Rous sarcoma virus mutant dlPA105 induces different transformed phenotypes in quail embryonic fibroblasts and neuroretina cells.

dlPA105 is a spontaneous variant of Rous sarcoma virus, subgroup E, which carries a deletion in the N-terminal portion of the v-src gene coding sequence. This virus was isolated on the basis of its ability to induce proliferation of quiescent quail neuroretina cells. The altered v-src gene encodes a phosphoprotein of 45,000 daltons which possesses tyrosine kinase activity. DNA sequencing of the mutant v-src gene has shown that deletion extends from amino acid 33 to 126 of wild-type p60v-src. We investigated the tumorigenic and transforming properties of this mutant virus. dlPA105 induced fibrosarcomas in quails with an incidence identical to that induced by wild-type virus. Quail neuroretina cells infected with the mutant virus were morphologically transformed and formed colonies in soft agar. In contrast, dlPA105 induced only limited morphological alterations in quail fibroblasts and was defective in promoting anchorage-independent growth of these cells. Synthesis and tyrosine kinase activity of the mutant p45v-src were similar in both cell types. These data indicate that the portion of the v-src protein deleted in p45v-src is dispensable for the mitogenic and tumorigenic properties of wild-type p60v-src, whereas it is required for in vitro transformation of fibroblasts. The ability of dlPA105 to induce different transformation phenotypes in quail fibroblasts and quail neuroretina cells is a property unique to this Rous sarcoma virus mutant and provides evidence for the existence of cell-type-specific response to v-src proteins.     

Attachment and multiplication,morphology and protein production of human fetal primary liver cells cultured in hormonally defined media

In Vitro Cellular &; Developmental Biology - Plant -     

Differentiation of newborn rat adipocyte precursors in defined serum-free medium

Summary Newborn rat adipocyte precursors, isolated from inguinal fat pads of 2 day-old NBR rats proliferate and undergo adipose differentiation in defined medium in the absence of serum when cultivated on polylysine coated dishes in DME-F12 medium supplemented with fibronectin, insulin, transferrin and FGF. After 7 days in culture in these conditions, 90% of the cells have undergone differentiation as measured by the increase of G3PDH specific activity and by the accumulation of triglycerides in their cytoplasm. In contrast, the cells cultivated in the presence of 10% fetal bovine serum, have a limited ability to differentiate. These results indicate that newborn rat adipocyte precursors from inguinal fat pads do not require the presence of an undefined adipogenic factor in order to differentiate in culture. In contrast, proliferation and differentiation are dependent on the presence of insulin in the culture medium. Moreover, the data presented in this paper show that the rat adipocyte precursor culture represents a rapid and reproducible system for investigating the processes of adipose tissue development and for studying the negative and positive regulators of the adipose differentiation in a controlled environment. This work was supported by grants from the Juvenile Diabetes Foundation, File #185221 and from the National Institutes of Health 1 PO1 CA37589. Editor’s Statement This paper extends to primary cultures the serum-free methods previously applied to studies of adipocyte differentiation in established lines. The observation that serum can block differentiation in this system suggests the existence of previously unrecognized circulating plasma or platelet factors affecting adipocyte differentiation, and the model developed provides an assay for the identification of these factors.     

Volatile constituents of the leaves of Cupressus dupreziana and Cupressus sempervirens

The essential oils of the leaves of Cupressus dupreziana and Cupressus sempervirens were compared. The composition of the hydrocarbon fraction showed a great similarity between the two species.     

Differentiation under the control of insulin of rat preadipocytes in primary culture: Isolation of homogeneous cellular fractions by gradient centrifugation

Using a density gradient medium (Percoll) we succeeded in isolating homogeneous cell populations from the stromal-vascular fraction of the inguinal tissue of 3-day-old rats. In primary culture, in medium 199 supplemented with 10% fetal calf serum and 5.5 mM glucose, almost complete differentiation (90%) of these fractions was obtained for the first time in presence of a physiological concentration of insulin (10^?9 M). During the adipose conversion, insulin markedly enhanced the activities of glycerol-3-phosphate dehydrogenase and acid:CoA ligase. When VLDL and heparin were added with insulin to the medium, this effect was not potentiated. On the contrary, VLDL and heparin in presence of insulin increased the triglyceride content of the cells. With VLDL and heparin only, the biochemical and morphological characteristics of the cells were very similar to those observed in control culture. The heavier fraction was morphologically heterogeneous and did not undergo the adipose conversion to the same extent as the two lighter fractions. It was concluded that this model could be helpful in studying the proliferation and the differentiation of preadipocytes at an early stage of development.     

Isolation of Cell Surface Membranes from Cultured C6 Glioblastoma Cells

Abstract: Plasma membranes were isolated from C6 glioblastoma cells by two methods. In the first method cells were treated with concanavalin A and lysed in hypotonic medium. After partial separation of plasma membranes from other cell material, the lectin was displaced with a-methyl-D-mannoside. In the second method untreated cells or cells iodinated in a lactoperoxidase-catalyzed reaction were homogenized in isotonic medium. Membrane fractions obtaincd by either homogenization procedure were further purified by rate zonal and equilibrium centrifugations into linear density gradients. Disruption of the glioblastoma cell membrane gives rise to heterogeneous assemblies of mem- brane fragments. Two populations of plasma membranes were isolated from untreated and from iodinated cells: a "lighter")membrane fraction characterized by relatively lower sedimentation velocity and buoyant density, and a "heavier" membrane fraction of relatively faster sedimentation velocity and higher buoyant density. Both fractions showed electrophoretic patterns similar to those of ¹²⁵I-labeled cell surface proteins. Their specific (Na⁺+ K⁺)-ATPase activity was seven- to eightfold the homogenate activity (recovery, 13.1%). Both fractions were, however, still contaminated by smooth endo- plasmic reticulum, as judged from the activity 0: NADPH-dependent cytochrome c reductase (recovery, 2.4%). It is suggested that plasma membrane fragments present in the two fractions might differ in the organization of their structures, e.g., membrane vesicle intactness and membrane orientation.     

Volatile constituents of Cupressus dupreziana and the sesquiterpenes of Cupressus sempervirens

Analysis of wood essential oil of Cupressus dupreziana revealed 26 components: 13 monoterpenes and 13 sesquiterpenes. The main components were carv     

Improvement in efficiency of microspore culture to produce doubled haploid lines of Ethiopian mustard

Factors affecting microspore embryogenesis of Ethiopian mustard (Brassica carinata A. Braun) were evaluated, including flower bud length, pollen developmental stage, and microspore density. An embryogenic frequency of 300 embryos per Petri plate was observed with NLN (Nitsch-Lichter-Nitsch) medium supplemented with 13% sucrose, 3.0–3.4-mm-long buds, and a plating density of 65,000 microspores/ml. About 65% of the microspores from buds 3.0–3.4-mm long were at the late uninucleate stage. Microspore-derived embryos were successfully transferred to solid medium for germination. After 4 wk, the resulting plantlets were transplanted to a soilless potting mixture and grew well under greenhouse conditions.