PCOS患者卵泡输出率预测IVF结局和脂联素及其受体与胚胎发育潜能的相关性研究
|
摘要
研究背景多囊卵巢综合征(polycystic ovary syndrome, PCOS)是常见的内分泌及代谢紊乱性疾病,发病率占生育年龄女性5%-10%。PCOS病因尚不清楚,临床表现异质性,可以表现为月经异常、排卵障碍、临床高雄等,同时可合并肥胖、胰岛素抵抗、血脂异常等代谢异常,是发生2型糖尿病、子宫内膜癌等疾病的高危因素。该疾病是生殖医学领域研究的热点和难点。
排卵障碍是PCOS患者不孕的主要原因,诱导排卵是PCOS合并不孕患者的一线治疗。但是对于接受了标准促排卵治疗后仍未妊娠,以及同时合并输卵管因素或者男性因素不孕的患者,需要借助体外受精-胚胎移植(in-vitro fertilization&embryo transfer, IVF-ET)及卵胞浆内单精子显微注射(intracytoplasmic sperm injection, ICSI)技术获得妊娠。IVF/ICSI技术为PCOS不孕患者提供了有效的治疗手段,控制性卵巢超促排卵(controlled ovarian hyperstimulation, COH)是辅助生殖技术的关键过程。但是,由于PCOS患者对于卵巢刺激的敏感性增加,与非PCOS患者相比,接受COH治疗的PCOS患者每周期的获卵率及未成熟卵率显著增加,但是受精率显著降低。在临床工作中,我们发现部分PCOS患者虽然获卵数多,但是卵子及胚胎质量差,影响患者治疗结局。优化促排卵方案,保证窦卵泡对卵泡刺激激素(follicular stimulating hormone, FSH)的适宜反应,获取高质量卵子,是获得良好妊娠结局的前提。迄今为止,尚没有一个指标可以同时预测卵巢对促排卵的反应性以及卵子发育潜能。2011年有学者创造性地提出了卵泡输出率(follicular output rate, FORT)的概念,对月经周期规律患者的研究认为,FORT是反应卵巢中卵泡潜能的量化指标,但是FORT对PCOS患者IVF/ICSI结局的预测价值尚不明确。
卵冠丘复合物(cumulus-oocyte complex, COCs)中,卵丘颗粒细胞直接包绕着卵母细胞,伴随着卵母细胞发育、成熟、排卵至受精的过程。卵丘颗粒细胞分解代谢胆固醇、葡萄糖,通过缝隙连接为卵母细胞提供必要的能量,对卵母细胞的发育是至关重要的。近年来的研究提示卵丘颗粒细胞基因表达的差异可以作为预测卵子及胚胎发育潜能的非侵入性方法。脂联素(Adiponectin)作为一种脂肪细胞因子,对于调节人胰岛素敏感性起到非常重要的作用。PCOS患者血清脂联素水平下降,而脂联素水平与胰岛素抵抗程度呈负相关,PCOS患者脂联素水平下降可能是导致其卵巢功能异常的重要原因。前期对山东汉族PCOS患者的研究显示,脂联素基因SNP rs1501299+276(A/C)与SNP rs2241766+45G15G(G/T)多态性与PCOS发病具有相关性,其中脂联素基因SNP+276(A/C)多态性具有调节PCOS患者胰岛素敏感性的作用,在PCOS的发病过程中提供了发生胰岛素抵抗的遗传易感性。近年来的研究认为脂联素直接参与调节生殖功能,体外培养小鼠卵冠丘复合物经脂联素作用后,可以显著增加受精率、胚胎卵裂率及囊胚形成率；人卵丘细胞中AdipoR1、AdipoR2的表达量与体外培养胚胎的囊胚形成率相关,提示卵丘细胞可能是脂联素作用的一个重要靶点,对胚胎的发育潜能具有一定的作用。但是,脂联素及其受体在PCOS患者卵丘颗粒细胞中的表达情况以及与PCOS患者胚胎发育潜能的相关性尚未见研究报道。
基于以上研究背景,本课题从提高PCOS患者胚胎发育潜能,改善妊娠结局着手,进行了相关临床及基础研究。第一部分通过分析IVF/ICSI周期促排卵方案及妊娠结局等临床资料,评估卵泡输出率在评价PCOS患者卵巢对FSH反应性、卵子质量及预测IVF/ICSI妊娠结局方面的价值。第二部分通过检测脂联素及其受体在不同质量胚胎所对应的卵丘颗粒细胞中的表达情况,分析脂联素及其受体与胚胎发育潜能的相关性,明确脂联素是否通过卵丘颗粒细胞发挥影响PCOS患者胚胎发育潜能的作用。
第一部分
多囊卵巢综合征患者卵泡输出率预测IVF/ICSI结局的研究
目的研究体外受精-胚胎移植/卵胞浆内单精子显微注射周期中,卵泡输出率在评估多囊卵巢综合征患者卵巢对FSH反应性及预测妊娠结局的价值。
方法以2010年1月至2011年12月在烟台毓璜顶医院生殖医学中心进行IVF/ICSI治疗的1643个周期作为研究对象。其中多囊卵巢综合征患者140例,非多囊卵巢综合征患者1503例。按照2003年ESHRE/ASRM建议的诊断标准作为PCOS患者的纳入标准。PCOS组与非PCOS组患者的中位年龄均为32岁。PCOS患者中,79例(56.43%)患者为稀发排卵和／或无排卵+高雄激素+卵巢多囊样改变；46例(32.86%)患者为稀发排卵和／或无排卵+高雄激素；11例(7.86%)为稀发排卵和／或无排卵+卵巢多囊样改变；4例(2.86%)为高雄激素+卵巢多囊样改变。所有患者均采用标准长方案进行控制性超促排卵(COH),开始应用促性腺激素(Gn)日通过阴道B超检测,记录双侧卵巢中直径3-10mm窦卵泡数(AFC),注射hCG日记录双侧卵巢中直径16-22mm成熟卵泡数(PFC),计算出卵泡输出率(FORT=PFC/AFC)。注射hCG后34-36h行阴道超声引导下穿刺取卵术,取卵当日根据男方精液情况行常规IVF或ICSI受精,IVF或ICSI后16-18小时观察受精情况,取卵后第3天进行胚胎移植,剩余可移植胚胎冷冻保存。自取卵日开始进行黄体支持,胚胎移植后14天查血hCG确定生化妊娠,移植后35天阴道B超检查探及妊娠囊确定为临床妊娠。
结果在PCOS患者的140个治疗周期中,临床妊娠66周期,未妊娠74周期,临床妊娠率为47.14%(66/140)。妊娠组FORT值显著低于未妊娠组(0.56±0.21vs.0.66±0.29,P<0.05)。
FORT的均数为0.65,根据FORT值,将PCOS患者分为三组,低FORT组(FORT<0.5, n=45),中等FORT组(FORT0.5～0.73, n=56)及高FORT组(FORT>0.73, n=39)。根据FORT分组后,胚胎种植率及临床妊娠率在低、中、高FORT三组间无统计学差异。随着FORT逐渐增高,成熟卵泡数及获卵数逐渐增加,然而值得注意的是,尽管在高FORT组,获卵数是最多的(12.04±7.54vs.15.63±7.08vs.17.21±7.66,P<0.05),但是2PN受精率(63.47%vs.69.49%vs.60.66%,P<0.01)以及优质胚胎率(68.06%vs.72.71%vs.64.99%,P<0.05)在中等FORT组最高,提示中等FORT值组的患者可以获得较好的IVF/ICSI结局。
非-PCOS患者的1503个治疗周期中,临床妊娠788周期,未妊娠715周期,临床妊娠率为52.43%(788/1503),PCOS患者与非PCOS患者临床妊娠率无统计学差异(P>0.05)。妊娠组与未妊娠组相比,两组间FORT (0.66±0.25vs.0.63±0.26,P>0.05)无统计学差异。妊娠组的2PN受精率(69.20%vs.64.79%,P<0.001)、优质胚胎率(75.77%vs.55.90%,P<0.001)显著高于未妊娠组。
根据FORT值,将非-PCOS患者分为三组,低FORT组(FORT<0.5,n=402),中等FORT组(FORT0.5～0.73,n=632)及高FORT组(FORT>0.73,n=469)。 FSH/LH比值(1.73±1.87vs.1.52±0.82vs.1.49±0.82,P=0.007)以及AFC(14.51±6.33vs.14.00±5.37vs.12.32±4.28,P<0.001)在低FORT组最高。从低FORT组至高FORT组,PFC(5.49士2.85vs.8,41±3.27vs.11.54士4.33,P<0.001),获卵数(8.45±5.64vs.11.52±6.37vs.13.30±6.34,P<0.001),可移植胚胎总数(4.94±3.22vs.6.37±3.69vs.7.33±3.89,P<0.001)逐渐增高。与此同时,优质胚胎率(65.98%vs.66.48%vs.68.91%,P<0.05)、胚胎种植率(29.68%vs.33.80%vs.35.06%,P<0.01)、临床妊娠率(46.27%vs.53.46%vs.55.65%,P<0.05)随着FORT的增高也是逐渐增高的,差异具有统计学意义,而2PN受精率(68.07%vs.66.95%vs.67.13%,P>0.05)在三组之间无统计学差异。
回归分析显示FORT与AFC(r=-0.162,P<0.001),血清基础卵泡刺激激素(r=-0.123,P<0.001)呈负相关,与PFC(r=0.607,P<0.001),可移植胚胎总数(r=0.259,P<0.001),以及优质胚胎总数呈正相关(r=0.197,P<0.001)。以优质胚胎数作为自变量,进行多元线性回归分析,结果显示在所有因变量中,FORT(t=1.982,P<0.05)以及获卵数(t=17.246,P<0.01)是主要影响变量,而不是AFC或者PFC。
结论中等FORT值的PCOS患者以及高FORT值的非PCOS患者可以获得较好的IVF/ICSI妊娠结局。PCOS患者进行超促排卵治疗时,应调整用药尽量得到适宜的卵泡输出率,从而改善IVF/ICSI妊娠结局。
第二部分
脂联素及其受体与多囊卵巢综合征患者胚胎发育潜能的相关性研究
目的检测脂联素及其受体基因在不同质量胚胎所对应的卵丘颗粒细胞中的表达情况,分析脂联素及其受体与胚胎发育潜能的相关性,明确脂联素是否通过卵丘颗粒细胞发挥影响PCOS患者胚胎发育潜能的作用。
方法PCOS组共27例,按照2003年ESHRE/ASRM建议的诊断标准作为PCOS患者的纳入标准。对照组为因输卵管因素接受IVF-ET治疗的患者,共28例。所有患者根据体重指数分为体重正常组(BMI<25kg/m2)及肥胖组(BMI≥25kg/m2)。行阴道超声引导下穿刺取卵术后获取卵冠丘复合物,用机械法分离每个卵子的卵丘颗粒细胞,采用单卵微滴短时授精,卵子与收集的卵丘颗粒细胞一一对应,根据每个卵子受精、胚胎发育及囊胚形成情况,回顾性将卵子所对应的卵丘颗粒细胞分为囊胚形成组(B+组)和未形成囊胚组(B-组)。采用real time RT-PCR和western blot的方法检测脂联素及其受体mRNA、蛋白表达情况,并比较B+组和B-组的mRNA表达量是否存在差异。
结果Real time RT-PCR和western blot方法检测到AdipoQ. AdipoR1、AdipoR2和T-cadherin基因mRNA及蛋白在卵丘颗粒细胞中有表达,但是以AdipoRl和AdipoR2的表达为主。体重正常PCOS患者中,AdipoR2基因mRNA的相对表达量在囊胚形成组(PB+组)所对应卵丘颗粒细胞中显著高于未形成囊胚组(PB-组)(0.1647±0.0161vs.0.0783±0.0385,P<0.05)；在肥胖PCOS患者中,AdipoRl及AdipoR2基因mRNA的相对表达量在囊胚形成组(PB+组)所对应卵丘颗粒细胞中均显著高于未形成囊胚组(PB-组)(0.5162±0.0371vs.0.2448±0.0333,P<0.01)(0.1948±0.0307vs.0.1057±0.0236,P<0.05)。在非-PCOS患者中,脂联素及其受体基因mRNA的相对表达量在不同质量胚胎组间无显著性差异。
结论脂联素、AdipoR1、AdipoR2和T-cadherin基因mRNA和蛋白在卵丘颗粒细胞中有表达,但是以AdipoRl和AdipoR2的表达为主。AdipoR2基因的mRNA表达水平与PCOS患者胚胎发育潜能具有相关性；AdipoR1基因的mRNA表达水平与肥胖PCOS患者的胚胎发育潜能相关。脂联素可能通过上调PCOS患者卵丘颗粒细胞AdipoR1和AdipoR2的表达而发挥影响胚胎发育潜能的作用。
Background Polycystic ovary syndrome (PCOS) is one of the most prevalent endocrinopathies, affecting5-10%of women of reproductive age. The characteristic clinical features of PCOS include menstrual irregularity such as oligomenorrhoea, signs of hyperandrogenaemia including hirsutism, acne, and/or obesity, and polycystic morphology on ovarian ultrasound. More recent studies have highlighted the link between PCOS and metabolic syndrome, insulin resistance and compensatory hyperinsulinemia appear to be a prominent feature of this phenotype. The pathophysiology of PCOS still remains largely uncharacterized and it has puzzled gynecologists and endocrinologists for many years and proven very difficult to define. Anovulation is the common cause of infertility in PCOS patients, the vast majority of women with PCOS will ovulate and conceive following ovulation induction therapy. Those who fail to be pregnancy often have an additional factor, such as male factor, fallopian tube factor or endometriosis. They should seek in-vitro fertilization&embryo transfer (IVF-ET) or intracytoplasmic sperm injection (ICSI) as a treatment for infertility. Controlled ovarian hyperstimulation (COH) is the key processor in assisted reproductive technology (ART). Women with PCOS will almost yield more oocytes than those without PCOS. However, the fertilization rate of the oocytes from PCOS is almost inevitably lower. A meta-analysis demonstrated an increased cycle cancellation rate, but more oocytes retrieved per retrieval and a lower fertilization rate in PCOS patients undergoing IVF. Although the regulatory mechanisms determining the extent of the sensitivity of individual antral follicles to follicle stimulating hormone (FSH) remains to be elucidated, the appropriate response of antral follicles to FSH and a high quality of oocytes may result in a good outcome after IVF/ICSI.
