Isolation and characterization of a novel bladder cancer cell line: inhibition by epidermal growth factor

A novel continuous cell line, designated BC3c, was established from a surgical biopsy of an invasive solid transitional cell carcinoma of the bladder derived from an 82-yr-old Caucasian female. BC3c cells were near-triploid bearing multiple structural and numerical chromosome anomalies. The epithelial origin of the cancer cells was indicated by the expression of cytokeratins 8 and 19 as well as by the absence of mesenchymal markers. Polymerase chain reaction-restriction-fragment length polymorphisms and single-strand conformation polymorphism mutation detection assays did not reveal any mutations in H-ras codon 12 and K-ras codons 12 and 13. In addition, no mutation in specific hot-spot codons of the p53 gene and no accumulation of the p53 protein were observed. BC3c cells grew rapidly in vitro, even in the absence of exogenous growth factors, because they were found to stimulate their growth in an autocrine manner. BC3c cells were found to express the epidermal growth factor-receptor (EGF-r) abundantly, but in contrast to other established bladder cancer cell lines, human recombinant epidermal growth factor inhibited the cells' proliferation in vitro. These features render the newly established bladder cancer cell line BC3c a useful tool for further experimentation.     

Isolation, cloning and characterization of a putative type-1 astrocyte cell line

The high prevalence of cholesterol gallstone disease in hypertriglyceridemic patients may be associated with frequent metabolic defects in cholesterol and bile acid syntheses and in the concomitant formation of bile supersaturated with cholesterol. This study had the two aims: 1) to assess whether the defects as well as the degree of biliary cholesterol supersaturation in patients with hyperlipoproteinemia (HLP) can be estimated by the simultaneous determination of plasma mevalonate (MVL) and 7alpha-hydroxy-4-cholesten-3-one (C4); and 2) to assess the possible application of an estimated cholesterol saturation index ([CSI]E) as a means of evaluating the clinical effects of simvastatin on biliary lipid composition. Biliary cholesterol supersaturation was observed in patients with both IIa and IV HLP types. Consistent with the high activity and steady-state messenger RNA level of 3-hydroxy-3 methylglutaryl coenzyme A (HMG-CoA) reductase, plasma MVL was significantly higher in 86 patients with HLP (38 type IIa, 44.1 +/- 2.4 nmol/L and 48 type IV, 56.7 +/- 2.3; P < .01) than in 41 normolipidemic subjects (34.2 +/- 1.5), closely correlating with the molar percentage of cholesterol in bile (r = .61, P = .0001; n = 86). On the other hand, consistent with the high activity and messenger RNA level of cholesterol 7alpha-hydroxylase, plasma C4 was significantly higher in patients with HLP (type IIa, 28.8 +/- 2.3 nmol/L and type IV, 38.3 +/- 2.7; P < .01) than in normolipidemic subjects (17.4 +/- 1.5). Plasma C4 was closely correlated with plasma MVL (r = .40, P = .0001; n = 86), but was inversely correlated with the molar percentage of bile acids in bile (r = .49, P = .0001; n = 86). Assuming that cholesterol supersaturation in patients with HLP may be governed by both an enhanced cholesterol secretion (closely reflected by plasma MVL) and a decreased secretion of bile acids (closely reflected by plasma C4), the multivariate linear regression-analyses revealed that an index defined as estimated CSI ([CSI]E) (%) in patients with HLP was given by the following equation using plasma MVL and C4 (nmol/L): [CSI]E = 1[MVL] + 0.7[C4] + 44.4. Biliary cholesterol supersaturation in patients treated with simvastatin improved in a manner parallel to the time course of decreases in plasma MVL and C4. The [CSI]E before and at the end of treatment were correlated with biliary CSI. These results indicate that defects of hepatic cholesterogenesis, and bile acid synthesis, and the degree of biliary cholesterol supersaturation in patients with HLP can be estimated exactly by the simultaneous determination of plasma MVL and C4; furthermore [CSI]E may be adopted for clinical use as a convenient index of biliary CSI.     

