Biomedical subjects
Publications and source records attributed to V Wunderlich.
Different methods of large-scale concentration of a type D retrovirus (PMFV) were compared, including continuous-flow centrifugation, ultrafiltration, and polyethylene glycol precipitation. Best results were obtained by a combination of continuous-flow centrifugation and subsequent purification by velocity gradient and isopycnic gradient centrifugation.
Extracts for different normal and malignant human tissues were checked for retrovirus antigens by Sepharose bead immuno-fluorescence assay. No convincing evidence could be obtained for the presence of type D and mammalian type C virus antigens in the tissues tested by three different immunoassays. It is concluded from these results that such viruses have no ubiquitous distribution in human beings.
The protein patterns of type D retroviruses isolated either from permanent human cells (HeLa virus, HeLa V, and HEp-2 virus, HEp-2V) or from spectacled langur (langur virus, LV), an Old World monkey, were investigated using sodium dodecyl sulfate polyacrylamide gel electrophoresis. Labeling with 14C-amino acids and 3H-glucosamine revealed for each of these isolates five polypeptides with molecular weights of 10,000, 12,000, 15,000, 25,000 and about 68,000 and identified the 68,000 D-protein as a glycoprotein. An additional glycoprotein with a molecular weight of 20,000 (gp20) was resolved in LV similarly as in previous studies with Mason-Pfizer monkey virus. However, in accordance with our recent analysis of a third type D isolate from human cells (PMFV), gp20 was not detectable in both HeLaV and HEp-2V regardless of the cell line in which they were grown. Thus, it appears that the type D virus isolates from human cells are slightly distinct from the presently known type D retroviruses of Old World monkeys. The relevance of this finding to the origin of the human isolates, however, remains to be shown.
Addition of the tumor promoter 12-0-Tetradecanoyl-phorbol-13-acetate (TPA) to the growth medium of certain human cell cultures persistently infected with simian retroviruses of type C (BaEV, SSV) or type D (MPMV) or a human cell line-derived type D isolate (PMFV), respectively, resulted in a considerable but transient stimulation of virus production. Enhanced virus expression was paralleled by striking morphological alterations of the cells. Among four infected cell types tested so far, ony one (A 204) failed to respond to TPA with significant virus stimulation or altered morphology.
Tumor viruses which in the past few years have been increasingly isolated from tissues of nonhuman primates are of considerable interest for contemporary investigations on putative human tumor viruses. This review covers the presently known isolates belonging to the families of papova, adeno, (gamma) herpes, pox, and retro viruses and originating from New World and Old World monkeys as well as from apes. Some of the isolates exhibit oncogenic activity in different simian species and are, therefore, particularly suited for the analysis of viral oncogenesis is primates.
The production of a type D retrovirus (PMFV) was analysed using different conditions of cultivation. Both in cell culture flasks and in roller culture bottles with discontinuous change of medium an increase of virus amount was determined within 12 h after seeding of virus-infected cells. Then the virus production considerably fluctuated around an apparent mean value until the end of the experiment at 56 h. In case of a continuous change of cell culture medium in roller bottles a maximum of virus production was measured at a retention time of 24 h of cell culture medium. The production of PMFV could be essentially increased when virus-producing cells were co-cultivated with non-infected cells of the same type.
The protein pattern of a type-D retrovirus (PMFV) isolated from and propagated in human cell lines has been investigated using sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Staining with Coomassie blue and labeling with 14C-leucine/14C-lysine revealed five viral polypeptides with molecular weights of 10,000, 12,000, 15,000, 25,000, and 68,000. The 68,000 D-protein was shown to be a glycoprotein by incorporation of 3H-glucosamine and the 15,000 D-protein was identified as a phosphoprotein. By comparing PMFV with the closely related Mason-Pfizer monkey virus (MPMV) in co-electrophoresis experiments no clear difference was detected in viral 14C-leucine/14C-lysine profiles. The viruses differ, however, with respect to their glycoprotein patterns. A glycoprotein corresponding to the gp20 of MPMV has not been detected in PMFV irrespective of the cell line used for propagation of viruses.
