Cellular site and mode of Fv-2 gene action

The Fv-2 genotype of erythroid progenitors directly determines whether they will undergo viral-induced transformation. This conclusion was reached from studies of allophenic mice compounded from congenic C57BL/6 strains differing at Fv-2 and an enzyme marker (GPI). Infection of these Fv-2ss in equilibrium Fv-2rr mosaic animals with the polycythemic strain of Friend virus results in the development of Friend disease. Concomitant with disease symptoms is a shift in the mosaic composition of the erythrocytes in favor of those of the susceptible strain. The observed viral-induced shift in the erythrocyte composition is paralleled by a similar change in the mosaic composition of the CFU-E pool but not the primitive (d8) BFU-E pool. Thus, with regard to this particular Fv-2 phenotype (susceptibility to FV-P-induced cellular hyperplasia), Fv-2 manifests itself specifically in the erythroid lineage, either in mature (d3) BFU-E or CFU-E.     

Testosterone action on BFU-E and CFU-E: correlation between enhancement of proliferative rate and amplification of pool size.

As previously reported, a single administration of testosterone propionate (TP) in ex-hypoxic polycythemic mice induces an 18–24 hr amplification of the erythroid burst-forming unit (BFU-E) pool and a 60-hr expansion of the erythroid colony-forming unit (CFU-E) compartment. Both phenomena are here shown to be temporally associated with an increase of the $\text{in vitro}$³H-TdR sensitivity of these compartments, thus indicating an elevation of their proliferative rate. On the other hand, no significant modification of both the DNA synthesis index and the pool size of BFU-E and CFU-E were observed at respectively 60 or 18 hr. At either time interval, both ³H-TdR sensitivity and compartment size were not modified at the level of the myeloid-macrophage colony-forming unit (CFU-C). It is therefore suggested that the early and late expansion of respectively BFU-E and CFU-E number after TP injection is at least partially mediated by enhancement of the proliferative rate within the respective compartments. Finally, mechanisms underlying TP action on BFU-E and CFU-E pools are discussed in the light of both present and previous observations.     

Defective Friend Spleen Focus-Forming Virus: Interfering Properties and Isolation Free from Standard Leukemia-Inducing Helper Virus

Defective Friend spleen focus-forming virus (SFFV) is able to interfere with the ability of its naturally occurring leukemia-inducing helper virus (LLV-F) to induce XC plaque formation in several different strains of mouse embryo cells. This interference has been observed by using two different SFFV preparations, one contained in an NB-tropic stock of Friend virus (FV) complex, and the second present in a C57BL-adapted strain of FV complex containing an associated B-tropic LLV-F helper. The LLV-F in NB-tropic FV complex effectively induced XC plaques in C57BL/6 (Fv-1^bb; Fv-2^rr) mouse embryo fibroblasts (MEF) only in the absence of coinfecting SFFV, indicating that Fv-2-associated resistance to SFFV-induced focus formation in vivo does not necessarily extend to the restriction of SFFV function(s) in vitro (i.e., in Fv-2^rr C57BL MEF). SFFV interference appears to be an intracellular event since LLV-F can adsorb onto, penetrate, and rescue defective murine sarcoma virus (MSV) from transformed 3T3FL S⁺L⁻ cells with equal efficiency in the presence and absence of SFFV. However, significantly fewer LLV-infected S⁺L⁻ cells released LLV-F progeny if SFFV was present. These observations suggest that Friend SFFV may be classified as a defective, interfering (DI) particle. Further support for this conclusion has come from studies designed to investigate two physical properties of defective SFFV particles. SFFV layered onto a 0 to 20% sucrose sedimentation gradient was recovered as a symmetrical band of virus that sedimented more slowly than standard LLV-F particles. Pooled SFFV-containing gradient samples contained visualizable type C virus particles and occasionally small amounts of detectable LLV-F. In an attempt to determine the buoyant density of sedimentation gradient-purified SFFV, pooled SFFV samples were layered onto a 25 to 50% sucrose equilibrium density gradient and were centrifuged to equilibrium. Greater than 50% of the infectious SFFV originally layered onto this gradient was recovered and seen as a narrow symmetrical band with peak SFFV infectivity at a sucrose density of 1.14 g/ml. The observed difference between SFFV and LLV-F buoyant densities appears to be related to an inherent physical property of each virus. Mixtures of these two viruses express the buoyant density of that virus population which is in excess in fabricated FV complexes probably due to the formation of SFFV-LLV aggregates. Finally, gradient-purified SFFV failed to induce XC plaques in MEF and did not function to rescue MSV as expected since SFFV itself is replication defective.     

