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E Spooncer

Publications and source records attributed to E Spooncer.

64 records · Page 4Linked to original sources
PubMed1984

Haemopoietic stem cells and the problem of self-renewal.

Haemopoiesis occurs in association with a complex stromal cell network in which all levels of haemopoietic cell development can be found. In order to understand the interaction between stromal cells and growth factors with the processes of self-renewal and differentiation, we have carried out a series of experiments attempting to define the circumstances in which self-renewal occurs in long-term marrow cultures. We have found that highly purified (FACS sorted) CFU-S do not undergo significant self-renewal in vitro when inoculated onto marrow stromal cells that can support self-renewal of unfractionated CFU-S. We have examined the effects of expression of the src oncogene on self-renewal of CFU-S. We have found that, following infection of long-term cultures with a retrovirus carrying the src oncogene, there is expression of src in certain of the stromal cells. There is also a selection for CFU-S that have an extended self-renewal capacity in vivo and in vitro. These CFU-S are non-leukaemic and can reconstitute haempoiesis in irradiated mice. Cells from src-infected cultures can also be induced to proliferate and form cell lines in vitro in the presence of interleukin 3 (IL-3). The cell lines produced are multipotential and non-leukaemic. From such data we conclude that expression of the src oncogene has (directly or indirectly) permanently altered the stem cells in such a way that they can undergo extensive self-renewal in situations that are unfavourable for growth and self-renewal of normal stem cells.

Animals↗
PubMed1983

Regulation of haemopoiesis in long-term bone marrow cultures. IV. Glycosaminoglycan synthesis and the stimulation of haemopoiesis by beta-D-xylosides.

Sulfated glycosaminoglycans (GAGs) are distributed in consistent and distinctive patterns between the cell surface and the growth medium of haemopoietically active long-term bone marrow cultures. Heparan sulfate is the main cell surface component and chondroitin sulfate is the major sulfated species in the medium. When the cultures are supplemented with beta-D-xylosides a significant increase in chondroitin sulfate synthesis is observed but no stimulation of heparan sulfate synthesis occurs. The chondroitin sulfate accumulates in the culture medium in beta-D-xyloside-treated cultures but the composition of sulfated GAGs in cell-surface derived material is unaffected. beta-D-xylosides also stimulate the production of haemopoietic cells without any apparent alteration in the adherent stromal cells of the marrow cultures. Equivalent increases are obtained in cells at all stages of development so that a fivefold increase in pluripotent stem cells (CFU-S) is matched by fivefold increase in the granulocyte-macrophage progenitors (GM-CFC) and in mature granulocytes. The stimulation persists for many weeks in beta-D-xyloside-treated cultures. These results indicate that the sulfated GAGs may play an important role in the regulation of haemopoiesis.

Animals↗
PubMed1983

Role of the cellular matrix in haemopoiesis. I. Synthesis of glycosaminoglycans by mouse bone marrow cell cultures.

Haemopoietically active mouse bone marrow cultures, incubated for 48 h with [3H]glucosamine and Na2(35)SO4, synthesized radiolabelled hyaluronic acid, heparan sulphate and chondroitin sulphate. Heparan sulphate was enriched in a trypsin extract of the adherent cells whereas hyaluronic acid and chondroitin sulphate were distributed mainly to the culture medium. Analysis of nitrous acid scission products of heparan sulphate by gel chromatography demonstrated the close association of N- and O-sulphate groups along the polysaccharide chain. Chondroitinase AC degradation established the copolymeric nature of chondroitin sulphate in which about 38% of the hexuronic acid residues were in the form of GlcUA. Studies on non-haemopoietic cultures, derived from W/Wv mice or from normal marrow cells maintained in foetal calf serum instead of horse serum, indicated that adherent stromal cells were the major source of glycosaminoglycans.

Animals↗
PubMed1980

The role of cells and their products in the regulation of in vitro stem cell proliferation and granulocyte development.

In long-term marrow cultures haemopoiesis can be maintained in vitro for up to 6 months. Critical analysis of the cell populations produced has shown that the stem cells and their committed progeny have characteristics in common with the corresponding cell types in vivo. The maintenance of haemopoiesis in vitro is associated with the development of an appropriate inductive environment provided by bone marrow derived adherent cells. Analysis of the interactions between environmental and haemopoietic cells has been facilitated by the development of vitro systems reproducing the naturally occurring genetic environmental defects and other systems where the development of a competent inductive environment shows a dependency upon corticosteroid hormones. Investigations have shown that stem cell proliferation may be controlled by production of opposing activities, one stimulatory for DNA synthesis, the other inhibitory. A model is proposed whereby modulation in the production of these factors is determined by the physical presence of stem cells in a proposed cellular milieu, within the adherent layer. The adherent layer, apart from acting at the level of stem cell proliferation, can also modify the response of differentiating cells (eg, GM-CFC) to exogenous stimulatory activities. Addition of GM-CSF or of CSF-antiserum has no effect on haemopoiesis in long-term cultures.

Animals↗
PubMedDate not supplied

Continuous in vitro generation of multipotential stem cell clones from src-infected cultures.

A molecular recombinant of Rous sarcoma virus and murine amphotropic leukaemia virus, src(MoMuLV), where the avian src oncogene has been placed under the influence of a murine virus promoter sequence, has been reported. Infection of long-term marrow cultures with this virus led to a dramatic change in the relative numbers of stem cells, granulocyte-macrophage progenitor cells and mature cells found in normal haematopoietic cell development. However, although the balance between self-renewal, differentiation and development was disturbed, injection of the cultured cells into irradiated syngeneic recipients did not lead to the development of leukaemia. Thus, although the control had been 'loosened', the host regulatory mechanisms were sufficient to impose a restraint on unlimited growth of the cells. We now show that the stem cells from the src-infected cultures show a remarkably increased capacity for self-renewal in vitro in situations which are inimical to the maintenance of self-renewal in normal uninfected stem cells and that self-renewal/differentiation can be modified by the culture conditions.

Animals↗
PubMedDate not supplied

Defective ability to self-renew in vitro of highly purified primitive haematopoietic cells.

Stromal cells play a critical role in haematopoiesis, both in a permissive and, probably, in a directive manner. Study of the interactions between stromal cells and haematopoietic stem cells, however, is difficult to perform using whole bone marrow, in which stem cells are indistinguishable from precursor cells and maturing haematopoietic cells, and where stromal and haematopoietic cells co-exist in a heterogeneous mixture. We have purified primitive haematopoietic spleen colony-forming cells (CFU-S) using fluorescence-activated cell sorting (FACS) and produced CFU-S populations which approach 100% purity (ref. 6 and B.I.L. and E.S., in preparation). This cell population is devoid of significant stromal cells and mature haematopoietic cells. Here, we report that when purified CFU-S are seeded onto a stromal adherent layer in vitro, foci of haematopoietic cells develop within the stroma followed by production of a wave of maturing and mature progeny. However, self-renewal of CFU-S does not occur and haematopoietic activity rapidly declines, indicating that caution should be applied in the use of highly purified stem cells for human bone marrow transplantation.

Bone Marrow Cells↗