Isolation and characterization of apoptosis-resistant mutants from a radiosensitive mouse lymphoma cell line

This paper reports the findings of a research study funded by the English National Board for Nursing, Midwifery and Health Visiting (ENB), which explored the impact of community care reforms on mental health and learning disability nurses and their practice. In this study we were struck by the divergent views of our respondents about the nature of mental health and learning disability nursing as practice disciplines and the implications of these views for the future shape of preregistration educational preparation. We noted, in particular, the debate between those who advocate what is referred to as specialist educational preparation and others who favour generic preparation. The specialist-generic debate is relevant to nursing as a whole but was magnified in the context of our study because genericism was perceived by many of our respondents as a threat to the minority branches and especially to those (arguably mental health and learning disability) that are not rooted in the biomedical tradition of general nursing. This paper seeks to contribute to this debate as it impinges on the two nursing specialties by developing models of future nurse education grounded in the empirical data from our research and interrogating them to draw out their central features. Two models are clearly independent: the 'specialist' and the 'generic' models. Another three models are partial in that they draw upon the first two: the 'pragmatic' model, the 'unity-of-nursing' model, and the 'social care' model. A common feature of the pragmatic and unity-of-nursing models is that they support the existing DipHE programme, which is now the dominant form of preregistration nurse preparation in the UK. The social care model is applicable principally to learning disability nursing.     

Isolation and characterization of a spontaneously transformed malignant mouse mammary epithelial cell line in culture

A method is described that permits the selection of spontaneously transformed mammary epithelial colonies from an untransformed mouse mammary epithelial cell line, NMuMG, and utilizes a long-term anchorage-independent growth of the transformants on soft agarose. These transformed cells (NMuMG-ST) are shown to be distinguishable from the untransformed cells by morphology, growth characteristics, induced carcinomas when transplanted into nude mice and ability to metastasize. This transformed phenotype displayed focal, multilayer growth and higher saturation density in comparison with the untransformed phenotype. Transplanted tumors as well as metastatic lung tumors in nude mice were adenocarcinomas morphologically similar to typical mammary tumors in humans. This selection procedure of mutant mammary cells from an immortalized cell line derived from normal mammary glands could be very useful to identify the genomic biomarkers in the growth regulation and malignant progression of breast cancer.     

Isolation and characterization of nocardia-like variants of Mycobacterium smegmatis

Orange-red-pigmented (OR) colonies were isolated from cream-yellow-pigmented Mycobacterium smegmatis after exposure to either mycobacteriophage MC4 or ultraviolet light; these variant strains were designated OR4 and ORuv, respectively. Early subculture of OR-colonies did not show any segregation of parental-type cells. However, colonies resembling the parental strains, possibly representing a back mutant (REV-OR4), were occasionally found during subculture of established OR-colonies or upon treatment with N-nitrose-N'-nitro-N-methylguanidine. The OR-variants were characterized by their lytic response to nocardiophage, but not to mycobacteriophages, presence of alpha-branched, beta-hydroxylated fatty acids of the Nocardia-type, and a guanine plus cytosine value of deoxyribonucleic acid (DNA) between 62 and 64 mol%. They were more resistant to the lethal action of both ultraviolet light and mitomycin C treatment than the parental and back mutant strains. Although the OR-variants in this study possess characteristics common to the genus Nocardia or some of the 'rhodochrous' mycobacteria, evidence is presented that they form a new class of mycobacterial variants.     

Isolation and characterization of an intracellular serine proteinase inhibitor from a monkey kidney epithelial cell line

A recent report described a thrombin inhibitory activity in the soluble fraction of human placenta and the cytosolic fraction of K562 cells. Isolation and characterization of the functionally inactive 35-38-kDa placental form of this protein revealed that it was a novel serine proteinase inhibitor (Coughlin, P. B., Tetaz, T., and Salem, H. H. (1993) J. Biol. Chem. 268, 9541-9547). In the present study, we observed a 67-kDa sodium dodecyl sulfate (SDS)-stable complex when 125I-thrombin was incubated with the cytosolic fraction of a monkey kidney epithelial cell line, BSC-1. This complex was not observed in either the particulate cell fraction extracted with 0.2% Triton X-100 or medium conditioned by cells, suggesting that the thrombin-complexing factor is confined to the cytoplasm. The cytoplasmic antithrombin activity was purified to apparent homogeneity from the cytosol of BSC-1 cells previously pulsed with [35S]methionine by a combination of heparin-agarose chromatography, Mono Q fast protein liquid chromatography, and anhydrotrypsin-Affi-Gel 10 affinity chromatography. Analysis of the affinity-purified preparation by SDS-polyacrylamide gel electrophoresis and fluorography revealed a single protein with an apparent molecular mass of 38 kDa. The purified 38-kDa protein inhibited the amidolytic activities of thrombin, trypsin, urokinase, and factor Xa but not that of elastase. Incubation of the 38-kDa protein with excess thrombin identified approximately 60% of the labeled 38-kDa protein in an SDS-stable 67-kDa complex. The purified 38-kDa inhibitor was cleaved with cyanogen bromide and the isolated peptides subjected to microsequencing. Amino acid sequence obtained for a region within this protein exhibited significant homology with human antithrombin III and plasminogen activator inhibitors 1 and 2. This homologous peptide contained the full complement of residues designated as highly conserved in helix F of the greater serine proteinase inhibitor superfamily. In addition, an internal sequence of GGGGDIHQGF was found in the monkey cytoplasmic inhibitor, which is identical to that reported for an internal sequence of the human placental inhibitor. These findings confirm the existence of a novel cytoplasmic serine proteinase inhibitor in mammalian cells and provide additional details of its molecular properties. The physiological function of this novel serine proteinase inhibitor in cytoplasm is unknown.     

