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  • 1
    Electronic Resource
    Electronic Resource
    Analysis of the Multistep Process of Carcinogenesis Using Human Fibroblasts (1994)
    Oxford, UK : Blackwell Publishing Ltd
    Risk analysis 14 (1994), S. 0 
    Details
    ISSN: 1539-6924
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Energy, Environment Protection, Nuclear Power Engineering
    Notes: Normal human cells in culture have never been neoplastically transformed by carcinogen exposure. One possible explanation is that the life span of such cells is too short for them to acquire the necessary changes. To test this hypothesis, we needed normal human cells with a greatly extended or an infinite life span. We transfected the v-myc gene and a selectable marker into normal human fibroblasts, identified a drug-resistant clone expressing v-myc protein, and passaged the progeny of the clone until they senesced. A few cells continued to proliferate and gave rise to a diploid, infinite life span cell strain, MSU-1.0, that has normal growth control and is nontumorigenic in athymic mice. Analysis showed that one more genetic change, in addition to unregulated expression of the v-myc gene, was involved in generating MSU-1.0 cells. They spontaneously gave rise to a variant strain, designated MSU-1.1, that grows more rapidly and is less dependent on exogenous growth factors. Analysis showed that at least two additional changes were involved in generating this cell strain. It has a stable karyotype composed of 45 chromosomes, including two markers. Transfection of specific oncogenes was used to determine the number and nature of additional changes required to transform MSU-1.1 cells into malignant cells. Analysis indicated that two changes were involved, but no change in karyotype. Exposure of MSU-1.1 cells to a single carcinogen treatment, followed by selection for cells with the characteristics of oncogene-trans-formed MSU-1.1 cells also yielded malignant human cells. We conclude that malignant transformation of normal human fibroblasts requires six or seven genetic changes, some of which involve suppressor genes.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1111/j.1539-6924.1994.tb00240.x
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  • 2
    Electronic Resource
    Electronic Resource
    Mutagenic Action of Aflatoxin B 1 on Transforming DNA and Inhibition of DNA Template Activity in vitro (1970)
    [s.l.] : Nature Publishing Group
    Nature 225 (1970), S. 68-70 
    Details
    ISSN: 1476-4687
    Source: Nature Archives 1869 - 2009
    Topics: Biology , Chemistry and Pharmacology , Medicine , Natural Sciences in General , Physics
    Notes: [Auszug] Fig. 1. Structures of the four aflatoxin components B1, G1, B1 and G2. Our previous studies (ref. 10 and unpublished work with J. A. Miller and E. C. Miller) with a series of metabolites and synthetic derivatives of five powerful liver carcinogens have shown that only forms considered bo be ...
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1038/225068a0
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  • 3
    Electronic Resource
    Electronic Resource
    Frequency of ultraviolet light-induced mutations is higher in xeroderma pigmentosum variant cells than in normal human cells (1976)
    [s.l.] : Nature Publishing Group
    Nature 261 (1976), S. 593-595 
    Details
    ISSN: 1476-4687
    Source: Nature Archives 1869 - 2009
    Topics: Biology , Chemistry and Pharmacology , Medicine , Natural Sciences in General , Physics
    Notes: [Auszug] Fig. 1 Comparison of the cytotoxic effect of ultraviolet irradiation as a function of dose, in normal cells (?) with that in the variant, XP4BE (?), as determined from the percentage survival of the colony-forming ability. Survival of the cloning ability of irradiated cells divided by that of ...
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1038/261593a0
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  • 4
    Electronic Resource
    Electronic Resource
    Synergistic effect of caffeine on the cytotoxicity of ultraviolet irradiation and of hydrocarbon epoxides in strains of Xeroderma pigmentosum (1975)
    [s.l.] : Nature Publishing Group
    Nature 258 (1975), S. 760-763 
    Details
    ISSN: 1476-4687
    Source: Nature Archives 1869 - 2009
    Topics: Biology , Chemistry and Pharmacology , Medicine , Natural Sciences in General , Physics
    Notes: [Auszug] Preliminary evidence, from alkaline sucrose gradient studies, that such XP variants suffer from a defect in their DNA synthesis following ultraviolet irradiation was reported12 and documented6 by Lehmann and his coworkers. They showed that of three variants tested after irradiation, all were ...