Characterization of the early and late stages of myelomonocytic leukemia induced by Friend helper-independent virus F-MuLV: isolation and induction of Friend myelomonocytic leukemic cell lines

With the use of cloned helper-independent Friend leukemia virus (F-MuLV), we have induced a high incidence (approximately 70%) of myelomonocytic leukemia in mice resistant (Fv-6rr or Fv-6rs) to erythroleukemia induction by this virus. The spleen cells from these mice (DBA/2 or BALB/c X DBA/2) were found to contain a high level of progenitor cells capable of forming granulocytic and macrophage colonies (CFU-GM). These CFU-GM, however, were different from those in the spleens of uninfected mice, as they were either very sensitive to or independent of conditioned medium. No erythroid progenitor bursts (BFU-E) or precursor (CFU-E) cells were detected in the spleens of these diseased animals. If these mice with myelomonocytic leukemia were kept alive by transfusion of red blood cells from uninfected mice, tumorigenic cell lines, capable of being transplanted, into adult mice can be isolated. Three such cell lines TTA-1, TTA-3, and TTA-9 have been established, and they retain their morphology of monocytes and macrophages as well as being positive for the monocyte-specific stain alpha-naphthyl-acetate esterase. These myelomonocytic leukemia cell lines can also be induced in culture by spleen cell-conditioned medium to differentiate into macrophages. Other conditioned media such as L-cell-conditioned medium, phytohemagglutinin-stimulated leukocyte-conditioned medium, and WEHI-3 conditioned medium were less effective in their abilities to stimulate differentiation in these myelomonocytic leukemia cell lines.     

Differentiation of lymphocytes in mouse bone marrow. II. Kinetics of maturation and renewal of antiglobulin-binding cells studied by double labeling

DNA labeling by ³H-thymidine in vitro and antiglobulin-¹³¹I binding in vitro were used to determine the development and turnover of immunoglobulin-bearing lymphocytes in mouse bone marrow.Bone marrow cells from CBA mice previously injected repeatedly with ³H-thymidine for 1–84 hr were exposed to ¹³¹I-labeled rabbit-antimouse globulin for 30 min at 0 °C, and examined radioautographically. The antiglobulin-binding cells in bone marrow were predominantly (97–98%) nondividing small lymphocytes. Some plasmacytoid and monocytoid cells, but not the proliferating large lymphoid cells, also bound antiglobulin. The ³H-thymidine labeling index of the small lymphocyte population showed a rapid exponential increase (50% in 32 hr). The first small lymphocytes to show ³H-thymidine labeling were those lacking antiglobulin-binding capacity, reaching approximately 90% ³H-thymidine labeling after 2 days. Small lymphocytes which bound antiglobulin-¹³¹I at a concentration of 1.0 μg/ml became labeled with ³H-thymidine only after a lag of approximately 1.5 days. More avid antiglobulinbinding cells were delayed a further 12 hr in ³H-thymidine labeling. During in vitro culture the proportion of antiglobulin-binding small lymphocytes increased progressively in bone marrow but decreased in spleen cell suspensions.The results demonstrate a continuous, rapid renewal of immunoglobulin-bearing small lymphocytes in adult mouse bone marrow. Surface immunoglobulin molecules are not detectable when marrow small lymphocytes are first formed, but they appear and increase progressively in density as the cells mature.     