Isolation and characterization of a microsomal arylaminopeptidase from rat kidney

The isolation and characterization of a microsomal arylaminopeptidase from rat kidney is reported. By treatment of a microsomal arylaminopeptidase-phosphatase-complex with trypsin and subsequent gel filtration of the solubilized proteins on Sepharose 6B a electrophoretic homogeneous arylaminopeptidase was obtained (yield, 3%; enrichment, 900 times). The following properties of the purified enzyme were determined: 1. Molecular weight: 182000 (gel filtration on Sepharose 6B) to 192000 (SDS-polyacrylamide gel electrophoresis). 2. Subunit structure: In the presence of 6 M guanidine - HC1 + 1% BETA-mercaptoethanol the enzyme dissociates into subunits (MW 46700, ESTIMATED BY SDS gel electrophoresis method). 3. Isoelectric point: 4,71 (agarose gel electrophoresis method). 4. UV characteristics: E 280nm/E260NM=1.3. 5. Substrate specifity: optimal substrates L-alanyl derivatives (anilide, beta-naphthyl amide, p-nitroanilide, 4-(phenylazo)-phenylamide and hydrazide). Among these compounds the anilide derivative was hydrolyzed most rapidly. Furthermore, di- and tripeptides, especially L-methionyl-L-leucine, were also split. No hydrolysis was observed with hemoglobin (pH 4.5 and 7.5) and amino acid- or peptide-ester substrates. 6. Optimal pH: 7.5 +/- 0,1; optimal temperature: 45 to 50 degrees C. 7. The enzyme has no transamidation activity with L-alanyl amide both as aminoacyl donator and -acceptor. 8. Influence of effectors: Heavy metal ions (Ni2+, Cd2+, Cu2+, Zn2+), chelating agents (EDTA, o-phenanthroline) and puromycin inhibit the enzyme significantly. SH-group reagents are without any influence. 9. L-alanyl-L-alanyl-4 (phenylazo)-phenylamide, a dipeptide aryl aminopeptidase substrate, is hydrolyzed by the purified enzyme preparation according to a consecutive or step by step mechanism.     

Purification and characterization of chondroitin 4-sulfotransferase from the culture medium of a rat chondrosarcoma cell line

Chondroitin 4-sulfotransferase, which transfers sulfate from 3'-phosphoadenosine 5'-phosphosulfate to position 4 of N-acetylgalactosamine in chondroitin, was purified 1900-fold to apparent homogeneity with 6.1% yield from the serum-free culture medium of rat chondrosarcoma cells by affinity chromatography on heparin-Sepharose CL-6B, Matrex gel red A-agarose, 3',5'-ADP-agarose, and the second heparin-Sepharose CL-6B. SDS-polyacrylamide gel electrophoresis of the purified enzyme showed two protein bands. Molecular masses of these protein were 60 and 64 kDa under reducing conditions and 50 and 54 kDa under nonreducing conditions. Both the protein bands coeluted with chondroitin 4-sulfotransferase activity from Toyopearl HW-55 around the position of 50 kDa, indicating that the active form of chondroitin 4-sulfotransferase is a monomer. Dithiothreitol activated the purified chondroitin 4-sulfotransferase. The purified enzyme transferred sulfate to chondroitin and desulfated dermatan sulfate. Chondroitin sulfate A and chondroitin sulfate C were poor acceptors. Chondroitin sulfate E from squid cartilage, dermatan sulfate, heparan sulfate, and completely desulfated N-resulfated heparin hardly served as acceptors of the sulfotransferase. The transfer of sulfate to the desulfated dermatan sulfate occurred preferentially at position 4 of the N-acetylgalactosamine residues flanked with glucuronic acid residues on both reducing and nonreducing sides.     