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1038/258760a0
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  • 5
    Electronic Resource
    Electronic Resource
    Kinds and locations of mutations arising spontaneously in the coding region of theHPRT gene of finite-life-span diploid human fibroblasts (1991)
    Springer
    Somatic cell and molecular genetics 17 (1991), S. 463-469 
    Details
    ISSN: 1572-9931
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Spontaneous thioguanine-resistant mutants were derived from populations of finite-life-span, diploid human fibroblasts by means of a fluctuation analysis. cDNA was prepared from mutantHPRT mRNA and amplified by the polymerase chain reaction, and the sequence of the product was analyzed. Exon deletions, which very likely arose from mutations in the intron splice site consensus sequences, were found in 10 of the 37 mutants examined (27% of the total). Among the 28 mutations in the coding sequence, base pair substitutions predominated (89%). With the exception of one base pair involved in a tandem mutation, all base pair substitutions resulted in alterations in the predicted amino acid sequence of the protein. In addition there were three frameshift mutations, consisting of the deletion of one or two base pairs. Although mutations occurred throughout the coding sequence, 50% (14/28) were found in the 5′ portion of exon 3.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF01233170
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  • 6
    Electronic Resource
    Electronic Resource
    Transfection of amyc gene as a means of generating infinite life span human fibroblast strains (1995)
    Springer
    Methods in cell science 17 (1995), S. 141-148 
    Details
    ISSN: 1573-0603
    Keywords: Human fibroblasts ; Simian virus 40 ; Transfection
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Human fibroblasts in culture have never been found to transform spontaneously into immortal cells. In an effort to generate an infinite life span cell strain from foreskin-derived normal diploid fibroblasts, we transfected the cells with a plasmid carrying a v-myc oncogene linked to theneo gene, or with a control vector carrying theneo gene, and selected drug-resistant clones. A clone that expressed the v-myc protein was propagated to the end of its life span, with periodic cryogenic storage of the progeny. The population went into crisis at the same time as cells from the control population and eventually senesced. However, while the cells were senescing, viable-appearing clones were noted. The cells of these clones continued to multiply, very slowly at first but eventually at a faster rate. Analysis showed that these cells have a diploid karyotype that has remained stable throughout more than 200 population doublings since their sibling cells senesced. Molecular analysis showed that the infinite life span cells are, indeed, derived from the cells used for transfection, and that they continue to express the v-myc protein.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF00986663
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  • 7
    Electronic Resource
    Electronic Resource
    Methods for quantifying carcinogen-induced transformation of diploid human fibroblasts to anchorage independence (1986)
    Springer
    Methods in cell science 10 (1986), S. 189-195 
    Details
    ISSN: 1573-0603
    Keywords: transformation ; human fibroblasts ; anchorage independence ; soft agar
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary We have determined the assay conditions necessary for quantifying the frequency of carcinogen-induced transformation of diploid human fibroblasts to anchorage independence. Emphasis is placed on the importance of using medium that supports clonal growth of the particular type of cells under investigation and titrating the percent of fetal bovine serum used in the assay to obtain a low, but measurable number of anchorage-independent colonies in the untreated control cultures. With this method, the target cells can be derived from adults as well as newborns and need not be in early passage, as long as they are able to grow vigorously.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1007/BF01405083
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  • 8
    Electronic Resource
    Electronic Resource
    Transformation of diploid human fibroblasts by DNA transfection with the v-sis oncogene (1986)
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 128 (1986), S. 313-321 
    Details
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The simian sarcoma virus (SSV) oncogene (v-sis) has a high degree of homology to the cellular gene coding for the B peptide of human platelet-derived growth factor (PDGF), a potent fibroblast mitogen. The cellular homolog of v-sis is activated in some mesenchymal human tumors and cell lines drived from them. To determine the phenotype produced by v-sis in diploid human fibroblasts, we constructed plasmids containing the SSV provirus and drugresistance markers and transfected them into early-passage human cells. Fibroblasts that had integrated the plasmid were selected for drug resistance and shown to contain and express the v-sis oncogene by DNA and RNA hybridization. The v-sis-expressing cells grew to higher saturation densities than control cells transfected with vector plasmid alone and formed large, well defined foci. This allowed selection of transfectants directly for focus formation. The v-sis transformed cells continued to grow well in the absence of serum, whereas age-matched, vector-transfected control cells ceased replicating under these conditions so that the final difference in density between the two populations was tenfold. Incorporation of thymidine in serum-free medium by the v-sis-transformed cells was independent of exogenous PDGF. In contrast, PDGF increased thymidine incorporation in such medium by the control cells to the level found in the v-sis-transformed cells with or without added PDGF. These results suggest that expression of the v-sis oncogene in diploid human fibroblasts causes sufficient endogenous synthesis of the B chain of PDGF to allow transformants to grow to abnormally high cell densities. When individual v-sis-transformed cells were grown on a background of normal cells, this higher cell density at confluence could be visualized as a focus.