There is no marker that can predict both ovarian response and oocyte competence. To evaluate follicular responsiveness to exogenous FSH, the use of the Follicular Output RaTe (FORT) as an innovative term has been suggested. FORT is assessed by the ratio of the pre-ovulatory follicle (16-22mm) counts (PFC) obtained in response to FSH administration on the day of hCG to the small antral follicle (3-10mm) count (AFC) observed after the complete suppression of endogenous gonadotropins by gonadotropin-releasing hormone agonist (GnRHa)(FORT=PFC/AFC). Gallot found that FORT might be a qualitative reflector of ovarian follicular competence only in patients with regular menstrual cycles. The values of FORT as a predictor of IVF/ICSI outcome in PCOS and non-PCOS patients were unknown.
In cumulus-oocyte complex (COCs), cumulus cells are the specialized granulosa cells that directly surround the oocyte during follicular development. Importantly, these cells accompany the oocyte throughout the development from an immature to a fully mature ovulated gamete, as well as beyond, as it awaits fertilization in the fallopian tube. Not surprisingly, cumulus cells thus serve a pivotal role in supporting the oocyte, whether in vivo or in vitro.
Adiponectin is the most abundant adipocytokine and it may have insulin-sensitizing property. Clinical evidences demonstrate that the reduction of plasma or serum adiponectin levels is commonly observed in the patients with insulin resistance in comparison with healthy control subjects. Similarly, hypoadiponectinemia is evident in PCOS, and adiponectin levels correlated inversely with insulin resistance. Our previous study confirmed that SNPs rs2241766+45G15(T/G) and rs1501299+276(G/T) in the ADIPOQ gene were strongly associated with PCOS among Han Chinese women. The susceptibility of these two polymorphisms was mainly conferred in insulin resistance and insulin action, which suggested that adiponectin genetic variations might play a common role in the pathogenesis of PCOS. Recently, few reports have summarized the effects of adiponectin on the reproductive organs, either at a molecular level or for clinical relevance. Studies have showed that adiponectin can modulate not only follicle growth but also embryo development in mice and humans. But, the role of adiponectin in embryo development of PCOS patients was unknown.
Therefore, the present study was divided into two parts. The first part was to assess the true accuracy of FORT as a prognostic indicator of the response to FSH and the reproductive competence reflected by the outcomes of oocytes and embryos after IVF/ICSI treatment, especially in PCOS patients. The second one was to identify the mRNA and protein expressions of adiponectin and adiponectin receptors in cumulus cells, and the role of adiponectin in embryo development of PCOS patients after IVF treatment.
PartⅠ
Prediction of IVF/ICSI outcome based on the values of the Follicular Output RaTe (FORT) in PCOS patients
Objective To assess the true accuracy of the Follicular Output RaTe (FORT) as a prognostic indicator of the response to FSH and reproductive competence after IVF/ICSI treatment, especially in PCOS patients.
Method A total of1643IVF/ICSI cycles, including140PCOS patients who underwent ovarian stimulation, were studied.140cases were diagnosed as PCOS based on the criteria of the Rotterdam ESHRE/ASRM-sponsored PCOS Consensus Workshop Group. The median age was32in both the PCOS and non-PCOS groups. Among the patients with PCOS, there were79cases (56.43%) with oligo-and/or anovulation+hyperandrogemism+PCO,46cases (32.86%) with oligo-and/or anovulation+PCO,11cases (7.86%) with oligo-and/or anovulation+ hyperandrogemism and4cases (2.86%) with hyperandrogemism+PCO. All patients underwent standard pituitary down-regulation protocol. FORT was calculated as the ratio of the pre-ovulatory follicle (16-22mm in diameter) count (PFC) on the day of hCG/small antral follicle (3-10mm in diameter) count (AFC) at baseline (the first day of rFSH use). Low, middle and high FORT groups were divided according to tertile values. Trans-vaginal oocyte retrieval was performed34-36hours after the administration of hCG. Oocytes were fertilized either via conventional insemination or ICSI based on the couple's history. Fertilization was assessed16-18hours after IVF or ICSI. Embryo transfers were performed3days after oocyte retrieval. No more than three embryos per patient were transferred; surplus embryos were cryopreserved. Progesterone vaginal tablets were administered as luteal support from the day of the oocyte retrieval. Clinical pregnancy was defined as the presence of a gestational sac confirmed5weeks after embryo transfer by ultrasonography.
Result Among the140PCOS cases, the clinical pregnancy rate was47.14%(66/140). Women with PCOS who achieved clinical pregnancy exhibited significantly lower FORT values (0.56±0.21vs.0.66±0.29, P<0.05) than those without clinical pregnancy.
The mean value of FORT was0.65. Low FORT (n=45), middle FORT (n=56) and high FORT (n=39) referred to FORT values below the33rd percentile (FORT<0.5), between the33rd and67th percentiles (FORT0.5～0.73) and above the67th percentile (FORT>0.73), respectively. In PCOS patients, there were no differences in embryo implantation or clinical pregnancy rates among the three groups according to FORT. Interestingly, although the number of retrieved oocytes was highest in the high FORT group (17.21±7.66vs.15.63±7.08vs.12.04±7.54, P<0.05), the rates of fertilization (69.49%vs.63.47%vs.60.66%, P<0.01) and good-quality embryos (72.71%vs.68.06%vs.64.99%, P<0.05) were significantly higher in middle FORT group. Furthermore, a better IVF/ICSI outcome was achieved in PCOS patients with middle FORT values.
Among the1503non-PCOS cases, the clinical pregnancy rate was52.43%(788/1503). FORT was similar in patients who were pregnant and in those who were not pregnant (0.66±0.25vs.0.63±0.26). The rates of2PN fertilization (69.20%vs.64.79%, P<0.001) and good-quality embryos (75.77%vs.55.90%, P<0.001) were significantly higher in the patients who achieved clinical pregnancy.
Low FORT (n=402), middle FORT (n=632) and high FORT (n-469) referred to FORT values, respectively. PFC (5.49±2.85vs.8.41±3.27vs.11.54±4.33, P<0.001), the number of retrieved oocytes (8.45±5.64vs.11.52±6.37vs.13.30±6.34, P<0.001) and the total number of embryos available (4.94±3.22vs.6.37±3.69vs.7.33±3.89, P<0.001) increased progressively from the low to high FORT groups. Meanwhile, the rates of good-quality embryos, embryo implantation and clinical pregnancy increased dramatically in accordance with the FORT values (P<0.05); the fertilization rate remained steady.
FORT was correlated with AFC (r=-0.162, P<0.001), PFC (r=0.607, P<0.001), serum basal FSH levels (r=-0.123, P<0.001), total number of embryos that could be transferred (r=0.259, P<0.001), and the number of good-quality embryos (r=0.197, P<0.001). Multiple linear regression analysis was performed to study the major independent factors for the number of good-quality embryos, which was used as the dependent variable. Among all of the independent variables, the FORT value (t=1.982, P<0.05) and number of retrieved oocytes (t=17.246, P<0.01) were influential factors but not the AFC and PFC.
Conclusion Data from the present study suggest that middle FORT values in PCOS patients and high FORT values in non-PCOS patients may predict better outcomes for IVF/ICSI. To improve the outcome of IVF/ICSI, we should modulate FORT according to the heterogeneity of follicle competence in PCOS patients.
Part II
The role of adiponectin and its receptors on early embryo development in PCOS patients
Objective To identify the mRNA and protein expressions of adiponectin and adiponectin receptors (AdipoRl, AdipoR2and T-cadherin) in cumulus cells isolated from oocytes according to blastocyst formation and to evaluate the role of adiponectin and its receptors on early embryo development in PCOS patients.
Method A total of55patients (27PCOS patients and28controls) were studied in this study. PCOS patients were diagnosed based on the presence of two out of three criteria of the Rotterdam ESHRE/ASRM-sponsored PCOS Consensus Workshop Group. In the control group, the patients were non-PCOS, who sought for IVF for the tubal factor. All the patients were divided according to the BMI index. The CCs surrounding a single oocyte were removed using a sharp needle. Oocytes were further inseminated and only mature and normal fertilized oocytes (2PN) were included in this research. CCs from the oocytes yielding blastocyst after5/6days in vitro culture was classified as "B+" group and CCs from oocytes could not develop to blastocyst stage after Day6were divided into "B-" groups. CCs according to the oocytes from the same type of embryos were collected into one pool, and at least three pools were analyzed for one group.8-15CCs per group were collected for RNA or protein extraction. Real time RT-PCR and western blot were performed to identify the mRNA and protein expressions of adiponectin and adiponectin receptors (AdipoRl, AdipoR2and T-cadherin) in cumulus cells.
Result mRNA and proteins of adiponectin and adiponectin receptors were expressed in cumulus cells of PCOS and non-PCOS patients by RT-PCR analysis and western blot. Adiponectin and receptor T-cadherin were weakly expressed in cumulus cells of human. The AdipoR2mRNA levels in cumulus cells were higher statistically in PB+group than in PB-group in non-obese or obese PCOS patients respectively (0.1647± 0.0161vs.0.0783±0.0385, P<0.05;0.1948±0.0307vs.0.1057±0.0236, P<0.05). And only in PCOS patients of obese, the AdipoRl mRNA levels in cumulus cells were higher statistically in PB`group than in PB-group (0.5162±0.0371vs.0.2448±0.0333, P<0.01).
Conclusion Our studies support the recent reports adiponectin can modulate embryo development in human. The development of early embryo was associated with the up-regulation of AdipoRl and AdipoR2expression in cumulus cells of PCOS patients. Further investigations should be done to clarify the mechanisms of the association of adiponectin and embryo development.
