Differential expression of subspecies of polyomavirus and murine leukemia virus enhancer core binding protein, PEBP2, in various hematopoietic cells.

The core sequence of the enhancer of murine leukemia virus (MuLV) long terminal repeat is highly conserved in a large number of MuLV strains and appears to play an essential role when SL3-3 or Moloney strains induce T cell lymphoma in mice. We found by using the electrophoretic mobility shift assay that a polyomavirus enhancer core-binding protein, PEBP2, bound to this core motif of MuLV. We also noted that PEBP2 in several hematopoietic cell lines derived from B lymphocyte, macrophage and myelocyte lineages migrated significantly faster than the authentic PEBP2 detected in NIH3T3 fibroblasts. Interestingly, PEBP2 detected in the cell lines of T lymphocyte lineage appeared to contain both types, which were indistinguishable in electrophoretic mobility from those of NIH3T3 and of B lymphocyte, macrophage and myelocyte lineages. The treatment of the nuclear extract containing PEBP2 with phosphatase generated PEBP3, which is a subcomponent of PEBP2 and retained the same DNA-binding specificity as PEBP2. The altered mobility of hematopoietic cell-derived or T lymphocyte-derived PEBP2 was found to be due to the alteration of the mobility of PEBP3. Based on the distinct mobility of PEBP2/3 of T lymphocytes from those of other hematopoietic cells, we discuss the implication of PEBP2 in MuLV-induced T cell leukemia and T cell-specific gene expression.

Microbial flora in the gastrointestinal tract abolishes cytostatic effects of alpha-difluoromethylornithine in vivo.

Although treatment with the ornithine decarboxylase inhibitor alpha-difluoromethylornithine (DFMO) leads to depletion of intracellular polyamines and to related growth inhibition in vitro, its cytostatic effects in vivo are disappointing. This may be due to abolition of DFMO-induced growth inhibition by polyamines released during normal body cell turnover, to dietary polyamines, or to putrescine synthesized by the microbial flora in the GI tract. We studied selectively (aerobic) and totally (aerobic + anaerobic) GI tract-decontaminated LI210-bearing mice fed with 3 types of diet differing in their polyamine and carbohydrate residue contents and treated with combinations of intraperitoneal DFMO and oral deuterium-labelled putrescine. Our data show that, irrespective of diet type, total decontamination markedly potentiates the moderate tumor growth inhibition that is caused by DFMO alone. During total decontamination, growth-inhibited L1210 cells accumulate in the G0/G1 phase of the cell cycle. Although orally administered deuterium-labelled putrescine gave rise to deuterium labelling of L1210 putrescine, spermidine and spermine, the polyamine levels in our diets played only a minor role.

Induction of tumor resistance in mice by L1210 leukemia cells persistently infected with HVJ (Sendai virus).

A temperature-sensitive strain of Sendai virus, HVJ-pi, showed little or no cytopathic effect and led to establishment of carrier cultures in several cell lines. By the use of this characteristic, L1210 leukemia cells persistently infected with HVJ-pi (L1210/c--HVJ-pi) was established, almost all of which were positively stained with fluorescent HVJ antibody. They are viable and grow almost equally as uninfected L1210 leukemia cells in vitro. Athymic nude mice (BALB/c, nu/nu), deficient of T-cells, died from intraperitoneal inoculation of L1210/c--HVJ-pi cells as well as by uninfected L1210 leukemia cells. However, viable L1210/c--HVJ-pi cells showed lower transplantability in normal syngeneic mice. This immunological mechanism of rejection was explained by the modification of cell surface membrane due to HVJ-pi infection. The mice which survived the inoculation of 10(5) L1210/c--HVJ-pi cells were able to reject 10(5) uninfected L1210 leukemia cells challenged subsequently. The induction of immune resistance was more prominent in (C57BL/6 x DBA/2)F1 mice or (BALB/c x DBA/2)F1 mice than in DBA/2 mice.

Vesicular stomatitis virus-infected L1210 murine leukemia cells: increased immunogenicity and altered surface antigens.

