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PubMed1992

An amphicrine pancreatic cell line: AR42J cells combine exocrine and neuroendocrine properties.

The permanent cell culture line AR42J, derived from a rat pancreatic acinar carcinoma, is widely used for functional studies of exocrine pancreatic acinar cells. We now present evidence that these cells are amphicrine in that they contain zymogen granules as well as small (40-80 nm) neuroendocrine (NE) vesicles and typical neurotransmitters. Using the small NE vesicle-specific markers synaptophysin and "protein S.V.2", including synaptophysin cDNA probes, we have found that AR42J cells synthesize these proteins and contain vesicles harboring these proteins with biophysical properties similar to those of small NE vesicles. NE properties of these cells are further indicated by the presence of considerable amounts of stored amino acids (gamma-aminobutyric acid (GABA), glycine, glutamate) and by the presence of the GABA-synthesizing enzyme, glutamic acid decarboxylase. Finally, intermediate filament (IF) protein typing showed only cytokeratins 8 and 18, indicating that AR42J cells possess an IF protein complement indistinguishable from that of acinar and islet cells. Our results document the unusual case of a permanent cell line with combined exocrine and neuroendocrine properties that may be indicative of a derivation from a cell with multipotential character.

Animals↗

PubMed2002

Antineoplastic effect of new boron compounds against leukemic cell lines and cells from leukemic patients.

Three new boron compounds, dihydroxy (oxybiguanido) boron (iii) hydrochloride monohydrate (HB), guanidine biboric acid adduct (GB) and hydroxosalicyl hydroxomato boron (iii) (SHB) were studied to observe their antineoplastic effect, if any. Leukemic cells isolated from acute lymphatic leukaemia (ALL) patients and chronic myeloid leukaemia patients (CML) and myeloid leukemia cell lines (HL 60 and U-937) showed cell growth inhibition after treatment with the boron compounds. MTT assay showed that the growth of metabolically active cells was inhibited by treatment with these drugs. The molecular mechanism by which SHB induced apoptosis in immature blast cells was also investigated by ladder formation in gel electrophoresis.

Acute Disease↗

PubMed1991

Ganglioside GM3 can induce megakaryocytoid differentiation of human leukemia cell line K562 cells.

The role of acidic glycosphingolipids in cell growth and differentiation was investigated using the multipotent leukemia cell line K562. When GM3 was added to cell culture media, the growth of K562 cells was remarkably inhibited and the cells were shown to have megakaryocytoid morphology. Ultrastructural study demonstrated that K562 cells treated with GM3 had platelet peroxidase-positive structures, which were considered to be the specific marker of megakaryocyte. Furthermore, AP-3 directed against an epitope present on membrane glycoprotein IIIa reacted with the GM3-treated cells. Free N-acetylneuraminic acid, GM1, GM2, GD1a, and a mixture of bovine brain gangliosides containing GD1a and GT1b did not affect growth of K562 cells or show morphological changes. According to chemical analyses, GM3 content increased in megakaryocytoid differentiation induced by tetradecanoylphorbol-13-acetate, whereas GM3 decreased in erythroid differentiation induced by hemin. Enzymatic analysis showed that the GM3 increase during megakaryocytoid differentiation was a result of the sialyltransferase activation. These results indicated that exogenous GM3 induced differentiation of K562 cells into a "GM3-rich" lineage, i.e., mainly megakaryocytoid lineage, and that GM3 accumulation in the GM3-rich lineage was the result of the activation of GM3 synthase. These findings strongly suggested that GM3 ganglioside, a minor membrane component, has a crucial role in not only the differentiation induction but also the determination of the differentiation direction in pluripotent K562 cells.

Cell Differentiation↗

PubMed1986

Continuous production of erythropoietin by an established human renal carcinoma cell line: development of the cell line.