排卵障碍是PCOS患者不孕的主要原因,诱导排卵是PCOS合并不孕患者的一线治疗。但是对于接受了标准促排卵治疗后仍未妊娠,以及同时合并输卵管因素或者男性因素不孕的患者,需要借助体外受精-胚胎移植(in-vitro fertilization&embryo transfer, IVF-ET)及卵胞浆内单精子显微注射(intracytoplasmic sperm injection, ICSI)技术获得妊娠。IVF/ICSI技术为PCOS不孕患者提供了有效的治疗手段,控制性卵巢超促排卵(controlled ovarian hyperstimulation, COH)是辅助生殖技术的关键过程。但是,由于PCOS患者对于卵巢刺激的敏感性增加,与非PCOS患者相比,接受COH治疗的PCOS患者每周期的获卵率及未成熟卵率显著增加,但是受精率显著降低。在临床工作中,我们发现部分PCOS患者虽然获卵数多,但是卵子及胚胎质量差,影响患者治疗结局。优化促排卵方案,保证窦卵泡对卵泡刺激激素(follicular stimulating hormone, FSH)的适宜反应,获取高质量卵子,是获得良好妊娠结局的前提。迄今为止,尚没有一个指标可以同时预测卵巢对促排卵的反应性以及卵子发育潜能。2011年有学者创造性地提出了卵泡输出率(follicular output rate, FORT)的概念,对月经周期规律患者的研究认为,FORT是反应卵巢中卵泡潜能的量化指标,但是FORT对PCOS患者IVF/ICSI结局的预测价值尚不明确。
卵冠丘复合物(cumulus-oocyte complex, COCs)中,卵丘颗粒细胞直接包绕着卵母细胞,伴随着卵母细胞发育、成熟、排卵至受精的过程。卵丘颗粒细胞分解代谢胆固醇、葡萄糖,通过缝隙连接为卵母细胞提供必要的能量,对卵母细胞的发育是至关重要的。近年来的研究提示卵丘颗粒细胞基因表达的差异可以作为预测卵子及胚胎发育潜能的非侵入性方法。脂联素(Adiponectin)作为一种脂肪细胞因子,对于调节人胰岛素敏感性起到非常重要的作用。PCOS患者血清脂联素水平下降,而脂联素水平与胰岛素抵抗程度呈负相关,PCOS患者脂联素水平下降可能是导致其卵巢功能异常的重要原因。前期对山东汉族PCOS患者的研究显示,脂联素基因SNP rs1501299+276(A/C)与SNP rs2241766+45G15G(G/T)多态性与PCOS发病具有相关性,其中脂联素基因SNP+276(A/C)多态性具有调节PCOS患者胰岛素敏感性的作用,在PCOS的发病过程中提供了发生胰岛素抵抗的遗传易感性。近年来的研究认为脂联素直接参与调节生殖功能,体外培养小鼠卵冠丘复合物经脂联素作用后,可以显著增加受精率、胚胎卵裂率及囊胚形成率；人卵丘细胞中AdipoR1、AdipoR2的表达量与体外培养胚胎的囊胚形成率相关,提示卵丘细胞可能是脂联素作用的一个重要靶点,对胚胎的发育潜能具有一定的作用。但是,脂联素及其受体在PCOS患者卵丘颗粒细胞中的表达情况以及与PCOS患者胚胎发育潜能的相关性尚未见研究报道。
基于以上研究背景,本课题从提高PCOS患者胚胎发育潜能,改善妊娠结局着手,进行了相关临床及基础研究。第一部分通过分析IVF/ICSI周期促排卵方案及妊娠结局等临床资料,评估卵泡输出率在评价PCOS患者卵巢对FSH反应性、卵子质量及预测IVF/ICSI妊娠结局方面的价值。第二部分通过检测脂联素及其受体在不同质量胚胎所对应的卵丘颗粒细胞中的表达情况,分析脂联素及其受体与胚胎发育潜能的相关性,明确脂联素是否通过卵丘颗粒细胞发挥影响PCOS患者胚胎发育潜能的作用。
第一部分
多囊卵巢综合征患者卵泡输出率预测IVF/ICSI结局的研究
目的研究体外受精-胚胎移植/卵胞浆内单精子显微注射周期中,卵泡输出率在评估多囊卵巢综合征患者卵巢对FSH反应性及预测妊娠结局的价值。
方法以2010年1月至2011年12月在烟台毓璜顶医院生殖医学中心进行IVF/ICSI治疗的1643个周期作为研究对象。其中多囊卵巢综合征患者140例,非多囊卵巢综合征患者1503例。按照2003年ESHRE/ASRM建议的诊断标准作为PCOS患者的纳入标准。PCOS组与非PCOS组患者的中位年龄均为32岁。PCOS患者中,79例(56.43%)患者为稀发排卵和／或无排卵+高雄激素+卵巢多囊样改变；46例(32.86%)患者为稀发排卵和／或无排卵+高雄激素；11例(7.86%)为稀发排卵和／或无排卵+卵巢多囊样改变；4例(2.86%)为高雄激素+卵巢多囊样改变。所有患者均采用标准长方案进行控制性超促排卵(COH),开始应用促性腺激素(Gn)日通过阴道B超检测,记录双侧卵巢中直径3-10mm窦卵泡数(AFC),注射hCG日记录双侧卵巢中直径16-22mm成熟卵泡数(PFC),计算出卵泡输出率(FORT=PFC/AFC)。注射hCG后34-36h行阴道超声引导下穿刺取卵术,取卵当日根据男方精液情况行常规IVF或ICSI受精,IVF或ICSI后16-18小时观察受精情况,取卵后第3天进行胚胎移植,剩余可移植胚胎冷冻保存。自取卵日开始进行黄体支持,胚胎移植后14天查血hCG确定生化妊娠,移植后35天阴道B超检查探及妊娠囊确定为临床妊娠。
结果在PCOS患者的140个治疗周期中,临床妊娠66周期,未妊娠74周期,临床妊娠率为47.14%(66/140)。妊娠组FORT值显著低于未妊娠组(0.56±0.21vs.0.66±0.29,P<0.05)。
FORT的均数为0.65,根据FORT值,将PCOS患者分为三组,低FORT组(FORT<0.5, n=45),中等FORT组(FORT0.5～0.73, n=56)及高FORT组(FORT>0.73, n=39)。根据FORT分组后,胚胎种植率及临床妊娠率在低、中、高FORT三组间无统计学差异。随着FORT逐渐增高,成熟卵泡数及获卵数逐渐增加,然而值得注意的是,尽管在高FORT组,获卵数是最多的(12.04±7.54vs.15.63±7.08vs.17.21±7.66,P<0.05),但是2PN受精率(63.47%vs.69.49%vs.60.66%,P<0.01)以及优质胚胎率(68.06%vs.72.71%vs.64.99%,P<0.05)在中等FORT组最高,提示中等FORT值组的患者可以获得较好的IVF/ICSI结局。
非-PCOS患者的1503个治疗周期中,临床妊娠788周期,未妊娠715周期,临床妊娠率为52.43%(788/1503),PCOS患者与非PCOS患者临床妊娠率无统计学差异(P>0.05)。妊娠组与未妊娠组相比,两组间FORT (0.66±0.25vs.0.63±0.26,P>0.05)无统计学差异。妊娠组的2PN受精率(69.20%vs.64.79%,P<0.001)、优质胚胎率(75.77%vs.55.90%,P<0.001)显著高于未妊娠组。
根据FORT值,将非-PCOS患者分为三组,低FORT组(FORT<0.5,n=402),中等FORT组(FORT0.5～0.73,n=632)及高FORT组(FORT>0.73,n=469)。 FSH/LH比值(1.73±1.87vs.1.52±0.82vs.1.49±0.82,P=0.007)以及AFC(14.51±6.33vs.14.00±5.37vs.12.32±4.28,P<0.001)在低FORT组最高。从低FORT组至高FORT组,PFC(5.49士2.85vs.8,41±3.27vs.11.54士4.33,P<0.001),获卵数(8.45±5.64vs.11.52±6.37vs.13.30±6.34,P<0.001),可移植胚胎总数(4.94±3.22vs.6.37±3.69vs.7.33±3.89,P<0.001)逐渐增高。与此同时,优质胚胎率(65.98%vs.66.48%vs.68.91%,P<0.05)、胚胎种植率(29.68%vs.33.80%vs.35.06%,P<0.01)、临床妊娠率(46.27%vs.53.46%vs.55.65%,P<0.05)随着FORT的增高也是逐渐增高的,差异具有统计学意义,而2PN受精率(68.07%vs.66.95%vs.67.13%,P>0.05)在三组之间无统计学差异。
回归分析显示FORT与AFC(r=-0.162,P<0.001),血清基础卵泡刺激激素(r=-0.123,P<0.001)呈负相关,与PFC(r=0.607,P<0.001),可移植胚胎总数(r=0.259,P<0.001),以及优质胚胎总数呈正相关(r=0.197,P<0.001)。以优质胚胎数作为自变量,进行多元线性回归分析,结果显示在所有因变量中,FORT(t=1.982,P<0.05)以及获卵数(t=17.246,P<0.01)是主要影响变量,而不是AFC或者PFC。
结论中等FORT值的PCOS患者以及高FORT值的非PCOS患者可以获得较好的IVF/ICSI妊娠结局。PCOS患者进行超促排卵治疗时,应调整用药尽量得到适宜的卵泡输出率,从而改善IVF/ICSI妊娠结局。
第二部分
脂联素及其受体与多囊卵巢综合征患者胚胎发育潜能的相关性研究
目的检测脂联素及其受体基因在不同质量胚胎所对应的卵丘颗粒细胞中的表达情况,分析脂联素及其受体与胚胎发育潜能的相关性,明确脂联素是否通过卵丘颗粒细胞发挥影响PCOS患者胚胎发育潜能的作用。
方法PCOS组共27例,按照2003年ESHRE/ASRM建议的诊断标准作为PCOS患者的纳入标准。对照组为因输卵管因素接受IVF-ET治疗的患者,共28例。所有患者根据体重指数分为体重正常组(BMI<25kg/m2)及肥胖组(BMI≥25kg/m2)。行阴道超声引导下穿刺取卵术后获取卵冠丘复合物,用机械法分离每个卵子的卵丘颗粒细胞,采用单卵微滴短时授精,卵子与收集的卵丘颗粒细胞一一对应,根据每个卵子受精、胚胎发育及囊胚形成情况,回顾性将卵子所对应的卵丘颗粒细胞分为囊胚形成组(B+组)和未形成囊胚组(B-组)。采用real time RT-PCR和western blot的方法检测脂联素及其受体mRNA、蛋白表达情况,并比较B+组和B-组的mRNA表达量是否存在差异。
结果Real time RT-PCR和western blot方法检测到AdipoQ. AdipoR1、AdipoR2和T-cadherin基因mRNA及蛋白在卵丘颗粒细胞中有表达,但是以AdipoRl和AdipoR2的表达为主。体重正常PCOS患者中,AdipoR2基因mRNA的相对表达量在囊胚形成组(PB+组)所对应卵丘颗粒细胞中显著高于未形成囊胚组(PB-组)(0.1647±0.0161vs.0.0783±0.0385,P<0.05)；在肥胖PCOS患者中,AdipoRl及AdipoR2基因mRNA的相对表达量在囊胚形成组(PB+组)所对应卵丘颗粒细胞中均显著高于未形成囊胚组(PB-组)(0.5162±0.0371vs.0.2448±0.0333,P<0.01)(0.1948±0.0307vs.0.1057±0.0236,P<0.05)。在非-PCOS患者中,脂联素及其受体基因mRNA的相对表达量在不同质量胚胎组间无显著性差异。
结论脂联素、AdipoR1、AdipoR2和T-cadherin基因mRNA和蛋白在卵丘颗粒细胞中有表达,但是以AdipoRl和AdipoR2的表达为主。AdipoR2基因的mRNA表达水平与PCOS患者胚胎发育潜能具有相关性；AdipoR1基因的mRNA表达水平与肥胖PCOS患者的胚胎发育潜能相关。脂联素可能通过上调PCOS患者卵丘颗粒细胞AdipoR1和AdipoR2的表达而发挥影响胚胎发育潜能的作用。
Background Polycystic ovary syndrome (PCOS) is one of the most prevalent endocrinopathies, affecting5-10%of women of reproductive age. The characteristic clinical features of PCOS include menstrual irregularity such as oligomenorrhoea, signs of hyperandrogenaemia including hirsutism, acne, and/or obesity, and polycystic morphology on ovarian ultrasound. More recent studies have highlighted the link between PCOS and metabolic syndrome, insulin resistance and compensatory hyperinsulinemia appear to be a prominent feature of this phenotype. The pathophysiology of PCOS still remains largely uncharacterized and it has puzzled gynecologists and endocrinologists for many years and proven very difficult to define. Anovulation is the common cause of infertility in PCOS patients, the vast majority of women with PCOS will ovulate and conceive following ovulation induction therapy. Those who fail to be pregnancy often have an additional factor, such as male factor, fallopian tube factor or endometriosis. They should seek in-vitro fertilization&embryo transfer (IVF-ET) or intracytoplasmic sperm injection (ICSI) as a treatment for infertility. Controlled ovarian hyperstimulation (COH) is the key processor in assisted reproductive technology (ART). Women with PCOS will almost yield more oocytes than those without PCOS. However, the fertilization rate of the oocytes from PCOS is almost inevitably lower. A meta-analysis demonstrated an increased cycle cancellation rate, but more oocytes retrieved per retrieval and a lower fertilization rate in PCOS patients undergoing IVF. Although the regulatory mechanisms determining the extent of the sensitivity of individual antral follicles to follicle stimulating hormone (FSH) remains to be elucidated, the appropriate response of antral follicles to FSH and a high quality of oocytes may result in a good outcome after IVF/ICSI.
There is no marker that can predict both ovarian response and oocyte competence. To evaluate follicular responsiveness to exogenous FSH, the use of the Follicular Output RaTe (FORT) as an innovative term has been suggested. FORT is assessed by the ratio of the pre-ovulatory follicle (16-22mm) counts (PFC) obtained in response to FSH administration on the day of hCG to the small antral follicle (3-10mm) count (AFC) observed after the complete suppression of endogenous gonadotropins by gonadotropin-releasing hormone agonist (GnRHa)(FORT=PFC/AFC). Gallot found that FORT might be a qualitative reflector of ovarian follicular competence only in patients with regular menstrual cycles. The values of FORT as a predictor of IVF/ICSI outcome in PCOS and non-PCOS patients were unknown.
In cumulus-oocyte complex (COCs), cumulus cells are the specialized granulosa cells that directly surround the oocyte during follicular development. Importantly, these cells accompany the oocyte throughout the development from an immature to a fully mature ovulated gamete, as well as beyond, as it awaits fertilization in the fallopian tube. Not surprisingly, cumulus cells thus serve a pivotal role in supporting the oocyte, whether in vivo or in vitro.
Adiponectin is the most abundant adipocytokine and it may have insulin-sensitizing property. Clinical evidences demonstrate that the reduction of plasma or serum adiponectin levels is commonly observed in the patients with insulin resistance in comparison with healthy control subjects. Similarly, hypoadiponectinemia is evident in PCOS, and adiponectin levels correlated inversely with insulin resistance. Our previous study confirmed that SNPs rs2241766+45G15(T/G) and rs1501299+276(G/T) in the ADIPOQ gene were strongly associated with PCOS among Han Chinese women. The susceptibility of these two polymorphisms was mainly conferred in insulin resistance and insulin action, which suggested that adiponectin genetic variations might play a common role in the pathogenesis of PCOS. Recently, few reports have summarized the effects of adiponectin on the reproductive organs, either at a molecular level or for clinical relevance. Studies have showed that adiponectin can modulate not only follicle growth but also embryo development in mice and humans. But, the role of adiponectin in embryo development of PCOS patients was unknown.
Therefore, the present study was divided into two parts. The first part was to assess the true accuracy of FORT as a prognostic indicator of the response to FSH and the reproductive competence reflected by the outcomes of oocytes and embryos after IVF/ICSI treatment, especially in PCOS patients. The second one was to identify the mRNA and protein expressions of adiponectin and adiponectin receptors in cumulus cells, and the role of adiponectin in embryo development of PCOS patients after IVF treatment.
PartⅠ
Prediction of IVF/ICSI outcome based on the values of the Follicular Output RaTe (FORT) in PCOS patients
Objective To assess the true accuracy of the Follicular Output RaTe (FORT) as a prognostic indicator of the response to FSH and reproductive competence after IVF/ICSI treatment, especially in PCOS patients.
