自泌性转化生长因子－β对人卵巢癌细胞生长的影响

目的：我们以往观察到，人卵巢癌细胞系ＣＯＣ１和ＣＯＣ２在无血清培养条件下，可以分泌转化生长因子－β（ＴＧＦ－β）类似物，本研究进一步观察该ＴＧＦ－β类似物对自身细胞生长的作用。方法：分别制备ＣＯＣ１和ＣＯＣ２ＲＰＭＩ１６４０无血清培养条件的培养液ＳＦＣＭ１和ＳＦＣＭ２；观察ＳＦＣＭ１和ＳＦＣＭ２对体外培养和接种ＢＡＬＢ／ｃ裸鼠ＣＯＣ１和ＣＯＣ２细胞生长的影响，并与ＴＧＦ－β标准品相比较。结果：ＳＦＣＭ１和ＳＦＣＭ２对体外培养的自身细胞ＣＯＣ１和ＣＯＣ２均表现为剂量依赖性促进作用，ＳＦＣＭ２明显促进ＣＯＣ２细胞在裸鼠体内生长。放射性受体分析结果显示，ＣＯＣ１和ＣＯＣ２细胞均表达ＴＧＦ－β受体（结合部位）；上述ＳＦＣＭ１和ＳＦＣＭ２的促进作用均可以被抗ＴＧＦ－β中和抗体部分阻断。结论：在ＣＯＣ１和ＣＯＣ２细胞存在ＴＧＦ－β自分泌环调节机制，该机制可能与ＣＯＣ１和ＣＯＣ２细胞自主性无止境生长有关。     

卵巢癌可溶性抗原和抗CD3单克隆抗体诱导细胞毒性T细胞抗人卵巢癌的实验研究

目的探讨卵巢癌过继细胞免疫治疗新方法。方法提取卵巢癌细胞（ＣＯＣ１和ＣＯＣ２ｎ）可溶性抗原（ＴＳＡ），用ＴＳＡ和抗ＣＤ３单克隆抗体（ＣＤ３ＭｃＡｂ）共同诱导正常人外周血单个核细胞（ＰＢＭＣ）产生细胞毒性Ｔ细胞（ＣＴＬ），用ＣＴＬ于体外杀伤ＣＯＣ１细胞和裸鼠体内抑制ＣＯＣ２ｎ移植瘤的生长，体外和淋巴因子激活杀伤细胞（ＬＡＫ）、淋巴因子和抗ＣＤ３单抗激活的杀伤细胞（ＣＤ３ＡＫ）细胞进行比较，体内和ＣＤ３ＡＫ细胞进行比较。结果ＣＴＬ、ＣＤ３ＡＫ和ＬＡＫ对ＣＯＣ１细胞的细胞毒作用分别为７９．４％、５２．１％和５１．７％（Ｐ＜０．０１）。在体内，ＣＴＬ与ＣＤ３ＡＫ和未经处理的对照组比较，卵巢癌细胞移植到裸鼠体内的第９天，肿瘤平均体积分别为４４．４±２４．２ｍｍ３、１１８．８±４０．０ｍｍ３和４４３．０±１５８．７ｍｍ３（Ｐ＜０．０１），裸鼠平均生存期分别为２８．５天、２５．５天和１７天（Ｐ＜０．０１）。结论本方法诱导产生的ＣＴＬ，在体内、体外对卵巢癌细胞均有较高的细胞毒作用，能明显抑制肿瘤生长，为卵巢癌治疗提供了新的思路。     