Establishment of a stable, transformed human renal carcinoma cell line that produces erythropoietin in vitro and has maintained this function continuously since 1981 and for greater than 150 passages in monolayer culture was accomplished by transplantation of human renal clear cell carcinoma tissue from a patient with erythrocytosis into an immunosuppressed athymic mouse. In addition to its immunocrossreactivity with native human urinary erythropoietin, the tumor erythropoietin demonstrates biological activity in the in vitro mouse erythroid colony-forming unit assay and in tumor-bearing nude mice. The cloned renal carcinoma cell line has an abnormal human karyotype and has ultrastructural features characteristic of human renal clear cell carcinoma. This cell line provides a reproducible model system for the production of an erythropoietin-like material and for the study of its synthesis and secretion.

Animals↗

PubMed1986

Transfection of DNA from a chloroethylnitrosourea-resistant tumor cell line (MER+) to a sensitive tumor cell line (MER-) results in a tumor cell line resistant to MNNG and CNU that has increased O-6-methylguanine-DNA methyltransferase levels and reduced levels of DNA interstrand crosslinking.

A subclone of a human glioma tumor cell strain which was deficient at O-6-alkylguanine repair was transfected with DNA from an O-6-alkylguanine DNA alkyltransferase-positive human colon tumor cell line (HT29). The transfected subclone, which was selected by its resistance to N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), acquired increased resistance to chloroethylnitrosourea (CNU), contained O-6-methylguanine-DNA methyltransferase activity, and failed to accumulate interstrand crosslinks in its DNA upon treatment with CNU. The subclone morphologically resembled its CNU-sensitive parental glioma cell strain. The close correlation between the increased cellular resistance, increased O-6-alkylguanine repair activity and the absence of crosslinking suggests that the formation of DNA crosslinks is one important mechanism of cytotoxicity produced by CNU, and that repair of DNA lesions by O-6-alkylguanine DNA alkyltransferase may partially prevent CNU-induced cytotoxicity.

Cell Line↗

PubMed2000

[Expression of Fas ligand in urogenital malignant cell lines and renal cell carcinoma].

OBJECTIVE: To explore the expression of Fas ligand in urogenital malignant cell lines and renal cell carcinoma. METHODS: With immunocytochemistry and reverse transcription-polymerase chain reaction (RT-PCR), we detected the expression of Fas ligand (FasL) in 6 urogenital malignant cell lines, bladder carcinoma cell lines (T(24), EJ, BIU-87), renal cell carcinoma cell lines (GRC-1, RCC-949), prostatic carcinoma cell line (PC-3M), and 10 cases of renal cell carcinoma. RESULTS: The expression of FasL was detected in 3 (BIU-87, RCC-949, GRC-1) of the 6 urogenital malignant cell lines with immunocytochemistry, and 4 (BIU-87, RCC-949, GRC-1, PC-3M) of 6 urogenital malignant cell lines with RT-PCR method. No expression of FasL was detected in bladder carcinoma cell lines T(24) and EJ. Eight of 10 renal cell carcinoma tissues positively expressed FasL mRNA. CONCLUSIONS: FasL can express in urogenital malignant cell lines and renal cell carcinoma.

Carcinoma, Renal Cell↗

PubMed2000

Extracellular matrix effects on a neuroblastoma cell line.

Cells of various lines assume similar shapes when grown attached to substrates like coverslips. In contrast, cells cultured in a collagen and/or laminin matrix often assume a relatively normal morphology in comparison with their in situ counterparts. During investigations of neuroblastoma SH-SY5Y cells, an attempt was made to identify culture conditions which would cause the cells to assume a more regular shape. SH-SY5Y cells cultured on bare coverslips, on coverslips coated with rat-tail collagen, and in approximately 1 mm thick gels containing extracellular matrix components were compared. Striking differences were apparent when comparing the gel-cultured cells with cells cultured on coverslips. Cells grown in the gel formed ganglia-like clusters which generated bundles of neurites which targeted other 'ganglia'. The same cells grown on coverslips, whether or not they were collagen-coated, appeared unaware of the presence of other cells, and did not cluster, nor did they generate neurites.

Axons↗

PubMed1989

Polio and rabies vaccines produced in continuous cell lines: a reality for Vero cell line.