Method A total of1643IVF/ICSI cycles, including140PCOS patients who underwent ovarian stimulation, were studied.140cases were diagnosed as PCOS based on the criteria of the Rotterdam ESHRE/ASRM-sponsored PCOS Consensus Workshop Group. The median age was32in both the PCOS and non-PCOS groups. Among the patients with PCOS, there were79cases (56.43%) with oligo-and/or anovulation+hyperandrogemism+PCO,46cases (32.86%) with oligo-and/or anovulation+PCO,11cases (7.86%) with oligo-and/or anovulation+ hyperandrogemism and4cases (2.86%) with hyperandrogemism+PCO. All patients underwent standard pituitary down-regulation protocol. FORT was calculated as the ratio of the pre-ovulatory follicle (16-22mm in diameter) count (PFC) on the day of hCG/small antral follicle (3-10mm in diameter) count (AFC) at baseline (the first day of rFSH use). Low, middle and high FORT groups were divided according to tertile values. Trans-vaginal oocyte retrieval was performed34-36hours after the administration of hCG. Oocytes were fertilized either via conventional insemination or ICSI based on the couple's history. Fertilization was assessed16-18hours after IVF or ICSI. Embryo transfers were performed3days after oocyte retrieval. No more than three embryos per patient were transferred; surplus embryos were cryopreserved. Progesterone vaginal tablets were administered as luteal support from the day of the oocyte retrieval. Clinical pregnancy was defined as the presence of a gestational sac confirmed5weeks after embryo transfer by ultrasonography.
Result Among the140PCOS cases, the clinical pregnancy rate was47.14%(66/140). Women with PCOS who achieved clinical pregnancy exhibited significantly lower FORT values (0.56±0.21vs.0.66±0.29, P<0.05) than those without clinical pregnancy.
The mean value of FORT was0.65. Low FORT (n=45), middle FORT (n=56) and high FORT (n=39) referred to FORT values below the33rd percentile (FORT<0.5), between the33rd and67th percentiles (FORT0.5～0.73) and above the67th percentile (FORT>0.73), respectively. In PCOS patients, there were no differences in embryo implantation or clinical pregnancy rates among the three groups according to FORT. Interestingly, although the number of retrieved oocytes was highest in the high FORT group (17.21±7.66vs.15.63±7.08vs.12.04±7.54, P<0.05), the rates of fertilization (69.49%vs.63.47%vs.60.66%, P<0.01) and good-quality embryos (72.71%vs.68.06%vs.64.99%, P<0.05) were significantly higher in middle FORT group. Furthermore, a better IVF/ICSI outcome was achieved in PCOS patients with middle FORT values.
Among the1503non-PCOS cases, the clinical pregnancy rate was52.43%(788/1503). FORT was similar in patients who were pregnant and in those who were not pregnant (0.66±0.25vs.0.63±0.26). The rates of2PN fertilization (69.20%vs.64.79%, P<0.001) and good-quality embryos (75.77%vs.55.90%, P<0.001) were significantly higher in the patients who achieved clinical pregnancy.
Low FORT (n=402), middle FORT (n=632) and high FORT (n-469) referred to FORT values, respectively. PFC (5.49±2.85vs.8.41±3.27vs.11.54±4.33, P<0.001), the number of retrieved oocytes (8.45±5.64vs.11.52±6.37vs.13.30±6.34, P<0.001) and the total number of embryos available (4.94±3.22vs.6.37±3.69vs.7.33±3.89, P<0.001) increased progressively from the low to high FORT groups. Meanwhile, the rates of good-quality embryos, embryo implantation and clinical pregnancy increased dramatically in accordance with the FORT values (P<0.05); the fertilization rate remained steady.
FORT was correlated with AFC (r=-0.162, P<0.001), PFC (r=0.607, P<0.001), serum basal FSH levels (r=-0.123, P<0.001), total number of embryos that could be transferred (r=0.259, P<0.001), and the number of good-quality embryos (r=0.197, P<0.001). Multiple linear regression analysis was performed to study the major independent factors for the number of good-quality embryos, which was used as the dependent variable. Among all of the independent variables, the FORT value (t=1.982, P<0.05) and number of retrieved oocytes (t=17.246, P<0.01) were influential factors but not the AFC and PFC.
Conclusion Data from the present study suggest that middle FORT values in PCOS patients and high FORT values in non-PCOS patients may predict better outcomes for IVF/ICSI. To improve the outcome of IVF/ICSI, we should modulate FORT according to the heterogeneity of follicle competence in PCOS patients.
Part II
The role of adiponectin and its receptors on early embryo development in PCOS patients
Objective To identify the mRNA and protein expressions of adiponectin and adiponectin receptors (AdipoRl, AdipoR2and T-cadherin) in cumulus cells isolated from oocytes according to blastocyst formation and to evaluate the role of adiponectin and its receptors on early embryo development in PCOS patients.
Method A total of55patients (27PCOS patients and28controls) were studied in this study. PCOS patients were diagnosed based on the presence of two out of three criteria of the Rotterdam ESHRE/ASRM-sponsored PCOS Consensus Workshop Group. In the control group, the patients were non-PCOS, who sought for IVF for the tubal factor. All the patients were divided according to the BMI index. The CCs surrounding a single oocyte were removed using a sharp needle. Oocytes were further inseminated and only mature and normal fertilized oocytes (2PN) were included in this research. CCs from the oocytes yielding blastocyst after5/6days in vitro culture was classified as "B+" group and CCs from oocytes could not develop to blastocyst stage after Day6were divided into "B-" groups. CCs according to the oocytes from the same type of embryos were collected into one pool, and at least three pools were analyzed for one group.8-15CCs per group were collected for RNA or protein extraction. Real time RT-PCR and western blot were performed to identify the mRNA and protein expressions of adiponectin and adiponectin receptors (AdipoRl, AdipoR2and T-cadherin) in cumulus cells.
Result mRNA and proteins of adiponectin and adiponectin receptors were expressed in cumulus cells of PCOS and non-PCOS patients by RT-PCR analysis and western blot. Adiponectin and receptor T-cadherin were weakly expressed in cumulus cells of human. The AdipoR2mRNA levels in cumulus cells were higher statistically in PB+group than in PB-group in non-obese or obese PCOS patients respectively (0.1647± 0.0161vs.0.0783±0.0385, P<0.05;0.1948±0.0307vs.0.1057±0.0236, P<0.05). And only in PCOS patients of obese, the AdipoRl mRNA levels in cumulus cells were higher statistically in PB`group than in PB-group (0.5162±0.0371vs.0.2448±0.0333, P<0.01).
Conclusion Our studies support the recent reports adiponectin can modulate embryo development in human. The development of early embryo was associated with the up-regulation of AdipoRl and AdipoR2expression in cumulus cells of PCOS patients. Further investigations should be done to clarify the mechanisms of the association of adiponectin and embryo development.
引文
1. Franks S. Polycystic ovary syndrome. N Engl J Med.1995,28,333:853-861.
2. Legro RS. Polycystic ovary syndrome:the new millenium. Mol Cell Endocrinol. 2001,26,184:87-93.
3. Ehrmann DA. Polycystic ovary syndrome. N Engl J Med.2005,24,352: 1223-1236.
4. Heijnen E, Eijkemans M, Hughes E, et al. A meta-analysis of outcomes of conventional IVF in women with polycystic ovary syndrome. Hum Reprod Update.2006,12:13-21.
5. Chang MY, Chiang CH, Hsieh TT, et al. Use of the antral follicle count to predict the outcome of assisted reproductive technologies. Fertil Steril.1998, 69:505-510.
6. de Carvalho BR, Rosa e Silva AC, Rosa e Silva JC, et al. Ovarian reserve evaluation:state of the art. J Assist Reprod Genet.2008,25:311-322.
7. Kline J, Kinney A, Kelly A, et al. Predictors of antral follicle count during the reproductive years. Hum Reprod.2005,20:2179-2189.
8. Scheffer GJ, Broekmans FJ, Dorland M, et al. Antral follicle counts by transvaginal ultrasonography are related to age in women with proven natural fertility. Fertil Steril.1999,72:845-851.
9. Bancsi LF, Broekmans FJ, Eijkemans MJ, et al. Predictors of poor ovarian response in in vitro fertilization:a prospective study comparing basal markers of ovarian reserve. Fertil Steril.2002,77:328-336.
10. Broer SL, Mol BW, Hendriks D, et al. The role of antimullerian hormone in prediction of outcome after IVF:comparison with the antral follicle count. Fertil Steril.2009,91:705-714.
11. Melo MA, Garrido N, Alvarez C, et al. Antral follicle count (AFC) can be used in the prediction of ovarian response but cannot predict the oocyte/embryo quality or the in vitro fertilization outcome in an egg donation program. Fertil Steril. 2009,91:148-156.
12. Genro VK, Grynberg M, Scheffer JB, et al. Serum anti-Mullerian hormone levels are negatively related to Follicular Output RaTe (FORT) in normo-cycling women undergoing controlled ovarian hyperstimulation. Hum Reprod.2011, 26:671-677.
13. Gallot V, Berwanger da Silva AL, Genro V, et al. Antral follicle responsiveness to follicle-stimulating hormone administration assessed by the Follicular Output RaTe (FORT) may predict in vitro fertilization-embryo transfer outcome. Hum Reprod.2012,27:1066-1072.
14. Genro VK, Matte U, De Conto E, et al. Frequent polymorphisms of FSH receptor do not influence antral follicle responsiveness to follicle-stimulating hormone administration as assessed by the Follicular Output RaTe (FORT). J Assist Reprod Genet.2012,29:657-663.
15. The Rotterdam ESHRE/ASRM-sponsored PCOS consensus workshop group. Revised 2003 consensus on diagnostic criteria and long-term health risks related to polycystic ovary syndrome (PCOS). Human Reproduction.2004,19:41-47.
16. Jurema MW, Bracero NJ, Garcia JE. Fine tuning cycle day 3 hormonal assessment of ovarian reserve improves in vitro fertilization outcome in gonadotropin-releasing hormone antagonist cycles. Fertil Steril. 2003;80:1156-1161.
17. Bancsi LF, Broekmans FJ, Mol BW, et al. Performance of basal follicle-stimulating hormone in the prediction of poor ovarian response and failure to become pregnant after in vitro fertilization:a meta-analysis. Fertil Steril. 2003,79:1091-1100.
18. Bancsi LF, Broekmans FJ, Looman CW, et al. Predicting poor ovarian response in IVF:use of repeat basal FSH measurement. J Reprod Med.2004,49:187-194.
19. Barad DH, Weghofer A, Gleicher N. Comparing anti-Mullerian hormone (AMH) and follicle-stimulating hormone (FSH) as predictors of ovarian function. Fertil Steril.2009,91:1553-1555.
20. Lekamge DN, Barry M, Kolo M, et al. Anti-Mullerian hormone as a predictor of IVF outcome. Reprod Biomed Online.2007,14:602-610.
21. Wunder DM, Guibourdenche J, Birkhauser MH, et al. Anti-Mullerian hormone and inhibin B as predictors of pregnancy after treatment by in vitro fertilization/intracytoplasmic sperm injection. Fertil Steril.2008,90:2203-2210.
22. Lee TH, Liu CH, Huang CC, et al. Serum anti-Mullerian hormone and estradiol levels as predictors of ovarian hyperstimulation syndrome in assisted reproduction technology cycles. Hum Reprod.2008,23:160-167.
23. Smeenk JM, Sweep FC, Zielhuis GA, et al. Antimullerian hormone predicts ovarian responsiveness, but not embryo quality or pregnancy, after in vitro fertilization or intracyoplasmic sperm injection. Fertil Steril.2007,87:223-226.
24. Broekmans FJ, Visser JA, Laven JS, et al. Anti-Mullerian hormone and ovarian dysfunction. Trends Endocrinol Metab.2008,19:340-347.
25. La Marca A, Broekmans FJ, Volpe A, et al. Anti-Mullerian hormone (AMH): what do we still need to know? Hum Reprod.2009,24:2264-2275.
26. Kaya C, Pabuccu R, Satiroglu H. Serum antimullerian hormone concentrations on day 3 of the in vitro fertilization stimulation cycle are predictive of the fertilization, implantation, and pregnancy in polycystic ovary syndrome patients undergoing assisted reproduction. Fertil Steril.2010,94:2202-2207.
27. Coffler MS, Patel K, Dahan MH, et al. Evidence for abnormal granulosa cell responsiveness to follicle-stimulating hormone in women with polycystic ovary syndrome. J Clin Endocrinol Metab.2003,88:1742-1747.
28. Franks S, Roberts R, Hardy K. Gonadotrophin regimens and oocyte quality in women with polycystic ovaries. Reprod Biomed Online.2003,6:181-184.
29. Fabregues F, Penarrubia J, Vidal E, et al. Oocyte quality in patients with severe ovarian hyperstimulation syndrome:a self-controlled clinical study. Fertil Steril. 2004,82:827-833.
30. Aboulghar MA, Mansour RT, Serour GI, et al. Oocyte quality in patients with severe ovarian hyperstimulation syndrome. Fertil Steril.1997,68:1017-1021.
1. Scherer PE, Williams S, Fogliano M, et al. A novel serum protein similar to C1q, produced exclusively in adipocytes. J Biol Chem.1995,270(45):26746-26749.
2. Nakano Y, Tobe T, Choi-Miura NH, et al. Isolation and characterization of GBP28, a novel gelatin-binding protein purified from human plasma. J Biochem. 1996,120(4):803-812.
3. Hu E, Liang P, Spiegelman BM. AdipoQ is a novel adipose-specific gene dysregulated in obesity. J Biol Chem.1996,271(18):10697-10703.
4. Maeda K, Okubo K, Shimomura I, et al. cDNA cloning and expression of a novel adipose specific collagen-like factor, apMI (adipose most abundant gene transcript 1). BiochemBiophys Res Commun.1996,221(2):286-289.
5. Ouchi N, Kihara S, Arita Y, et al. Novel modulator for endothelial adhesion molecules:adipocyte-derived plasma protein adiponectin. Circulation.1999, 100(25):2473-2476.
6. Shapiro L, Scherer PE. The crystal structure of a complement-lq family protein suggests an evolutionary link to tumor necrosis factor. Curr Biol.1998,8(6): 335-338.
7. Arita Y, Kihara S, Ouchi N, et al. Paradoxical decrease of an adipose-specific protein, adiponectin, in obesity. BiochemBiophys Res Commun.1999,257(1): 79-83.