人绒毛膜促性腺激素和胰岛素对多囊卵巢综合征卵泡内膜细胞雄激素分泌的影响

目的检验多囊卵巢综合征（ＰＣＯＳ）患者卵巢卵泡内膜细胞（简称内膜细胞）雄激素合成是否异常。方法对来自８例ＰＣＯＳ的卵巢卵泡与１８例正常妇女卵巢卵泡内膜细胞进行单层培养，采用放射免疫法测定两者在基础状态、人绒毛膜促性腺激素（ｈＣＧ）和胰岛素（ＩＮＳ）刺激作用下培养液中１７α羟孕酮（ＯＨＰ）、雄烯二酮（Ａ）、孕酮（Ｐ）。结果在基础状态、ｈＣＧ与ＩＮＳ刺激作用下，ＰＣＯＳ患者的卵巢内膜细胞分泌Ａ、ＯＨＰ和Ｐ的量与正常妇女相比，除在基础状态下Ｐ无明显增加外（Ｐ＞０．０５），其余均显著增加，分别为几倍到十几倍（Ｐ＜００５～０００１）。此外，ｈＣＧ和ＩＮＳ还有协同刺激作用。ＰＣＯＳ患者３种激素分泌的持续时间比正常妇女长。结论ＰＣＯＳ患者的卵巢内膜细胞存在雄激素的合成异常，ＩＮＳ可能起关键作用。     

多囊卵巢综合征促排卵失败的相关因素探讨

探讨多囊卵巢综合征（ＰＣＯＳ）妇女促排卵失败的危险因素。方法：将 １０３名ＰＣＯＳ病人,随机分为 ３组,分别以 ＣＣ／ ｈＣＧ、ＣＣ／ｈＭＧ／ｈＣＧ及 ＧｎＲＨ－ａ／ｈＭＧ／ｈＣＧ三种促排卵方案治疗,并以 ３１名正常妇女作对照,应用放免法测定４组对象血清性激素水平。应用糖耐量试验及胰岛素释放试验测定糖负荷后 ０ ｍｉｎ、 ６０ ｍｉｎ、 １２０ ｍｉｎ血糖及胰岛素水平。结果：促排卵治疗失败者卵巢过度刺激综合征（ＯＧＴＴ）后 １２０ ｍｉｎ胰岛素水平明显增高,与排卵者相比,Ｐ＜０．０５,ＯＲ＝１．０１３;而 ＣＣ／ｈＭＧ／ｈＣＧ治疗方案失败率较低,ＯＲ＝０． ３１１０。结论：ＯＧＴＴ后 １２０ ｍｉｎ胰岛素是促排卵失败的危险因素, ＣＣ／ｈＭＧ／ｈＣＧ方案是较为成功的促排卵治疗方案。     

在绒毛膜促性腺激素β亚基羧基端37肽四聚体基因在E,Coli中的表达

人绒毛膜促性腺激素β亚基（ｈＣＧβ）羧基端１０９～１４５残基组成的３７肽（ｈＣＧβＣＴＰ３７）是ｈＣＧ分子所特有的片段，并存在ｈＣＧ的特异表位抗原决定簇）。为提高ｈＣＧβ－ＣＴＰ３７肽的的性，本研究构建ｈＣＧβ－ＣＴＰ３７四聚体ｃＤＮＡ，克隆人载体ｐＱＥ６０，获得含（ｈＣＧβ－ＣＴＰ３７），ｃＤＮＡ的表达质业ｐＱＥ６０－（ｈＣＧβ－ＣＴＰ３７），在大肠 力Ｘ－Ｂｌｕｅ中进行表达，得到ｈＣＧβ－ＣＴ     

应用hMG/hCG加DIPI结合应用GnRHa治疗原发不明原因不孕症可显著提高妊娠率

为了评价ＧｎＲＨａ在应用超促排卵方案治疗不孕症中的功效,作者于１９９５年５月至１９９６年１２月期间,对用克罗米芬／ｈＣＧ或ｈＭＧ／ｈＣＧ促排卵并行宫腔内人工授精（ＩＵＩ）治疗至少７个周期未孕之不明原因不孕夫妇,采用ｈＭＧ／ｈＣＧ加ＤＩＰＩ（直接腹腔内授精）法随机分２组研究治疗效果。Ａ组为ｈＭＧ／ｈＣＧ／ＤＩＰＩ加ＧｎＲＨａ,Ｂ组为ｈＭＧ／ｈＣＧ／ＤＩＰＩ（未用ＧｎＲＨａ）。患者年龄、不孕年限、ＩＵＩ治疗周期及ＤＩＰＩ注入的活精子数,两组无明显差别。行ＤＩＰＩ术之前之基本检查如ＢＢＴ测定,血Ｅ２峰…     