Eosinophilic cystitis: a rare case which presented as an invasive bladder tumor

Eosinophilic cystitis is a rare inflammatory lesion of the bladder of unknown etiology. It can be confused clinically and cytoscopically with invasive bladder cancer. We report a case in which the lesion presented as an invasive bladder tumor and we discuss the literature in brief.     

Isolation and characterization of rat primary lung cells

Lung cell culture may be useful as an in vitro alternative to study the susceptibility of the lung to various toxic agents. Lungs from female Wistar rats were enzymatically digested by recirculating perfusion through the pulmonary artery with a sequence of solutions containing deoxyribonuclease, chymopapain, pronase, collagenase, and elastase. Lung tissue was microdissected and resuspended and the cells obtained were washed by centrifugation. By this isolation method, 2 x 10(8) cells per rat lung were obtained with an average viability of 97%. Lung cells cultured in medium containing antibiotics and serum maintained a viability of > 70% for 5 d. Rat primary lung cells were exposed to various toxic agents and their viability was assessed by formazan production capacity after 18 h of incubation. Compared to rat and mouse hepatocyte cultures (EC50 = 5.8 mM), rat primary lung cells were much more susceptible to hydrogen peroxide (EC50 = 0.6 mM). All cell types were equally sensitive to the more potent toxicant tert-butylhydroperoxide (EC50 = 0.1 mM). Paraquat was more toxic to lung cells (EC50 = 0.03 mM) than to rat (EC50 = 2.8 mM) and mouse (EC50 = 0.2 mM) hepatocytes. In contrast, rat lung cells were less sensitive to sodium nitroprusside (EC50 = 2.6 mM) compared to rat (EC50 = 0.2 mM) and mouse (EC50 = 0.03 mM) hepatocytes. Nitrofurantoin and menadione (at EC50 = 0.04 mM and 0.006 mM, respectively) were more toxic to rat lung and liver cells than to murine hepatocytes (EC50 = 0.2 mM and 0.04 mM, respectively). Our findings demonstrate the applicability of this rat primary lung cell culture for studying the effects of lung toxicants.     

Isolation and characterization of brain-specific transglutaminases from rat

Angiotensin II (AT2) has been implicated in the growth and/or differentiation of its target tissues. In the present study, testicular AT2 receptor and its subtypes in hypophysectomized rats were examined using quantitative in vitro autoradiography and Northern blot analysis in an attempt to determine possible involvement of pituitary hormones in their expression. Prepubescent (3 weeks of age) male Sprague-Dawley rats underwent hypophysectomy or sham operation. From 10 days thereafter, they were treated with vehicle, growth hormone, human chorionic gonadotrophin or human menopausal gonadotrophin for 10 days. Testicular AT2 receptors were labelled with 125I-[Sar1,Ile8] AT2 and differentiated into its subtypes (AT1 and FAT2) according to their susceptibility to AT1 (losartan, 5 microM) and AT2 (CGP42112B, 1 microM) antagonists. Hypophysectomy led to a marked increase in AT2 receptor concentration (sham-operated rats: 0.7 +/- 0.2 fmol/mg protein, hypophysectomized rats: 2.5 +/- 0.6 fmol/mg protein, mean +/- SEM, n = 11-12, p < 0.01) with predominant occurrence of AT1 receptors. Both human chorionic gonadotrophin and human menopausal gonadotrophin decreased testicular AT2 receptor concentration, whereas growth hormone did not affect AT2 receptor expression. Northern blot analysis revealed both testicular AT1 and AT2 receptor mRNA expression to be significantly increased after hypophysectomy and reduced by gonadotrophin treatment. These results suggest that the expression of testicular AT2 receptors is regulated by pituitary gonadotrophins and that AT2 may play a role in testicular growth and/or differentiation.     

Isolation and characterization of rat olfactory marker protein

The olfactory marker protein was isolated and characterized from rat olfactory bulbs. Its properties and those of the olfactory marker protein isolated from the mouse are described. The rat protein was less acidic (pI = 5.0) than the mouse protein (pI = 4.7). However, the amino acid compositions were very similar: in both proteins arginine plus lysine accounted for 13 mol% and glutamate plus aspartate for 30 mol% of the total residues. Molecular weights of both proteins estimated by sodium dodecyl sulfate gel electrophoresis were indistinguishable and estimated to be 16,500. The molecular weight of the native rat olfactory marker protein estimated by gel filtration techniques was 30,000, which is identical to the molecular weight of the native mouse and garfish olfactory marker proteins. This suggested a dimeric structure. The purified rat and mouse proteins behaved like species of 35,000 molecular weight on gel filtration.     

