子宫内膜蜕膜化的特征及其调节因素

子宫内膜向蜕膜的转化是正常着床和妊娠的一个重要特征,对于胚泡着床是必不可少的。在蜕膜化过程中,子宫内膜基质细胞在形态和生理等方面都发生了很大的变化。蜕膜化过程受多种因素的调节,包括cAMP、胰岛素样生长因子结合蛋白-1(IGFBP-1)、自然杀伤细胞、同源盒基因-10(HOXA10)、激活素等。但对蜕膜化的机制及调节等仍不清楚。     

PCOS易感基因Hmga2在小鼠子宫蜕膜化中的表达与调节

目的研究PCOS易感基因Hmga2在子宫内膜容受性和蜕膜化中的表达与调节。方法通过早期妊娠、延期着床与激活、人工蜕膜化、卵巢类固醇激素处理等实验,利用q PCR、Western blot技术,阐述Hmga2在子宫内膜容受性中的作用。结果 Hmga2随着妊娠表达量逐渐增加,着床点与非着床点相比表达量显著升高,胚胎激活组比延迟着床表达量显著增高,人工诱导蜕膜化与非蜕膜化比较表达显著升高,Hmga2的表达与雌激素和孕激素呈正相关,体内受雌孕激素调节。结论表明Hmga2的表达与小鼠早期妊娠胚胎着床过程密切相关,参与子宫内膜蜕膜化过程,受活化胚泡和类固醇激素的影响。     

孕酮对人早孕子宫蜕膜细胞活性肾素分泌的调节

子宫蜕膜是肾素产生的主要部位,肾素包括活性与非活性肾素,活性肾素可使血管紧张素原水解生成血管紧张素Ⅰ,继而调节血管紧张素Ⅱ的表达。实验表明：（１）妊娠早期（孕５～９周）,人子宫蜕膜活性肾素的含量随妊娠周龄增加而升高,孕８周时可达６３３７±１２８４ＡⅠｎｇ／ｇｗｗ·ｈ－１;（２）早孕子宫蜕膜组织活性肾素占总肾素量的１／４;（３）孕酮可调节蜕膜细胞活性肾素的合成与分泌。人早孕子宫蜕膜组织中存在高水平的活性肾素,性类固醇激素可调节蜕膜细胞活性肾素的表达,可以认为,子宫局部肾素血管紧张素系统在妊娠过程中发挥了重要作用。     

双炔失碳酯对兔移植胚泡着床的影响

本文用胚泡移植技术验证了双炔失碳酯对子宫内膜的影响是胚泡不能着床的重要原因。将 29只胚龄4d的兔胚泡移植到给药后假孕4d的子宫,结果没有一只胚泡着床;而对照组的5只受卵兔,移植了41只胚泡,有26只着床成功。 为了了解双炔失碳酯对黄体的影响,在5只妊娠兔上,从交配后1—7d与对照组比较血清孕酮浓度的变化,结果表明:对照组的血清孕酮浓度随着妊娠天数显著升高,妊娠第7天时达 17.35±2.12ng/ml,而给药组升高不明显,第7天仅 1.83±1.03ng/ml,为对照组的1/9。已知足够的孕酮浓度对妊娠的维持和胚泡的成功着床具有十分重要的作用。由此推测,双炔失碳酯抑制孕酮的分泌势必影响子宫内膜的发育,从而阻碍胚泡的着床。     

MK基因对小鼠子宫基质细胞蜕膜化进程的影响

为探索肿瘤坏死因子相关凋亡诱导配体(TNF related apoptosis inducing ligand,TRAIL)的死亡受体(mouse killer,MK)对小鼠子宫基质细胞蜕膜化进程的影响,构建MK基因过表达和siRNA干扰重组腺病毒.原代培养的小鼠子宫基质细胞感染MK过表达或者干扰重组腺病毒并诱导蜕膜化,72 h后用免疫细胞化学与流式细胞术分别检测蜕膜细胞的标志物催乳素(prolactin,PRL)与蜕膜细胞凋亡率的变化情况.妊娠d4小鼠子宫角注射MK重组腺病毒,观察胚胎植入点的数量变化.实验结果表明,与对照组相比,在诱导的蜕膜细胞中过表达MK使得催乳素的含量显著降低(P<0.05),同时,蜕膜细胞的凋亡率明显升高(P<0.05),而siRNA干扰之后催乳素的含量显著升高,凋亡率明显下降(P<0.05),但是,宫角注射MK基因过表达和siRNA干扰重组腺病毒之后,胚胎植入数量均显著减少(P<0.01).提示MK基因通过参与小鼠子宫内膜基质细胞的蜕膜化进程,调节蜕膜细胞增殖与凋亡之间的平衡从而影响胚胎的植入.     