8. Yamauchi T, Kamon J, Ito Y, et al. Cloning of adiponectin receptors that mediate antidiabetic metabolic effects. Nature.2003,423(6941):762-769.
9. Hug C, Wang J, Ahmad NS, et al. T-cadherin is a receptor hexameric and high-molecular-weight forms of Acrp30/adiponectin. NatlAcadSci USA,2004. 101(28):10308-10313.
10. Ivanov D, Philippova M, Tkachuk V, et al. Cell adhesion molecule T-cadherin regulates vascular cell adhesion, phenotype and motility. Exp Cell Res.2004, 293(2):207-218.
11. Kubota N, Terauchi Y, Yamauchi T, et al. Disruption of adiponectin causes insulin resistance and neointimal formation. J Biol Chem.2002,277(29):25863-25866.
12. Yamauchi T, Kamon J, Waki H, et al. Globular adiponectin protected ob/ob mice from diabetes and ApoE-deficient mice from atherosclerosis. J Biol Chem.2003, 24; 278(4):2461-2468.
13. Brakenhielm E, Veitonmaki N, Cao R, et al. Adiponectin-induced antiangiogenesis and antitumor activity involve caspase-mediated endothelial cell apoptosis. ProcNatlAcadSci USA.2004,101(8):2476-2481.
14. Schulze MB, Shai I, Rimm EB, et al. Adiponectin and future coronary heart disease events among men with type 2 diabetes. Diabetes.2005,54(2):534-539.
15. Iglseder B, Mackevics V, Stadlmayer A, et al. Plasma adiponectin levels and sonographic phenotypes of subclinical carotid artery atherosclerosis:data from the SAPHIR Study. Stroke.2005,36(12):2577-2582.
16. Yamamoto Y, Hirose H, Saito I, et al. Correlation of the adipocyte-derived protein adiponectin with insulin resistance index and serum high-density lipoprotein-cholesterol, independent of body mass index, in the Japanese population. ClinSci (Lond).2002,103(2):137-142.
17. Lindsay RS, Funahashi T, Hanson RL, et al. Adiponectin and development of type 2 diabetes in the Pima Indian population. Lancet.2002,360(9326):57-58.
18. Snehalatha C, Mukesh B, Simon M, et al. Plasma adiponectin is an independent predictor of type 2 diabetes in Asian indians. Diabetes Care.2003,26(12): 3226-3229.
19. Pellme F, Smith U, Funahashi T, et al. Circulating adiponectin levels are reduced in nonobese but insulin-resistant first-degree relatives of type 2 diabetic patients. Diabetes.2003,52(5):1182-1186.
20. Yamamoto Y, Hirose H, Saito I, et al. Adiponectin, an adipocyte-derived protein, predicts future insulin resistance:two-year follow-up study in Japanese population. J ClinEndocrinolMetab.2004,89(1):87-90.
21. Hojlund K, Frystyk J, Levin K, et al. Reduced plasma adiponectin concentrations may contribute to impaired insulin activation of glycogen synthase in skeletal muscle of patients with type 2 diabetes. Diabetologia.2006,49(6):1283-1291.
22.张宁,陈子江,石玉华,等.血清脂联素水平测定在多囊卵巢综合征中的意义.实用妇产科杂志.2007,23(3)：154-156.
23. Ducluzeau, PH, Cousin, P, Malvoisin, E, et al. Glucose-to-insulin ratio rather than sex hormone-binding globulin and adiponectin levels is the best predictor of insulin resistance in nonobese women with polycystic ovary syndrome. Journal of Clinical Endocrinology & Metabolism.2003,88(8):3626-3631.
24. Spranger J, Mohlig M, Wegewitz U, et al. Adiponectin is independently associated with insulin sensitivity in women with polycystic ovary syndrome. Clinical Endocrinology.2004,61(6):738-746.
25. Aroda V, Ciaraldi, TP, Chang, SA, et al. Circulating and cellular adiponectin in polycystic ovary syndrome:Relationship to glucose tolerance and insulin action. Fertility & Sterility.2008,89(5):1200-1208.
26. Jensterle M, Weber M, Pfeifer M, et al. Assessment of insulin resistance in young women with polycystic ovary syndrome. International Journal of Gynecology and Obstetrics.2008,102(2):137-140.
27. Ehrmann DA. Medical progress:Polycystic ovary syndrome. New England Journal of Medicine.2005,24; 352(12):1223-1236.
28. Escobar-Morreale HF, Villuendas G, Botella-Carretero JI, et al. Adiponectin and resistin in PCOS:A clinical, biochemical and molecular genetic study. Human Reproduction.2006,21(9):2257-2265.
29. Glintborg D, Andersen M, Hagen C, et al. Evaluation of metabolic risk markers in polycystic ovary syndrome (PCOS). Adiponectin, ghrelin, leptin and body composition in hirsute PCOS patients and controls. European Journal of Endocrinology.2006,155(2):337-345.
30. Ardawi MS, Rouzi AA, Ardawi M SM,&Rouzi AA. Plasma adiponectin and insulin resistance in women with polycystic ovary syndrome. Fertility & Sterility. 2005,83(6):1708-1716.
31. Toulis KA, Goulis DG, Farmakiotis D, et al. Adiponectin levels in women with polycystic ovary syndrome:a systematic review and a meta-analysis. Hum Reprod Update.2009,15(3):297-307.
32. Campos DB, Palin MF, Bordignon V, et al. The beneficial adipokines in reproduction and fertility. Int J Obes.2008,32(2):223-231.
33. Ledoux S, Campos DB, Lopes FL, et al. Adiponectin induces periovulatory changes in ovarian follicular cells. Endocrinology.2006,147(11):5178-5186.
34. Bersinger NA, Birkhauser MH, Wunder DM. Adiponectin as a marker of success in intracytoplasmic sperm injection/embryo transfer cycles. GynecolEndocrinol. 2006,22(9):479-483.
35. Chabrolle C, Tosca L, Rame C, et al. Adiponectin increases insulin-like growth factor I induced progesterone and estradiol secretion in human granulose cells. FertilSteril.2008,92(6):1988-1996.
36. Chabrolle C, Tosca L, Crochet S, et al. Expression of adiponectin and its receptors (AdipnRl and AdipoR2) in chicken ovary:potential role in ovarian steroidogenesis. DomestAnimEndocrinol.2007,33(4):480-487.
37. Chabrolle C, Tosca L, Dupont J. Regulation of adiponectin and its receptors in rat ovary by human chorionic gonadotrophin treatment and potential involvement of adiponectin in granulosa cell steroidogenesis. Reproduction.2007,133(4): 719-731.
38. Chappaz E, Albornoz MS, Campos D, et al. Adiponectin enhances in vitro development of swine embryos. DomestAnimEndocrinol.2008,35(2):198-207.
39. Tabandeh MR, Hosseini A, Sae b M, et al. Changes in the gene expression of adiponectin and adiponectin receptors (AdipoRl and AdipoR2) in ovarian follicular cells of dairy cow at different stages of development. Theriogenology. 2010,73(5):659-669.
40. Maillard V, Uzbekova S, Guignot F, et al. Effect of adiponectin on bovine granulosa cell steroidogenesis, oocyte maturation and embryo development. Reproductive Biology and Endocrinology.2010,10; 8:23-37.
41. Richards JS, Liu Z, Kawai T, et al.Adiponectin and its receptors modulate granulosa cell and cumulus cell functions, fertility, and early embryo development in the mouse and human. FertilSteril.2012,98(2):471-479.
42. Kim ST, Marquard K, Stephens S, et al. Adiponectin and adiponectin receptors in the mouse preimplantation embryo and uterus. Human Reproduction.2011,26(1): 82-95.
43. The Rotterdam ESHRE/ASRM-sponsored PCOS consensus workshop group. Revised 2003 consensus on diagnostic criteria and long-term health risks related to polycystic ovary syndrome (PCOS). Human Reproduction.2004,19:41-47.
44. Psilopanagioti A, Papadaki H, Kranioti EF, et al. Expression of adiponectin and adiponectin receptors in human pituitary gland and brain. Neuroendocrinology. 2009,89(1):38-47.
45. Wen JP, Lv WS, Yang J, et al. Globular adiponectin inhibits GnRH secretion from GT1-7 hypothalamic GnRH neurons by induction of hyperpolarization of membrane potential. Biochem Biophys Res Commun.2008,11,371(4):756-761.
46. Wilkinson M, Brown R, Imran SA, et al. Adipokine gene expression in brain and pituitary gland. Neuroendocrinology.2007,86(3):191-209.
47. Lu M, Tang Q, Olefsky JM, et al. Adiponectin activates adenosine monophosphate-activated protein kinase and decreases luteinizing hormone secretion in LbetaT2 gonadotropes. Mol Endocrinol.2008,22(3):760-771.
48. Rodrigues-Pacheco F, Martinez-Fuentes AJ, Tovar S, Pinilla L, et al. Regulation of pituitary cell function by adiponectin. Endocrinology.2007,148(1):401-410.
49. Mitchell M, Armstrong DT, Robker RL, et al. Adipokines:implications for female fertility and obesity. Reproduction.2000,130(5):583-597.
50. Lagaly DV, Aad PY, Grado-Ahuir JA, et al. Role of adiponectin in regulating ovarian theca and granulosa cell function. Molecular and Cellular Endocrinology. 2008,12,284(1-2):38-45.
51. Kadowaki T, Yamauchi T. Adiponectin and Adipohectin receptors. Endocrine reviews.2005,26(3):439-451.
52. Liu YH, Tsai EM, Wu LC, et al. Higher basal adiponectin levels are associated with better ovarian response to gonadotropin stimulation during in vitro fertilization. GynecolObstet Invest.2005,60(3):167-170.
53. Gutman G, Barak V, Maslovitz S, et al. Recombinant luteinizing hormone induces increased production of ovarian follicular adiponectin in vivo:implications for enhanced insulin sensitivity. FertilSteril.2008,91(5):1837-1841.
54. Pierre P, Froment P, Negre D, et al. Role of adiponectin receptors, AdipoRl and AdipoR2, in the steroidogenesis of the human granulosa tumor cell line, KGN. Hum Reprod.2009,24(11):2890-2901.
1. Scherer PE, Williams S, Fogliano M, et al. A novel serum protein similar to Clq, produced exclusively in adipocytes. J Biol Chem.1995,270(45):26746-26749.
2. Nakano Y, Tobe T, Choi-Miura NH, et al. Isolation and characterization of GBP28, a novel gelatin-binding protein purified from human plasma. J Biochem. 1996,120(4):803-812.
3. Hu E, Liang P, Spiegelman BM. AdipoQ is a novel adipose-specific gene dysregulated in obesity. J Biol Chem.1996,271(18):10697-10703.
4. Maeda K, Okubo K, Shimomura I, et al. cDNA cloning and expression of a novel adipose specific collagen-like factor, apMI (adipose most abundant gene transcript 1). Biochem Biophys Res Commun.1996,221(2):286-289.
5. Ouchi N, Kihara S, Arita Y, et al. Novel modulator for endothelial adhesion molecules:adipocyte-derived plasma protein adiponectin. Circulation.1999, 100(25):2473-2476.
6. Shapiro L, Scherer PE. The crystal structure of a complement-1q family protein suggests an evolutionary link to tumor necrosis factor. Curr Biol.1998,8(6): 335-338.
7. Arita Y, Kihara S, Ouchi N, et al. Paradoxical decrease of an adipose-specific protein, adiponectin, in obesity. Biochem Biophys Res Commun.1999,257(1): 79-83.
8. Escobar-Morreale HF, Peral B, Villuendas G, et al. Common single nucleotide polymorphisms in intron 3 of the calpain-10 gene influence hirsutism. Fertil Steril.2002,77(3):581-587.
9. Gonzalez A, Abril E, Roca A, et al. Comment:CAPN10 alleles are associated with polycystic ovary syndrome. J Clin Endocrinol Metab.2002,87(8): 3971-3976.
10. Haddad L, Evans JC, Gharani N, et al. Variation within the type 2 diabetes susceptibility gene calpain-10 and polycystic ovary syndrome. J Clin Endocrinol Metab.2002,87(6):2606-2610.
11. Milner CR, Craig JE, Hussey ND, et al. No association between the -308 polymorphism in the tumour necrosis factor alpha (TNF alpha) promoter region and polycystic ovaries. Mol Hum Reprod.1999,5(1):5-9.
12. Yamauchi T, Kamon J, Ito Y, et al. Cloning of adiponectin receptors that mediate antidiabetic metabolic effects. Nature.2003,423(6941):762-769.
13. Yamauchi T, Nio Y, Maki T, et al. Targeted disruption of AdipoRl and AdipoR2 causes abrogation of adiponectin binding and metabolic actions. Nature.2007, 13(3):332-339.
14. Hug C, Wang J, Ahmad NS, et al. T-cadherin is a receptor hexameric and high-molecular-weight forms of Acrp30/adiponectin. Natl Acad Sci USA,2004. 101(28):10308-10313.
15. Angst BD, Marcozzi C, Magee Al. The cadherin superfamily:diversity in form and function. J Cell Sci.2001,114(pt 4):629-641.
16. Ivanov D, Philippova M, Tkachuk V, et al. Cell adhesion molecule T-cadherin regulates vascular cell adhesion, phenotype and motility. Exp Cell Res.2004, 293(2):207-218.
17. Kubota N, Terauchi Y, Yamauchi T, et al. Disruption of adiponectin causes insulin resistance and neointimal formation. J Biol Chem.2002,277(29):25863-25866.
18. Yamauchi T, Kamon J, Waki H, et al. Globular adiponectin protected ob/ob mice from diabetes and ApoE-deficient mice from atherosclerosis. J Biol Chem.2003, 24; 278(4):2461-2468.