Radio-sensitive murine thymoma cell line 3SB: characterization of its apoptosis-resistant variants induced by repeated X-irradiation

This paper describes public health nurses' perceptions of changes in their practice. The participants were 28 public health staff nurses from six Alberta, Canada health units serving urban and rural populations. Data were collected in 1993-94 using individual and focus group interviews. Content analysis was used to identify the following themes: "pulling back", "from hands on to arms length", "handing over responsibility", "developing working partnerships", and "doing less surveillance". These themes are discussed in terms of their implications for population health and for public heath nursing, using as a point of reference the principles of Primary Health Care. Continuing research is needed to chronicle further changes in public health nursing practice that will result from health care restructuring and health system reform.     

Overexpression of transforming growth factor beta type I receptor abolishes malignant phenotype of a rat bladder carcinoma cell line

The disposition of ISIS 2922, a phosphorothioate oligonucleotide for treatment of cytomegalovirus associated retinitis, was evaluated in rabbits. Vitreous humor and retina samples were collected from rabbits that received a single intravitreal injection of 66 microg [14C]-labeled ISIS 2922 and were analyzed using anion exchange HPLC. Four hr postdosing, the concentration of ISIS 2922 in vitreous humor was 3.3 microM. The elimination of ISIS 2922 from the vitreous humor exhibited first-order kinetics with a t1/2 of 62 hr. By 10 days postdosing, the mean concentration of ISIS 2922 in rabbit vitreous humor had decreased to 0.17 microM, which represented 22% of the total radioactivity remaining in the vitreous. The remaining 78% coeluted on anion exchange HPLC with shorter oligonucleotides. In retina, ISIS 2922 accumulated over the first 5 days postdosing, reaching a maximum concentration of 3.5 microM, and then declined thereafter with an estimated t1/2 of 79 hr. By 10 days postdosing when only 24% of the total radioactivity in the retina was parent compound, the concentration of ISIS 2922 remained at 1.6 microM, which was 10 times higher than the concentration in the vitreous humor. Whereas the elimination of full-length ISIS 2922 and total radioactivity from the vitreous humor occurred at nearly equal rates, ISIS 2922 disappeared more rapidly than did total radioactivity from the retina, suggesting a greater role for metabolism in the clearance process from retina than the vitreous. Alternatively, the results are consistent with metabolites being cleared from the vitreous at approximately the same rate as parent compound while in the retina metabolites may be cleared more slowly. The data were analyzed with a user-defined pharmacokinetic model, which was then used to predict the potential for accumulation of ISIS 2922 during clinical dosing.     

Isolation and characterization of a novel gene from human glioblastoma multiforme tumor tissue

The therapeutic application of high-dose interleukin (IL) 2 in human malignancy is limited by severe multiorgan toxicities that are mediated, in part, by tumor necrosis factor (TNF) and IL-1. CT1501R (lisofylline; LSF) is one of several methyl xanthine congeners that inhibit the effects of TNF by the interruption of specific signal transduction pathways. This randomized, placebo-controlled trial was designed to assess the activity of LSF in reducing the toxicities of high-dose IL-2 therapy. Fifty-three patients with metastatic renal cancer or malignant melanoma were treated with i.v. bolus IL-2, 600, 000 IU/kg every 8 h for 5 days (14 doses), followed by 9 days of rest and another 5-day course of IL-2. Patients were randomly assigned to LSF, 1.5 mg/kg i.v. bolus, or placebo every 6 h during IL-2 therapy. All patients were to be treated to individual maximum tolerance of IL-2 at the intensive care unit level of support. The end points for statistical analysis were the number of IL-2 doses administered during the first cycle of treatment (maximum, 28) and the toxicities experienced by each group after the first 8 planned IL-2 doses. There was no difference between the LSF and placebo groups in the mean number of IL-2 doses tolerated in the entire first cycle of therapy (19.6 +/- 5.4 versus 19.5 +/- 5.8, P = 0.86) or in the first or second 5-day course of IL-2. The only significant difference in toxicities occurring through the eighth dose of IL-2 was in the maximum elevation of serum creatinine (mean, 1.7 +/- 0.8 for placebo versus 1.5 +/- 0.6 mg/dl for LSF, P = 0.013). A Monte Carlo analysis of major toxicities over the first 14-dose course of therapy showed a statistically significant difference favoring the LSF-treated group (P = 0.025). LSF was well tolerated, associated only with mildly increased nausea (P = 0.006 after eight IL-2 doses, but not significant for the entire first cycle). The antitumor activity was comparable in both groups (objective responses, 2 of 28 with LSF versus 4 of 24 with placebo). The mean peak plasma concentrations of LSF on days 1, 5, and 19 were 6.24, 3.83, and 5.04 micromol/liter, respectively. In conclusion, with this dose and schedule, LSF did not alter the toxicities of high-dose i.v. IL-2 sufficiently to impact the overall dose intensity of IL-2. Successful IL-2 toxicity modulation may require the use of higher doses of LSF, the development of agents with more potent anti-TNF activity, and/or combined modulating agents that function via distinct mechanisms to interrupt cytokine-mediated signaling.     