Stuuyon thesecondrycytoreductivesurgeryforptientswithdvncedovrinepithelilcncer

１ＩＭＰＡＣＴＯＦＰＲＩＭＡＲＹＣＹＴＯＲＥＤＵＣＴＩＶＥＳＵＲＧＥＲＹＯｖɑｒɑｎｅｐｉｔｈｅｌｉɑｌｃɑｎｃｅｒ（ＯＥＣ）ｉｓｔｈｅｍｏｓｔｌｅｔｈɑｌｇｙｎｅｃｏｌｏｇｉｃｎｅｏｐｌɑｓｍ,ｌɑｒｇｅｌｙｄｕｅｔｏｆｒｅｑｕｅｎｔｐｒｅｓｅｎｃ...     

促性腺激素释放激素激动剂超短方案在超促排卵中的应用

目的：探讨促性腺激素释放激素激动剂（ＧｎＲＨ－ａ）超短方案在促排卵中的作用。方法：以采用克罗米芬联合人绒毛膜促性腺激素（ＣＣ／ｈＣＧ组，５０个周期、３１例），及克罗米芬联合人绝经期促性腺激素、绒毛膜促性腺激素（ＣＣ／ｈＭＧ／ｈＣＧ组，１６个周期、１６例）方案者为对照，对比ＧｎＲＨ－ａ超短方案联合人绝经期促性腺激素、绒毛膜促性腺激素方案者（ＧｎＲＨ－ａ超短方案／ｈＭＧ／ｈＣＧ组，１５个周期、１５例）ｈＣＧ注射日激素水平、优势卵泡个数、子宫内膜厚度、宫颈评分及妊娠率。ＧｎＲＨ－ａ超短方案／ｈＭＧ／ｈＣＧ组全部来自采用ＣＣ助孕失败或采用ＣＣ／ｈＭＧ／ｈＣＧ方案显示卵巢反应性差的患者。结果：ＣＣ／ｈＭＧ／ｈＣＧ组有３例（１８．８％）发生过早黄素化。ＧｎＲＨ－ａ超短方案／ｈＭＧ／ｈＣＧ组ｈＣＧ注射日血清黄体生成素（ＬＨ）水平明显低于对照组，其优势卵泡个数、子宫内膜厚度及宫颈评分都明显高于对照组，差异均具有显著性（Ｐ＜０．０５）。３组周期妊娠率相近。结论：ＧｎＲＨ－ａ超短方案／ｈＭＧ／ｈＣＧ方案为一种较好的促超排卵方案，对ＣＣ助孕失败及ＣＣ／ｈＭＧ／ｈＣＧ方案卵巢反应性差的患者仍有较好的效果。     

输精管结扎家兔血清总胆固醇、酯质过氧化物及一些酶系的变化

应用日本大耳白兔复制输精管结扎模型，分为输精管结扎２５月组（ＶＧ２５）和６月组（ＶＧ６），并设同龄假手术对照组（ＳＯＧ２５和ＳＯＧ６）．实验结果表明，１．增龄可使血清总胆固醇（Ｃｈ）和甘油三脂（ＴＧ）含量增高，肌酸磷酸激酶（ＣＰＫ）、乳酸脱氢酶（ＬＤＨ）、谷草转氨酶（ＧＯＴ）、ａ－羟丁酸脱氢酶（ＨＢＤＨ）的血清水平略增高，但ＶＧ与ＳＯＧ比较无差异；２．血清酸性磷酸酶（ＡＣＰ）水平，２５月组明显高于６月组（Ｐ＜０．０５），ＶＧ与ＳＯＧ比较无差异（Ｐ＞０．０５）；３．血清脂质过氧化物（ＬＰＯ）含量，２５月组明显地高于６月组（Ｐ＜０．０５），而输精管结扎对其无影响；４．相关检验表明，血清ＬＰＯ含量与血清ＣＰＫ和ＡＣＰ水平均呈明显正相关关系，ｒ值分别为０．４７２（Ｐ＜０．０５），０．６８０（Ｐ＜０．００１）。提示血清酶水平的增高与膜的脂质过氧化有关。输精管结扎对老化过程、细胞及亚细胞膜结构无不良影响。     