19. Brakenhielm E, Veitonmaki N, Cao R, et al. Adiponectin-induced antiangiogenesis and antitumor activity involve caspase-mediated endothelial cell apoptosis. Proc Natl Acad Sci USA.2004,101(8):2476-2481.
20. Schulze MB, Shai I, Rimm EB, et al. Adiponectin and future coronary heart disease events among men with type 2 diabetes. Diabetes.2005,54(2):534-539.
21. Iglseder B, Mackevics V, Stadlmayer A, et al. Plasma adiponectin levels and sonographic phenotypes of subclinical carotid artery atherosclerosis:data from the SAPHIR Study. Stroke.2005,36(12):2577-2582.
22. Tsao TS, Tomas E, Murrey HE, et al. Role of disulfide bonds in Acrp30/adiponectin structure and signaling specificity. Different oligomers activate different signal transduction pathways. J Biol Chem.2003,278(50): 50810-50817.
23. Tomas E, Tsao TS, Saha AK, et al. Enhanced muscle fat oxidation and glucose transport by ACRP30 globular domain:acetyl-CoA carboxylase inhibition and AMP-activated protein kinase activation. Proc Natl Acad Sci USA.2002,10(99): 16309-16313.
24. AMPK. Carling D. Curr Biol.2004,23; 14(6):R220.
25. Yamauchi T, Kamon J, Minokoshi Y, et al. Adiponectin stimulates glucose utilization and fatty-acid oxidation by activating AMP-activated protein kinase. Nat Med.2002,8(11):1288-1295.
26. Schoonjans K, Staels B, Auwerx J. The peroxisome proliferator activated receptors (PPARS) and their effects on lipid metabolism and adipocyte differentiation. Biochim Biophys Acta.1996,26; 1302(2):93-109.
27. Barger PM, Browning AC, Garner AN, et al. J Biol Chem.2001,30; 276(48): 44495-44501.
28. Mao X, Kikani CK, Riojas RA, et al. APPL1 binds to adiponectin receptors and mediates adiponectin signalling and function. Nat Cell Biol.2005,8(5):516-523.
29. Hosch SE, Olefsky JM, Kim JJ. APPLied mechanics:uncovering how adiponectin modulates insulin action. Cell Metab.2006,4(1):5-6.
30. Xin X, Zhou L, reyes CM, et al. APPL1 mediates adiponectin-stimulated p38 MAPK activation by scaffolding the TAK1-MKK3-p38MAPK pathway. Am J Physiol Endocrinol Metab.2011,300(1):E103-E110.
31. Nechamen CA, Thomas RM, Dias JA. APPL1, APPL2, AKt2 and FOXO1a interact with FSHR in a potential signaling complex. Mol Cell Endocrinol.2007, 260-262:93-99.
32. Awazawa M, Ueki K, Inabe K, et al. Adiponectin enhances insulin sensitivity by increasing hepatic IRS-2 expression via a macrophage-derived IL-6-dependent pathway. Cell Metabolism,2011,13(4):401-412.
33. Weyer C, Funahashi T, Tanaka S, et al. Hypoadiponectinemia in obesity and type 2 diabetes:close association with insulin resistance and hyperinsulinemia. J Clin Endocrinol Metab.2001,86(5):1930-1935.
34. Yamamoto Y, Hirose H, Saito I, et al. Correlation of the adipocyte-derived protein adiponectin with insulin resistance index and serum high-density lipoprotein-cholesterol, independent of body mass index, in the Japanese population. Clin Sci (Lond).2002,103(2):137-142.
35. Lindsay RS, Funahashi T, Hanson RL, et al. Adiponectin and development of type 2 diabetes in the Pima Indian population. Lancet.2002,360(9326):57-58.
36. Snehalatha C, Mukesh B, Simon M, et al. Plasma adiponectin is an independent predictor of type 2 diabetes in Asian indians. Diabetes Care.2003,26(12): 3226-3229.
37. Pellme F, Smith U, Funahashi T, et al. Circulating adiponectin levels are reduced in nonobese but insulin-resistant first-degree relatives of type 2 diabetic patients. Diabetes.2003,52(5):1182-1186.
38. Yamamoto Y, Hirose H, Saito I, et al. Adiponectin, an adipocyte-derived protein, predicts future insulin resistance:two-year follow-up study in Japanese population. J Clin Endocrinol Metab.2004,89(1):87-90.
39. Hojlund K, Frystyk J, Levin K, et al. Reduced plasma adiponectin concentrations may contribute to impaired insulin activation of glycogen synthase in skeletal muscle of patients with type 2 diabetes. Diabetologia.2006,49(6):1283-1291.
40.张宁,陈子江,石玉华,等.血清脂联素水平测定在多囊卵巢综合征中的意义.实用妇产科杂志.2007,23(3)：154-156.
41. Ducluzeau, PH, Cousin, P, Malvoisin, E, et al. Glucose-to-insulin ratio rather than sex hormone-binding globulin and adiponectin levels is the best predictor of insulin resistance in nonobese women with polycystic ovary syndrome. Journal of Clinical Endocrinology & Metabolism.2003,88(8):3626-3631.
42. Spranger J, Mohlig M, Wegewitz U, et al. Adiponectin is independently associated with insulin sensitivity in women with polycystic ovary syndrome. Clinical Endocrinology.2004,61(6):738-746.
43. Aroda V, Ciaraldi, TP, Chang, SA, et al. Circulating and cellular adiponectin in polycystic ovary syndrome:Relationship to glucose tolerance and insulin action. Fertility & Sterility.2008,89(5):1200-1208.
44. Jensterle M, Weber M, Pfeifer M, et al. Assessment of insulin resistance in young women with polycystic ovary syndrome. International Journal of Gynecology and Obstetrics.2008,102(2):137-140.
45. Ehrmann DA. Medical progress:Polycystic ovary syndrome. New England Journal of Medicine.2005,24; 352(12):1223-1236.
46. Escobar-Morreale HF, Villuendas G, Botella-Carretero JI, et al. Adiponectin and resistin in PCOS:A clinical, biochemical and molecular genetic study. Human Reproduction.2006,21(9):2257-2265.
47. Glintborg D, Andersen M, Hagen C, et al. Evaluation of metabolic risk markers in polycystic ovary syndrome (PCOS). Adiponectin, ghrelin, leptin and body composition in hirsute PCOS patients and controls. European Journal of Endocrinology.2006,155(2):337-345.
48. Ardawi MS, Rouzi AA, Ardawi M SM,& Rouzi AA. Plasma adiponectin and insulin resistance in women with polycystic ovary syndrome. Fertility & Sterility. 2005,83(6):1708-1716.
49. Carmina E, Orio F, Palomba S, et al. Evidence for altered adipocyte function in polycystic ovary syndrome. European Journal of Endocrinology.2005,152(2): 389-394.
50. Orio F, Palomba S, Cascella T, et al. Adiponectin levels in women with polycystic ovary syndrome. Journal of Clinical Endocrinology & Metabolism.2003,88(6): 2619-2623.
51. Panidis D, Kourtis A, Farmakiotis D, et al. Serum adiponectin levels in women with polycystic ovary syndrome. Human Reproduction.2003,18(9):1790-1796.
52. Toulis KA, Goulis DG, Farmakiotis D, et al. Adiponectin levels in women with polycystic ovary syndrome:a systematic review and a meta-analysis. Hum Reprod Update.2009,15(3):297-307.
53. Xu A, Chan KW, Hoo RL, et al. Testosterone selectively reduces the high molecular weight form of adiponectin by inhibiting its secretion from adipocytes. J Biol Chem.2005,280(18):18073-18080.
54. Bik W, Baranowska-Bik A, Wolinska-Witort E, et al. The relationship between metabolic status and levels of adiponectin and ghrelin in lean women with polycystic ovary syndrome. Gynecol Endocrinol.2007,23(6):325-331.
55. Carmina E, Bucchieri S, Mansueto P, et al. Circulating levels of adipose products and differences in fat distribution in the ovulatory and anovulatory phenotypes of polycystic ovary syndrome. Fertil Steril.2009,91(4 Suppl):1332-1335.
56. Stumvoll M, Tschritter O, Fritsche A, et al. Association of the T-G polymorphism in adiponectin (exon 2) with obesity and insulin sensitivity:interaction with family history of type 2 diabetes. Diabetes.2002,51(1):37-41.
57. Hara K, Boutin P, Mori Y, et al. Genetic variation in the gene encoding adiponectin is associated with an increased risk of type 2 diabetes in the Japanese population. Diabetes.2002,51(2):536-540.
58. Fumeron F, Aubert R, Siddiq A, et al. Adiponectin gene polymorphisms and adiponectin levels are independently associated with the development of hyperglycemia during a 3-year period:the epidemiologic data on the insulin resistance syndrome prospective study. Diabetes.2004,53(4):1150-1157.
59. Zacharova J, Chiasson JL, Laakso M, et al. The common polymorphisms (single nucleotide polymorphism [SNP]+45 and SNP +276) of the adiponectin gene predict the conversion from impaired glucose tolerance to type 2 diabetes:the STOP-NIDDM trial. Diabetes.2005,54(3):893-899.
60. Jang Y, Lee JH, Kim OY, et al. The SNP276G>T polymorphism in the adiponectin (ACDC) gene is more strongly associated with insulin resistance and cardiovascular disease risk than SNP45T>G in nonobese/nondiabetic Korean men independent of abdominal adiposity and circulating plasma adiponectin. Metabolism.2006,55(1):59-66.
61. Qj L, Doria A, Manson JE, et al. Adiponectin genetic variability, plasma adiponectin, and cardiovascular risk in patients with type 2 diabetes. Diabetes. 2006,55(5):1512-1516.
62. Panidis D, Kourtis A, Kukuvitis A, et al. Association of the T45G polymorphism in exon 2 of the adiponectin gene with polycystic ovary syndrome:role of Delta4-androstenedione. Hum Reprod.2004,19(8):1728-1733.
63. Xita N, Georgiou I, Chatzikyriakidou A, et al. Effect of adiponectin gene polymorphisms on circulating adiponectin and insulin resistance indexes in women with polycystic ovary syndrome. Clin Chem.2005,51(2):416-423.
64. Haap M, Machicao F, Stefan N, et al. Genetic determinants of insulin action in polycystic ovary syndrome. Exp Clin Endocrinol Diabetes.2005,113(5): 275-281.
65. Heinonen S, Korhonen S, Helisalmi S, et al. Associations between two single nucleotide polymorphisms in the adiponectin gene and polycystic ovary syndrome. Gynecol Endocrinol.2005,21(3):165-169.
66. Ranjzad F, Mahmoudi T, Irani Shemirani A, et al. A common variant in the adiponectin gene and polycystic ovary syndrome risk. Mol Biol Rep,2012,39(3): 2313-2319.
67. Yoshihara K, Yahata T, Kashima K, et al. Association of single nucleotide polymorphisms in adiponectin and its receptor genes with polycystic ovary syndrome. J Reprod Med.2009,54(11/12):669-674.
68. Ning Zhang, Yu-Hua Shi, Cui-Fang Hao, et al. Association of +45G15G(T/G) and +276(G/T) polymorphisms in the ADIPOQ gene with polycystic ovary syndrome among Han Chinese women. European Journal of Endocrinology.2008,158(2): 255-260.
69. Li L, Yun JH, Lee JH, et al. Association study of +45G15G(T/G) and +276(G/T) polymorphisms in the adiponectin gene in patients with polycystic ovary syndrome. Int J Mol Med.2011,27(2):283-287.
70. Demirci H, Yilmaz M, Ergun MA, et al. Frequency of adiponectin gene polymorphisms in polycystic ovary syndrome and the association with serum adiponectin, androgen levels, insulin resistance and clinical parameters. Gynecol Endocrinol.2010,26(5):348-355.
71. Campos DB, Palin MF, Bordignon V, et al. The beneficial adipokines in reproduction and fertility. Int J Obes.2008,32(2):223-231.
72. Ledoux S, Campos DB, Lopes FL, et al. Adiponectin induces periovulatory changes in ovarian follicular cells. Endocrinology.2006,147(11):5178-5186.
73. Bersinger NA, Birkhauser MH, Wunder DM. Adiponectin as a marker of success in intracytoplasmic sperm injection/embryo transfer cycles. Gynecol Endocrinol. 2006,22(9):479-483.
74. Chabrolle C, Tosca L, Rame C, et al. Adiponectin increases insulin-like growth factor I induced progesterone and estradiol secretion in human granulose cells. Fertil Steril.2008,92(6):1988-1996.
75. Chabrolle C, Tosca L, Crochet S, et al. Expression of adiponectin and its receptors (AdipnRl and AdipoR2) in chicken ovary:potential role in ovarian steroidogenesis. Domest Anim Endocrinol.2007,33(4):480-487.
76. Chabrolle C, Tosca L, Dupont J. Regulation of adiponectin and its receptors in rat ovary by human chorionic gonadotrophin treatment and potential involvement of adiponectin in granulosa cell steroidogenesis. Reproduction.2007,133(4): 719-731.
77. Chappaz E, Albornoz MS, Campos D, et al. Adiponectin enhances in vitro development of swine embryos. Domest Anim Endocrinol.2008,35(2):198-207.
78. Tabandeh MR, Hosseini A, Sae b M, et al. Changes in the gene expression of adiponectin and adiponectin receptors (AdipoR1 and AdipoR2) in ovarian follicular cells of dairy cow at different stages of development. Theriogenology. 2010,73(5):659-669.
79. Maillard V, Uzbekova S, Guignot F, et al. Effect of adiponectin on bovine granulosa cell steroidogenesis, oocyte maturation and embryo development. Reproductive Biology and Endocrinology.2010,10; 8:23-37.
80. Lord E, Ledoux S, Murphy BD, et al. Expression of adiponectin and its receptors in swine. J Anim Sci.2005,83(3):565-578.
