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Lymph node and spleen cells from normal mice were cultured for 3 days with polyoma virus-induced tumour, Ehrlich's ascites turnout or leukaemia L 1210 cells. This resulted in in vitro immunization of the lymphocytes, which were then transferred to irradiated target cells labelled with 51Cr. Normal, i.e. non-immune thioglycollate-stimulated peritoneal macrophages were also added to some tubes. Non-immune macrophages mixed with immunized lymphocytes showed a significantly increased ability to destroy tumour ceils as compared with macrophages in the absence of immunized lymphocytes. The immunized lymphocytes were almost entirely inactive alone. When the number of macrophages was kept constant the cytotoxicity was dependent on the number of viable immunized lymphocytes placed on the target cells. Immunized lymphocytes, in the presence of macrophages, only exhibited strong killing of the target cells against which they had been immunized; some lysis of ‘bystander’ cells was, however, seen provided specific target cells were present. Macrophage monolayers exposed to immunized lymphocytes upon contact with specific antigen became ‘armed’ and showed a significant cytotoxicity for specific target cells. When immunized lymphocytes and normal macrophages were treated with actinomycin D and puromycin, cytotoxicity was inhibited in the immunized lymphocytes but not in the macrophages. The possible mechanism of normal macrophage cooperation with immunized lymphocytes in the cytotoxic killing reaction is discussed. Results presented in this paper favour the view that immunologically specific cytophilic factor (presumptive cytophilic antibody) is involved in the macrophage-mediated cytotoxicity in the system studied. [ABSTRACT FROM AUTHOR]

This paper confirms results showing that TA3 tumour cells coated in vivo with immunoglobulins, lose some of the coat upon transfer to in vitro conditions. By labelling IgG isolated from the ascitic fluid of TA3 tumours it was found that the immunoglobulin coating TA3 cells is dynamically exchanged with immunoglobulin in the corresponding ascitic fluid. Some of the released immunoglobulin is in a degraded state as judged from the fact that most of the released material lost the antigenicity of intact immunoglobulin and has a lower capacity to precipitate with ammonium sulphate at 50% saturation. The degraded immunoglobulin has a higher binding efficiency to tumour cells. In vivo experiments demonstrated that the ascitic fluid of mice bearing TA3 and MCIM tumours contains a material which has some of the physicochemical properties of IgG but which does not precipitate with antisera directed aguinst IgG. [ABSTRACT FROM AUTHOR]

Various techniques were used to obtain cell cultures from benign and malignant tumors removed from human breasts. Cell suspensions were prepared by trypsinizing finely minced tissue or by the "spilling" technique or Lasfargues and Ozzello, and T-flasks or T-flasks coated with collagen were inoculated. Trypsinized tissue yielded predominantly fibroblastic cultures, whereas those obtained by the "spilling" technique were rich in epithelial cells. The mixed cell populations were separated into pure suspensions by density gradient centrifugation. Most epithelial cell cultures grew best when they were collected by the "spilling" technique and inoculated into collagen-coated T-flasks with Eagle's medium enriched with 30% fetal calf serum. However, tears developed in the collagen coat with subsequent peeling which led to the loss of many epithelial cell colonies. Primary epithelial cells did not remain viable for prolonged periods, and attempts to obtain subcultures were unsuccessful. In this Paper the work of several investigators is reviewed and their methods of growing breast tumor cell are discussed. [ABSTRACT FROM AUTHOR]

Within solid neoplasms. the population of tumor cells and the population of capillary endothelial cells constitute a highly integrated ecosystem. Tumor cells release an endothelial mitogen. Tumor-Angiogenesis-Factor (T.A.F.) which continually stimulates new capillaries to grow into the tumor. When T.A.F. is blocked, neovascularization is prevented and tumor nodules stop expanding at a diameter less than 2.5 mm. They enter a dormant phase because they are forced to live by simple diffusion of nutrients and wastes. Thus "anti-angiogenesis can force a population of tumor cells to become dormant at a tiny diameter. In this paper an analogy is drawn between tumor angiogenesis and the angiogenesis which accompanies psoriasis. if the relationship between psoriatic epithelium and its capillary endothelium turns out to be similar to the integration of capillaries by solid tumors, then "anti-angiogenesis' may eventually become a useful therapeutic approach in psoriasis. [ABSTRACT FROM AUTHOR]