卵巢过度刺激综合征防治的研究进展

近２０年来，由于助孕技术的发展和促排卵药物（包括ＣＣ、ＦＳＨ、ｈＭＧ、ｈＣＧ和ＧｎＲＨ－ａ）的广泛应用，卵巢过度刺激综合征（ｏｖａｒｉａｎｈｙｐｅｒｓｔｉｍｕｌａｔｉｏｎｓｙｎｄｒｏｍｅ，ＯＨＳＳ）发生率有增高趋势。典型的ＯＨＳＳ以双侧卵巢增大，高雌激素血症、腹水、电解质酸碱失衡、血液高凝状态、血液浓缩和少尿为特征，严重者可出现肾功能衰竭、血管栓塞、ＤＩＣ和死亡。因此，了解ＯＨＳＳ发病机理、预防和治疗的研究进展十分重要。     

The relationship between trophoblast differentiation and the production of bioactive hCG

We previously showed that a significant number of failing pregnancies are associated with production of human chorionic gonadotropin (hCG) having relatively low bioactivity. The present study was designed to compare the secretion of intact, immunoreactive hCG to the secretion of bioactive hCG during trophoblast differentiation, and to test the hypothesis that the lower bioactive: immunoreactive hCG ratios in failing pregnancies are related to reduced or impaired trophoblast differentiation. Cytotrophoblast cells were isolated from term placentas and cultured under conditions that induced or did not induce syncytiotrophoblast formation. Culture media were collected at regular intervals up to 72 h and levels of immunoreactive and bioactive hCG were measured. The differentiation of cytotrophoblast cells to multinucleated syncytiotrophoblast was monitored by immunocytochemistry and electron microscopy. During the 72 h culture period, concentrations of immunoreactive and bioactive hCG increased in both differentiating and non-differentiating cells. However, the concentrations of immunoreactive and bioactive hCG were higher under culture conditions that promoted trophoblast differentiation. Furthermore, the ratio of bioactive hCG to immunoreactive hCG was higher in differentiating cultures. When differentiation was inhibited by dimethyl sulfoxide, the secretion of bioactive hCG was reduced and the bioactive: immunoreactive hCG ratio did not change. These findings are consistent with the idea that production of bioactive hCG accompanies syncytiotrophoblast formation.     

Effects of prolactin on secretion and synthesis of human chorionic gonadotropin in human term placentas in vitro: short-term increase in secretion, followed by medium-term suppression of synthesis and secretion.

We studied the influence of human prolactin on the secretion and de novo synthesis of human chorionic gonadotropin (hCG) in the human term placenta in culture. Placental tissue from 14 patients with uncomplicated pregnancies and deliveries was prepared mechanically, with addition of a Percoll gradient step. hCG levels were determined in the culture media and in the cytosolic fraction of cells by means of an enzyme immunoassay with coated beads. The amount of newly synthesized hCG was measured by the extent of incorporation of 35S-methionine into the hCG molecule. Our results showed that human prolactin had two different effects in vitro: between 1/2 and 1 h, prolactin slightly increased secretion of hCG into the culture medium without affecting de novo synthesis; after 2 h, prolactin began to cause a significant decrease in both secretion and de novo synthesis of hCG over several hours. It appears that both effects are receptor mediated, for ovine prolactin failed to produce any response. We conclude that prolactin is one of the main factors regulating the synthesis and secretion of hCG in the human trophoblast at term.     

Oestrogen synthetase (aromatase) and hormone secretion in primary cultures of human placental trophoblast cells. Effects of cyclic AMP addition at the start of culture in attached and unattached cell populations

