Interaction Between X-Irradiated Plateau-Phase Bone Marrow Stromal Cell Lines and Co-Cultivated Factor-Dependent Cell Lines Leading to Leukemogenesis in Vitro

Plateau-phase mouse clonal bone marrow stromal cell lines D2XRII and C3H c1 II produce decreasing levels of M-CSF (CSF-I), a specific macrophage progenitor cell humoral regulator, following x-irradiation in vitro. The decrease did not go below 40% of control levels, even after irradiation doses of 50,000 rad (500 Gy). In contrast, a distinct humoral regulator stimulating growth of GM-CSF/IL-3 factor-dependent (FD) hematopoietic progenitor cell lines was detected following radiation to doses above 2,000 rad. This humoral factor was not detectable in conditioned medium from irradiated cells, weakly detected using factor-dependent target cell populations in agar overlay, and was prominently detected by liquid co-cultivation of factor-dependent cells with irradiated stromal cell cultures. Subclonal lines of FD cells, derived after co-cultivation revealed karyotypic abnormalities and induced myeloblastic tumors in syngeneic mice. Five - eight weeks co-cultivation was required for induction of factor independence and malignancy and was associated with dense cell to cell contact between FD cells and stromal cells demonstrated by light and electron microscopy. Increases in hematopoietic to stromal cell surface area, total number of adherent cells per flask, total non-adherent cell colonies per flask, and cumulative non-adherent cell production were observed after irradiation. The present data may prove very relevant to an understanding of the cell to cell interactions during x-irradiation-induced leukemia

Multiple biologic activities of a cloned inducer T-cell population.

The mouse T-cell clone Ly1+2(-)/9, belonging to the Ly1 set, displays the following functions in vitro: (i) augmentation of immunoglobulin output by B cells; (ii) stimulation of bone marrow cells to produce colonies composed of granulocytes, macrophages, or both; and (iii) proliferative stimulation of T-cell clones belonging to other Ly sets. These functions are induced by Ly1+2(-)/9 cells themselves and by supernatants of Ly1+2(-)/9 cultures and are not evinced by tested clones belonging to other Ly sets. The agent or agents responsible for colony formation and for B-cell stimulation had an apparent molecular weight of 45,000-50,000 and could not be physically separated. The T-cell stimulating agent(s) had an apparent molecular weight of 30,000 and could be separated from the agent(s) that acts upon colony formation and B cells. Thus, clone LY1+2(-)/9 produces at least two soluble products that induce or augment activities of at least three other differently programmed cell sets

Enhancer sequences of a retroviral vector determine expression of a gene in multipotent hematopoietic progenitors and committed erythroid cells.

To analyze the transcriptional activity of retroviral enhancer sequences in hematopoietic lineages, we determined the effect of enhancer sequences on the expression of the neomycin resistance gene transferred by two retroviral vectors to primary hematopoietic lineages. We constructed the vector pFr-SV(X). The Moloney murine leukemia virus enhancer region of a vector, pZIP-SV(X), was replaced by a 380-nucleotide-long fragment containing the enhancer sequences of the Friend murine leukemia virus. The enhancer sequences of Friend murine leukemia virus were used because these sequences have been shown to target the disease specificity of the virus to the erythroid lineage. Hematopoietic progenitors in murine continuous marrow cultures were infected with identical numbers of pure defective, infectious viral vector particles of either pFr-SV(X) or pZIP-SV(X). Expression of the transferred neomycin resistance gene in multipotential stem cells and their differentiated progeny was assayed as the ability of infected progenitors to form colonies (greater than 50 cells) in G418. Expression of the neomycin resistance gene in multipotential progenitor cells during the entire 11 weeks of the cultures was independent of the vector used to transfer the gene. Conversely, committed hemoglobinized erythroid bursts and myeloid colonies resistant to G418 were consistently produced by pFr-SV(X)-infected cultures but not pZIP-SV(X)-infected cultures. These results demonstrate that both pFr-SV(X) and pZIP-SV(X) were stably integrated and expressed in more primitive, multilineage, hematopoietic progenitor cells and suggest that the enhancer sequences of a vector affects expression of the transferred neomycin resistance gene when these cells differentiate to committed myeloid and erythroid cells