the cells by immunohistochemical techniques. Chromosome study revealed the chromosome number in a range from 61 to 74 without mode. The tumorigenicity of KYN-1 cells was identified by the tumor formation after subcutaneous inoculation of the cells into nude mice. The developed tumor showed compact growth of the tumor cells with gland formations containing mucicarmin-positive materials. Features of adenocarcinoma were identified by electron microscopy. The tumor cells were also identified to contain albumin, AFP, CEA, ferritin, and AAT by immunohistochemical techniques. AFP, CEA, and BMG were detected in the sera of nude mice. Thus, KYN-1 cells represented the morphologic features of adenocarcinoma, retaining some characteristics of original HCC. These findings suggest that KYN-1 is a new human HCC cell line with transformation to adenocarcinoma, which will provide useful information to clarify the histogenesis of combined hepatocellular and cholangiocellular carcinoma. This study was supported in part by the Sarah Cousins Fund.