Elastolytic enzymes were isolated from the extract of the lysosome-like granules of human leukaemic myeloid cells by chromatography on Sephadex G-75, εAhx-Sepharose 4 B, elastin-Sepharose 4 B and preparative agarose-gel electrophoresis. The elastase were cationic proteins but appeared as three proteins with a slight difference in electrophoretic mobility. The three elastases were inactivated by α1-antitrypsin and α2-macroglobin. The amino-acid composition of the three elastases was the same except for a variation in arginine content. Dodecylsulfate-electrophoresis suggested molecular weights in the range 33 000–36 000. The s20,w0 value determined for one of the elastases was 2.6 S. The three elastases gave the reaction of identity on immunodiffusion according to Ouchterlony and crossed immunoelectrophoresis. They showed the same type of reaction with elastase partly purified from normal human granulocytes, which also contained three separate elastolytic enzymes. [ABSTRACT FROM AUTHOR]

A number of parameters concerning the interaction of highly purified elongation factor 1 (EF-1) from ascites tumor cells with ribosomes and elongation factor 2 (EF-2) were investigated. The following results were obtained : 1. The EF-l-directed attachment of aminoacyl-tRNA to "run-off" ribosomes is stimulated by GTP and to a somewhat lesser extent by Guo-5'-P2-CH2-P. 2. During prolonged incubations, EF-1 can be shown to function catalytically rather than stoichiometrically. 3. Although a specific interaction between aminoacyl-tRNA and EF-1 from ascites cells has previously been shown to occur in the presence of GTP, preincubation of aminoacyl-tRNA with EF-1 and GTP does not enhance the rate of its subsequent binding to ribosomes. 4. In a cell-free system, containing "run-off" ribosomes, poly(uridylic acid), Phe-tRNAPhe and elongation factors, the rate of poly(phenylalanine) synthesis is strongly enhanced when EF-1 is added before EF-2. This requirement for sequential addition of the factors led to the suggestion that both factors bind to the same region of the ribosome. 5. EF-1 binds Phe-tRNAPhe to P-site-occupied ribosomes at a significantly higher rate than to empty ("run-off", ribosomes. The conclusions which can be drawn from these results are discussed in conjunction with our previous findings and with data from the literature relating to the structure and function of mammalian elongation factor 1. [ABSTRACT FROM AUTHOR]

An IF-M3-deficient ascites cell-free system is described. An IF-M3 species of high molecular weight has been isolated and partially purified from a high-KCl ascites ribosomal wash fluid. Translation of encephalomyocarditis virus RNA and globin mRNA is completely dependent upon the presence of the ascites IF-M3 while the factor is not required with either poly(A, U, G) or poly(U) as messengers. The sparsomycin technique for the generation of short initiation peptides has been employed to demonstrate that IF-M3 acts at the level of chain initiation. Rabbit reticulocyte IF-M3 and IF-E3 as well as rabbit liver IF-M3, can substitute for ascites IF-M3 in EMC-RNA and globin mRNA translation. The translation of EMC-RNA in the ascites cell-free system is, therefore, dependent upon the presence of two discrete initiation factors, a high-molecular-weight species, IF-M3, and a smaller component, IFEMC. [ABSTRACT FROM AUTHOR]