81. Lagaly DV, Aad PY, Grado-Ahuir JA, et al. Role of adiponectin in regulating ovarian theca and granulosa cell function. Molecular and Cellular Endocrinology. 2008,12; 284(1-2):38-45.
82. Chabrolle C, Jeanpierre E, Tosca L, et al. Effects of high levels of glucose on the steroidogenesis and the expression of adiponectin receptors in rat ovarian cells. Reproductive Biology and Endocrinology.2008,19; 6:11.
83. Chabrolle C, Tosca L, Rame C, et al. Adiponectin increase insulin-like growth factor 1 induced progesterone and estradiol secretion in human granulose cells. Fertil Steril.2009,92(6):1988-1996.
84. Pierre P, Froment P, Negre D, et al. Role of adiponectin receptors, AdipoRl and AdipoR2, in the steroidogenesis of the human granulosa tumor cell line, KGN. Hum Reprod.2009,24(11):2890-2901.
85. Kim ST, Marquard K, Stephens S, et al. Adiponectin and adiponectin receptors in the mouse preimplantation embryo and uterus. Human Reproduction.2011,26(1): 82-95.
86. Liu YH, Tsai EM, Chen YL, et al. Serum adiponectin levels increase after human chorionic gonadotropin treatment during in vitro fertilization. Gynecol Obstet Invest.2006,62(2):61-65.
87. Liu YH, Tsai EM, Wu LC, et al. Higher basal adiponectin levels are associated with better ovarian response to gonadotropin stimulation during in vitro fertilization. Gynecol Obstet Invest.2005,60(3):167-170.
88. Bersinger NA, Birkhauser MH, Wunder DM. Adiponection as a marker of success in intracytoplasmic sperm injection/embryo transfer cycles. Gynecol Endocrinol.2006,22(9):479-483.
89. Gutman G, Barak V, Maslovitz S, et al. Recombinant luteinizing hormone induces increased production of ovarian follicular adiponectin in vivo:implications for enhanced insulin sensitivity. Fertil Steril.2008,91(5):1837-1841.
90. Richards JS, Liu Z, Kawai T, et al. Adiponectin and its receptors modulate granulosa cell and cumulus cell functions, fertility, and early embryo development in the mouse and human. Fertil Steril.2012,98(2):471-479.
2. Legro RS. Polycystic ovary syndrome:the new millenium. Mol Cell Endocrinol. 2001,26,184:87-93.
3. Ehrmann DA. Polycystic ovary syndrome. N Engl J Med.2005,24,352: 1223-1236.
4. Heijnen E, Eijkemans M, Hughes E, et al. A meta-analysis of outcomes of conventional IVF in women with polycystic ovary syndrome. Hum Reprod Update.2006,12:13-21.
5. Chang MY, Chiang CH, Hsieh TT, et al. Use of the antral follicle count to predict the outcome of assisted reproductive technologies. Fertil Steril.1998, 69:505-510.
6. de Carvalho BR, Rosa e Silva AC, Rosa e Silva JC, et al. Ovarian reserve evaluation:state of the art. J Assist Reprod Genet.2008,25:311-322.
7. Kline J, Kinney A, Kelly A, et al. Predictors of antral follicle count during the reproductive years. Hum Reprod.2005,20:2179-2189.
8. Scheffer GJ, Broekmans FJ, Dorland M, et al. Antral follicle counts by transvaginal ultrasonography are related to age in women with proven natural fertility. Fertil Steril.1999,72:845-851.
9. Bancsi LF, Broekmans FJ, Eijkemans MJ, et al. Predictors of poor ovarian response in in vitro fertilization:a prospective study comparing basal markers of ovarian reserve. Fertil Steril.2002,77:328-336.
10. Broer SL, Mol BW, Hendriks D, et al. The role of antimullerian hormone in prediction of outcome after IVF:comparison with the antral follicle count. Fertil Steril.2009,91:705-714.
11. Melo MA, Garrido N, Alvarez C, et al. Antral follicle count (AFC) can be used in the prediction of ovarian response but cannot predict the oocyte/embryo quality or the in vitro fertilization outcome in an egg donation program. Fertil Steril. 2009,91:148-156.
12. Genro VK, Grynberg M, Scheffer JB, et al. Serum anti-Mullerian hormone levels are negatively related to Follicular Output RaTe (FORT) in normo-cycling women undergoing controlled ovarian hyperstimulation. Hum Reprod.2011, 26:671-677.
13. Gallot V, Berwanger da Silva AL, Genro V, et al. Antral follicle responsiveness to follicle-stimulating hormone administration assessed by the Follicular Output RaTe (FORT) may predict in vitro fertilization-embryo transfer outcome. Hum Reprod.2012,27:1066-1072.
14. Genro VK, Matte U, De Conto E, et al. Frequent polymorphisms of FSH receptor do not influence antral follicle responsiveness to follicle-stimulating hormone administration as assessed by the Follicular Output RaTe (FORT). J Assist Reprod Genet.2012,29:657-663.
15. The Rotterdam ESHRE/ASRM-sponsored PCOS consensus workshop group. Revised 2003 consensus on diagnostic criteria and long-term health risks related to polycystic ovary syndrome (PCOS). Human Reproduction.2004,19:41-47.
16. Jurema MW, Bracero NJ, Garcia JE. Fine tuning cycle day 3 hormonal assessment of ovarian reserve improves in vitro fertilization outcome in gonadotropin-releasing hormone antagonist cycles. Fertil Steril. 2003;80:1156-1161.
17. Bancsi LF, Broekmans FJ, Mol BW, et al. Performance of basal follicle-stimulating hormone in the prediction of poor ovarian response and failure to become pregnant after in vitro fertilization:a meta-analysis. Fertil Steril. 2003,79:1091-1100.
18. Bancsi LF, Broekmans FJ, Looman CW, et al. Predicting poor ovarian response in IVF:use of repeat basal FSH measurement. J Reprod Med.2004,49:187-194.
19. Barad DH, Weghofer A, Gleicher N. Comparing anti-Mullerian hormone (AMH) and follicle-stimulating hormone (FSH) as predictors of ovarian function. Fertil Steril.2009,91:1553-1555.
20. Lekamge DN, Barry M, Kolo M, et al. Anti-Mullerian hormone as a predictor of IVF outcome. Reprod Biomed Online.2007,14:602-610.
21. Wunder DM, Guibourdenche J, Birkhauser MH, et al. Anti-Mullerian hormone and inhibin B as predictors of pregnancy after treatment by in vitro fertilization/intracytoplasmic sperm injection. Fertil Steril.2008,90:2203-2210.
22. Lee TH, Liu CH, Huang CC, et al. Serum anti-Mullerian hormone and estradiol levels as predictors of ovarian hyperstimulation syndrome in assisted reproduction technology cycles. Hum Reprod.2008,23:160-167.
23. Smeenk JM, Sweep FC, Zielhuis GA, et al. Antimullerian hormone predicts ovarian responsiveness, but not embryo quality or pregnancy, after in vitro fertilization or intracyoplasmic sperm injection. Fertil Steril.2007,87:223-226.
24. Broekmans FJ, Visser JA, Laven JS, et al. Anti-Mullerian hormone and ovarian dysfunction. Trends Endocrinol Metab.2008,19:340-347.
25. La Marca A, Broekmans FJ, Volpe A, et al. Anti-Mullerian hormone (AMH): what do we still need to know? Hum Reprod.2009,24:2264-2275.
26. Kaya C, Pabuccu R, Satiroglu H. Serum antimullerian hormone concentrations on day 3 of the in vitro fertilization stimulation cycle are predictive of the fertilization, implantation, and pregnancy in polycystic ovary syndrome patients undergoing assisted reproduction. Fertil Steril.2010,94:2202-2207.
27. Coffler MS, Patel K, Dahan MH, et al. Evidence for abnormal granulosa cell responsiveness to follicle-stimulating hormone in women with polycystic ovary syndrome. J Clin Endocrinol Metab.2003,88:1742-1747.
28. Franks S, Roberts R, Hardy K. Gonadotrophin regimens and oocyte quality in women with polycystic ovaries. Reprod Biomed Online.2003,6:181-184.
29. Fabregues F, Penarrubia J, Vidal E, et al. Oocyte quality in patients with severe ovarian hyperstimulation syndrome:a self-controlled clinical study. Fertil Steril. 2004,82:827-833.
30. Aboulghar MA, Mansour RT, Serour GI, et al. Oocyte quality in patients with severe ovarian hyperstimulation syndrome. Fertil Steril.1997,68:1017-1021.
1. Scherer PE, Williams S, Fogliano M, et al. A novel serum protein similar to C1q, produced exclusively in adipocytes. J Biol Chem.1995,270(45):26746-26749.
2. Nakano Y, Tobe T, Choi-Miura NH, et al. Isolation and characterization of GBP28, a novel gelatin-binding protein purified from human plasma. J Biochem. 1996,120(4):803-812.
3. Hu E, Liang P, Spiegelman BM. AdipoQ is a novel adipose-specific gene dysregulated in obesity. J Biol Chem.1996,271(18):10697-10703.
4. Maeda K, Okubo K, Shimomura I, et al. cDNA cloning and expression of a novel adipose specific collagen-like factor, apMI (adipose most abundant gene transcript 1). BiochemBiophys Res Commun.1996,221(2):286-289.
5. Ouchi N, Kihara S, Arita Y, et al. Novel modulator for endothelial adhesion molecules:adipocyte-derived plasma protein adiponectin. Circulation.1999, 100(25):2473-2476.
6. Shapiro L, Scherer PE. The crystal structure of a complement-lq family protein suggests an evolutionary link to tumor necrosis factor. Curr Biol.1998,8(6): 335-338.
7. Arita Y, Kihara S, Ouchi N, et al. Paradoxical decrease of an adipose-specific protein, adiponectin, in obesity. BiochemBiophys Res Commun.1999,257(1): 79-83.
8. Yamauchi T, Kamon J, Ito Y, et al. Cloning of adiponectin receptors that mediate antidiabetic metabolic effects. Nature.2003,423(6941):762-769.
9. Hug C, Wang J, Ahmad NS, et al. T-cadherin is a receptor hexameric and high-molecular-weight forms of Acrp30/adiponectin. NatlAcadSci USA,2004. 101(28):10308-10313.
10. Ivanov D, Philippova M, Tkachuk V, et al. Cell adhesion molecule T-cadherin regulates vascular cell adhesion, phenotype and motility. Exp Cell Res.2004, 293(2):207-218.
11. Kubota N, Terauchi Y, Yamauchi T, et al. Disruption of adiponectin causes insulin resistance and neointimal formation. J Biol Chem.2002,277(29):25863-25866.
12. Yamauchi T, Kamon J, Waki H, et al. Globular adiponectin protected ob/ob mice from diabetes and ApoE-deficient mice from atherosclerosis. J Biol Chem.2003, 24; 278(4):2461-2468.
13. Brakenhielm E, Veitonmaki N, Cao R, et al. Adiponectin-induced antiangiogenesis and antitumor activity involve caspase-mediated endothelial cell apoptosis. ProcNatlAcadSci USA.2004,101(8):2476-2481.
14. Schulze MB, Shai I, Rimm EB, et al. Adiponectin and future coronary heart disease events among men with type 2 diabetes. Diabetes.2005,54(2):534-539.
15. Iglseder B, Mackevics V, Stadlmayer A, et al. Plasma adiponectin levels and sonographic phenotypes of subclinical carotid artery atherosclerosis:data from the SAPHIR Study. Stroke.2005,36(12):2577-2582.
16. Yamamoto Y, Hirose H, Saito I, et al. Correlation of the adipocyte-derived protein adiponectin with insulin resistance index and serum high-density lipoprotein-cholesterol, independent of body mass index, in the Japanese population. ClinSci (Lond).2002,103(2):137-142.
17. Lindsay RS, Funahashi T, Hanson RL, et al. Adiponectin and development of type 2 diabetes in the Pima Indian population. Lancet.2002,360(9326):57-58.
18. Snehalatha C, Mukesh B, Simon M, et al. Plasma adiponectin is an independent predictor of type 2 diabetes in Asian indians. Diabetes Care.2003,26(12): 3226-3229.
19. Pellme F, Smith U, Funahashi T, et al. Circulating adiponectin levels are reduced in nonobese but insulin-resistant first-degree relatives of type 2 diabetic patients. Diabetes.2003,52(5):1182-1186.
20. Yamamoto Y, Hirose H, Saito I, et al. Adiponectin, an adipocyte-derived protein, predicts future insulin resistance:two-year follow-up study in Japanese population. J ClinEndocrinolMetab.2004,89(1):87-90.
21. Hojlund K, Frystyk J, Levin K, et al. Reduced plasma adiponectin concentrations may contribute to impaired insulin activation of glycogen synthase in skeletal muscle of patients with type 2 diabetes. Diabetologia.2006,49(6):1283-1291.
22.张宁,陈子江,石玉华,等.血清脂联素水平测定在多囊卵巢综合征中的意义.实用妇产科杂志.2007,23(3)：154-156.
23. Ducluzeau, PH, Cousin, P, Malvoisin, E, et al. Glucose-to-insulin ratio rather than sex hormone-binding globulin and adiponectin levels is the best predictor of insulin resistance in nonobese women with polycystic ovary syndrome. Journal of Clinical Endocrinology & Metabolism.2003,88(8):3626-3631.
24. Spranger J, Mohlig M, Wegewitz U, et al. Adiponectin is independently associated with insulin sensitivity in women with polycystic ovary syndrome. Clinical Endocrinology.2004,61(6):738-746.
25. Aroda V, Ciaraldi, TP, Chang, SA, et al. Circulating and cellular adiponectin in polycystic ovary syndrome:Relationship to glucose tolerance and insulin action. Fertility & Sterility.2008,89(5):1200-1208.
