PURPOSE: To gain insight on transcriptional repression of Topo II a in HL-60 cells arrested to G2/M and M phase, the levels of Topo IIa mRNA and the binding activity of ATF have been investigated with Northern blot hybridization and DNA mobility shift assay, respectively. MATERIALS AND METHODS: HL-60 cells were grown in RPMI 1640 medium supplemented with 10% heat-mactivated fetal bovine serum and antibiotics in a humidified 5% CO2 at 37C degree. Total RNA was prepared by a modification of the method of Karlinsey et al. Northern blot hybridization was performed by the method of Virca et al. A Xho I-Mlu I fragment of phTOP2 was used as probe for Northern blot analysis of Topo II a mRNA. DNA mobility shift assay was performed by the method of Lim et al. End labeled DNA oligomer (upper strand, 5-TCTCCGCTATGACGCCGAGTGGTG-3) for ATF binding activity was mixed with nuclear extracts in a 20 pl reaction volume containing 60 mM KC1, 12 mM HEPES, pH 7.9, 5 mM MgCl2, 0.2 mM EDTA, 0.2 mM DTT, 12% glycerol, and 2 ug of poly [dI-dC]. RESULTS: HL-60 cells were arrested at G2/M phase and M phase after taxol or nocodazole treatment. The levels of Topo II a mRNA were reduced at 24 hours after exposure with nocodazole or taxol but the unknotting activities were not changed. DNA mobility shift assay using oligonucleotide containing the ATF binding site showed that ATF binding activity was reduced after pretreatment of nododazole or taxol. CONCLUSIONS: These results suggest that the reduction of ATF binding activity may be important to transcriptional repression of Topo II a gene by nocodazole and taxol in HL- 60 cells.
Anti-Bacterial Agents ; Binding Sites ; Blotting, Northern ; Cell Division* ; DNA Topoisomerases, Type I* ; DNA Topoisomerases, Type II* ; DNA* ; Edetic Acid ; Electrophoretic Mobility Shift Assay ; Genes, vif ; Glycerol ; HEPES ; HL-60 Cells ; Humans ; Hydrogen-Ion Concentration ; Magnesium Chloride ; Nocodazole ; Paclitaxel ; Repression, Psychology ; RNA ; RNA, Messenger

Objective To explore the correlation of caries susceptibility of patients with the immediate tooth polish,application of fluoride,and the patients' oral hygiene habits after interproximal enamel reduction (IER) in clinical practice.Methods The present study retrospectively collected 45 cases of IER (18 males and 27 females) in 196 tooth sites.Each sample was examined clinically and radiologically to obtain the incidence of caries,and the outcome was correlated with amount of IER,tooth polish,application of fluoride,and the brushing method and frequency for statistical analysis.Results The present study showed that there was no statistical significance among the amount of IER,the site of IER,the patients' brushing method,and the gender with the susceptibility of caries.However,there was statistical significance among the application of fluoride,tooth polish and brushing frequency with the susceptibility of caries.Conclusions The current research suggests that it is desirable to polish tooth and apply fluoride for patients with IER in clinical practices.In the meanwhile,the clinician should pay attention to patients' oral hygiene habits,especially brushing frequency,due to the positive correlation between the brushing frequency and incidence of caries.