THE ROLE OF ERYTHROPOIETIN IN REGULATION OF POPULATION SIZE AND CELL CYCLING OF EARLY AND LATE ERYTHROID PRECURSORS IN MOUSE BONE MARROW

This study was designed to determine the stage in haemopoietic cell differentiation from multipotential stem cells at which erythropoietin becomes physiologically important. The responses of haemopoietic precursor cells were monitored in the bone marrow of mice under conditions of high (after bleeding) and low (after hypertransfusion) ambient erythropoietin levels. The number of relatively mature erythroid precursors (CFU-E), detected by erythroid colony formation after 2 days of culture, increased three-fold in marrow by the fourth day after bleeding, and decreased three-fold after hypertransfusion. Assessed by sensitivity to killing by a brief exposure to tritiated thymidine (³H-TdR) in vitro, the proliferative activity of CFU-E was high (75% kill) in untreated and bled animals, and was slightly lower (60% kill) after hypertransfusion. The responses of more primitive erythroid progenitors (BFU-E), detected by erythroid colony formation after 10 days in culture, presented a contrasting pattern. After hypertransfusion they increased slightly, while little change was noted until the fourth day after bleeding, when they decreased in the marrow. The same response pattern was observed for the progenitors (CFU-C) detected by granulocyte/macrophage colony formation in culture. The sensitivity of BFU-E to ³H-TdR was normally 30%, and neither increased after bleeding nor decreased after hypertransfusion. However, in regenerating marrow the ³H-TdR sensitivity of BFU-E increased to 63%, and this increase was not affected by hypertransfusion. These results are interpreted as indicating (1) that physiological levels of erythropoietin do not influence the decision by multipotential haemopoietic stem cells to differentiate along the erythroid pathway as opposed to the granulocyte/macrophage pathway; (2) that early erythroid-committed progenitors themselves do not respond to these levels of erythropoietin, but rather are subject to regulation by erythropoietin-independent mechanisms; and (3) that physiological regulation by erythropoietin commences in cells at a stage of maturation intermediate between BFU-E and CFU-E.     

Erythropoietin responses and physical characterization of erythroid progenitor cells in Rauscher virus infected BALB/c mice.

Infection of BALB/c mice with Rauscher leukemia virus (RLV) gives rise to pronounced erythrocytopoiesis manifesting in splenomegaly and is associated with progressive development of anemia. In the spleen erythroid colony forming units (CFU-E) increase exponentially up to 800-fold that of normal levels by the third week of infection. In vitro these CFU-E are dependent on erythropoietin for colony formation, their erythropoietin requirements being higher than that of CFU-E from normal mice. Numbers of CFU-E in spleen and degree of splenomegaly in anemic RLV infected mice were also shown to be modified by red blood cell transfusion, but progression of the disease was not stopped. Erythroid burst forming units (BFU-E) were also responsive to erythropoietin. However, a small proportion of cells also formed BFU-E colonies at concentrations which did not support growth of normal marrow BFU-E. When compared to normal, CFU-E found in RLV-infected spleen have similar velocity sedimentation rates. However, buoyant density separation of leukemic spleen cells indicated that CFU-E were more homogeneous (modal density 1.0695 g/cm3) than CFU-E from normal spleen. Analysis of physical properties of CFU-E and the nonhemoglobinized erythroblast-like cells, which accumulate in the spleen showed that they differed mainly in their distribution of cell diameter. Our findings show that erythroid progenitor cells in RLV infected mice are responsive to erythropoietin in vitro. Also in vivo erythropoiesis appears to be under control of erythropoietin but other factors which lead to progression of RLV disease apparently exist. Most proerythroblast-like cells, which are characteristic of this disease, apparently lack the potential to form colonies and may be more mature than CFU-E.     