人子宫内膜纤蛋白溶酶元激活因子及其抑制因子的分布与调控

人子宫内膜中存在组织型(tPA)及尿激酶型(uPA)两类纤蛋白溶酶元激活因子,其含量在增殖期高于分泌期。本文应用免疫组织化学定位证实uPA及tPA两类抗原存在于子宫内膜的腺体细胞和间质细胞中。应用SDS-PAGE分高蛋白质,继而应用纤蛋白-琼脂糖铺盖技术测得离体培养下间质细胞仅释放tPA,腺体细胞仅释放uPA,但两种细胞均分泌PA的抑制因子(PAI)。培液中加入孕酮,明显抑制PA和刺激PAI生成。雌二醇作用与孕酮相反。某些肽类激素hCG、PRL、GnRH及cAMP作用基本与雌二醇相同。但福司克林(FK)则刺激间质、腺体两种细胞产生tPA及少量uPA,抑制PAI生成。本工作表明人子宫内膜中存在PA及PAI作用相反的酶,受激素调控,其生理意义尚待进一步探讨。     

幼龄大鼠子宫内膜蜕膜化的研究

出生10—14天的Wistar大鼠,在雌二醇和孕酮一定剂量和时程的作用下,以油剂作为蜕膜化刺激,成功地诱发了蜕膜瘤。并在亚显微结构水平上,系统地观察了基质细胞转化为蜕膜细胞的过程。结果表明,出生10—14天的幼龄大鼠子宫内膜,已能对卵巢甾体作出迅速反应。孕酮加微量雌激素处理第三天,基质细胞已处于敏感时期,具备了向蜕膜细胞转化的条件,只是在蜕膜化刺激作用后12小时,才实现了这一转化。子宫内膜上皮在子宫腔内注油后开始解体,基膜处积聚大量无定形物质,它在基质细胞蜕膜化中可能起重要作用。     

孕酮调节途径在着床过程中的作用

孕酮对于哺乳动物排卵、受精、着床、蜕膜化及妊娠维持均起到重要作用.孕酮主要通过孕酮受体(PR)起作用,PR基因敲除的小鼠表现为多种生殖障碍.此外,PR作用的发挥还需要类固醇受体辅助激活因子(SRC)和FK506结合蛋白4(Fkbp52)等辅助因子的参与.目前采用高通量基因芯片技术,筛选出许多直接或间接受孕酮调节的基因,其中同源盒基因、免疫反应基因-1、Indian hedgehog、12 / 15脂肪氧合酶、钙离子结合蛋白等在胚泡着床过程中起重要作用.本文就孕酮调节途径在胚泡着床研究方面的进展作一简要综述.     

15甲-前列腺素F_(2α)对家兔子宫收缩及血浆孕酮水平的影响

本文报道 15甲-前列腺素PG F_(2α)对家兔子宫收缩及血浆孕酮水平的影响。15甲-PGF_(2α)0.4mg/kg、0.2mg/kg 对大鼠抗早孕均有明显作用,能使蜕膜细胞变性,胎盘剥离。15甲-PGF_(2α)剂量为0.4mg/kg时,对家兔在位子宫内压及收缩频率有显著的加强作用。15甲-PGF_(2α)剂量为0.4mg/kg时,对妊娠家兔有明显的降低血浆孕酮作用。     

DEK对人子宫内膜蜕膜化的调节作用

为探讨DNA结合蛋白果蝇Eph激酶(Drosophila Eph kinase,DEK)对人子宫内膜基质蜕膜化的调节作用和途径,该研究采用qPCR(Real-time quantitative polymerase chain reaction)、免疫组化(immunohistochemical)和蛋白印迹(Western blot)分别检测人子宫内膜增生期、分泌期和蜕膜组织中DEK基因和蛋白的表达;利用siRNA抑制基质细胞和蜕膜细胞的DEK,再用细胞流式技术、细胞免疫荧光、qPCR、细胞碱性磷酸酶脂显色和Western blot检测DEK沉默后细胞的变化。结果显示,蜕膜组织中DEK mRNA表达水平低于增生期和分泌期(P0.05),蜕膜组织中DEK蛋白表达水平高于增生期和分泌期(P0.05);抑制基质细胞DEK会使细胞增殖和分化能力降低,从而抑制基质细胞蜕膜化;抑制蜕膜细胞DEK可使细胞凋亡增加,DNA损伤情况加剧,导致蜕膜细胞的维持和发展受到影响。综上,该研究初步证实,DEK可能通过调控细胞蜕膜化而参与胚胎着床过程,其可能途径与其通过调控细胞增殖、分化、凋亡和核损伤有关。     

