MOLECULAR ANALYSIS OF RADIATION-INDUCED MUTATION IN EXON 7/8 OF RAT HPRT GENE

Objective To investigate the relationship between the radiation dose and the HPRT gene lo-cus mutation in rat smooth muscle cells, and provide the molecular basis for prevention of restenosis after percutaneous transluminal coronary angioplasty (PTC4). Methods The smooth muscle cells cultured in vitro were irradiated by radionuclide 188Re in different doses. HPRT gene mutation colonies were selected and isolatedby 6-thioguanine. Analysis of mutation in exon 7/8 of HPRT gene were accomplished by polymerase chain reaction and single-strand conformation polymorphism. Results The HPRT gene mutation frequency of rat smooth muscle cells that were irradiated by radionuclide 188Re ranged from 5.5× 10-6 to 13 ×10-6. Of 91 HPRT gene mutation colonies, 13 (14.3%) contained exon 7/8 deletion and 15(16.5%) had point mutation.The exon 7/8 mutation frequency was 30.8% . There were significant relationships between radiation dose and mutation frequency of HPRT gene and exon 7/8 . Conclusion The DNA damage and gene mutation inducedby radiation has positive relationship with radiation dose, and is a basis of proliferation inhibition and apopto-sis of smooth muscle cells.

Genotoxic and Nongenotoxic Effects of Glycidyl Methacrylate on Human Lung Fibroblast Cells

Objective To evaluate the genotoxic and nongenotoxic effects of short-term exposure to glycidyl mathacrylate (GMA) on human lung fibroblast cells (2BS cells) in vitro. Methods DNA strand breakage was determined by single cell gel electrophoresis, and DNA ladder formation assay and flow cytometric analysis were carried out to detect apoptic responses of cells to GMA exposure. The HPRT gene mutation assay was used to evaluate the mutagenicity, and the effect of GMA on gap junctional intercellular communication (GJIC) in the exposed cells was examined with the scrape loading/dye transfer technique. The ability of GMA to transform 2BS cells was also tested by an in vitro cell transformation assay. Results Exposure to GMA resulted in a dose-dependent increase in DNA strand breaks but not apoptic responses. GMA was also shown to significantly induce HPRT gene mutations and morphological transformation in 2BS cells in vitro. In contrast, GMA produced a concentration-dependent inhibition of GJIC. Conclusions GMA elicits both genotoxic and nongenotoxic effects on 2BS cells in vitro. The induction of DNA damage and gene mutations and inhibition of GJIC by GMA may casually contribute to GMA-induced cell transformation.

Response of Lymphocytes to Radiation in Untreated Breast Cancer Patients as Detected with Three Different Genetic Assays

Objective To detect the response of lymphocytes to radiation in untreated breast cancer patients with three different genetic assays. Methods Blood samples were collected from 25 untreated patients and 25 controls. Each blood sample was divided into two parts： one was irradiated by 3-Gy X-ray （irradiated sample）, the other was not irradiated （non-irradiated sample）. The radiosensitivity of lymphocytes was assessed by comet assay, cytokinesis-block micronucleus （CBMN） assay and 6-TG-resistant cells scored （TG） assay. Results The baseline values of micronucleated cell frequency （MCF） and micronucleus frequency （MNF） in the patients were significantly higher than those in the controls （P〈0.01）, and 3-Gy X-ray induced genetic damage to lymphocytes in the patients increased significantly as compared with that in the controls as detected with the three genetic assays （P〈0.01）. The proportion of radiosensitive cases in the patient group was 48% for the mean tail length （MTL）, 40% for the mean tall moment （MTM）, 40% for MCF, 44% for MNF, and 48% for mutation frequencies of the hprt gene （Mfs-hprt）, respectively, whereas the proportion of radiosensitive cases in the control group was only 8% for all the parameters. Conclusion The difference in the lymphocyte radiosensitivity between the breast cancer patients and the controls is significant. Moreover, there are wide individual variations in lymphocyte radiosensitivity of patients with breast cancer. In some cases, the radiosensitivity of the same patient may be different as detected with the different assays. It is suggested that multiple assays should be used to assess the radiosensitivity of patients with breast cancer before therapy.