Effects of low-dose mifepristone on apoptosis in human ovarian luteinized granulosa cells

AIM: To investigate the effect of different low-dose mifepristone on apoptosis in granulosa cells and to test low-dose mifepristone as an orally contraceptive drug. METHODS: By using immunofluorescence, terminal deoxynucleotidyl transferase-mediated nick end labelling (TUNEL) and flow cytometry technique, the nuclear morphologic features and ratio of apoptosis and fluorescent intensity of caspase-3 in granulosa cells cultured in vitro treated with different low-doses of mifepristone were observed, respectively. RESULTS: By the display of immunofluorescence, the granulosa cells in treatment group were classified as apoptotic cells on the basis of their morphologic features contained a single condensed chromatin, multiple nuclear fragments. The results of TUNEL showed significant difference between control group and groups treated with different concentration of mifepristone (P<0.01). A significant difference (P<0.01) was also observed between the treatment groups with 1.25 μmol/L and 2.50 μmol/L mifepristone. The fluorescent intensity of caspase-3, observed by flow cytometry showed significant difference (P<0.01) between control group and treatment groups. CONCLUSION: Granulosa cells are induced to apoptosis by low-dose mifepristone, which may be regulated by the activation of caspase-3.     

The effect of mifepristone on natural killer subsets of peripheral blood and decidua in early pregnancy

AIM: To investigate the effect of mifepristone on natural killer(NK) subpopulations of peripheral blood and decidua in early pregnancy. METHODS: Flow cytometry was used to detect the expression of CD56 and CD16 on lymphocytes of decidua and peripheral blood in early pregnancy, mifepristone-treated pregnancy. RESULTS: The percentages of different natural killer subsets in peripheral blood between early pregnancy and mifepristone-treated pregnancy were almost identical. The serum levels of estradiol and progesterone in mifepristone-treated pregnancy were slightly higher than in early pregnancy. The percentage of decidual CD56⁺NK cell in early pregnancy was significantly higher than in mifepristone-treated pregnancy. The CD56⁺NK cells were predominant lymphocyte population of decidua in mifepristone-treated pregnancy, but in which the CD56⁺CD16⁺ and CD16⁺NK cells were major lymphocyte subpopulations of peripheral blood. CONCLUSION: Mifepristone acted principally on feto-maternal interface, it blocked the proliferation and differentiation of decidual CD56⁺NK cells and induced embryo immune rejection.     

Study of establishment of K562/HHT cell line and reversal of multidrug resistance by antiprogestin drug mifepristone

AIM: To study the mechanism of multidrug resistance (MDR) of leukemia cells induced by homoharringtonine (HHT) and the reversal effect of mifepristone on MDR.METHODS: Human leukemia cell line K562 was induced into MDR cell line by intermittent administration of high dose of HHT.MTT assay was used to detect the sensitivity of these MDR cells to all sorts of chemotherapeutic agents with or without mifepristone.The cytotoxicity of mifepristone was also observed.RT-PCR was used to detect the expression of MDR1 gene and glucosylceramide synthase (GCS) gene.Flow cytometry was used to detect the expression of P-glucoprotein and the accumulative value of intracellular daunorubicin (DNR) in these MDR cells with or without mifepristone.Immunohistochemistry was used to detect the expression of Bcl-2,Bax and caspase-3 in these MDR cells with or without mifepristone.RESULTS: MDR cell line K562/HHT was acquired after induced by HHT for 2 months.This MDR cell line possessed the ability of 462.6 fold resistance to HHT and cross-resistance to adriamycin,vincristine and etoposide.The expression of MDR1 gene,GCS gene,P-glucoprotein and Bcl-2/Bax ratio in K562/HHT cells were significantly higher than those in K562 cells (P<0.05).The caspase-3 expression and the accumulative value of intracellular DNR in K562/HHT cells were significantly lower than those in K562 cells (P<0.05).10 μmol/L mifepristone reversed the resistance of K562/HHT cells to HHT,adriamycin,vincristine and etoposide at different levels.The Bcl-2/Bax ratio,caspase-3 expression and accumulative value of intracellular DNR in K562/HHT cells treated with RU486 were significantly different compared with K562/HHT cells without RU486 treatment (P<0.05).CONCLUSIONS: Leukemia cell line K562 can be induced into MDR cell line K562/HHT by HHT.P-glucoprotein,GCS,Bcl-2/Bax ratio and caspase-3 may play an important role in K562/HHT cells.Mifepristone can reverse MDR in K562/HHT cells by decreasing the accumulative value of intracellular drug and regulating the expression of Bcl-2,Bax and caspase-3.     