Target cells for Friend virus-induced erythroid bursts in vitro

Erythropoietin (Epo) acts on mouse bone marrow cells in vitro in plasma clot or methyl cellulose culture systems to induce the formation of single erythroid colonies, or clusters of erythroid colonies termed bursts. Our laboratory has recently reported the observation that infection of mouse bone marrow cells in vitro with the polycythemia-inducing strain of Friend virus (FV) resulted in the formation of erythroid bursts after 5 days in plasma clot culture in the absence of added Epo. We have now used this system to characterize the target cells for this FV-induced erythroid transformation. The greatest number of FV bursts were observed when marrow cells were obtained from mice whose erythropoiesis had been stimulated by bleeding or phenylhydrazine treatment. Bleeding also resulted in an increase in the number of FV bursts following the infection of spleen cells in vitro. Hypertransfusion of mice, which results in decreased erythropoiesis, yielded a reduced number of FV bursts in vitro, as did prior treatment with actinomycin D. Cell separation studies using velocity sedimentation at unit gravity showed that the cells, which give rise to FV bursts, sedimented with a modal sedimentation velocity between 5.1–8.5 mm/hr. The Epo-dependent colony-forming unit erythroid (CFU-E), which gives rise to a single erythroid colony, also sediments with a modal velocity between 5.1–8.5 mm/hr, while the Epo-dependent day 8 burst-forming unit erythroid (day 8 BFU-E) sediments with a modal velocity between 3.0–6.0 mm/hr. A 20 min incubation of marrow cells with high specific activity ³H-thymidine, prior to virus infection, resulted in a 75–80% reduction in the number of FV bursts. Mixing cells from the upper portion of the gradient, which yielded no FV bursts, with cells from an area in which high numbers of FV bursts were observed did not result in the inhibition of burst formation. These experiments indicate that the primary target cells for FV bursts in vitro are most probably erythroid precursor cells that have matured beyond the day 8 BFU-E and are closely related to the CFU-E.     

Myeloproliferative sarcoma virus: its effects on erythropoiesis in adult DBA/2J mice

Adult susceptible mice (DBA/2J) infected with MPSV (myeloproliferative sarcoma virus), a defective RNA tumour virus, develop splenomegaly and progressive disruption of the haematologic system culminating in death. The present study was specifically directed toward determining the effects of the virus on erythroid differentiation. Early and late precursor cells (erythroid burst-forming units; BFU-E and colony-forming units; CFU-E, respectively) were evaluated by the ability of bone marrow and spleen cells to form colonies of fully differentiated erythroid cells in vitro. MPSV caused substantial modification of both the BFU-E and CFU-E populations in the bone marrow and spleen of infected animals. Changes were detected in the CFU-E population preceding any significant increase in spleen weight. In the bone marrow, the proportion of CFU-E cells increased almost twofold by days 5-10 after virus infection but decreased by day 15. In the spleen, CFU-E frequency rose 40-fold by days 10-15 and then declined steadily prior to death. At the peak of CFU-E expansion, a small proportion of the population appeared to be erythropoietin (Ep) independent, although there was no evidence of a complete switch to Ep-independence which occurs in Friend virus-induced erythroleukemia. Dose-response curves showed that none of these data could be explained in terms of a changing responsiveness to Ep. However, evidence is presented that indicates that BFU-E from MPSV-infected animals lose or have a reduced requirement for burst-promoting activity (BPA) relative to normal cells although their progeny still need Ep for terminal erythroid differentiation.     

Plasmodium chabaudi AS: erythropoietic responses during infection in resistant and susceptible mice.