Term placental trophoblast cells, released by trypsin digestion of placental villi, purified on a Percoll gradient and grown in serum-containing medium, differentiate within 24 to 48 h in culture from mononucleated cytotrophoblast-like cells at the start of culture to highly multinucleated giant (syncytiotrophoblast-like) cells that are more active in hormonogenesis. To determine the changes in hormone biosynthesis and secretion that occur early in the trophoblast differentiation process in vitro, freshly isolated placental cells were cultured in Dulbecco's Modified Eagle's Medium (DMEM) containing 20 per cent FBS, in the presence and absence of cAMP for up to 48 h. Cell attachment and growth, oestrogen synthetase (aromatase) activity in attached and unattached cells, and secretion of human chorionic gonadotropin (hCG) and progesterone were studied. The aromatase specific activity, low in freshly isolated cells, increased fourfold in attached cells by 3 h, and achieved a 10- to 15-fold increase by 40 to 48 h. In attached cells grown with cAMP, aromatase activity was further stimulated by about fourfold, relative to the control. The aromatase activity of the unattached cells removed from the culture dishes at various times up to 48 h showed a biphasic response: the activity decreased by 18 h and then increased back to the fresh cell levels. The effect of cAMP on aromatase in these unattached cells was manifested by a two-fold stimulation of activity by 18 h, relative to control unattached cells. Secretion of hCG from both attached and unattached cells remained at a low level (less than 200 ng/mg protein) in control cells; in the presence of cAMP, hCG secretion was stimulated by tenfold after 40 h.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of leptin on the regulation of placental hormone secretion in cultured human placental cells.

Placenta is an important source of leptin during pregnancy that contributes to the high plasma leptin levels in pregnant women. Leptin and its functional receptors are synthesized in trophoblast cells that, in turn, secrete gestational hormones supporting a paracrine or autocrine role for leptin in the endocrine activity of the placenta. In the present study we examined the effect of leptin on in vitro release of gestational hormones (human chorionic gonadotropin (hCG), human placental lactogen (hPL), progesterone, estrogens and testosterone) by human term placental cells in culture. Placentas at term were obtained immediately after delivery from mothers with uncomplicated pregnancies. Progesterone, hCG, hPL, estradiol, estrone, estriol and testosterone levels were measured by different assays in culture media of cells maintained in monolayer culture after incubation for 12, 24, 48 or 72 h with leptin or placebo. Incubation with leptin did not modify hCG, hPL, progesterone, estriol and estrone secretion for any of the doses and times assayed. However, leptin led to a dose-dependent decrease in estradiol release. This effect was observed when treatment with recombinant human leptin spanned from 12 to 72 h. At this time an increase in testosterone levels was observed in leptin-treated cells versus placebo. These results indicate that leptin can be considered a gestational hormone implied in the endocrine function of the placenta, with an important role in control of the production of steroid reproductive hormones in placental cells in vitro.     

Human chorionic gonadotropin expression in human trophoblasts from early placenta: comparative study between villous and extravillous trophoblastic cells

Human trophoblast differentiates into two pathways: extravillous cytotrophoblasts (EVCT) that invade the uterus wall and villous cytotrophoblasts (VCT) that fuse to form the syncytiotrophoblast (ST) involved in placental exchanges and endocrine function. It is established that hCG is produced and secreted by the ST into the maternal compartment where it plays a key endocrine role and stimulates ST formation in an autocrine manner. Herein, we investigated hCG expression in early placentas by immunohistochemistry using different antibodies. We then compared hCG secretion by primary cultures of VCT and EVCT isolated from the same first trimester human chorionic villi. In situ hCG was immunodetected in EVCT all along their invasive differentiating pathway except in cells near the stromal core of the proximal column. hCG expression was confirmed in vitro by immunocytochemistry and hCG secretion quantified in cell supernatants. Interestingly, whereas hCG secretion increased during VCT differentiation into ST (from 60 to 350UI/L/microg DNA), EVCT secretion remained constant and at a high level during the same culture period (160UI/L/microg DNA). Our data demonstrated that in addition to the ST, invasive EVCT also expressed and secreted high levels of hCG, suggesting a specific paracrine and/or autocrine role for hCG from EVCT origin.     

