Biomedical subjects

J Maio

Publications and source records attributed to J Maio.

5 recordsLinked to original sources

PubMed1990

DNA base and strand damage in X-irradiated monkey CV-1 cells: influence of pretreatment using small doses of radiation.

Base damage in alpha DNA from irradiated monkey CV-1 cells was determined by measuring release of 5'-32P-end labelled DNA fragments after digestion with endonuclease III of E. coli. The frequency and base sequence locations of the enzyme-sensitive sites were determined. Fragments were released from irradiated DNA at sequence sites of pyrimidines and guanines. The time for repair of half the single strand breaks was approximately 1.5 h. Repair of base damage as judged from loss of enzyme-sensitive sites in DNA was slower, with more than half of the damaged bases still detectable after 4 h of repair. Two important changes in the pattern of fragment release from DNA were produced when small radiation doses preceded the large ones needed to produce measurable DNA strand breaks and base damage. 5 Gy to cells incubated several hours before 320 Gy increased by five-fold the abundance of small DNA fragments with 3'-phosphoryl termini detected in high-resolution denaturing gels. These increases were detectable with doses as small as 0.2 Gy and were accompanied by the appearance of new species of DNA fragments of intermediate mobility at specific locations in the base sequence. The patterns resemble those produced by digesting DNA from heavily irradiated cells with endonuclease III.

Animals↗

PubMed1988

Base sequence damage in DNA from X-irradiated monkey CV-1 cells.

Two kinds of 3'-ends were detected in DNA scission fragments of highly repetitive primate component alpha DNA which were isolated from irradiated monkey CV-1 cells. The fragments' 3'-ends were characterized by 5'-32P-end labelling the DNA, followed by examination in high-resolution polyacrylamide gels under denaturing conditions. Hydrolysis of the labelled fragments' termini with exonuclease III of E. coli or by the 3'-phosphatase activity of T4 polynucleotide kinase generated a third, slowest migrating species in each mobility size class. Reference to mobility size class standards makes it highly probable that the fragment ends generated by X-rays in cells are 3'-phosphoryl and 3'-phosphoglycolate, and that they are converted to slower migrating fragments with 3'-OH ends, similar to results obtained with DNA irradiated in water (Henner et al. 1982, 1983 a, b). Densitometer measurements of gel autoradiograms showed that X-ray induction of DNA fragments with 3'-phosphoryl and 3'-phosphoglycolate ends was dose-dependent over a range 100-900 Gy. In CV-1 cells the frequency of single-strand breaks in alpha DNA was 8.6 x 10(-7) breaks/nt/Gy. The two kinds of ends disappeared in post-radiation incubation with a half-time of 1.6 h. These results provide a new means to study X-ray damage and repair of specific sequences in animal cells.

Animals↗

PubMed1987

Expression of prokaryotic genes after transfection of X-irradiated plasmids into primate cells.

Expression of the prokaryotic gene for chloramphenicol acetyltransferase (EC 2.3.1.28) (CAT) in primate cells transfected with X-irradiated plasmid pSV2CAT was determined in transient expression assays. CAT expression did not depend upon the presence of supercoiled plasmids, but relaxed circular forms were essential. X-ray conversion of relaxed circles to linear forms paralleled the loss of CAT expression, with identical D0's in the first part of dose-response curves. X-ray-induced loss of supercoiled forms was complete at much lower doses. The D0 for inactivation of CAT expression by X irradiation of the plasmids in 1 mM Tris buffer was 270 Gy; it was 13 Gy for plasmids irradiated in water. The D0's for conversion of pSV2CAT to relaxed circle forms were only one-seventh as large as the D0's for CAT inactivation after X-ray in water or in 1 mM Tris buffer. Expression of the CAT gene in some representative repair-deficient human fibroblasts transfected with X-irradiated pSV2CAT was less than in monkey CV-1 cells or cell lines from normal human subjects. These results demonstrate a novel means to study low levels of X-ray damage in DNA correlating specific X-ray damage in the DNA with expression of the gene in unirradiated primate cells.

Acetyltransferases↗

PubMed1986

X-ray-induced base sequence damage in primate alphoid DNA.

Radiation-induced single-strand breaks were found throughout the 172 bp repeat units of African green monkey component alpha DNA. Two kinds of 3'-ends of 5'-32P-labeled restriction fragments were found, as previously described by others. After irradiation in vitro, the yield of single-strand breaks was 4 X 10(-5) breaks/nucleotide/Gy, as determined by analyses in DNA sequencing type gels. Protection from X-ray damage was found when the DNA received 150 Gy in the presence of 2-mercaptoethanol. The results demonstrate a very sensitive quantitative means to study the role of indirect effects of ionizing radiation on strand-break induction and protection at the base sequence level. Component alpha DNA was isolated from irradiated CV-1 cells and was analyzed for single-strand breaks. Under these conditions the frequency of breaks was less than the frequency obtained when purified DNA was irradiated. The methodology is presented because of its relevance to the study of DNA strand breakage in living cells.

Animals↗

PubMedDate not supplied

Concomitant treatment with mild hyperthermia, cisplatin and low dose-rate irradiation in human ovarian cancer cells sensitive and resistant to cisplatin.

Hyperthermia has been shown to be an effective radiation sensitizer. Cisplatin has also been shown to cause radiosensitization. In part, the sensitization is through the inhibition of repair of radiation damage. In this study we have set out to combine low dose-rate irradiation (during which extensive repair occurs) with both cisplatin and hyperthermia to maximize the radiation sensitizing effect. Two human ovarian carcinoma cell lines, one parental (A2780S) and the other a cisplatin resistant derivative (A2780CP) cell line were used in these experiments. Long duration hyperthermia at 40 degrees C was combined with low concentrations of cisplatin (0.5-3 microg/ml) and low dose-rate irradiation (LDRI). The responses to the individual treatments showed that there was cross resistance in the two cell lines for cisplatin and radiation, but for hyperthermia the opposite effect was found. When all treatments were given concurrently the response was greater than the calculated response of all three individual treatments, indicating a synergistic interaction. The effect was greater in the cisplatin resistant cell line. The combination of mild hyperthermia, low dose cisplatin and LDRI are a good combination for potential clinical application. In addition, this could be a good approach to deal with cisplatin resistance.

Cell Survival↗