The course of anemia and the erythropoietic response in the bone marrow, spleen, and blood were studied during Plasmodium chabaudi AS infection in resistant C57BL/6 (B6) and susceptible A/J (A) mice. Infections in B6 mice were characterized by moderate levels of both parasitemia and anemia and survival. In contrast, A mice experienced high parasitemia, severe anemia, and high mortality rates. During the period of anemia, erythropoiesis, as measured by in vivo 59Fe incorporation, was significantly more depressed in bone marrow and more increased in the spleen in resistant B6 mice. The increase in splenic 59Fe incorporation was a function of the size of the spleen. Bone marrow CFU-E were decreased to 50% of control in both strains, while splenic CFU-E were increased twofold greater in B6 mice compared to those in A mice. However, the absolute numbers of CFU-E per spleen in the two strains were not significantly different during peak parasitemia. Bone marrow BFU-E were transiently increased before peak parasitemia whereas splenic BFU-E peaked during peak parasitemia. A mice had significantly lower numbers of BFU-E per spleen on all days except at peak parasitemia. The frequency of blood-borne BFU-E and plasma erythropoietin titers was increased earlier and to a greater extent in A mice. These results suggest that an impaired amplification of late-stage splenic erythropoiesis may be an important determinant in the severity of anemia and lethality of infection with P. chabaudi AS in A mice. Moreover, these results demonstrate that the defective amplification of splenic erythropoiesis in A mice is neither caused by a defect in the mobilization of BFU-E from the bone marrow to the spleen nor caused by a defect in erythropoietin production.     

Fate of Unintegrated Viral DNA in Fv-1 Permissive and Resistant Mouse Cells Infected with Murine Leukemia Virus

We have found that levels of unintegrated linear viral DNA were nearly identical in several Fv-1 resistant cell lines, whereas levels of closed circular viral DNA are markedly reduced in these resistant cells, to the same extent as virus production (P. Jolicoeur and E. Rassart, J. Virol. 33:183-195, 1980). To determine the fate of linear viral DNA made in resistant cells we performed pulse-chase experiments, labeling viral DNA with 5-bromodeoxyuridine and following it with a thymidine chase. 5-Bromodeoxyuridine-labeled viral DNA (HH) recovered by banding on cesium chloride gradients was sedimented on neutral sucrose density gradients or separated by the agarose gel-DNA transfer procedure and detected by hybridization with complementary DNA. Levels of linear viral DNA made in Fv-1^b/b (JLS-V9 and SIM.R) and Fv-1^n/n (NIH/3T3 and SIM) cells were found to decrease during the chase period at about the same rate in permissive and nonpermissive conditions, indicating that linear viral DNA is not specifically degraded in Fv-1 resistant cells. Levels of the two species of closed circular viral DNA made in Fv-1 permissive cells increased relative to the levels of linear DNA during the chase period. This confirmed the precursor-product relationship between linear DNA and the two species of circular DNA. In Fv-1 resistant cells, this apparent conversion of linear viral DNA into circular forms was not seen, and no supercoiled viral DNA could be detected. To determine whether the transport of linear viral DNA from the cytoplasm into the nucleus was prevented by the Fv-1 gene product, SIM.R cells were fractionated into cytoplasmic and nuclear fractions, and viral DNA was detected in each fraction by the agarose gel-DNA transfer procedure. Levels of linear viral DNA were nearly identical in both cytoplasmic and nuclear fractions of permissive or resistant cells. Circular viral DNA could be detected in the nuclear fraction of permissive cells, but not in that of resistant cells. A pulse-chase experiment was also performed with SIM.R cells. During the thymidine chase period, linear viral DNA was seen to accumulate in nuclei of both permissive and resistant cells, whereas supercoiled viral DNA accumulated only in nuclei of permissive cells. These results indicate that the Fv-1 gene product does not interfere with the transport of linear viral DNA into the nucleus. Our data also suggest that the Fv-1 restriction does not operate through a degradation process. Therefore, the Fv-1 gene product could either block the circularization of linear viral DNA directly or promote the synthesis of a faulty linear viral DNA whose defect (yet undetected) would prevent its circularization.     