内皮素—1对大鼠排卵前卵泡颗粒细胞产生孕酮的影响

本文用离体细胞体外孵育法研究了内皮素－１（ＥＴ）对大鼠排卵前卵泡颗粒细胞孕酮生成的影响及其作用机理。结果发现,ＥＴ能显著抑制ｈＣＧ刺激下的孕酮产生,抑制作用在浓度为１０－８ｍｏｌ／Ｌ时,即有显著意义（Ｐ＜０．０５,ｎ＝６）,至１０－７ｍｏｌ／Ｌ时则有非常显著的意义（Ｐ＜０．０１,ｎ＝６）;不同浓度ＥＴ（１０－７—１０－７ｍｏｌ／Ｌ）,对颗粒细胞基础孕酮的产生无明显影响。进一步研究表明,ＥＴ对ｈＣＧ刺激下孕酮生成的抑制作用,在用免抗人内皮素抗血清（ＥＴ－Ａ）１：１０００及ｃＡＭＰ后能明显被逆转。实验中还观察到,ＥＴ使颗粒细胞ＬＨ／ｈＣＧ受体数下降,亲和力降低。本文结果提示,ＥＴ可能为卵巢内的一种局部调节肽,通过作用于ＥＴ受体,干扰ＬＨ／ｈＣＧ受体功能和ｃＡＭＰ生成而抑制颗粒细胞孕酮的产生。     

Prolactin, progesterone, and dexamethasone coordinately and adversely regulate glucokinase and cAMP/PDE cascades in MIN6 beta-cells

Islet cells undergo major changes in structure and function to meet the demand for increased insulin secretion during pregnancy, but the nature of the hormonal interactions and signaling events is incompletely understood. Here, we used the glucose-responsive MIN6 beta-cell line treated with prolactin (PRL), progesterone (PRG), and dexamethasone (DEX, a synthetic glucocorticoid), all elevated during late pregnancy, to study their effects on mechanisms of insulin secretion. DEX alone or combined with PRL and PRG inhibited insulin secretion in response to 16 mM glucose-stimulating concentrations. However, in the basal state (3 mM glucose), the insulin levels in response to DEX treatment were unchanged, and the three hormones together maintained higher insulin release. There were no changes of protein levels of GLUT2 or glucokinase (GK), but PRL or PRG treatment increased GK activity, whereas DEX had an inhibitory effect on GK activity. alpha-Ketoisocaproate (alpha-KIC)-stimulated insulin secretion was also reduced by DEX alone or combined with PRL and PRG, suggesting that DEX may inhibit distal steps in the insulin-exocytotic process. PRL treatment increased the concentration of intracellular cAMP in response to 16 mM glucose, suggesting a role for cAMP in potentiation of insulin secretion, whereas DEX alone or combined with PRL and PRG reduced cAMP levels by increasing phosphodiesterase (PDE) activity. These data provide evidence that PRL and to a lesser extent PRG, which increase in early pregnancy, enhance basal and glucose-stimulated insulin secretion in part by increasing GK activity and amplifying cAMP levels. Glucocorticoid, which increases throughout gestation, counteracts only glucose-stimulated insulin secretion under high glucose concentrations by dominantly inhibiting GK activity and increasing PDE activity to reduce cAMP levels. These adaptations in the beta-cell may play an important role in maintaining the basal hyperinsulinemia of pregnancy while limiting the capacity of PRL and PRG to promote glucose-stimulated insulin secretion during late gestation.     

Lefty inhibits in vitro decidualization by regulating P57 and cyclin D1 expressions

Endometrial decidualization is highly important for successful construction and maintenance of embryo implantation and pregnancy. Lefty gene at different menstrual cycle phases has different expressions, indicating its regulatory significance. To study the mechanism of Lefty in decidualization, human endometrial stromal cells (hESCs) were cultured and induced with medroxyprogesterone acetate (MPA) and 8‐bromoadenosine‐cAMP (8‐Br‐cAMP) in vitro as a research model. Our results showed that Lefty1 overexpression inhibited MPA‐ and 8‐Br‐cAMP‐induced hESC decidualization and significantly reduced the secretion of prolactin (PRL) and insulin‐like growth factor‐binding protein 1 (IGFBP‐1). With the inhibition of Lefty1 expression, hESC decidualization induced by MPA and 8‐Br‐cAMP became more remarkable, and the secretions of PRL and IGFBP‐1 were higher too. Further tests indicated that during the process of decidualization, P57 expression increased, whereas cyclin D1 expression decreased. Although Lefty1 overexpression did not significantly change the expressions of P57 and cyclin D1, inhibition of Lefty1 expression resulted in more evident changes in P57 and cyclin D1 expressions. Meanwhile, cell cycle examination showed that Lefty1 overexpression reduced the cell cycle arrest at G1/S phase in the in vitro hESC decidualization model. Therefore, Lefty1 could regulate the cell cycle via modulating the expressions of P57 and cyclin D1 and then inhibit the decidualization in vitro. Copyright © 2014 John Wiley & Sons, Ltd.     