In the nucleus of L5178 Y cells, a lymphocytic mouse leukemia cell strain, newly synthesized 5-S ribosomal RNA first appears mainly in the nucleoplasm and then diffuses into the nucleotus. Kinetic studies have revealed that the appearance rates of 5-S RNA and ‘45-S fraction’ ribosomal precursor RNA are identical. During the first 30 rain, 5-S RNA binds to proteins and constitutes a nuclear pool of independent ribonucleoprotein particles which sediment at approximately 11 S. After this time, it begins to integrate with the nucleolar ‘80-S nascent’ particle. This ribonucleoprotein particle contains ‘45-S fraction’ RNA, the direct product of ribosomal DNA genes and has been shown to represent the first stage of ribosome formation. 5-S RNA remains associated to preribosomal particles during the nuclear maturation process leading to the large ribosomal subunit, located in the cytoplasm. In labeling experiments, the molar ratio of labeled 5-S RNA compared to high-molecular-weight ribosomal RNA in ribonucleoprotein particles, continues to increase until reaching a value of 1:1 for steady-state labeling. The association of 5-S RNA to preribosomal particles is more labile than it is to the mature ribosomal subunit. Chemical products such as heparin or sarkosyl used to extract nucleolar ribonucleoprotein particles, were shown to detach 5-S RNA from these particles; but dithiothreitol keeps the particles intact. [ABSTRACT FROM AUTHOR]

The three major forms of elongation factor 1 (EF-1) occurring in Krebs II mouse ascites tumor cells were investigated with respect to their affinities for guanosine nucleotides and aminoacyltRNA. The binding of GTP, GDP and 5'-guanyly-methylene-diphosphonate (Guo-5'-P2-CH2-P) to a pure preparation of the tetramer form of EF-1 and to partially purified dimer and hexamer preparations of the same factor was studied by equilibrium dialysis techniques. Another series of experiments was designed to describe the interaction of the tetramer and dimer form of EF-1 with aminoacyl-tRNAs. The following results were obtained. 1. When measured at 2 °C, the association constants for the complexes between pure EF-1 and GTP or GDP were both 92 mM-1. 2. There is one guanosine nucleotide binding site per molecule of the EF-1 tetramer which can be occupied by GTP, GDP or Guo-5'-P2-CH2-P. 3. The affinities of the dimer and hexamer forms of EF-1 for GTP and GDP were of the same order of magnitude as the corresponding values relating to the tetramer. 4. Large (tetramer) and small (dimer) aggregates of EF-1 provide equal protection of aminoacyl-tRNA against hydrolytic deacylation. This protective effect depends on the previous interaction of the factor with GTP. Although GDP and Guo-5'-P2-CH2-P bind to EF-1, they cannot replace GTP in this reaction. 5. The protective effect of EF-1 on aminoacyl-tRNAs is limited to tRNA species which can function in polypeptide chain elongation. Met-tRNAfMet and the chemically formylated MettRNAMet are only poorly protected by EF-1 indicating that these tRNAs are not well recognized by the factor. The significance of these results with respect to our understanding of the function of elongation factor 1 in mammalian cells is discussed. [ABSTRACT FROM AUTHOR]

Immunoglobulin-coated ascites tumour cells lose some of their immunoglobulin coat following their transfer to in vitro culture conditions. The uncoating process is metabolism-dependent and related (o the turning over of cell-surface components. Uncoated tumour cells bind globulin isolated from tumour eluates more efficiently than coated cells. Analysis of the spent medium of short-term tumour cell cultures revealed the presence of both immunoglobulin and antigenic cell surface components. [ABSTRACT FROM AUTHOR]

It has been shown that CBA mouse lymphocytes cultured in the presence of allogeneic fibroblasts or syngeneic polyoma virus-induced tumour cells became specifically immunized. These immunized lymphocytes, however, are by themselves non-cytotoxic and only cause target cell killing when mixed with normal macrophages. The reaction is immunologically specific and seems to depend on the antigenic differences between lymphocytes and target cells. The mechanism of the interaction between the immunized lymphocytes and the normal macrophages in the cytotoxic killing of target cells is discussed. [ABSTRACT FROM AUTHOR]