莫沙必利的妊娠安全性研究

为准确判断孕鼠妊娠日期及给药后流产数和探究莫沙必利对小鼠的妊娠安全性及相关作用机制,将99只小鼠按雌雄比2∶1随机分组进行妊娠造模,根据母鼠体重、外形及阴部变化情况综合判断小鼠妊娠日期,造模成功后将部分妊娠13 d小鼠分为空白对照组、米非司酮加米索前列醇阳性对照组、莫沙必利组,给药后观察整体子宫、胎儿及胎盘发育情况判断莫沙必利对妊娠后期小鼠流产率的影响;将剩余造模成功小鼠随机分为空白对照组、米非司酮加米索前列醇阳性对照组、莫沙必利组,并于妊娠15 d给药后采血,测定血清中缩宫素、孕酮、雌激素、前列腺素含量。结果显示：合笼30 d内共怀孕小鼠60只,其中56只可准确判断其妊娠日期并用于后续试验,占总试验母鼠的84.84%;给药后莫沙必利组胚胎均正常发育,阳性组流产率为100%,空白组流产率为1.79%;激素测定试验中莫沙必利组各项数据与空白组差异均不显著,与阳性对照组相比,血清雌激素,孕酮,前列腺素含量差异极显著。结果表明：试验中所建立的判断小鼠妊娠日期及流产数的方法准确度高且操作简便;初步认为莫沙必利对妊娠后期小鼠的胚胎发育无影响;莫沙必利对部分潜在致孕鼠子宫收缩而造成流产的性激素分泌无影响。     

Inhibitory effect of mifepristone on growth and stemness of olaparib-resistant ovarian epithelial cancer cells

AIM: To investigate the effect of mifepristone on the growth and stemness of olaparib-resistant ovarian epithelial cancer (OEC) cells. METHODS: The primary cells of OEC were isolated from 5 OEC tumor samples and respectively cultured. OEC/olaparib cells resistant to olaparib, which were paired with OEC cells isolated from the 5 samples, were established by successive increments of drug concentration. The sensitivity of OEC cells and paired OEC/olaparib cells to olaparib was evaluated by MTT assay. Five groups of OEC/olaparib cells were constructed and the effect of mifepristone on the viability of OEC/olaparib cells was measured by MTT assay. One group of OEC/olaparib cells was selected to detect the effect of mifepristone on the colony formation ability of OEC/olaparib cells by colony formation assay. The expression levels of stemness markers in the OEC/olaparib cells were determined by RT-qPCR and flow cytometry. The effect of mifepristone on the growth ability of OEC/olaparib cells transplanted into nude mice was tested by tumorigenesis assay. RESULTS: The OEC cells and olaparib-resistant OEC/olaparib cells were successfully established in this study. Mifepristone inhibited the growth of OEC/olaparib cells in vitro and in vivo, and reduced the stemness of OEC/olaparib cells. CONCLUSION: Mifepristone inhibits the growth and stemness of olaparib-resistance OEC cells.     

