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  • 1
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 91 (1977), S. 335-344 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: A liquid culture system is described whereby proliferation of haemopoietic stem cells (CFU-S), production of granulocyte precursor cells (CFU-C), and extensive granulopoiesis can be maintained in vitro for several months. Such cultures consist of adherent and non-adherent populations of cells. The adherent population contains phagocytic mononuclear cells, “epithelial” cells, and “giant fat” cells. The latter appear to be particularly important for stem cell maintenance and furthermore there is a strong tendency for maturing granulocytes to selectively cluster in and around areas of “giant fat” cell aggregations. By “feeding” the cultures at weekly intervals, between 10 to 15 “population doublings” of functionally normal CFU-S regularly occurs. Increased “population doublings” may be obtained by feeding twice weekly. The cultures show initially extensive granulopoiesis followed, in a majority of cases, by an accumulation of blast cells. Eventually both blast cells and granulocytes decline and the cultures contain predominantly phagocytic mononuclear cells.Culturing at 33°C leads to the development of a more profuse growth of adherent cells and these cultures show better maintenance of stem cells and increased cell density.When tested for colony stimulating activity (CSA) the cultures were uniformly negative. Addition of exogenous CSA caused a rapid decline in stem cells, reduced granulopoiesis and an accumulation of phagocytic mononuclear cells.
    Additional Material: 4 Ill.
    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 111 (1982), S. 177-186 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: We have isolated continuously growing cell lines derived from mouse bone marrow stroma. These cell lines were independently obtained, and though they showed morphologies ranging from the epithelioid to the fibroblastoid patterns, they all differentiated into adipocytes. Subclones obtained from two cell lines had a very high frequency (90-100%) of differentiation into adipocytes after two or three weeks of arrested growth. Though extensive accumulation of lipid often mechanically impaired mitosis, the cells committed to adipocytes did not suffer an irreversible loss of proliferative capacity. Adipogenesis was obtained in conditions similar to those required for fat cell formation in long-term bone marrow culture. The cell lines were found to be insensitive to insulin as a signal of adipocyte differentiation. The ultrastructural characteristics of the preadipocytes and fat cells are also similar to those of the fat cells developing in long-term bone marrow culture. As such, these cell lines should prove useful for analysing cell/cell interactions in haemopoiesis.
    Additional Material: 5 Ill.
    Type of Medium: Electronic Resource
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  • 3
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: With the aid of a multipotent stem cell line (FDCP-mix cells) co-cultured with either normal or irradiated Swiss 3T3, cellular interactions between stromal cells and haemopoietic stem cells were studied by electron microscopy and time-lapse video microscopy. When cultured in the presence of interleukin 3 (IL-3) but in the absence of stromal cells, the FDCP-mix cells have a characteristic blast morphology. In the absence of IL-3, the cells die unless they are co-cultured with marrow stromal cells or 3T3 cells. In the latter case, they attach, proliferate, and differentiate on both normal and irradiated Swiss 3T3 cell layers without the addition of extrinsic growth factor (IL-3). At the initial attachment sites of these two cell lines, cellular recognition seemed to be mediated by the formation of microvillus cytoplasmic projections and extracellular matrix. These areas may well be the sites of plasma-membrane-bound signalling/adhesional molecules between the interacting cells.
    Additional Material: 16 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 82 (1973), S. 461-473 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: A liquid culture system, for haemopoietic cells, has been developed using bone marrow cells alone, or co-cultures of thymus and bone marrow cells, inoculated into four ounce medical bottles. After several days growth, such cultures consisted of an attaching population of cells, forming discrete colonies, and a non-attaching population. In the (co-cultures) there was a 2 X enhancement of monolayer colony development compared with the combined total present in the (marrow alone) plus (thymus alone) cultures. Also, better maintenance of non-attaching cells was seen in the (co-cultures). Normal CFUS and CFUC were present in both the (marrow alone) and the (co-cultures) for at least 14 days.In the (marrow alone) cultures, granulocytes in all stages of development were present for the first week, but by 12 days the culture consisted mainly of mono-nuclear cells. In the (co-cultures), however, at 12 days more than 60% of the cells were granulocytes, in all stages of differentiation. (Co-cultures) established using lethally irradiated thymus cells were not able to support this prolonged myeloid differentiation.By feeding the (co-cultures) it was possible to maintain production of (granulocytic) cells for at least ten weeks, although no fully mature granulocytes were observed. After the second feeding, no CFUS were detectable, but variable numbers of agar colony forming cells (not classical CFUC) were present at least for ten weeks.
    Additional Material: 10 Ill.
    Type of Medium: Electronic Resource
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  • 5
    ISSN: 0002-9106
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: The introductory review amplifies the finding that simply holding lymphocytes in vitro reversibly compromises their ability to enter lymph nodes from the blood, although entry into the spleen is unaffected. The differential migration of T and B lymphocytes from the blood, lymphocyte traffic in athymic rats, and the secretion of a sulphated glycoconjugate by high endothelial cells in lymph nodes are also discussed.Original data are presented concerning the effects of varying the conditions under which lymphocytes are held in vitro (time, temperature, medium, centrifugation) on their ability to enter lymph nodes and also on their surface morphology. In general, conditions that reduced the number of microvilli and induced surface blebbing also tended to affect the delicate function of crossing specialized vascular endothelium; but there was no simple relationship between morphology and migratory behavior. The localization of lymphocytes to the bone marrow was augmented by holding them in vitro, and this effect was greater after holding at room temperature (RT) than at 0°C, in contrast to impaired entry into lymph nodes. Small amounts of heparin (10 units) injected along with lymphocytes significantly reduced early localization in lymph nodes. These findings have practical implications for the design of lymphocyte traffic experiments and are relevant to the mechanism of lymphocyte attachment to vascular endothelium, since the well-known effect of trypsinizing lymphocytes can be reproduced by maintenance in vitro.
    Additional Material: 4 Ill.
    Type of Medium: Electronic Resource
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  • 6
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: When highly enriched multipotential spleen colony forming cells (CFU-S) obtained following fluorescence activated cell sorting (FACS-CFU-S) are cultured on marrow stromal cells, they undergo proliferation and development to produce mature haemopoietic cells (Spooncer et al., Nature, 316:62-64, 1985). We now show that FACS-CFU-S behave in a similar way when cultured on monolayers of 3T3 cells, indicating that the 3T3 cells can supply at least part of the environment which is representative of marrow stromal cells and provide, therefore, a system for studying stromal cell: haemopoietic cell interactions. We also demonstrate that IL-3-dependent multipotential stem cell lines (FDCP-Mix), but not a variety of other “committed” IL-3-dependent cell lines, resemble FACS-CFU-S in terms of their ability to proliferate and differentiate when cultured on 3T3 cells in the absence of IL-3. In this system, attachment of the FDCP-Mix to the 3T3 cells is critical for the subsequent maintenance of viability and stimulation of development of the cells. When the FDCP-Mix cells are physically separated from the 3T3 cells, they die and their death cannot be prevented by using 3T3-cell-conditioned medium. The extracellular matrix generated by 3T3 cells is not sufficient for promoting attachment or viability of the FDCP-Mix cells, indicating the importance of integral membrane components. However, attachment and development of FDCP-Mix cells occurs on 3T3 cells that have been lightly fixed with glutaraldehyde indicating that active metabolism is not essential for the effects promoted by the 3T3 cells. We suggest that the ability of FACS-CFU-S and FDCP-Mix cells to respond to 3T3 cells involves specific ligand/receptor interactions.
    Additional Material: 11 Ill.
    Type of Medium: Electronic Resource
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