Cloning of a polymorphic sequence from the nontranscribed spacer of horse rDNA.
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Biomedical subjects
Publications and source records attributed to L Bertoni.
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We have isolated eight PALA-resistant mutants from CHO-PV cells and have shown that the CAD gene was amplified. We then localized the CAD genes with fluorescence in situ hybridization followed by G-banding and identified 10 different marker chromosomes carrying amplified DNA. TTAGGG repetitions, which normally map to the telomeres and centromeres, have also been localized on the 10 marker chromosomes. The organization of amplified genes and of TTAGGG sequences suggests that dicentrics were formed during amplification and that breakage-fusion-bridge cycles may have generated 7 marker chromosomes. One isochromosome was probably derived from abnormal centromere segregation at anaphase. The most striking observation was that TTAGGG sequences of centromeric origin surrounded the amplified regions and were always localized at the telomeres of the chromosome arms carrying amplified DNA. These results indicate that the recombination events that accompanied gene amplification frequently involved centromeric DNA. Moreover, breakage within centromeric TTAGGG repeats may produce telomere-like structures that stabilize the ends of rearranged chromosomes.
The trifunctional enzyme CAD catalyzes the first three steps of pyrimidine biosynthesis. By using fluorescence in situ hybridization we have localized the Chinese hamster CAD gene on chromosome 7q11-q13 of diploid fibroblasts. Other genes previously assigned to chromosome 7 include acid phosphatase-1, the M2 subunit of ribonucleotide reductase and ornithine decarboxylase. These genes are also syntenic with CAD on human chromosome 2p. We have then mapped CAD on the pericentromeric region of two different rearranged chromosomes (Z8p and R2q) in a cell line derived from Chinese hamster ovary. The presence of CAD on Z8 and R2 indicates that they derive from rearrangements involving chromosome 7.
A new cell line (BRC-230) was established from surgical material of primary ductal infiltrating breast carcinoma. The epithelial nature of this cell line was confirmed by ultrastructural analysis and demonstrated the retention of structural properties characteristic of the original tumor. The BRC-230 cell line induced tumor in athymic Cr1:nu/nu(CD-1)BR nude mice, it possessed an abnormal karyotype with a modal chromosome number between 60-61 with eight recurrent marker chromosomes, and it presented a doubling time of 30.5 hr. Scatchard analysis demonstrated that both primary tumor and BRC-230 cells were estrogen and progesterone receptor negative. Immunoenzymatic and radioimmunoassays showed a production of marker antigens (CEA, TPA, CA125, CA15-3, CA19-9) which was similar in the patient's serum and BRC-230 cells. The in vitro drug sensitivity assay of the cell line and of the parental tumor tissue showed overlapping results to all tested antiblastic drugs. BRC-230 cells were resistant to 4-Idroperoxy-cyclophosphamide, Idarubicinol, Mitoxantrone, Etoposide, 4'Epidoxorubicin, and Doxorubicin, showing a multiple drug resistance phenotype. Amplification or rearrangement of Her-2neu, Ha-ras, and C-myc genes was observed neither in the original tumor nor in BRC-230 cells; the mdr-1 gene was also present in a single copy. We conclude from these studies that the BRC-230 cell line maintains the same characteristics as the original tumor and may provide us with a good model to study in vitro the biology of drug resistance of breast cancer.
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Four cell lines (MP1, -4, -5, -7), isolated from baby hamster kidney cells after simultaneous selection with N-(phosphonacetyl)-L-aspartate and methotrexate, have previously been shown to amplify their DNA at an increased rate. We now show that all four lines are hypersensitive to killing by UV light and mitomycin C. At high doses of UV light or mitomycin C, the MP lines survived less than 10% or less than 5% as well as parental cells, respectively. After UV irradiation, inhibition of DNA and RNA synthesis was greater in MP than in parental cells, and recovery was slower or absent. A 2- to 3.5-fold increase in the frequency of UV-induced sister chromatid exchange was also seen in the four cell lines. In MP5, unscheduled DNA replication after treatment with UV light was only approximately 70% as great as in parental cells and the other MP lines. In MP4 and MP7 cells S phase was elongated. Although their individual properties confirm that the four cell lines are independent, their common properties suggest a relationship between tolerance of DNA damage and gene amplification.
An open multicentre trial to study the efficacy and safety of ceftazidime in elderly patients has been conducted in four geriatric units on 135 subjects suffering from urinary-tract or respiratory-tract infections. Sixty-two patients were cured (45.9%), and 60 improved (44.4%). Of the evaluable cases bacteriological eradication was achieved in 91.3%. No adverse events were recorded.
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