Isolation and characterization of an alpha 1,2-mannosidase cDNA from the lepidopteran insect cell line Sf9

Norepinephrine (NE) (2.5 micrograms/kg/min) was administered to 5-week-old male Sprague Dawley rats by subcutaneous osmotic mini pumps for 14 days to generate an in vivo cardiac hypertrophy model and the responses with respect to aging examined. In the model, ventricles were significantly hypertrophied without myocardial necrosis and without significant increases in heart rate or blood pressure; the beta adrenergic system was down-regulated. In 37-week-old rats receiving 1.0 microgram/kg/min NE, there was a tendency towards heart failure, and myocardial necrosis and interstitial fibrosis were revealed by histological examinations. The density of beta adrenergic receptors and adenylyl cyclase activity was lower in the older rats. The excess stimulation of adrenergic receptors caused severe cardiac injury in old rats regardless of down regulation of beta adrenergic receptors.     

Relationship of tumor angiogenesis and nuclear p53 accumulation in invasive bladder cancer

The purpose of this investigation was to evaluate the relationship between tumor angiogenesis and nuclear p53 accumulation in invasive bladder cancer. We studied 161 patients with invasive transitional cell carcinoma of the bladder who had previously undergone radical cystectomy. Analysis was performed to determine the presence of p53 nuclear accumulation and extent of tumor-associated angiogenesis. p53 status identified a group of patients at high risk for tumor progression (p53-altered tumors), and microvessel density determinations added additional prognostic information by identifying a subset of aggressive tumors within the wild-type p53 subgroup. At 5 years, patients with tumors exhibiting no evidence of p53 alterations and low microvessel counts demonstrated 3% recurrence and 88% survival, compared to 43% recurrence and 59% overall survival for patients with intermediate vessel counts and 61% recurrence and 43% overall survival for patients with the highest vessel counts (P < 0.001 and P = 0.003, respectively). Angiogenesis also provides additional prognostic information to patients with tumors that demonstrate p53 alterations. An association between angiogenesis and p53 status did exist (P = 0. 05); however, 27% of the tumors that showed no evidence of p53 alterations exhibited high microvessel counts, and 26% of tumors with evidence of p53 alterations had low microvessel counts. Tumor-associated angiogenesis adds additional useful prognostic information to that which is obtained from p53 status in patients with invasive transitional cell carcinoma of the bladder. Although an association between p53 status and the degree of angiogenesis was identified, other factors appear to play a role in the regulation of tumor-induced neovasularization.     

Establishment and characterization of high- and low-lung-metastatic cell lines derived from murine colon adenocarcinoma 26 tumor line

We established and characterized high- (LuM1) and low-lung-metastatic (NM11) cell lines derived from murine colon adenocarcinoma 26 tumor line. LuM1 cell line was established as a clonal cell line from a cultured cell mixture derived from a lung-metastatic nodule after 7 sequential subcutaneous transplantations of lung-metastatic tumors in the abdominal wall of BALB/c mice. NM11 cell line was established from a cultured cell mixture derived from a subcutaneous transplant of murine colon adenocarcinoma 26 tumor cells. LuM1 cells showed marked spontaneous lung metastases, but NM11 cells rarely did. High invasive potential of LuM1 cells was revealed by in vitro invasion assay using Matrigel reconstituted membranes. Rapid retraction was observed in monolayers of human umbilical vein endothelial cells and bovine aortic endothelial cells when LuM1 cells were added on the monolayers. Gelatin zymography and immunochemical examinations with monoclonal antibodies against gelatinase B (Mr 95,000 type IV collagenase) showed secretion of large amounts of the gelatinase by LuM1 cells.