催乳素对培养的绵羊睾丸间质细胞睾酮分泌的影响

在绵羊睾丸间质细胞体外无血清长期培养的条件下,研究了催乳素对睾丸间质细胞睾酮分泌的调节作用。实验结果表明,催乳素可增强细胞对人绒毛膜促性腺激素(hCG)刺激的反应。催乳素的这种作用呈双相调节。睾酮分泌量显著高于hCG和催乳素单独作用时的总和。在hCG存在下,不同的底物转化为睾酮的量不同。其中雄烯二酮和孕酮转化为睾酮的方式存在着双相性。脱氢表雄酮转为睾酮的量少,不存在双相性,而与其剂量成正比。催乳素在hCG存在下可调节底物转化为睾酮。低剂量的催乳素(1ng/ml)可使一定剂量的孕酮(10～30ng/ml)转化为睾酮的量明显增加,而高剂量的催乳素(>10ng/ml)却明显地抑制孕酮转化为睾酮。催乳素可明显地抑制雄烯二酮转化为睾酮,与剂量无关。可见催乳素对于孕酮和雄烯二酮这两个关键底物转化为睾酮的调节是不同的。催乳素增强hCG刺激睾酮分泌的作用可能部分是通过其促进孕酮转化为睾酮来实现的。     

Hmgn5 functions downstream of Hoxa10 to regulate uterine decidualization in mice

Although Hmgn5 is involved in the regulation of cellular proliferation and differentiation, its physiological function during decidualization is still unknown. Here we showed that Hmgn5 was highly expressed in the decidual cells. Silencing of Hmgn5 expression by specific siRNA reduced the proliferation of uterine stromal cells and expression of Ccnd3 and Cdk4 in the absence or presence of estrogen and progesterone, whereas overexpression of Hmgn5 exhibited the opposite effects. Simultaneously, Hmgn5 might induce the expression of Prl8a2 and Prl3c1 which were 2 well-known differentiation markers for decidualization. In the uterine stromal cells, cAMP analog 8-Br-cAMP and progesterone could up-regulate the expression of Hmgn5, but the up-regulation was impeded by H89 and RU486, respectively. Attenuation of Hmgn5 expression could block the differentiation of uterine stromal cells in response to cAMP and progesterone. Further studies found that regulation of cAMP and progesterone on Hmgn5 expression was mediated by Hoxa10. During in vitro decidualization, knockdown of Hmgn5 could abrogate Hoxa10-induced upregulation of Prl8a2 and Prl3c1, while overexpression of Hmgn5 reversed the inhibitory effects of Hoxa10 siRNA on the expression of Prl8a2 and Prl3c1. In the stromal cells undergoing decidualization, Hmgn5 might act downstream of Hoxa10 to regulate the expression of Cox-2, Vegf and Mmp2. Collectively, Hmgn5 may play an important role during mouse decidualization.     

The inhibition of progesterone secretion and the regulation of cyclic nucleotides by atrial natriuretic factor in gonadotropin responsive murine Leydig tumor cells

We have found that atrial natriuretic factor (ANF) has a profound effect on testicular cells in altering intracellular cyclic nucleotide levels as well as progesterone secretion. Using clonal cultured Leydig tumor cells we found that 1 X 10(-8)M ANF caused a two thousand-fold elevation in the accumulation of cellular cGMP and inhibited cAMP in treated cells by more than 90% as compared to the controls. ANF (1 X 10(-8)M) also significantly inhibited gonadotropin-stimulated accumulation of cAMP in response to bovine luteinizing hormone (bLH) or human chorionic gonadotropin (hCG). Gonadotropin-stimulated progesterone secretion was inhibited by ANF (1 X 10(-10) - 1 X 10(-9)M) in these cultured Leydig tumor cells. Approximately 50% inhibition of progesterone secretion was observed at the peptide concentration of 1 X 10(-9) M.     

