| Objective The aim of the research was not only to detect thefunction of the of the EM65-2 on the endometrial carcinoma cell line RL95-2 by in vivo and in vitro test, but also to study the regulation factors of the VEGF expression in endometrial carcinoma.Materials and MethodsThe function of EM65-2 on the RL95-2 was detected by in vivo and in vitro test. We established the protocol of making the human RL95-2 endometrial carcinoma xenografts in the ovariectomized nude mice at first. Secondly we detect the function of EM65-2 on the subcutaneous enografts with or without benzoate estradiol and estrone. The function was evaluated by the change of tumor volume and the weight of the tumor and the uterine at the end of the test. The action of the EM65-2 and estradiol on the RL95-2 cell was determined by the MTT method, plate clone formation test, and micro-plate alkaline phosphatase assay.The regulation factors of VEGF was also detected by the in vivo and in vitro test. We detect the VEGF expression in the previously acquired xenografs and uterine through immumohistochemistry method. The in vitro gregulation factors was evaluated by detecting the VEGF protein level in the condition culture medium.ResultsK We randomizedly divided the balb/c nude mice intonon-ovariectomized , ovariectomized , and ovariectomized supplied with benzoate estradiol groups. The successful rate of tumor transplantation was one hundred percenht. The mean tumor volume and the weight of xenografts among these three groups were significantly different, OVXE2 >NOVX >OVX.2-. The alive cell number was reflected by the MTT method. Thenumber was invarious in the first three days after plantation in respect of the estradiol conditions. The number of the cell increased rapidly from the fourth day to the sixth day. From the seventh day, the cell number began to decrease in the medium without estradiol and still increase slowly in themedium with estradiol. The cell number among different conditions were not significantly different in the first five days and were significantly different from the sixth to the eighth day. The cell number in the 10nmol/l estradiol medium was the highest. The cell number in the medium without estradiol was not changed after different dosage of EM65-2 or 4-hydroxy tamoxifen was added. In the medium with lOnM estradiol, the cell number were not different when different concentration , 103, 10, 10\ 10'3 and OnM, EM65-2 or 4-hydroxy tamoxifen was supplied.3> In varius concentration of estradil and antiestrogen, EM65-2and 4-hydroxy tamoxifen, cultur medium, the result of plating clone formation test was not different.4> The VEGF titration in the conditioned culture medium werecontinuously increase through the course of the test, expect for the instantly decrease at 24th hour. The lowest VEGF titration 250pg/ml occurred at the 2nd hour, and the highest 1725pg/ml present at the 72th hour in the low pH conditionded culture medium. There is significant difference among the 3 different conditioned medium at the 2nd ,8th ,24th, and 72th hour. The VEGF tirtration in the 48th conditioned medium. From the results of the q test, we found that there is significant difference among the 3 different conditioned medium at the 2nd hour, there is significant difference between the low pH conditioned culture medium and the two normal conditioned culture medium with or without estradiol and 8th, 24th and 72th hour. There is no significant diference between the two normal pH conditioned culture medium at 8th, 24th and 72th hour.5-. VEGF was positive in the tissue surround the vascularity inthe muscle of the uterine, but the vascular wall and the endothelial cell were negative. The secretary phase endometrial epithelial cell of the NOVX mice were positive, the epithelial cell of the endometrium of all other groups were all negative. The stromal cell of the uterine endometrium were positive in the...
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