Radioactive thymidine

Ohvieri, G., Bodycote, J., and Wolff, S., 1984, Adaptive response ofhuman lymphocytes to low concentrations of radioactive thymidine, Science 223 594-597. 

The mitotic index is the fraction or percentage of cells in mitosis within a given cell population. The thymidine labeling index is the fraction of cells incorporating radioactive thymidine. They represent cells in M-phase and S-phase and define the proliferative characteristics of normal and tumor cells. 

The graph shows E. coli labeled with radioactive thymidine for a short pulse (10 s) followed by a chase with an excess of nonradioactive thymidine. The DNA is then extracted and centrifuged in alkaline sucrose gradients (under high pH conditions the DNA denatures). Explain what these data imply, and interpret these results in light of our current model for DNA replication. 

Cell Proliferation. Increased liver weight is a typical response in rodents exposed to DEHP and other peroxisome proliferators. This response is largely due to a transient increase in replicative DNA synthesis and cell division. Although considered a weak inducer of cell proliferation, DEHP causes an almost immediate increase in cell division in rats and mice (Busser and Lutz 1987 Smith-Oliver and Butterworth 1987). A single dose of 664 mg/kg DEHP produced a significant increase in DNA synthesis in the rat liver, as indicated by the incorporation of radioactive thymidine into polynucleotides during the first 24 hours (Busser and Lutz 1987). In mice, a dose of 500 mg/kg stimulated mitosis within 24 hours of... 

With the introduction of radiochemical methods, DNA biosynthesis and its inhibition is usually followed either by measuring the incorporation of radioactive thymine into thymine auxotrophs of bacteria or the incorporation of radioactive thymidine into prototrophic organisms. In the latter instance, it is practical to include in the experimental medium a large excess of non-radioactive deoxyadenosine in order... 

Adding radioactive thymidine before the OD550 reaches 0.5 results in a higher final specific activity. 

Cell Division Inhibition. If the mitotic index (% of cells in mitosis) decreases to zero during the first few hours of treatment (approximately 8 h), the compound is inhibiting some metabolic process in interphase (Gi, S, or G2) The location and possible cause of the inhibition, which can be determined by autoradiographic techniques, involves treating root meristems with radioactive thymidine after herbicide treatment. The details of this procedure have been published by Van t Hof (33). An alternative to the above procedure is to treat root tissue with herbicide plus radioactive thymidine or uridine and follow the increase of radioactivity precipitable by trichloroacetic acid (34). 

Some of the chromosomal regions boxmd to the nuclear membrane may be the sites where DNA synthesis is initiated (Comings and Kakefuda, 1968). When the site of incorporation of radioactive thymidine was studied at the start of the S phase in synchronized human amnion cell cultures, the label was found to be localized at the nuclear membranes (Comings and Kakefuda, 1968), suggesting that the chromosomal sites at which DNA synthesis was initiated were attached to the nuclear membrane. It was... 

The earliest studies using radioactive precursors to measure the overall rates of synthesis of nucleic acids showed that in growing rabbit kidney cells infected with pseudorabies virus (PRV) the rate of DNA synthesis declined steadily over the first 5 hr and then increased but the total quantity of DNA in the cells did not change significantly (Kaplan and Ben-Porat, 1960). Newton et al. (1962) observed a similar decline in incorporation of radioactive thymidine during the first 6 hr of infection with HSV followed by a gradual rise to 10 hr after infection. The decline was accompanied by release of radioactive material from cells prelabeled before infection with [ HJthymidine (Newton, 1964). 

In the continuous labeling method, which is more frequently used, the cells are in contact with radioactive thymidine from a certain point of the S-phase (first, second, third, etc. hours) until mitosis, in some cases loci synthesizing DNA at the end of the S-phase being labeled, in other cases those synthesizing DNA at the end and in the middle of the S-phase being labeled, in a third group the chromosomes are totally labeled —for the whole period of DNA synthesis, and so on in other variations. Partial incorporation... 

---