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  • 1980-1984  (4)
  • 1
    ISSN: 1572-9931
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology , Medicine
    Notes: Abstract Thymidine auxotrophic mutants were selectively isolated from mutagenized mouse FM3A cells by resistance to methotrexate in the presence of thymidine and 5-methyl-tetrahydrofolate with a frequency of 10−5−10−6. In most of the thymidine auxotrophs the activity of thymidylate synthase was very low or undetectable, but dihydrofolate reductase activity was normal. Upon starvation of thymidine, the mutant cells immediately stopped growing and started to lyse within one day. In the presence of thymidine, the mutant cells grew quite normally. This phenotype behaved recessively in cell—cell hybrids, and the segregation profile of its marker indicated that the lesions in the mutants are not linked to the X chromosome. Prototrophic revertants could be isolated from these mutants, and they showed almost the normal level of thymidylate synthase activity. The selection method described here should be useful for isolating large numbers of thymidylate synthase-negative mutants from various mammalian cell lines.
    Type of Medium: Electronic Resource
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  • 2
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 112 (1982), S. 83-88 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: We established a clonal preadispose cell line from newborn mouse calvaria. Cells of this cell line, designated MC3T3-G2/PA6, had the capacity to convert to adipose cells, to accumulate triglycerides in their cytoplasm, and to mature to differentiated fat cells in a resting state. This adipose conversion was markedly accelerated by addition of dexamethasone, which was the most potent inducer among the steroid hormones tested. The presence of dexamethasone was needed during logarithmic growth phase for maximal conversion. The frequency of adipose conversion was dependent on exposure time to the hormone, but cells already committed to differentiation continued to accumulate lipid and developed into mature adipose cell even in its absence. This indicates that the hormone accelerates the initiation of the adipose conversion, but is not required for the ongoing conversion process. In fact, it was rather inhibitory for the process of fat accumulation. Insulin alone slightly inhibited the adipose conversion, but its combination with dexamethasone neutralized the above inhibitory effect of dexamehasone. The responsiveness of this cell line is consistent with that observed for mouse bone marrow preadipocytes in primary culture but differs from that for preadipose cell lines derived from extramedullary tissues. These results strongly suggest that the MC3T3-G2/PA6 cell line was derived from bone marrow.
    Additional Material: 5 Ill.
    Type of Medium: Electronic Resource
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  • 3
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 112 (1982), S. 89-95 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: A clonal preadipose cell line MC3T3-G2/PA6, established from newborn mouse calvaria, responds to glucocorticoids and converts to adipose cells in a fashion similar to bone marrow preadipocytes. We investigated the effect of the cells on in vitro hemopoiesis of mouse bone marrow cells by cocultivation. When bone marrow cells were inoculated into confluent cultures of MC3T3-G2/PA6 cells (104-106 cells/25-cm2 flask), the number of hemopoietic stem cells (CFU-S) significantly increased during 7-day cultivation in proportion to inoculum size. Under these conditions, active replication of CFU-S was maintained for several weeks until MC3T3-G2/PA6 cell layers detached from the substratum. This capacity of the MC3T3-G2/PA6 line was unique because other established cell lines, including the MTF preadipose line, failed to support CFU-S growth. When bone marrow cells were not allowed to contact the MC3T3-G2/PA6 cell layer, only a small number of CFU-S survived for 7 days. Moreover, MC3T3-G2/PA6 cell-conditioned medium did not show any growth-promoting activity for CFU-S. These results indicate that the MC3T3-G2/PA6 cell line has the ability to promote the proliferation of CFU-S through a short range cell-to-cell interaction by providing an in vitro microenvironment probably similar to that for in vivo hemopoiesis.
    Additional Material: 5 Ill.
    Type of Medium: Electronic Resource
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  • 4
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 118 (1984), S. 233-240 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The clonal preadipose cell line, MC3T3-G2/PA6, has the capacity to differentiate into adipocytes in response to glucocorticoids and to support in vitro growth of hemopoietic stem cells (CFU-S). To study the relationship between these capacities, we precultured the MC3T3-G2/PA6 cells for varying days in the presence or absence of dexamethasone and then cocultured them with mouse bone marrow cells. Logarithmically growing cultures contained no detectable adipocytes and showed the highest growth-supporting activity for CFU-S, whereas cultures containing the largest number of adipocytes showed the lowest activity. When bone marrow cells were seeded onto 3-day-old MC3T3-G2/PA6 preadipocyte layers at 1 × 105 cells/35-mm dish, day 12 CFU-S grew with a population doubling time of about 37 hr, and at least 75% of them were associated with the cell layer between days 2 and 7. In the absence of the preadipocytes, CFU-S were not detected in the adherent cell fraction and decreased with a half-life of about 18 hr. More than 80% of CFU-C were also found to be associated with the preadipocyte layer, and they increased about 24-fold in number during 7 days in culture. Morphologically, hemopoietic cells developing into mature granulocytes and macrophages were distributed between the layers of preadipocytes. Dendritic processes of preadipocytes were frequently in close alignment with the hemopoietic cells. However, adipocytes failed to show such an intimate association with hemopoietic cells. These results indicate that MC3T3-G2/PA6 cells in the preadipocyte stage, but not in the adipocyte stage, have the capacity to support CFU-S growth, and that hemopoiesis in our cocultivation system proceed within the microenvironmental milieu provided by MC3T3-G2/PA6 preadipocytes.
    Additional Material: 6 Ill.
    Type of Medium: Electronic Resource
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