Survival of erythroid burst-forming units and erythroid colony-forming units in canine bone marrow cells exposed in vitro to 1 MeV neutron radiation or X rays

Conditioned media (CM) from allogeneic stimulated cultures of light density cells (less than 1.08 g/cm3) from the peripheral blood of normal dogs were used to stimulate the growth of erythroid burst-forming units (BFU-E) in bone marrow from normal dogs. Maximum numbers of BFU-E were obtained when 5% (vol/vol) 3 X CM and 2 U/ml erythropoietin were added to plasma clot cultures of bone marrow cells. In addition, the radiation sensitivity (D0 value) was determined for CFU-E and for BFU-E in bone marrow cells exposed in vitro to 1 MeV fission neutron radiation or 250 kVp X rays. BFU-E were more sensitive than CFU-E to the lethal effects of both types of radiation. For bone marrow cells exposed to 1 MeV neutron radiation, the D0 for CFU-E was 0.27 +/- 0.01 Gy, and the D0 for BFU-E was 0.16 +/- 0.03 Gy. D0 values for CFU-E and BFU-E were, respectively, 0.61 +/- 0.05 Gy and 0.26 +/- 0.09 Gy for cells exposed to X rays. The neutron RBE values for the culture conditions described were 2.3 +/- 0.01 for CFU-E and 1.6 +/- 0.40 for BFU-E.     

Effect of Danggui Buxue Tang on immune-mediated aplastic anemia bone marrow proliferation mice

To investigate the pharmacological effects of Danggui Buxue Tang (DBT) on immune-mediated aplasia anemia mice. The model of immune-mediated aplasia anemia mice was induced by means of ⁶⁰Co γ-ray irradiation and mixed cells of thymus and lymphnode of DBA/2 mice infusion through tail vein, the parameters tested indices were as following: blood picture, bone marrow nucleated cell count (BMNC), murine colony-forming unit-megakaryocytes (CFU-GM) of bone marrow cells, murine colony-forming unit-erthroid (CFU-E) and burst forming unit-erythroid (BFU-E). The results showed that DBT could not only withstand significantly decreation of blood cells by immune-mediated, but also stimulate on the growth of bone marrow colony cell and increase the weight of hemopoietic progenitor of bone marrow. Therefore, DBT had an obvious treat effect on immune-mediated aplasia anemia models mice.     

Inheritance of susceptibility to the myeloproliferative sarcoma virus: effect of the Fv-2 locus and evidence for a myeloproliferative sarcoma virus resistance locus.

Myeloproliferative sarcoma virus (MPSV) causes a generalized stem cell leukemia with erythroid and myeloid hyperplasia in adult mice. MPSV also transforms fibroblasts. Mice congenic for the Fv-2 locus showed marked differences in susceptibility to MPSV according to the Fv-2 genotype. MPSV was injected into C57BL/6 Fvs and C57BL/6 Fv-2r mice congenic except for the Fv-2 locus. C57BL/6 mice with the Fvs genotype were much more susceptible to MPSV than were those with the Fvr genotype. Both DDD Fv-2r mice congenic with DDD Fv-2s mice except for the Fv-2 locus and DDD Fv-2s mice, however, were sensitive to spleen focus formation by MPSV. These data indicate that at least one additional resistance locus to MPSV is present in C57BL/6 mice but not in DDD mice. Both the Fv-2 locus and the putative MSPV resistance locus (loci) Mpsvr appear to be epistatic to either of the sensitivity loci. Fibroblast focus formation by MPSV was obtained well in C57BL/6 Fv-2r and C57BL/6 Fvs fibroblasts, indicating that the genes for MPSV resistance (Fv-2r and Mpsvr) were not operating in fibroblast cells. A model is proposed which may account for the differences in response of genetically different mice to MPSV and Friend spleen focus-forming virus.     

Fate of H2-activated T lymphocytes in syngeneic hosts: II. Residence in recirculating lymphocyte pool and capacity to migrate to allografts

T.TDL—a purified population of lymph-borne H2-activated T lymphocytes—were transferred to syngeneic mice to examine their capacity to remain in the recirculating lymphocyte pool (RLP). Experiments with cells labelled with ³H-thymidine (³HTdR) or ⁵¹Chromium showed that although a considerable proportion of T.TDL joined the RLP (i.e., were mobilizable through a thoracic duct fistula) for several days after transfer, most of the cells soon left the pool. This applied whether the cells were transferred to normal mice or to “B” mice. Normal thoracic duct lymphocytes, by contrast, joined the RLP for long periods post-transfer.Studies with ³HTdR-labelled T.TDL showed that a small number of heavily labelled cells remained in the RLP for at least 3 months. Experiments with the θ-antigen as a cell marker suggested that a further small proportion of cells underwent division at some stage after transfer and then rejoined the RLP in expanded numbers.T.TDL showed a tendency to home to specific allografts of either skin, tumor cells or lymphoid cells. Although homing was specific it was very limited in extent.     

