Development of Model System for Radiation Induced Congenital Malformations by Whole Embryo Culture

Park, Hyoung Woo; Seung, Jin Sil; Kim, Myung Hee; Chung, Eun Ji; Seol, Eun Young; Yoo, Byoung Ki

Abstract

It is known that radiation can induce various kind of mutations, cancers and congenital malformations. Day 9 rats embryos were cultured by whole embryo culture method and irradiated 0.1, 0.5, 2 and 5 Gy at day 10 to study the effect of radiation on the development of rat embryos. Haversting after 48 hours culture, the morphological changes and apoptosis were investigated. In addition, we assessed the expression patterns of p53, WAF1, Bcl -2, Bcl -x and Bax. Compared to the control group, no remarkable morphological changes were observed in the low dosage group (0.1 and 0.5 Gy). But at high dosage group (2 and 5 Gy), growth was retarded and the heart beats were weak. The crown rump length, the number of somites, and branchial arch were decreased and the rotation of embryo and development of otic pit and lens pit, and upper limb bud was delayed significantly. Especially in the 5 Gy group the most of external morphology were difficult to discriminate. In histological observations, high dosage group showed marked increase in the number of apoptotic bodies in the optic cup, trigeminal ganglion, pharyngeal arches, heart and small intestine. In western blot analysis, Bcl -x and Bax were not expressed in all groups. Expression level of Bcl -2 was constant regardless of the amount of radiation. Expression level of p53 and WAF1 increased significantly in accordance with the increasing amount of radiation. Especially, WAF1 was expressed 7.2, 6.3, 9.9 and 11.3 folds more than the control groups when embryos were exposed to 0.1, 0.5, 2 and 5 Gy, respectively. Considering WAF1 arrests cell cycle, we concluded that cell cycle was affected most sensitively to radiation injury. From these results, radiation showed growth retardation, decrease in protein synthesis, increase in apoptosis and expression of related genetic materials. These results may be used as a standard to test the effect of drugs for reducing and protecting agents against deterious effect of radiation in developing embryo and fetus.