The Vero cell line was applied to the production of Enhanced-Inactivated Polio Vaccine (E-IPV), Oral Polio Vaccine (OPV) and Rabies Vaccine, also called Purified Vero Rabies Vaccine (PVRV) for humans. The cell line was expanded through the cell bank systems. Using a microcarrier culture technique, a large-scale production combined with an efficient purification process was developed. An extensive study of purity and biological safety was applied to the cell line to validate several Manufacturer's Working Cell Banks. Special emphasis was laid on the absence of any virus in the cells or any trace of viral sequence included in the Vero cell genome. Sensitive tests were applied to check that residual cellular DNA was as low as possible per dose of vaccine. E-IPV was licensed in July 1982 and since 1983 more than 20 million doses have been inoculated into children, without any side-effects. The production capacity of this E-IPV is presently more than 60 million doses per year. For PVRV, the license was delivered in June 1985 and over 350,000 doses have been injected without any problem. For the OPV, 27 monovalent lots were produced, 9 lots for each serotype. The studies of RNA sequences in variable regions and viral proteins gave results similar to the OPV produced from primary monkey kidney cells. All the other tests, and especially the neurovirulence tests in live monkeys showed that the OPV produced in Vero cell lines was acceptable. Since the completion of clinical studies, the license has been pending. The quality of the Vero cell line, as produced and tested by the "Institut Mérieux", combined with the safety afforded by the virus purification, has helped to prevent potential problems.

Animals↗

PubMed1991

Ligand binding and functional characterization of muscarinic acetylcholine receptors on the TE671/RD human cell line.

Cells of the TE671/RD human clonal line express a finite number (Bmax) of about 350 fmol/mg of membrane protein) of apparently noninteracting, high-affinity binding sites (KD of 0.07 nM and a Hill coefficient close to unity, nH = 0.94) for the muscarinic acetylcholine receptor (mAChR) radio antagonist, tritium-labeled quinuclidinyl benzilate [( 3H]QNB). The rank order potency of selective antagonists that inhibit specific [3H]QNB binding is: atropine greater than 4-DAMP (4-diphenylacetoxy-N-methylpiperidine methiodide) greater than pirenzepine greater than methoctramine greater than AFDX-116 (11-2[[2-[(diethylamino)methyl]-1-[piperidinyl] acetyl]-5,11-dihydro-6H-pyrido[2,3-b][1,4]benzodiazepin-6-one). Functional studies indicate that phosphoinositide (PIns) hydrolysis in TE671/RD cells is increased by carbachol (EC50 of 10 microM), but not by nicotine (to concentrations as high as 1 mM). Agonist-stimulated PIns metabolism is inhibited by antagonists with the same rank order potency as for inhibition of [3H]QNB binding. Functional responses are augmented in the presence of a nonhydrolyzable GTP analog, are strongly inhibited after 24-hr exposure to cholera toxin, but are only slightly inhibited after long-term exposure to pertussis toxin or forskolin. These studies identify a pharmacologically-defined M3-subtype of mAChR strongly coupled via a cholera toxin-sensitive mechanism to PIns hydrolysis in these cells. Within 1 hr of treatment of TE671/RD cells with 1 mM dibutyryl cyclic AMP or with 10 microM phorbol-12-myristate-13-acetate (PMA), there is a 30 to 50% decrease in carbachol-stimulated PIns responsiveness that recovers to control values after 5 days of continued drug treatment. However, a comparable and more persistent inhibition of mAChR function is observed on cell treatment with 20 nM PMA.(ABSTRACT TRUNCATED AT 250 WORDS)

Cell Line↗

PubMed1994

Comparison of thermoradiosensitization in two human melanoma cell lines and one fibroblast cell line by concurrent mild hyperthermia and low-dose-rate irradiation.

