Studies on the Activities of Glutathione-S-Transferases and the Metabolism of Benzo($\alpha$)pyrene in the Primary Culture of Mouse Embryonic Fibroblast.

국문 초록 (Abstract)

NIH(GP) 마우스 태아 세포 일차배양계에서 GST활성도와 B($\alpha$)P 대사에 미치는 BHA의 영향에 관한 실험을 수행하였다. 1-Chloro-2,4-dinitrobenzene (CDNB)을 기질로 하였을 때 GST의 최적 pH는 6.5 이었으며, 이때 반응액은 1.0 mM CDNB, 1.0 mM GSH 그러고 $50-100\;{\mu}g$의 세포질 단백으로 조성 하였다. 1,2-Epoxy-3-(p-nitrophenoxy)propane (alkyl epoxide)를 기질로 하였을 때 GST의 최적 pH는 5.8이었으며, 이때 반응액은 0.5 mM alkyl epoxide, 10 mM GSH, $50-100\;{\mu}g$의 세포잘 단백으로 조성하였다. CDNB를 기질로 하는 GST 활성도는 배양 5일에 의미있게 증가되었으며 그 활성도는 배양 7일까지 유지되었다. BHA배지에서 7 일간 배양된 세포내 GST 활성도는 대조군에 비하여 의미있게 증가하였다. $4\;{\mu}M$ $^3H-B(\alpha)P$ (500 mCi/m mol) 첨가하여 24시간 배양한 후 회수한 완전 배지와 BHA배지에서 분리되는 수용성, B(a)P 대사산물의 분획은 배양 3일에 비하여 배양 7일에 현저히 증가하였으며, 이때 BHA 첨가로 인한 영향은 없었다. B($\alpha$)P-DNA, B($\alpha$)P-RNA 그리고 B($\alpha$)P-protein adducts 양은 배양 일수나 배양액의 조건에 의하여 차이가 없었다.

다국어 초록 (Multilingual Abstract)

We examined the effect of BRA on the activities of glutathione-S-transferase(GST) and the metabolism of benzo($\alpha$)pyrene in the primary culture of embryonic fibroblasts derived from the NIH(GP) mice. Optimum pR of GST was 6.5 with 1.0 mM of 1-chloro-2, 4-dinitrobenzene, 1.0 mM of reduced glutathione, and $50-100\;{\mu}g$ of cytosol proteins in 0.1 M potassium phosphate buffer. Optimum pH of GST with 0.5 mM 1,2-epoxy-3 (p-nitrophenoxy) propane, 10 mM of GSH and $50-100\;{\mu}g$ of cytosol proteins in 0.1 M potassium phosphate buffer was found to be 5.8. The activities of GST catalyzing CDNB were increased on the 5th day of the culture and maintained on the 7th day. The culture medium containing 100\;${\mu}M$ BRA significantly increased the activities of GST with CDNB on the 7th day as compared with those of the control. The fractions of the conjugated B($\alpha$)P metabolites in both of the control and the BRA media were increased on the 7th day as compared with those on the 3rd day of the culture, after incubation with $4\;{\mu}M$ $^3H-B(\alpha)P$ (500 mCi/m mol) for 24 hours. However, the binding levels of B($\alpha$)P to the cellular DNA, RNA and proteins were not changed depending on the culture days and the culture medium. BHA had no trans-placental effect on the activities of glutathione-S-transferases and the metabolism of B($\alpha$)P in our experiment.