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Enzyme neutralization experiments by using an antiserum to reverse transcriptase (RT) of Mason-Pfizer monkey virus (MPMV) demonstrated the presence of common antigenic determinants on the RTs of all known type D retroviruses and of determinants discriminating different isolates. The RT of MPMV is antigenically closely related to the RTs of type D viruses isolated from human cell lines although significant differences are also detectable. It shows a lesser degree of relationship to the RTs of squirrel monkey retrovirus (SMRV) and of langur virus (PO-1-Lu). Since the major structural proteins of MPMV and PO-1-Lu are much more related to each other than to the corresponding protein of SMRV it can be concluded that there is no correlation in the degree of relatedness of different proteins of different type D viruses.
PMFV, a type D retrovirus isolated from a malignant human embryo cell line, was compared with Mason-Pfizer monkey virus (MPMV) in a sensitive tannic acid enhanced indirect immunodiffusion test. In addition to the previously shown common antigens, both viruses contain identical group-specific antigenic determinants on their p 10--12 as demonstrated with a specific p 10--12 MPMV test system. Interspecies mammalian type C virus antigens were not detected in highly concentrated PMFV preparations.
Disrupted primate retroviruses of type C (baboon endogenous virus, BaEV) and type D (human cell line-derived isolate PMFV) considerably suppressed Concanavalin A - induced blastogenic response of human lymphocytes. Rauscher mouse leukemia virus (RLV) displayed a suppressive activity on murine splenic lymphocytes when tested under analogous conditions. The immunosuppressive activities were shown not to result from cytotoxicity or from virus-mitogen binding.
This paper describes the detection of viral particles in a human amnion cell line (FL). These particles belong to the group of type D retroviruses, because of their characteristic morphology, their major antigenic determinants, and the presence of particle-associated Mg++-preferring RNA-dependent DNA polymerase. In addition to budding and mature type D virus particles, intracytoplasmic type A structures have also been found. Immunological analysis provided evidence of cross-reactivity between the particles described and the Mason-Pfizer monkey virus as well as type D retroviruses from other human cell lines (PMF, HEp-2). Particles isolated from FL cell medium were shown to infect type D virus-susceptible human TU 197 cells.
The neuroleptic drug haloperidol (Hal) shows, when administered in multiple intraperitoneal or intravenous injections beginning at 5 hrs post inoculation of Rauscher leukemia virus to male NMRI mice, a marked activity in inhibiting virus-induced splenomegaly and prolonging mean survival time. Evidence is presented that a direct action of the drug on the virus is involved in its inhibitory effect.
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Sodium dodecylsulfate-polyacrylamide gel electrophoresis of bovine leukemia virus (BLV) grown in fetal lamb kidney cells in the presence of 32P-phosphoric acid showed that the slightly basic 15000 D protein (pp 15) is the major phosphorylated component of the virus. This result further distinguishes BLV from mammalian type C viruses which possess acidic major phosphorproteins.
The mechanism of carcinogenesis is not yet understood. There is increasing evidence justifying the assumption that an unifying concept of carcinogenesis should be possible at the molecular level. New insights into the molecular mechanism of carcinogenesis were mainly obtained in studies on both chemical and viral carcinogenesis. In the present paper, selected results of these studies are reviewed. It is concluded that the interaction of different carcinogenic agents with the cellular DNA results in alterations of DNA. Only some of these alterations, however, seem to be relevant to carcinogenesis. Alterations of DNA can be caused by reaction of electrophilic agents with DNA constituents, by increased infidelity of DNA replication, by integration of viral genomes or by recombination events involving integrated proviruses.
The present paper describes a competitive immunoassay using antiserum to FeLV p 27 and SSV-conjugated Sepharose beads. The assay is applied to compare the interspecies-specific antigenic determinants of the major structural proteins of type-C viruses of different mammalian species. The test proves to be highly sensitive and specific and may be used for the demonstration of viral proteins in crude cellular extracts.
The type D-retraviruses PMF virus (PMFV) and Mason-Pfizer moneky virus (MPMV) show RNA-dependent DNA polymerase (revertase) activity after treatment of the nondisrupted virions with the chelating agent ethylenediaminetetraacetate (EDTA). In the range of 1 to 5 mM, the effect is dependent on the concentration of EDTA. As compared with Nonidet P40 disrupted particles treatment of PMFV with EDTA results in a revertase activity of about 65%. Simultaneous addition of Mg- or Ca-ions prevents the EDTA effect. Other divalent cation-binding agents (o-phenathroline, thiosemicarbazide) do not induce an EDTA-like effect. It is suggested that EDTA chelates divalent ions responsible for the structural organization of the viral membrane which, after chelation, gets permeable for the exogenous revertase template.