Two human melanoma cell lines, one radioresistant (SK-MEL-3) and one radiosensitive (HT-144), and a normal human fibroblast line (AG1522) were evaluated for thermoradiosensitization of low-dose-rate irradiation by concurrent mild hyperthermia (39-41 degrees C). None of the cell lines expressed chronic thermotolerance during heating at 39-41 degrees C. The SK-MEL-3 cells were the most heat sensitive, while AG1522 and HT-144 cells had the same sensitivity at 39 and 40 degrees C but HT-144 cells were more sensitive at 41 degrees C. All cell lines expressed thermal enhancement of radiosensitivity with heating during irradiation which increased with heating temperature. The SK-MEL-3 cells, which were the most resistant to radiation and demonstrated the greatest repair of sublethal damage (SLD) during low-dose-rate irradiation, had the greatest thermal enhancement of radiosensitivity, while the HT144 cells, which were the most sensitive and expressed little repair of SLD during low-dose-rate irradiation, had the smallest thermal enhancement of radiosensitivity. These data show that concurrent mild hyperthermia during low-dose-rate irradiation may be most efficacious in radiation-resistant tumor cells which express resistance through an enhanced capacity for repair of SLD.

Cell Line↗

PubMedDate not supplied

Establishment and characterization of cisplatin-resistant human epidermoid carcinoma cell line, A431 cell.

Cisplatin, cis-diamminedichloroplatinum(II) (CDDP) is one of the most important anticancer agents, initially producing good responses in various tumors. However, resistance to this drug often develops in various tumors, and additional administration decreases its chemotherapeutic efficacy. The precise mechanism of acquisition of resistance to this drug is still uncertain. However in the present study, we established two CDDP-resistant sublines A431/CDDP1 and A431/CDDP2 from human epidermoid carcinoma cell line A431. These resistant sublines were constituted by exposing A431 cells to a gradually increasing dose of CDDP (A431/CDDP1), and by mutagenic induction with mutagen (A431/CDDP2). A431/CDDP1 and A431/CDDP2 have developed 3.1 and 2.7 times more resistance to CDDP than the original A431 cell in terms of IC50. The two CDDP-resistant sublines showed cross-resistance to the CDDP analogue, carboplatin (CBDCA), but not to other chemotherapeutic drugs such as Adriamycin (ADR) and 5-fluorouracil (5-FU). These CDDP-resistant sublines were transplanted into nude mice to demonstrate the resistance to CDDP treatment in vivo. According to the in vitro assay, the mechanism of resistance in A431/CDDP1 and A431/CDDP2 seems to be based on a reduction of intracellular accumulation of CDDP, because their platinum concentration, which is the major component of CDDP, significantly declined. The established CDDP-resistant sublines may be used in further trials to improve the understanding of the mechanisms of resistance to CDDP.

Animals↗

PubMed1997

Recombinant HIV envelope expressed in an alpha-glucosidase I-deficient CHO cell line and its parental cell line in the presence of 1-deoxynojirimycin is functional.

alpha-Glucosidase inhibitors-e.g., 1-deoxynojirimycin (DNM)-interfere with HIV infectivity in CD4+ cell cultures but have proven unsuccessful in clinical trials. In vitro, several HIV Env properties, including the cleavage of the Env precursor gp 160, the immunoreactivity of the third variable domain (V3) of Env, the binding to the CD4 receptor, and the induction of the membrane fusion between the virus and the host cell, have been reported to be altered by such inhibitors. We have studied these properties for Env expressed via a recombinant vaccinia virus in two Chinese hamster ovary cell lines, an alpha-glucosidase I-deficient cell line and its parental cell line, treated with DNM under conditions that have been reported to alter Env properties. The glycosylation of Env, but not the quantity produced, varied in accordance with the experimental conditions. However, irrespective of these conditions, Env cleavage, V3 immunoreactivity, CD4 binding, membrane expression, and ability to induce syncytium formation were similar. Thus, neither the alpha-glucosidase I deficiency nor DNM treatment had a significant effect on the properties of Env produced here. Cellular mechanisms that may allow the normal expression of Env are discussed and may offer an explanation for the many discrepant results obtained to date on the effects of DNM on HIV Env.