In HeLa S3 cells grown in suspension and synchronized with thymidine, while nuclear DNA is replicated during the S-phase, cytoplasmic DNA is synthesized with two peaks in the S- and G2- phases: the first has its maximum 1–2 h before that of nuclear DNA ; the second takes place after that of nuclear DNA, approximately at the 9th hour from the removal of synchronizing thymidine from the culture medium. Newly synthesized nuclear and G2-phase-cytoplasmic DNAs in CsCl gradients have buoyant density of 1.698 and 1.688 g/cm3, respectively. The buoyant density of S-phase-cytoplasmic DNA is 1.701 g/cm3, probably corresponding to the closed circular duplex form of mitochondrial DNA. The kinetics of synthesis of nuclear DNA is exponential, differing from that of S- and G2-phase-cytoplasmic DNAs which show an asymptotic shape of [3H]thymidine labeling. Ethidium bromide inhibits the synthesis in vivo of both S and G2 peaks of cytoplasmic DNA, but does not appreciably influence the labeling of nuclear DNA. [ABSTRACT FROM AUTHOR]

[35S]Cystine-labelled MOPC21 (IgG1:ℵ) myeloma protein was prepared by incubating myeloma cells, grown as a suspension culture, in medium containing [35S]cystine. The radioactive protein was used as a tracer in conjunction with carrier myeloma protein. A peptic-tryptic peptide of 36 residues was prepared, which contained ten half-cystine residues and included the heavy-light, and all the heavy-heavy inter chain disulphide bridges of the molecule. The peptide was digested with thermolysin and subjected to partial acid hydrolysis. A large number of small peptides was isolated and, when possible, their amino acid compositions were determined. From 170 possible arrangements of the disulphide bonds a unique arrangement, was deduced ill which all the heavy-heavy interchain bonds were parallel. This is the first case in which the arrangement of all the interchain bonds of any of the human or mouse IgG subclasses has been established. [ABSTRACT FROM AUTHOR]

New methods are presented for the isolation of nucleoli and the extraction of intact preribosomes under mild conditions avoiding EDTA and high salt buffers. These nucleolar preribosomes were characterized by an improved technique of polyacrylamide-gel electrophoresis which loves better resolution than sedimentation analysis for both ribonucleoprotein particles anti free-RNA molecules. Thus, the 50-S native-ribosomal subunit could be separated from its 55-S precursor containing 32-S pre-RNA, and another nucleolar precursor, a "70-S" particle, could be identified. The separation technique newly developed is disc electrophoresis using exponential polyacry1amide gels. The resolution with these gels is gloater than that with uniform gels over a wider range of molecular weights. The final resolution is independent of the volume in which the sample is loaded and the diffusion of molecules during migration is eliminated. Molecular weights of RNA molecules can be estimated in exponential gels as in uniform gels. Electrophoresis in quartz tubes allows the direct scanning of ultraviolet, absorbance during and after the run. As all handling is thus avoided, gels of very low concentration can be used. [ABSTRACT FROM AUTHOR]

RNA synthesis and metabolism were investigated in Hela cells heated to 42°. At this temperature cellular growth is arrested. After prolongued periods of preheating the initial rate of uridine incorporation into RNA is only slightly affected but no new stable RNA accumulates. After a shift down to 37° the normal pattern of RNA formation resumes. 45 S precursor to rRNA is still formed but its normal metabolism to functional rRNA is blocked. No new rRNA appears in cytoplasmic ribosomes. On the contrary, the synthesis of nascent messenger-like RNA is only slightly affected and non-ribosomal RNA appears in the cytoplasm at a normal rate. The temperature sensitive event seems thus to be selectively located in the metabolic chain producing functional rRNA. [ABSTRACT FROM AUTHOR]

This communication describes the use of micro-electrophoresis in studying the changes in ascites tumour cells exposed to antibodies and complement. Treatment of the cells with rabbit antibody led to a change in electrophoretic mobility consistent with a surface adsorption of γ-globulin. The addition of complement led to a reduction in this electrophoretic effect of antibody. Treatment of the cells with neuraminidase, which produced a marked fall in their electrophoretic mobility, did not alter the effect of rabbit antibody and complement on the cells. Incubation with iso-antibody, in the presence or absence of complement, did not alter the mobility of the ascites cells measured at pH 7.0. The addition of 0.01 M calcium chloride to the electrophoresis medium produced a fall in mobility of the cells exposed to either antibody preparation in the presence of complement. Therefore, although iso-antibody and complement did not produce direct changes in cell mobility, changes in the cell surface could be detected by electrophoresis in the presence of calcium ions. The possibility is discussed that, during immune cytolysis, unmasking of phosphate groups of phospholipids might take place in the cell surface. [ABSTRACT FROM AUTHOR]