Homogenates of L1210 cells infected in vitro with vesicular stomatitis virus (VSV) were immunogenic agains a tumor graft of 100 times the LD50 dose of L1210 cells" whereas those of uninfected cells were not. The immunogenicity of intact X-irradiated L1210 cells was distinguishable from that of VSV-infected cell homogenates on the basis of the susceptibility of immunogenicity to experimental procedures used in preparation of the immunogenic homogenates: Homogenization of intact X-irradiated cells or their infection with VSV prior to irradiation led to loss of immunogenicity. In addition, uninfected cell homogenates were not made immunogenic nor was the immunogenicity of VSV-infected cell homogenates eliminated by X-irradiation. At the time of tumor challenge, sera from mice that were effectively immunized with VSV-infected cell homogenate showed a high VSV-neutralizing titer but no complement-dependent cytotoxicity for L1210 cells. Quantitative absorption studies demonstrated that VSV infection led to a marked reduction in L1210 surface antigens recognized by cytotoxic alloantibody; spatial association between these antigens and VSV antigens was not demonstrable on VSV-infected cells. Antigens recognized by heterologous antiserum to L1210 cells were also reduced following VSV infection.

Murine leukemia: depressed response to interferon induction correlated with a serum hyporeactive factor.

Mice injected by the intraperitoneal route with either L1210 or P388 leukemic cells progressively developed a state of hyporeactivity to interferon induction that was dependent upon inducer and time of administration. A circulating factor was detected in the serum of both L1210 and P388 leukemic mice that could transfer hyporeactivity to normal murine cells in vitro. A direct relationship existed between the concentration of serum factor and development of hyporeactivity to interferon induction in vivo. Characterization of the serum hyporeactive factor from P388 or L1210 leukemic mice indicated that both were similar to a hyporeactive factor previously detected in serum from virus-infected mice. These results suggest a cause-effect relationship between the serum hyporeactive factor and development of hyporeactivity in vivo.

Ultrastructure and virus particles of L1210/Mes. A hamster-adapted murine leukemia.

The ultrastructural features and virus particle content of L1210/Mes., the hamster-adapted strain of murine leukemia L1210, has been studied in cheek pouch tumors and cell cultures. In comparison with the parental murine tumor in culture, L1210/Mes. has experienced certain morphologic alterations, such as a striking increase in cell size, enlarged and more bizarre nuclei, and an increase in the number of lysosome-related structures. Type A (intracytoplasmic and intracisternal) and type C virus particles could be demonstrated in cell cultures of murine leukemia L1210, and in cheek pouch grafts and cell cultures of L1210/Mes. No. "H" (or "R") hamster-associated virus particles, however, could be distinguished in L1210/Mes. tumors or in vitro cultivated cells. It appears that a murine lymphoma capable of serial propagation in a xenogeneic host permits the replication of virus particles structurally indistinguishable from those indigenous to it.

Identification of the virions in the in vitro L1210(V) leukemia cell lines by morphological, virological, and immunological techniques.

In vitro L1210 (V) cell lines contained abundant intracytoplasmic A-particles, numerous C-type particles, a small number of B-type particles, and occasional intracisternal A-particles. The intracytoplasmic A-particles were incorporated into both spiked (B-type) and smooth-surfaced (C-type) particles formed at the budding site. Both B-and C-type particles also developed by gradual accululation of neucleooid material. The particles, particularly the C-type, exhibited a wide range of densities. The cells showed strong surface immunofluoresence for both murine mammary tumor virus and Gross murine leukemia virus antigens and variable degrees of cytoplasmic immunoflurescence for the protein antigens (p1 to p6) of Rauscher leukemia virus. The cells, the culture supernatant, and the purified virus each gave positive reactions with murine mammary tumor virus and murine leukemia virus antisera by immunodiffusion. The viral particles failed to infect C57BL, C57BL/6 X DBA/2F1 (hereafter called BD2F1), BALB/c, Af,and RIIIf mice. Howver, the cells were highly tumorigenic in BD1F-1 mice, moderately tumorigenic in BALB/c mice, but not tumorigenic in C57BL, Af, and RIIIf mice.