1-Deoxynojirimycin↗

PubMed1999

Induced p21waf expression in H1299 cell line promotes cell senescence and protects against cytotoxic effect of radiation and doxorubicin.

The CDK inhibitor p21waf is a principal mediator of p53 function but can also be transactivated by many p53-independent stimuli leading to cell growth arrest or differentiation. In order to study the function of p21waf in a p53-deficient environment, we established an inducible expression of p21waf in the p53-null lung cancer cell line H1299, based on the muristerone-regulated system. Overexpression of p21waf led cells to growth arrest which after several days became irreversible and the arrested cells acquired a senescent phenotype as judged by cell shape, the senescence-associated beta-gal marker and inhibition of colony formation. The effect of p21waf overexpression, in the absence of p53, on the cytotoxicity caused by irradiation, doxorubicin and taxol was studied. Expression of p21waf provided protection against the cytotoxic effect of radiation and doxorubicin but not of taxol. These results are relevant to treatment of cancer when p53 is inactive.

Adenocarcinoma↗

PubMed2003

Contrasting effects of hydroxyurea on cell growth and reduction in Epstein-Barr virus genomes in EBV-infected epithelioid cell lines vs Burkitt's lymphoma cell lines.

Eliminating Epstein-Barr virus (EBV) genomes from infected cells is an intriguing theoretical strategy in therapy for EBV-associated malignant diseases. Respective patterns were characterized for hydroxyurea (HU)-promoted loss of EBV genomes from EBV-infected epithelioid cell lines derived from the noncancerous portion of gastric carcinoma tissues and Burkitt's lymphoma (BL) cell lines. Epithelioid cell lines GT38 and PN were less sensitivity to HU than BL cell lines Akata, Raji, and Daudi in terms of cell growth inhibition and cell cycle arrest. On passage in medium with 50 microM HU, the fraction of EBV nuclear antigen (EBNA)-positive cells was reduced substantially in the BL cell lines, but only slightly in the epithelioid cell lines. EBV DNA was reduced in Akata, Raji, and Daudi cells upon passage in 50 microM HU by 95%, 70%, and 50%, respectively, but by only 10% in GT38 cells, in which EBV DNA reduction was enhanced at increased concentrations of HU. This indicates that EBV genome is more easily lost from BL cell lines than from epithelioid cell lines upon culturing in HU. These findings support the view that the elimination of EBV could be therapeutically effective in EBV-associated BL by HU.

Burkitt Lymphoma↗

PubMed1976

Terminal deoxynucleotidyl transferase activities in human blood leukocytes and lymphoblast cell lines: high levels in lymphoblast cell lines and in blast cells of some patients with chronic myelogenous leukemia in acute phase.

Terminal deoxynucleotidyl transferase, an enzyme which catalyzes the polymerization of deoxyribonucleoside triphosphates, elongating oligo- or polydeoxynucleotide chains, but without direction from a nucleic acid template, is thought to be specific for thymus gland and thymus-derived cells. We have confirmed the observations that high levels are characteristic of thymus gland with both human and calf tissue and that elevated levels may be found in some cases of acute lymphocytic leukemia. High levels were also found in human lymphoblast cell lines with T-cell characteristics, and insignificant activity was observed in leukocytes of patients with chronic myelogenous leukemia not in acute blast phase of the disease, chronic lymphocytic leukemia, human B-cells, and normal human blood lymphocytes even after stimulation with phytohemagglutinin. However, high levels (approximately 200 nmoles/hr/10(9) cells) equivalent to those in thymus tissue and lymphoblast cell lines with T-cell characteristics were found in the peripheral blood blast cells of four patients with chronic myelogenous leukemia in an acute blast phase of their disease. One hypothesis that may explain the present results is that in chronic myelogenous leukemia in acute blast phase of the disease the proliferative blast response may not always be myeloblasts but in some cases it may be lymphoblasts.

Adult↗

PubMed1985

Cytostasis of tumor cell lines by promyelocytic leukemia cell line HL60 differentiated to granulocyte lineage.