Effect of cryopreservation on human cytotrophoblast cells in culture: hCG and PALP production

Term villous cytotrophoblasts differentiate into syncytiotrophoblast during culture exhibiting characteristic changes in cellular morphology and protein expression profiles. Measurement of human chorionic gonadotropin (hCG) and placental alkaline phospatase (PALP) is often used to assess viability and syncytialisation of cultured cells. The objective of this study was to assess the effect of cryopreservation of isolated cytotrophoblasts on the expression hCG and PALP by cells during subsequent culture. Villous cytotrophoblasts isolated from term placentae from uncomplicated pregnancies were either cultured immediately after isolation or were cryopreserved (liquid nitrogen) prior to culture. Cells were cultured in identical conditions (5% CO(2) in air) for 96 h. Protein and DNA content of cells and HCG and PALP levels in culture medium were measured at 24 h intervals. Cryopreservation had no significant effect on the protein or DNA content of cultured cells but hCG levels in culture medium were significantly reduced after 72 h (P=0.025) compared to cultures of fresh cells. PALP levels were unchanged. Cryopreservation of cytotrophoblast cells prior to culture resulted in a decrease in basal secretion of hCG possibly caused by a failure or delay in the morphological and functional differentiation of cells.     

Studies on influence of synthetic LH-RH and dbcAMP on normal chorionic villi and BeWo cells (author's transl)

The influence of synthetic LH-RH and dbcAMP on human chorionic villi and BeWo cells was studied through measurement of cAMP, hCG and estradiol. The results obtained were as follow: 1) There was a rapid increase of cAMP in chorionic villi after the 7th week of gestation, marking a maximum value at the 9th week, and gradually decreasing thereafter. 2) When dbcAMP was added to chorionic villi, there was a significant increase estradiol production in chorionic villi and hCG secretion in media. 3) The stimulation of cAMP in chorionic villi and hCG secretion into medial resulting from the addition of synthetic LH-RH to chorionic villi showed significant increases at 5 minutes and 30 minutes, respectively. 4) When dbcAMP or synthetic LH-RH was added to BeWo cells, increased secretion of hCG and estradiol into media was seen. It therefore is concluded that there is a relationship between the activity of synthetic LH-RH in which cAMP participates and a mechanism involving hCG secretion and estradiol production originating from human chorionic villi and BeWo cells. This is turn suggest the possibility of the existence in chorionic villi of a LH-RH like substance which plays a role in the production or secretion of hormone in the placenta.     

Modulation of ectopic secretion of human chorionic gonadotropin by cultured ovarian adenocarcinoma cells

Modulation of ectopic human chorionic gonadotropin (hCG) secretion by a human nontrophoblastic ovarian papillary cystadenocarcinoma cell line maintained in monolayer culture was studied. Exposure of cells to methotrexate (MTX, 0.1 microM) significantly enhanced hormone secretion while actual cell replication was decreased. In contrast, exposure of cells to actinomycin D (25 pM) for 24 hr completely abolished hormone secretion and resulted in death of all cells. Exposure of the cells to hypothalamic peptides (thyrotropin-releasing hormone, gonadotropin-releasing hormone, and somatostatin) did not alter hCG production. hCG secretion was stimulated after 24-hr incubation with dibutyryl cAMP (100 microM) and by prostaglandin F1 alpha (10 microM). Two separate mechanisms of modulation of ectopic hCG by these cells are possible: a cAMP-mediated stimulation independent of cell-growth kinetics after exposure to dibutyryl cAMP and prostaglandin F1 alpha, and a selective inhibition of DNA synthesis which results in slowing of cell replication and concomitant increase in hCG production per cell.     

Variations of luteinizing hormone serum concentrations after exogenous human chorionic gonadotropin administration during ovarian hyperstimulation

Changes in luteinizing hormone (LH), estradiol, and progesterone (P) serum levels before and after preovulatory administration of human chorionic gonadotropin (hCG) were assayed in 30 patients stimulated with clomiphene citrate (CC) and human menopausal gonadotropin (hMG) and compared with LH variations in 43 patients submitted to pharmacological hypophysectomy with a gonadotropin-releasing hormone agonist (GnRH-a) and stimulation with hMG. In CC + hMG-treated patients, an endogenous LH surge occurred systematically 4.25 +/- 2.75 hours after hCG injection. Multiparametric analysis indicated an inverse correlation between the delay in the initial rise of the LH surge and the increase in P levels during the 6 hours after hCG administration. Gonadotropin-releasing hormone agonist + hMG treatment did not lead to an LH surge after hCG but to a significant fall in LH levels. Thus, exogenous hCG, administered before ovulation, induces an endogenous LH surge if pituitary function is not blocked by a GnRH-a, probably through an increase in P secretion.