Trypan blue가 흰쥐 태아의 기형발생 발육지연 및 H3-thymidine섭취율에 미치는 영향

Abstract

[한글]

[영문]

Since the discovery that a wide variety of malformations have occurred in the offspring of thypan blue-treated mammals, this teratogen has been used extensively and its site and mode of action have also been discussed in these investigations. Waddington and Carter (1953) suggested that the main efforts of trypan blue were firstly, a general inhibition of development and secondly, a more specific effect on body fluids and circulatory organs in embryo.

There is no doubt that some teratogens can affect maternal protein metabolism in rats. Beaudoin(1968) reported that, among the six diazo dyes trypan blue caused the greater number of malformations. Runner (1968) reported the effect of trepan blue

on chick embryos. Thymidine incorporation rate and protein nitrogen levels were significantly reduced.

Previous investigations have emphasized the teratogenic activities of trypan blue with special reference to possible correlations between the dye and thyroid function.

The present experiment studied the teratogenic and growth retarding effects of trypan blue on the offspring of treated does and the incorporation rate of H**3-thymidine in the several fetal tissues such as the liver, spleen, kidney, skin and cartilage in albino rats.

17 female albino rats weighing 200 to 250 Gm. and 112 fetuses of their offspring were uses in the present study. The animals were divided into experimental and control groups. They were maintained on a normal diet ad libitum. Day 1 of pregnancy was considered to begin on the morning after sperm were found in the

vaginal smear. On day 8, pregnant rats, which consisted of 11 does in the experimental group, received a subcutaneous injection of 50 mg/ kg of 2% trypan blue dissolved in normal saline solution.

The fetuses obtained by a cesarean section on day 20 of pregnancy were examined for external malformations such as vertebral and cranial abnormalities, club-foot, short and curled tail, and absence of tail, etc. They were also measured for body

weight, crown-rump length, length of forelimb, hindlimb, and tail.

In order to find talc effect of trypan blue on the rate of H**3-thymidine incorporation in several fetal tissues such as the limier, spleen, kidney, skin, and cartilage in the lumbosacral region, tail and forelimb, radioautography was done by the stripping and dipping method using methyl-H**3-thymidine (1μCim/Gm. of body weight). The radioisotope was injected intraperitoneally into the pregnant rats on day 19 of pregnancy. Twelve hours later the fetuses were obtained. The frequency of each type, biometrical results of the malformations produced, and the rate of the H**3-thymidine incorporation were studied. Labeled nuclei among 500 cells of the above mentioned organs and tissues were used in the calculations(%).

On fetuses obtained from trypan blue treated·rats, the teratogenic effect, growth retardation and inhibitory result concerning the rate of the H**3-thymidine incorporation in several fetal tissues were obvious:

A. Frequency of Each Malformation.

No malformation was observed in talc control group, but there was a high frequency of malformation(incidence of 34.2%) in the experimental group. The most frequent type of malformation was spina bifida aperta, an incidence of 23. 8%. This abnormality usually appeared in the lumbosacral region of the vertebral column. The high incidence of club-foot (incidence of 12.7%) was also remarkable. The curled tail malformation occurred least with an incidence of 1.6%.

B. Biometrical Observations on Growth Retardation.

The inhibitory Infect of trypan blue on the fetal growth was also very conspicuous in the experimental group compared with that of the control one. The mean body weight and crown-rump length of the experimental fetuses which were 4.0Gm. and 35.7mm. respectively were less than those of the control group which

were 5.2Gm. and 41.7mm. The greatest inhibition of growth was found in the tail.

C. Rate of H**3-thymidine Incorporation.

The inhibiting effects of trypan blue on H**3-thymidine incorporation into the fetal organs were usually quite remarkable in the experimental group, The incorporation rate (6.6%) of the spleen of the experimental group was less than that (10.7%) of the control one. In the kidney there were also significant derceases in the incorporation rate in both the glomeruli and the uriniferous tubules in the experimental group. In the skin, remarkable decreases were also observed in the lumbosacral region, tail and forelimb. No significant decreases in the incorporation rate were observed in the liver.

A general decrease of the H**3-thymidine incorporation rate was observed in the cartilage of the lumbosacral region, tail and fore limb in the experimental group.

Significant decreases of the incorporation rate appeared in the lumbosacral region and tail in the experimental group.

Consequently, results of the teratological studies show that the teratogenic effect of trypan blue on fetuses, growth retarding effects and the rate of H**3-thymidine incorporation are very remarkable. Also there was a High frequency of malformation. The tint frequent type of malformation, spina bifida aperta, occurred in the lumbosacral region of the vertebral column. Also the growth retarding effect of the dye was demonstrated in the body weight, crown·rump length and tail length. Also the inhibitory effects of trypan blue on the H**3-thymidine incorporation rate are easily seen in the skin and cartilage.