In vitro decidualization of human endometrial stromal cells.

Stromal cells isolated from proliferative human endometrium undergo morphologic and biochemical changes when exposed to a mixture of ovarian hormones, acquiring characteristics of decidual cells. In addition to the previously reported progestin-induced secretion of prolactin (PRL) by explants of human proliferative endometrium, and of PRL and laminin by stromal cells in culture, "in vitro" induction of several other decidual cell products was demonstrated in the present study, using cultures of stromal cells isolated from proliferative endometrium. Incubation of stromal cells with a mixture of estradiol, medroxyprogesterone acetate and relaxin, at a concentration reported to yield maximal stimulation of PRL production, resulted in changes from elongated to rounder cells, approx. 90% of which showed immunostaining for PRL under these conditions. Immunocytochemical procedures were carried out on cytospins of decidual cells isolated from decidual tissue adherent to fetal membranes collected at delivery (positive controls), and on stromal cells cultured in Lab-Tek chamber-slides, in the absence (negative controls) or in the presence of added hormones. Antibodies to 24K (a heat-shock protein also named HRP27), desmin (present in intermediate filaments), p29 (a protein associated with the estrogen receptor), and PP12 (an insulin growth factor-1 binding protein), did not react with stromal cells isolated from proliferative endometrium but showed immunostaining of the rounder cells obtained after hormonal treatment when tested with the peroxidase-labeled second antibody complex. In another series of similar experiments, in which the same decidualization end-points were employed, changes in 24K, desmin and PP12 expression were obtained by adding to the insulin-containing medium PRL instead of the hormonal mixture, a finding suggesting sequential steps during the decidualization process.     

Estrogen and uterine sensitization for the decidual cell reaction in the rat: role of prostaglandin E2 and adenosine 3':5'-cyclic monophosphate

The possibility that estrogen affects uterine sensitization for decidualization by altering the ability of E-series prostaglandins (PGs) to increase adenosine 3':5'-cyclic monophosphate (cAMP) concentrations was investigated. To determine if increased endometrial vascular permeability, a response which precedes decidualization, could be obtained in nonsensitized uteri by treatments designed to increase endometrial intracellular cAMP concentrations, cholera toxin, an activator of adenylate cyclase, was injected into the uterine lumen of immature rats pretreated with progesterone and either 0, 0.5 or 10 micrograms estrone with indomethacin to inhibit endogenous PG synthesis. Endometrial vascular permeability, determined using 125I-labeled bovine serum albumin, was assessed 8 h later. Cholera toxin produced a dose-dependent increase in endometrial vascular permeability in all groups; the uteri of rats pretreated with the optimal hormone regimen (0.5 micrograms estrone plus 2 mg progesterone) responded to a lower dose of the toxin. As determined by uterine weights and histologic examination 5 days after the intrauterine administration of cholera toxin or its vehicle, the toxin induced decidualization in rats pretreated with progesterone and 0 or 0.5 micrograms estrone, but not in those receiving 10 micrograms estrone. Cholera toxin had no detectable effect on uterine cAMP concentrations in animals sacrificed 15 min or 3 h after intrauterine treatment. The intrauterine injection of 8-Br-cAMP, with or without 3-isobutyl-1-methyl-xanthine, did not increase endometrial vascular permeability in indomethacin-treated animals pretreated with the different hormone regimens.(ABSTRACT TRUNCATED AT 250 WORDS)     

The effect of aging on prolactin secretion in rat pituitary monolayer culture.

A high basal rate of prolactin (PRL) secretion (.16±.03 μg/well/hr) was produced for over four weeks by pre-confluent male rat pituitary monolayer cell cultures. When the media was changed, a rapid release of microgram quantities of PRL occurred followed by a return to the basal PRL secretory rate by seven hours. Theophylline (3.8×10^?3M), but not dibutyrl cAMP (1×10^?3M), produced a significant (p<.02) increase in PRL secretion, and simultaneous addition of these agents potentiated the PRL secretory rate. TRH (2×10^?8M) had no effect on PRL release by six hours, whereas dopamine (4.9×10^?5M) produced a significant suppression (p<.002) of PRL secretion. In addition, the effects of theophylline, TRH, and dopamine on PRL secretion were similar in cultures of various ages. Ovine prolactin in concentrations up to 50 μg per ml produced no change in PRL secretion during 72 hours of incubation suggesting that PRL feedback control of its own secretion may be transmitted via the hypothalamus. These studies show that a high rate of PRL secretion can be maintained by pre-confluent monolayer cultures for extended periods of time, permitting repeated experimentation on the same wells.