【作者】 陈波；

【导师】 徐克惠； 岳焕勋；

【作者基本信息】 四川大学 ， 妇产科学， 2007， 硕士

【摘要】 目的：米非司酮可延迟、抑制卵泡发育和排卵，停药后卵泡发育及排卵在短时间内得到恢复。本实验通过给予大鼠不同剂量米非司酮灌胃，初步探讨不同剂量米非司酮对大鼠卵巢卵泡发育、雌激素分泌的影响，特别是对卵巢储备功能的影响，为临床安全用药提供理论依据。方法：选用体重230±20g雌性未孕Sprague-Dawley（SD）大鼠40只，每日取阴道细胞学，巴氏染色，有连续两个规律动情周期者纳入研究，随机分为米非司酮低剂量组、米非司酮中剂量组、米非司酮高剂量组及空白对照组共四组，于动情间期早期（相当于早卵泡期）开始分别予米非司酮1．04（mg／kg／d）、2．604（mg／kg／d）、10．4（mg／kg／d）及生理盐水灌胃，连续四周。用药结束后于动情间期处死每组动物各5只，剩余大鼠停药后每日取阴道细胞学，在停药后第三个动情间期早期（相当于早卵泡期）处死。所有大鼠卵巢常规石蜡包埋、HE染色，显微镜观察卵巢组织形态学改变，并采用体视学计数始基卵泡和初级卵泡卵泡数目；化学发光法检测血清雌二醇（E₂）水平变化；用药4周结束时，采用免疫组化检测caspase～3在生长卵泡中的表达变化及半定量分析。结果：1．卵巢形态学观察：停药后对照组可见大量生长卵泡，实验组各级生长卵泡减少，闭锁卵泡增多；停药后第三个动情周期，各组均可见大量生长卵泡，闭锁卵泡较少：所有实验动物均可见大量始基卵泡及初级卵泡，分布于卵巢皮质下。2．体视学分析：停药后，对照组与各实验组始基卵泡和初级卵泡数目组间无差异（P＞0．05）；停药后第三个动情周期，对照组与各实验组始基卵泡和初级卵泡数目组间无差异（P＞0．05）。3．血清雌二醇水平变化：停药后各实验组血清E₂水平较对照组下降，随用药剂量的增加呈下降趋势，对照组与低剂量组、中剂量组组间差异无统计学意义（P＞0．05），中剂量组与低剂量组、高剂量组组间差异无统计学意义（P＞0．05），高剂量组与对照组、低剂量组有差异（P＜0．05）：停药后第三个动情周期，低剂量组血清E₂水平较前升高不明显，中剂量组及高剂量组血清E₂水平较前升高，各实验组与对照组E₂水平组间无差异（P＞0．05）。4．caspase-3免疫组化：各级生长卵泡颗粒细胞中活化caspase-3蛋白表达，正常对照组颗粒细胞胞浆呈浅棕黄色、低剂量组、中剂量组颗粒细胞胞浆呈棕黄色表达，高剂量组绝大多数颗粒细胞胞浆呈棕黄至棕褐色表达，部分出现胞核阳性表达。随着用药剂量增加，蛋白表达增强，各组IOD值呈上升趋势，组间差异有统计学意义（P＜0．05）。结论：1．动物实验表明用不同剂量米非司酮四周可抑制卵泡生长、不同程度降低血清E₂水平，随着药物剂量的增加对卵巢抑制作用增强，停药后第三个动情周期，卵泡生长发育及血清E₂水平可得以恢复，不同剂量米非司酮不引起始基卵泡和初级卵泡数目减少。提示米非司酮可能作用于生长卵泡，抑制卵泡发育、引起卵泡闭锁，而对始基卵泡及初级卵泡影响较小，表明对卵巢储备功能没有影响，临床用药安全；随着米非司酮剂量增加，对卵巢抑制作用增强，如何选择最佳药物剂量，在临床应用中取得较好治疗效果同时产生较小副作用，还需进一步研究。2．动物实验表明用不同剂量米非司酮四周后可引起生长卵泡中颗粒细胞的凋亡，药物剂量增加，促凋亡作用增强：调节caspase-3蛋白表达是米非司酮诱导颗粒细胞凋亡的途径之一。颗粒细胞凋亡可能是米非司酮引起卵泡闭锁、抑制卵泡生长及雌激素分泌的机制。更多还原

【Abstract】 Objective:The development of follicular and ovulation can be delayed or suppressed by mifepristone. When the treatment finishied, ovarian function will recover soon. Different doses of mifepristone were administered to rats to study the effects on the ovarian function, especially on the ovarian reserve function, to supply safety evidence for clinical medication.Method:Fourty Sprague-Dawley(SD) female rats,weight 230±20g,were divided into four groups randomly:low dose of mifepristone group,middle dose of mifepristone group,high dose of mifepristone group and blank group. Daily vaginal smears were taken and stained using Quick Pap staining to ascertain cyclicity and only the rats that showed two consecutive estrous cycles were used for the experiments. Mifepristone tablets were given through gastromy for three different groups, 1.04(mg/kg/d),2.604(mg/kg/d), 10.4(mg/kg/d) , respectively in dioestrus for four weeks. Five rats of each group were sacrificed after treatment in dioestrus,the rests were killed in the third dioestrus after treatments. The ovarians of all rats were paraffin imbedded and serially sectioned at 8μm and stained with hematoxylin and eosin stain. Histology of every ovarian were observed and the total number of primordial and primary folliculars were couted by Stereology. The estradiol lever of serum were detected by chemiluminescence. The immunohistochemical SP and gray scales scan quantity analysis for the expression of active caspase-3 protein in the granulosa cells of developing follicles of the rats sacrificed after treatment in dioestrus.Results:1. The number of developing follicles decreased while corpora atreticas increased in mifepristone groups which were sacrificed after treatment in dioestrus. A great quantity of developing follicles were observed in other groups. Considerable of primordial and primary folliculars, which located down of ovarian cortex, were observed in various groups.2. The number of primordial and primary follicles in disparate groups have no significant differences.3. After treatment, Estrogen level decreased with the accrescenced dosage of mifepristone, there were differences between blank group and high dose group. Estrogen level of mifepristone groups were recovered in the third dioestrus after treatments.4. The active caspase-3 proteins, which were detected by immunohistochemistry, were expressed in granulosa cells of developing follicles,the positive expression increased with the accrescenced dosage of mifepristone. Conclusion:1. Four weeks using of mifepristone can inhibin the growth of developing follicles and depress the surum estrogen lever with dose dependent, ovarian function will recover in the third dioestrus after treatments. Four weeks using of mifepristone can’t induce atresia of primordial and primary folliculars, have no influence on the ovarian reserve of rats.2. Modulating the expression of caspase-3 protein may be one of pathway correlate with apoptosis of granular cells in ovarian induced by mifepristone, which can induce atresia of developing folliculars.更多还原