Biochemical and histochemical methods were used to study the interaction of antibodies and complement with mouse Ehrlich ascites tumour cells. In the presence of complement, both iso- and hetero-antibodies caused cell lysis with penetration of antibodies into the damaged cells, as detected by immunofluorescence; the cells were then unable to support aerobic glycolysis though they retained their ability to consume oxygen in the presence of succinate. Under these conditions there was unmasking of phospholipid particularly at the cell surface, together with lysosomal changes resulting in diffuse staining for lysosomal acid-phosphatase. In the absence of complement, antibodies did not appear to penetrate the cells which respired normally and were not lysed. However, in these cells there was intense lysosomal activation accompanied by unmasking of cytoplasmic phospholipid; it appeared that an immune reaction confined to the cell surface was able to induce changes in the cytoplasm without acutely impairing the viability of the cell. [ABSTRACT FROM AUTHOR]

Rabbit antisera against mouse cells have been absorbed with normal mouse serum and used to detect mouse cell-bound antigens. Mouse ascites tumor cells and their products have mainly been used as source of antigens but those described here are normal mouse components. Seven antigens have been found by agar diffusion precipitin methods and data are given to show that each is a separate molecular entity. Information is provided for the identification of each component. [ABSTRACT FROM AUTHOR]

A method for the quantitation of cytotoxic action of antiserum and complement is described, measuring the incorporation of trypan blue into Ehrlich ascites tumour cells. The incorporation of the dye into the cells in the presence of antiserum and complement was maximum at ionic strength between 010 and 015, and decreased with increasing ionic strength higher than 015. The permeability barrier of the cell against the dye was stable at pH between 7.2 and 7.4. The cytotoxic reaction required Mg ++ and Ca++ ion, but was inhibited with concentrations higher than 5 mM Mg+ + and 1 M NCa. The incorporation of the dye in the presence of antiserum and complement was more rapid with increasing concentrations of the dye, and depended on temperature. From these findings we obtained optimal condition for the assay of cytotoxic antiserum. [ABSTRACT FROM AUTHOR]

The results of spleen transplantation from a healthy individual to his leukaemic identical twin are reported. The rationale for the procedure and its significance in relation to present hypotheses of tumour growth and spread are reviewed. Although lymphocytes of both the donor and recipient showed specific immune capacity to destroy tumour cells in vitro, there was evidence for partial inhibition of the immunological effector process that destroys the tumour cells in the diminished lymphocyte phytoagglutinin stimulation, hypoglobulinaemia, and marked deficiency of IgA in the leukaemic twin. These abnormalities and the final relapse in spite of transplantation of the spleen as a source of immunocompetent cells could also be considered the result of growth potential in the tumour, capable of overriding any immunological control. [ABSTRACT FROM AUTHOR]

This article presents an analysis of the results of cancer therapy, based on the histories of treated and untreated cancers taking into account the biology of immunity and drug action. The administration of cancer chemotherapeutic drugs (CCD) or irradiation produces remission in certain cases of cancer. It is generally believed that these remissions are due to the direct action of the treatment on the tumor cells. Reasons for the acceptance of this opinion are its simplicity and its analogy with antibiotics. CCD have diverse actions and there are many ways in which these drugs could act on, or in association with, the immune system to affect the tumor.

The article focuses on the interrelationships between molecular biology and cancer chemotherapy. It talks about the need for more effective drugs and the challenge to find means for effective prevention, detection and treatment of cancer. The presence of drug receptors in cancer cells and the ability of drugs acting upon quantitative metabolic differences to kill cancer cells are discussed. The article also discusses the use of drugs as probes for metabolic dissection and the applications of anticancer drugs to other fields of biology.