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Biomedical subjects

P Bentvelzen

Publications and source records attributed to P Bentvelzen.

At least 19 recordsLinked to original sources
PubMed1988

Flow cytometric detection of ribosomal RNA in suspended cells by fluorescent in situ hybridization.

A method using flow cytometry and fluorescent in situ hybridization (ISH) to detect RNA in cells is described. L1210 murine leukemia cells were fixed with 1% formaldehyde in HEPES buffered Hank's balanced salt solution (HH) followed by 70% ethanol. Endogenous RNAses were blocked by diethylpyrocarbonate treatment. Single-stranded sense and antisense RNA probes, labeled with biotin-11-UTP, were transcribed from a 2.1 kb 28S ribosomal RNA (rRNA) gene fragment subcloned into the pGEM2 plasmid. For good results, it was essential that the probes were degraded to 100-150 nucleotides before use. Hybridization was performed at 45 degrees C in 50% formamide, 5 x SSC, 0.5% SDS. Hybrids were detected with streptavidin-FITC by flow cytometry. Antisense rRNA probe signal was 100 times higher than the background. The hybrids were largely resistant to RNAse and melted at high temperature. The sense probe also gave a signal (5 times background), which was not RNAse resistant and was attributed to the presence of internal inverted repeats in the ribosomal RNA. When sufficient background reduction can be achieved, it is expected that as few as ten mRNA molecules per cell can be detected with the fluorescent in situ hybridization method.

Animals↗
PubMed1988

Transactivating potential of the 3' open reading frame of murine mammary tumor virus.

The procaryotic chloramphenicol acetyltransferase (CAT) gene controlled by the murine mammary tumor virus (MMTV) promoter shows reduced activity in a rat mammary tumor cell line after infection with MMTV but to a considerably lesser extent than the CAT gene controlled by a heterologous promoter, indicating trans-acting regulation of promoter activity by MMTV. Cotransfection of vectors capable of expressing RNA from the 3' open reading frame (orf) of MMTV with the CAT-MMTV construct resulted in enhanced CAT activity, suggesting that orf carries a transactivating potential. Since transactivation was also found with a vector containing only orf and part of the viral env gene, it was concluded that a separate transcriptional unit exists for the orf message.

Animals↗
PubMed1984

Presence of fibroblast-transforming genes in normal DNA of several mouse and rat strains.

Cellular DNA of the inbred mouse strains BALB/c, C3Hf, C57BL, CBA, DBAf , GRS and ND2 and the inbred rat strains BN, SD and WAG was shown to oncogenically transform murine fibroblasts of the continuous cell line BALB/3T3. Subcutaneous inoculation of transformed cells into BALB/c mice led to the rapid development of sarcomas. For transformation the DNA had to be fragmented to a size smaller than 23 kilobase pairs (kbp), with a lower limit of 6.5 kbp. No transforming virus could be rescued from the transformed lines by infection with a murine leukemia virus.

Animals↗
PubMed1983

Sera from irradiated rats contain antibodies to a ubiquitous tumour-associated antigen.

Female rats of the inbred strains BN/BiRij and WAG/Rij were irradiated with 30 kV X-rays, or 15 MeV or 0.5 MeV fast neutrons. Sera were collected several months after irradiation and found to be negative for antibodies reacting with the murine mammary tumour virus as tested by a solid-phase radioimmunoassay and an immunofluorescence absorption test. We found, however, that in an immunofluorescence assay several sera from irradiated rats reacted with a cytoplasmic antigen in rat mammary, ureter and skin carcinoma cell lines as well as a mouse mammary tumour and a transformed BALB/3T3 mouse fibroblast line. No reaction was found with normal fibroblast cell lines of rat or murine origin. Endpoint titres of the sera on tumour cells ranged from 1:20 to 1:160. Of 48 sera from unirradiated rats 18 also stained tumour cells, but usually at the low dilution of 1:10. Irradiation seems to enhance antibody activity to a ubiquitous tumour-associated antigen.

Animals↗
PubMed1982

The concept of gene transfer-misrepair mechanism of radiation carcinogenesis may challenge the linear extrapolation model of risk estimation for low radiation doses.

The recent demonstration that transformation of cultured cells can be induced by exposure to DNA fragments prepared from normal mouse tissues provides experimental support to the gene transfer-misrepair hypothesis of radiation carcinogenesis. Employing various assumptions with regard to the generation of oncogenic DNA fragments and of cells which are susceptible to incorporate these fragments into their genome, it is predicted that the proposed mechanism implies a non-linear extrapolation model for the calculation of cancer risks caused by very low doses of ionizing radiation of low LET. It also follows from this hypothesis that X- and gamma-radiation delivered at an extremely low dose rate will be less carcinogenic than at high dose rate, in particular where low total doses are concerned. Several aspects of the hypothesis can be verified experimentally by the use of in vitro cultured cell systems.

Cell Transformation, Neoplastic↗
PubMed1980

Ubiquity of natural antibodies to the mammary tumour virus in mice.

Sera of female and male mice from eleven inbred mouse strains collected at either 4, 12, 36 or 60 weeks of age were tested for the presence of natural antibodies to the murine mammary tumour virus by means of the Sepharose bead immunofluorescence assay. Antibodies to the virus proved to be ubiquitous, but pronounced strain differences were found in titer and onset of antibody production. These differences were related to neither release of virus in the milk nor susceptibility to spontaneous mammary tumour development of a given strain. Immunological specificity of the observed reactions was concluded from a) the failure to block the reaction by absorption with fetal calf serum, mouse milk or sheep erythrocytes, while absorption with purified virus abolished the reactivity; b) the lack of reactivity of rat sera with the mouse mammary tumour virus in this system; c) the negative response of mouse sera with Sepharose beads coated with ovalbumin; d) the lack of correlation between antibody titers to Rauscher murine leukemia virus and mammary tumour virus in this system; e) the retaining of activity to highly purified viral polypeptides; f) blocking of the reaction by preincubation with rabbit anti-mouse immunoglobulin serum or Protein A from Staphylococcus aureus. Since germfree mice of various strains also have such antibodies, it is concluded that the reactions are not due to horizontal transmission of the virus. From the lack of correlation between antibody titers and tumour incidences, it is concluded that various systems overshadow the potential immunosurveillance role of such natural antiviral antibodies.

Aging↗
PubMed1979

Characterization of a type-C virus produced by co-cultures of human leukemic bone-marrow and fetal canine thymus cells.

The putative human helper virus SKA-21/A204V, isolated by Nooter et al. in 1977 from human leukemic bone-marrow cells following co-culture with normal fetal canine thymus cells, Cf2th, has been characterized with respect to its major viral core protein, reverse transcriptase, and nucleic acid sequences. The results of these analyses show that this virus is not distinguishable from the woolly monkey type-C virus, SSAV-1, by the techniques employed.

Animals↗