    Additional Material: 5 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/jcp.1041280225
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  • 9
    Electronic Resource
    Electronic Resource
    Platelet-derived growth factor or basic fibroblast growth factor induce anchorage-independent growth of human fibroblasts (1988)
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 137 (1988), S. 588-592 
    Details
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Anchorage-independent growth, i.e., growth in semi-solid medium is considered a marker of cellular transformation of fibroblast cells. Diploid human fibroblasts ordinarily do not exhibit such growth but can grow transiently when medium contains high concentrations of fetal bovine serum. This suggests that some growth factor(s) in serum is responsible for anchorage-independent growth. Much work has been done to characterize the peptide growth factor requirements of various rodent fibroblast cells for anchorage-independent growth; however, the requirements of human fibroblasts are not known. To determine the peptide growth factor requirements of human fibroblasts for anchorage-independent growth, we used medium containing serum that had had its peptide growth factors inactivated. We found that either platelet-derived growth factor (PDGF) or the basic form of fibroblast growth factor (bFGF) induced anchorage-independent growth. Epidermal growth factor (EGF) did not enhance the growth induced by PDGF, or did so only slightly. Transforming growth factor beta (TGF-β) decreased the growth induced by PDGF. EGF combined with TGF-β induced colony formation in semi-solid medium at concentrations at which neither growth factor by itself was effective, but the combination was much less effective in stimulating anchorage-independent growth than PDGF or bFGF. This work showed that PDGF, or bFGF, or EGF combined with TGF-β can stimulate anchorage-independent growth of nontransformed human fibroblasts. The results support the idea that cellular transformation may reduce or eliminate the need for exogenous PDGF or bFGF.
    Additional Material: 1 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/jcp.1041370328
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  • 10
    Electronic Resource
    Electronic Resource
    Failure of infinite life span human cells from different immortality complementation groups to yield finite life span hybrids (1994)
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 159 (1994), S. 151-160 
    Details
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The observation that fusion of infinite life span cells with finite life span cells produces hybrid cells with finite life spans led to the conclusion that an infinite life span in culture is a recessive trait resulting from loss of the function of a gene or genes that contribute to an active program for cellular senescence. Furthermore, finding that certain pairs of infinite life span cells, when fused to one another, can complement each other to yield finite life span hybrids allowed 30 infinite life span cell lines to be assigned to four immortality complementation groups (Pereira-Smith and Smith, 1988, Proc. Natl. Acad. Sci. U.S.A., 85:6042). In the present study, we fused a chromosomally stable, near diploid, morphologically normal, infinite life span cell strain, designated MSU-1.1, with its normal, finite life span, precursor cell strain and obtained finite life span hybrids, as expected if infinite life span in culture is a recessive trait. However, 14 of the 14 hybrids from our fusions of MSU-1.1 cells with representative cell lines from each of the four immortality complementation groups, and 38 of the 39 hybrids from our fusions of infinite life span cells that have been reported to complement each other, failed to exhibit finite life spans. This result suggests that infinite life span cells cannot complement each other to yield finite life span hybrids. In examining this unexpected result, we obtained evidence that long-term dual drug selection can be deleterious to hybrid cells even though they carry resistance markers for both drugs, indicating that the cell death of such hybrids observed in other studies may have resulted from the cytotoxic effect of long-term drug selection, rather than from senescence. © 1994 wiley-Liss, Inc.
    Additional Material: 3 Ill.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1002/jcp.1041590119
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