Suicide gene transfer has been study extensively for therapies in various human diseases. We can evaluate cellular activity of thymidine kinase and cytotoxic effect in colon cancer cells after suicide gene transfer. We observed cellular expression of green fluorescence protein after transfer with adenovirus into colon adenocarcinoma HCT-15 cells. After transfer HSVtk, we also estimated thymidine kinase activity using [3H]-penciclovir and cellular cytotoxicity by MTT assay. After transfer green fluorescence protein into HCT-15 cells, we could observed fluorescence expression in 10 moi concentration. Expression level of green fluorescence protein markedly increased in 30 moi and most of HCT-15 cells expressed green fluorescence protein in 100 moi. By infection with HSVtk in HCT-15 cells and HT-29 cells, thymidine kinase activity in HCT-15 cells was about two fold higher than that HT-29 cells. Thymidine kinase activity at 1 moi concentration makes no difference with 0 moi in both cells. At 10 moi concentration, thymidine kinase activity increased about three fold compared with 1moi in HCT-15 cells, but not observed high increase in HT-29 cells. Thymidine kinase activity at 100 moi showed about three fold increase in HCT-15 cells and one and a half fold in HT-29 cells compared with 10 moi. By treatment of HSVtk at various mois and ganciclovir to HCT-15 cells, we could find that increased cytotoxic effect according to HSVtk concentration. Cellular cytotoxic effect was slightly appeared at 5 moi concentration and intensively increased at 30 moi concentration, dead colon cancer cells were reached about 30% of total colon cancer cells. Cellular cytotoxic effect was consistently increased until 50 moi, and about 50% of cells at 100 moi and less then 50% of HCT-15 cells at 200 moi were survived. Finally, we can identify that suicide gene transfer into HCT-15 cells is performed according to concentration of suicide gene and thymidine kinase activity also increase with HSVtk concentration in both HCT-15 cells and HT-29 cells. Additionally, we also find that suicide gene therapy by HSVtk with ganciclovir intensively increase cellular cytotoxicity in colon cancer cells. Therefore, our findings suggest that suicide gene therapy by HSVtk can affect cytotoxicy for colon cancer cells and eventually seems to influence therapeutic efficacy.

The objective of this study was to examine the effect of thymidine treatment during in vitro maturation (IVM) of porcine follicular oocytes on blastocyst development. Porcine oocytes were treated with thymidine (10 mM, 20 mM and 30 mM) for 2 or 6 hr in the preiods of IVM I and/or II. The survival rates of the blastocysts in the 6 hr treatment groups of 10 mM and 20 mM during IVM I period were significantly higher than those of control group (p<0.05). However, the survival rate of the blastocysts in the 2 hr treatment group of 20 mM during IVM II period was significantly higher than control group (p<0.05). Furthermore, the survival rate of the blastocysts in the 6 hr treatment group of 30 mM during IVM II period was significantly lower than control group (p<0.05). Consistent with the previous result, blastocyst development of both IVM I and II treatment group was also showed as similar pattern. Total and apoptotic cell numbers of blastocysts derived from thymidine treated porcine oocytes were examined by using Tunel assay. The results showed that there was no significant differences in total cell number of blastocysts between thymidine treated and untreated groups. However, apoptosis-positive cells in the thymidine treated group (6 hr IVM I) were significantly lower than those of other groups (p<0.05). Taken together, these results indicate that high quality oocytes were selected by DNA synthesis mechanism according to high concentration thymidine treatment during porcine oocyte maturation. Therefore, we concluded that presumptive selected oocytes by thymidine treatment during maturation periods improved the further embryo development and embryonic quality of IVF embryos by decreasing the incidence of apoptosis in preimplantation porcine embryos.