The human promyelocytic leukemia cell line HL60, when cultured in medium containing dimethyl sulfoxide (DMSO) or granulocyte colony-stimulating factor (G-CSF), stopped dividing and differentiated into cells with granulocyte characteristics. We found that differentiated HL60 cells have no detectable cytolytic activity against cultured human bladder cell line (T24 cell), as measured by release of (3H) thymidine, or against K562 cells, as measured by release of chromium-51. Differentiated HL60 cells inhibited incorporation of (3H) thymidine into the DNA of adherent T24 cells. Decreased incorporation was not caused by detachment of the target T24 cells from the culture wells. The degree of cytostasis was dependent on the E/T ratio, with a 70%-80% inhibition usually reached at the E/T ratio of 200:1. A wide variety of target cells was also shown to be sensitive to differentiated HL60 cell-mediated cytostasis.

Cell Differentiation↗

PubMed2000

Caveolin-1 levels are down-regulated in human colon tumors, and ectopic expression of caveolin-1 in colon carcinoma cell lines reduces cell tumorigenicity.

Caveolin-1 expression and function were investigated in human colon cancer. Low levels of caveolin-1 mRNA and protein were detected in several colon carcinoma cell lines. Moreover, caveolin-1 protein levels were significantly reduced in human tumor epithelial mucosa (3.6 +/- 1.4-fold) when compared with normal colon mucosa for a majority (10 of 15) of the patients characterized. To directly assess the role of caveolin-1 in tumor development, caveolin-1 was reexpressed in the HT29 and DLD1 colon carcinoma cells, and the resulting HT29-cav-1 or DLD1-cav-1 cells were tested for tumorigenicity in nude mice. In most experiments, tumor formation was either blocked or retarded for HT29-cav-1 cells (10 of 13 mice) and DLD1-cav-1 cells (5 of 7 mice), as compared with both mock-transfected and parental HT29 or DLD1 cells. Interestingly, basal caveolin-1 levels were significantly reduced in HT29-cav-1 and DLD1-cav-1 cells isolated from tumors. Likewise, endogenous caveolin-1 mRNA and protein levels were found to be reduced in NIH-3T3 cells recovered from tumors after injection into nude mice. Thus, reexpression of caveolin-1 in colon carcinoma lines reduced the probability of tumor formation in vivo, and when tumors did develop from either HT29-cav-1, DLD1-cav-1, or NIH-3T3 cells, lower basal levels of caveolin-1 were detected. Finally, evidence was obtained indicating that initial caveolin-1 down-regulation in colon cancer cells need not be an entirely irreversible process because cell survival on selection for either drug resistance or increased metastatic potential correlated with increased caveolin-1 expression levels.

3T3 Cells↗

PubMed1999

CD9 is involved in invasion of human trophoblast-like choriocarcinoma cell line, BeWo cells.

The CD9 molecule is expressed on human extravillous trophoblasts, which invade the endometrium during implantation and placentation. To elucidate the role of CD9 in trophoblastic function, we investigated the expression of CD9 protein and mRNA in BeWo cells, a human trophoblast-like choriocarcinoma cell line, using immunohistochemistry, Western blotting and reverse transcription-polymerase chain reaction (RT-PCR). When BeWo cells were cultured with anti-CD9 monoclonal antibodies (mAb), their invasion through the extracellular matrices was significantly enhanced in a dose-dependent manner. Cell proliferation and human chorionic gonadotrophin production were unaffected. On the other hand, culture in the presence of mAb against integrins alpha3, alpha5 and beta1, which partially block the interaction with the extracellular matrices, inhibited BeWo cell invasion. Anti-CD9 monoclonal antibody had a stimulatory effect on BeWo cell invasion in the presence of anti-integrin alpha3 antibody. In contrast, it had no effect in the presence of mAb against integrins alpha5 and beta1, which were also highly expressed on BeWo cells. These findings suggest that CD9 has a function connected with the invasive properties of BeWo cells, which is partially mediated by integrin alpha5beta1. This may relate to the involvement of CD9 in trophoblastic invasion.

Antibodies, Monoclonal↗