Effects of mifepristone on ultrastructure of human endometrium in the early secretory phase

AIM:To investigate the influence of mifepristone on ultrastucture of human endometrium in the early secretory phase. METHODS: Endometrial tissue was obstained from 10 patients of reproductive age, who underwent a hysterectomy within 1 week postovulatory for gynecologic diseases not involving the endometrium. Patients were divided into mifepristone group (n=5) and control group (n=5) randomly. Each patient in the mifepristone group had taken 25 mg mifepristone per os 24 h before the operation was performed, while none of the control group had taken mifepristone. After removal of uterus, endometrial tissue was immediately acquired and prepared for electron microscopic examination. RESULTS:In comparison with the control group, the endometrial tissue in mifepristone group displayed the following distinctly morphological changes: (1) In the endometrial epithelium neither nucleolar channel system nor giant mitochondrium was seen, and subnuclear glycogen accumulation was seldom observed, but giant lysosomes were frequently found. (2) The intercellular spaces of the epithelium were narrow and straight, the indigitations of lateral plasma membranes were rarely visible. (3) Cytolysis and karyopyknosis of stroma cells and extravasal red cells were repeatedly observed.CONCLUSION:The above ment ioned morphological changes in endometrium in the early secretory phase caused by mifepristone are undoubtedly sufficient to prevent implantation.Consequently, mifepristone may have a contraception effect.     

米非司酮药物流产后发生恶性滋养细胞肿瘤2例临床分析

目的探讨应用米非司酮行药物流产后发生恶性滋养细胞肿瘤的临床特点及其与不全药物流产的鉴别诊断.方法对2例应用米非司酮行药物流产后发生恶性滋养细胞肿瘤的患者进行临床分析.结果药物流产后发生恶性滋养细胞肿瘤患者与不全药物流产一样,表现为不规则阴道出血,β-hCG高于正常,但前者动态监测或刮宫术后β-hCG呈明显上升趋势;彩色多普勒血流图显示子宫动脉扩张,子宫旁血管有滋养细胞浸润,可见肿瘤染色、动静脉漏等.结论米非司酮药物流产后,阴道异常出血者有早期恶性滋养细胞肿瘤发生的可能,动态监测β-hCG、彩色多普勒血流图及血管减影术是诊断药物流产后发生恶性滋养细胞肿瘤及其与不全流产鉴别诊断的有效方法.     

Influence of Radix scutellariae on Th1/Th2 cytokine balance in RU486-induced abortion in mice

The aim of this study is to investigate the significance of Th1/Th2 cytokine balance in the uterus in the early embryo loss (or resorption), and to elucidate immunological modulation at the maternal-fetal interface with Chinese herbal medicine Radix scutellariae (Huang Qin) and its constituents (Baicalin and Baicalein). Mifepristone (RU486) was given via subcutaneous injection in the scapular area to induce abortion in mice at day 7 of gestation. The levels of uterine Th1 cytokines (IFN-γ, IL-2) and Th2 cytokines (IL-4, IL-10) were analyzed by enzyme-linked immunosorbent assay (ELISA), respectively. The mean values of Th1 cytokines in the uterus of RU486-treated abortion mice were significantly higher (P < 0.05) than that of the control mice, but no significant difference was observed regarding to the contents of Th2 cytokines of different groups (P > 0.05). However, when the Radix scutellariae and its constituents were used to prevent RU486-induced abortion, the levels of IFN-γ and IL-2 decreased while that of IL-4 and IL-10 increased. The embryo loss induced by RU486 was closely related to the Th1/Th2 immune balance at the maternal-fetal interface. Radix scutellariae and its constituents have an anti-abortive effect through restoring the Th1/Th2 balance at the maternal-fetal interface.     

米非司酮配伍米索终止孕11～18周妊娠的观察

目的探讨米非司酮配伍米索在终止孕11～18周的妊娠的可行性、安全性及优缺点,方法对2001年12月-2003年12月收治的妊娠11～18周且自愿要求终止妊娠的妇女162例进行回顾性分析,结果米非司酮配伍米索终止孕11～18周妊娠成功率95％，且较以往的钳刮术及小月份中引术痛苦小、并发症少、出血量少,结论米非司酮配伍米索终止孕11～18周妊娠安全、有效，且痛苦小、并发症少，但需住院观察