Effect of thiopyrimidine ribonucleosides on DNA and RNA synthesis in synchronized mammalian cell cultures

The uptake of ³H-uridine into RNA and of ³H-thymidine into DNA was investigated in synchronized Chinese hamster cells which had been exposed to thiopyrimidine ribonucleosides. The cells were synchronized at metaphase by reversal of colcemid inhibition; these cells were then labeled with either ³H-thymidine or ³H-uridine at selected times, and analyzed in autoradiographs. Incorporation of ³H-thymidine into DNA was not inhibited by administration to the cells of 2-thiouridine or 4-thiouridine (4 × 10⁻³ M). Exposure of the cells to the anti-metabolites for over 15 h significantly reduced the incorporation of ³H-uridine into nuclear RNA and completely blocked the labeling of cytoplasmic RNA. This finding is interpreted as an indication that RNA synthesis was inhibited in cells which continued to synthesize DNA. The inhibition of RNA synthesis hindered cell division and decreased cell viability. This lethal effect is similar to the “unbalanced growth” induced by inhibitors of DNA synthesis. The thiopyrimidine ribonucleosides, however, killed mammalian cells without inhibiting DNA synthesis.     

Growth of Pseudotypes of Vesicular Stomatitis Virus with N-Tropic Murine Leukemia Virus Coats in Cells Resistant to N-Tropic Viruses

Formation of pseudotypes between murine RNA tumor viruses and vesicular stomatitis virus (VSV) has been confirmed. Pseudotypes of VSV genomes coated by the surface envelope from an N-tropic tumor virus grew equally well in cells homozygous for either the Fv-1ⁿ or Fv-1^b alleles. Therefore, the product of the Fv-1 locus, which restricts growth of murine RNA tumor viruses, must act on an intracellular aspect of tumor virus replication, a step after attachment and penetration.     

Stimulatory effects of androgens on normal children's bone marrow in culture: effects on BFU-E, CFU-E, and uroporphyrinogen I synthase activity

We studied the effect of natural and synthetic androgens on children's erythropoietic precursor cells in culture. Cultures of normal marrow were carried out according to a miniaturized methylcellulose method in the presence of erythropoietin. We then evaluated the effects of testosterone, nortestosterone, fluoxymesterone and etiocholanolone (10(-9)-10(-6) M) on erythroid colony-forming units (CFU-E) and burst-forming units (BFU-E). Androgen-induced growth of erythroid progenitors was quantified by directly scoring colonies and by a biochemical determination of the uroporphyrinogen I synthase activity (UROS). We observed a significant increase (p less than or equal to 0.05) in the number of CFU-E and BFU-E and in the UROS activity of derived colonies in the presence of androgens (10(-8) or 10(-7)M). This microculture assay could be useful not only to study the effect of androgens on erythroid progenitor cells in culture, but also to predict the best androgenic treatment of anemia in children and adults.     

In vitro effect of glucan on mouse hemopoietic committed progenitor cells

In vitro methylcellulose clonal cell culture assays of granulopoietic and erythropoietic colonies were used to study the primary effect of glucan on the hematopoietic stem cells. Addition of glucan to the cultures inhibits the formation of colony-forming units-erythrocytic (CFU-E), enhances the production of burst forming units-erythrocytic (BFU-E) and has no effect on colony-forming unit-culture (CFU-C). These results indicate that glucan has a direct effect on late and early erythroid precursor cells.