A Study for Cardiovascular Malformations and Functional Changes Induced by Tetrachloroethylene in Developing Chick Embryos

Kim, Wan; Kim, Won Kyu; Choi, Tae Yeal; Cho, Ja Yeon

Abstract

Chlorinated solvents are recognized as well-known drinking water contaminants and potent teratogens for developing embryos. Among theses compounds, tetrachloroethylene was studied to evaluate the teratogenic effects in detail. This study was also undertaken to understand the teratogenetic mechanisms by assessing the relationship between the physiological impairments and cellular degeneration including apoptosis induced by tetrachloroethylene, and the cardiovascular malformations. Fertilized white leghorn eggs (n=530) were incubated for 90~96 hours up to Hamburger-Hamilton stage 27~28. To the experimental group, tetrachloroethylene was dissolved in mineral oil at concentrations of 40 micrometer and 50 micrometer and was injected into the chorioallantoic membrane. One half of the control was injected with saline and the other half with mineral oil. The experimental animals are studied for ECG changes, and light and electron microscopic observation. Two silver thread electrodes were used to record the ECG. ECG changes were recorded for 6~30 minutes after tetrachloroethylene injections. In parallel studies, blood velocity through the dorsal aorta was measured by a pulse Doppler velocity meter with 20 MHz probe. The specimens were stained with hematoxylin and eosin for light microscopic study and the Hitachi H-800 EM was used for transmission electron microscopic evaluation. The embryos underwent formalin fixation on day 14, and the presence of malformations was observed by a microscopy. The results obtained are as follows: Repeated injections of 40~50 micrometer tetrachloroethylene appeared to have a strong influence on the formation of cardiovascular malformations. Various types of cardiac malformations were observed in the experimental group compared to control group, that were statistically significant (p<0.01) and many embryos also died. The malformations observed were large ventricular septal defect, endocardial cushion defect, double outlet right ventricle and truncus arteriosus associated with single ventricle. Large ventricular septal defect was most common type of cardiovascular abnormalities. Arrythymias including ventricular fibrillations were evident on ECG 6~30 minutes after the injections. Blood flow through the dorsal aorta was markedly decreased in concordance with the ECG changes. A large amount of individual cell death suggesting apoptosis was seen on light microscopic evaluation and the electron microscope revealed cellular degeneration of the embryonic myocardium. We concluded that tetrachloroethylene, like other chloride compounds has potent teratogenic effects. The cardiac malformations were probably caused by decreased blood flow due to physiofunctional changes and cellular necrosis associated with apoptosis. By including birds and mammals in further experiments, we hope to elucidate an embry-ological clue into how early cardiovascular malformations are caused.