26. Jensterle M, Weber M, Pfeifer M, et al. Assessment of insulin resistance in young women with polycystic ovary syndrome. International Journal of Gynecology and Obstetrics.2008,102(2):137-140.
27. Ehrmann DA. Medical progress:Polycystic ovary syndrome. New England Journal of Medicine.2005,24; 352(12):1223-1236.
28. Escobar-Morreale HF, Villuendas G, Botella-Carretero JI, et al. Adiponectin and resistin in PCOS:A clinical, biochemical and molecular genetic study. Human Reproduction.2006,21(9):2257-2265.
29. Glintborg D, Andersen M, Hagen C, et al. Evaluation of metabolic risk markers in polycystic ovary syndrome (PCOS). Adiponectin, ghrelin, leptin and body composition in hirsute PCOS patients and controls. European Journal of Endocrinology.2006,155(2):337-345.
30. Ardawi MS, Rouzi AA, Ardawi M SM,&Rouzi AA. Plasma adiponectin and insulin resistance in women with polycystic ovary syndrome. Fertility & Sterility. 2005,83(6):1708-1716.
31. Toulis KA, Goulis DG, Farmakiotis D, et al. Adiponectin levels in women with polycystic ovary syndrome:a systematic review and a meta-analysis. Hum Reprod Update.2009,15(3):297-307.
32. Campos DB, Palin MF, Bordignon V, et al. The beneficial adipokines in reproduction and fertility. Int J Obes.2008,32(2):223-231.
33. Ledoux S, Campos DB, Lopes FL, et al. Adiponectin induces periovulatory changes in ovarian follicular cells. Endocrinology.2006,147(11):5178-5186.
34. Bersinger NA, Birkhauser MH, Wunder DM. Adiponectin as a marker of success in intracytoplasmic sperm injection/embryo transfer cycles. GynecolEndocrinol. 2006,22(9):479-483.
35. Chabrolle C, Tosca L, Rame C, et al. Adiponectin increases insulin-like growth factor I induced progesterone and estradiol secretion in human granulose cells. FertilSteril.2008,92(6):1988-1996.
36. Chabrolle C, Tosca L, Crochet S, et al. Expression of adiponectin and its receptors (AdipnRl and AdipoR2) in chicken ovary:potential role in ovarian steroidogenesis. DomestAnimEndocrinol.2007,33(4):480-487.
37. Chabrolle C, Tosca L, Dupont J. Regulation of adiponectin and its receptors in rat ovary by human chorionic gonadotrophin treatment and potential involvement of adiponectin in granulosa cell steroidogenesis. Reproduction.2007,133(4): 719-731.
38. Chappaz E, Albornoz MS, Campos D, et al. Adiponectin enhances in vitro development of swine embryos. DomestAnimEndocrinol.2008,35(2):198-207.
39. Tabandeh MR, Hosseini A, Sae b M, et al. Changes in the gene expression of adiponectin and adiponectin receptors (AdipoRl and AdipoR2) in ovarian follicular cells of dairy cow at different stages of development. Theriogenology. 2010,73(5):659-669.
40. Maillard V, Uzbekova S, Guignot F, et al. Effect of adiponectin on bovine granulosa cell steroidogenesis, oocyte maturation and embryo development. Reproductive Biology and Endocrinology.2010,10; 8:23-37.
41. Richards JS, Liu Z, Kawai T, et al.Adiponectin and its receptors modulate granulosa cell and cumulus cell functions, fertility, and early embryo development in the mouse and human. FertilSteril.2012,98(2):471-479.
42. Kim ST, Marquard K, Stephens S, et al. Adiponectin and adiponectin receptors in the mouse preimplantation embryo and uterus. Human Reproduction.2011,26(1): 82-95.
43. The Rotterdam ESHRE/ASRM-sponsored PCOS consensus workshop group. Revised 2003 consensus on diagnostic criteria and long-term health risks related to polycystic ovary syndrome (PCOS). Human Reproduction.2004,19:41-47.
44. Psilopanagioti A, Papadaki H, Kranioti EF, et al. Expression of adiponectin and adiponectin receptors in human pituitary gland and brain. Neuroendocrinology. 2009,89(1):38-47.
45. Wen JP, Lv WS, Yang J, et al. Globular adiponectin inhibits GnRH secretion from GT1-7 hypothalamic GnRH neurons by induction of hyperpolarization of membrane potential. Biochem Biophys Res Commun.2008,11,371(4):756-761.
46. Wilkinson M, Brown R, Imran SA, et al. Adipokine gene expression in brain and pituitary gland. Neuroendocrinology.2007,86(3):191-209.
47. Lu M, Tang Q, Olefsky JM, et al. Adiponectin activates adenosine monophosphate-activated protein kinase and decreases luteinizing hormone secretion in LbetaT2 gonadotropes. Mol Endocrinol.2008,22(3):760-771.
48. Rodrigues-Pacheco F, Martinez-Fuentes AJ, Tovar S, Pinilla L, et al. Regulation of pituitary cell function by adiponectin. Endocrinology.2007,148(1):401-410.
49. Mitchell M, Armstrong DT, Robker RL, et al. Adipokines:implications for female fertility and obesity. Reproduction.2000,130(5):583-597.
50. Lagaly DV, Aad PY, Grado-Ahuir JA, et al. Role of adiponectin in regulating ovarian theca and granulosa cell function. Molecular and Cellular Endocrinology. 2008,12,284(1-2):38-45.
51. Kadowaki T, Yamauchi T. Adiponectin and Adipohectin receptors. Endocrine reviews.2005,26(3):439-451.
52. Liu YH, Tsai EM, Wu LC, et al. Higher basal adiponectin levels are associated with better ovarian response to gonadotropin stimulation during in vitro fertilization. GynecolObstet Invest.2005,60(3):167-170.
53. Gutman G, Barak V, Maslovitz S, et al. Recombinant luteinizing hormone induces increased production of ovarian follicular adiponectin in vivo:implications for enhanced insulin sensitivity. FertilSteril.2008,91(5):1837-1841.
54. Pierre P, Froment P, Negre D, et al. Role of adiponectin receptors, AdipoRl and AdipoR2, in the steroidogenesis of the human granulosa tumor cell line, KGN. Hum Reprod.2009,24(11):2890-2901.
1. Scherer PE, Williams S, Fogliano M, et al. A novel serum protein similar to Clq, produced exclusively in adipocytes. J Biol Chem.1995,270(45):26746-26749.
2. Nakano Y, Tobe T, Choi-Miura NH, et al. Isolation and characterization of GBP28, a novel gelatin-binding protein purified from human plasma. J Biochem. 1996,120(4):803-812.
3. Hu E, Liang P, Spiegelman BM. AdipoQ is a novel adipose-specific gene dysregulated in obesity. J Biol Chem.1996,271(18):10697-10703.
4. Maeda K, Okubo K, Shimomura I, et al. cDNA cloning and expression of a novel adipose specific collagen-like factor, apMI (adipose most abundant gene transcript 1). Biochem Biophys Res Commun.1996,221(2):286-289.
5. Ouchi N, Kihara S, Arita Y, et al. Novel modulator for endothelial adhesion molecules:adipocyte-derived plasma protein adiponectin. Circulation.1999, 100(25):2473-2476.
6. Shapiro L, Scherer PE. The crystal structure of a complement-1q family protein suggests an evolutionary link to tumor necrosis factor. Curr Biol.1998,8(6): 335-338.
7. Arita Y, Kihara S, Ouchi N, et al. Paradoxical decrease of an adipose-specific protein, adiponectin, in obesity. Biochem Biophys Res Commun.1999,257(1): 79-83.
8. Escobar-Morreale HF, Peral B, Villuendas G, et al. Common single nucleotide polymorphisms in intron 3 of the calpain-10 gene influence hirsutism. Fertil Steril.2002,77(3):581-587.
9. Gonzalez A, Abril E, Roca A, et al. Comment:CAPN10 alleles are associated with polycystic ovary syndrome. J Clin Endocrinol Metab.2002,87(8): 3971-3976.
10. Haddad L, Evans JC, Gharani N, et al. Variation within the type 2 diabetes susceptibility gene calpain-10 and polycystic ovary syndrome. J Clin Endocrinol Metab.2002,87(6):2606-2610.
11. Milner CR, Craig JE, Hussey ND, et al. No association between the -308 polymorphism in the tumour necrosis factor alpha (TNF alpha) promoter region and polycystic ovaries. Mol Hum Reprod.1999,5(1):5-9.
12. Yamauchi T, Kamon J, Ito Y, et al. Cloning of adiponectin receptors that mediate antidiabetic metabolic effects. Nature.2003,423(6941):762-769.
13. Yamauchi T, Nio Y, Maki T, et al. Targeted disruption of AdipoRl and AdipoR2 causes abrogation of adiponectin binding and metabolic actions. Nature.2007, 13(3):332-339.
14. Hug C, Wang J, Ahmad NS, et al. T-cadherin is a receptor hexameric and high-molecular-weight forms of Acrp30/adiponectin. Natl Acad Sci USA,2004. 101(28):10308-10313.
15. Angst BD, Marcozzi C, Magee Al. The cadherin superfamily:diversity in form and function. J Cell Sci.2001,114(pt 4):629-641.
16. Ivanov D, Philippova M, Tkachuk V, et al. Cell adhesion molecule T-cadherin regulates vascular cell adhesion, phenotype and motility. Exp Cell Res.2004, 293(2):207-218.
17. Kubota N, Terauchi Y, Yamauchi T, et al. Disruption of adiponectin causes insulin resistance and neointimal formation. J Biol Chem.2002,277(29):25863-25866.
18. Yamauchi T, Kamon J, Waki H, et al. Globular adiponectin protected ob/ob mice from diabetes and ApoE-deficient mice from atherosclerosis. J Biol Chem.2003, 24; 278(4):2461-2468.
19. Brakenhielm E, Veitonmaki N, Cao R, et al. Adiponectin-induced antiangiogenesis and antitumor activity involve caspase-mediated endothelial cell apoptosis. Proc Natl Acad Sci USA.2004,101(8):2476-2481.
20. Schulze MB, Shai I, Rimm EB, et al. Adiponectin and future coronary heart disease events among men with type 2 diabetes. Diabetes.2005,54(2):534-539.
21. Iglseder B, Mackevics V, Stadlmayer A, et al. Plasma adiponectin levels and sonographic phenotypes of subclinical carotid artery atherosclerosis:data from the SAPHIR Study. Stroke.2005,36(12):2577-2582.
22. Tsao TS, Tomas E, Murrey HE, et al. Role of disulfide bonds in Acrp30/adiponectin structure and signaling specificity. Different oligomers activate different signal transduction pathways. J Biol Chem.2003,278(50): 50810-50817.
23. Tomas E, Tsao TS, Saha AK, et al. Enhanced muscle fat oxidation and glucose transport by ACRP30 globular domain:acetyl-CoA carboxylase inhibition and AMP-activated protein kinase activation. Proc Natl Acad Sci USA.2002,10(99): 16309-16313.
24. AMPK. Carling D. Curr Biol.2004,23; 14(6):R220.
25. Yamauchi T, Kamon J, Minokoshi Y, et al. Adiponectin stimulates glucose utilization and fatty-acid oxidation by activating AMP-activated protein kinase. Nat Med.2002,8(11):1288-1295.
26. Schoonjans K, Staels B, Auwerx J. The peroxisome proliferator activated receptors (PPARS) and their effects on lipid metabolism and adipocyte differentiation. Biochim Biophys Acta.1996,26; 1302(2):93-109.
27. Barger PM, Browning AC, Garner AN, et al. J Biol Chem.2001,30; 276(48): 44495-44501.
28. Mao X, Kikani CK, Riojas RA, et al. APPL1 binds to adiponectin receptors and mediates adiponectin signalling and function. Nat Cell Biol.2005,8(5):516-523.
29. Hosch SE, Olefsky JM, Kim JJ. APPLied mechanics:uncovering how adiponectin modulates insulin action. Cell Metab.2006,4(1):5-6.
30. Xin X, Zhou L, reyes CM, et al. APPL1 mediates adiponectin-stimulated p38 MAPK activation by scaffolding the TAK1-MKK3-p38MAPK pathway. Am J Physiol Endocrinol Metab.2011,300(1):E103-E110.
31. Nechamen CA, Thomas RM, Dias JA. APPL1, APPL2, AKt2 and FOXO1a interact with FSHR in a potential signaling complex. Mol Cell Endocrinol.2007, 260-262:93-99.
32. Awazawa M, Ueki K, Inabe K, et al. Adiponectin enhances insulin sensitivity by increasing hepatic IRS-2 expression via a macrophage-derived IL-6-dependent pathway. Cell Metabolism,2011,13(4):401-412.
33. Weyer C, Funahashi T, Tanaka S, et al. Hypoadiponectinemia in obesity and type 2 diabetes:close association with insulin resistance and hyperinsulinemia. J Clin Endocrinol Metab.2001,86(5):1930-1935.
34. Yamamoto Y, Hirose H, Saito I, et al. Correlation of the adipocyte-derived protein adiponectin with insulin resistance index and serum high-density lipoprotein-cholesterol, independent of body mass index, in the Japanese population. Clin Sci (Lond).2002,103(2):137-142.
35. Lindsay RS, Funahashi T, Hanson RL, et al. Adiponectin and development of type 2 diabetes in the Pima Indian population. Lancet.2002,360(9326):57-58.
36. Snehalatha C, Mukesh B, Simon M, et al. Plasma adiponectin is an independent predictor of type 2 diabetes in Asian indians. Diabetes Care.2003,26(12): 3226-3229.
37. Pellme F, Smith U, Funahashi T, et al. Circulating adiponectin levels are reduced in nonobese but insulin-resistant first-degree relatives of type 2 diabetic patients. Diabetes.2003,52(5):1182-1186.
38. Yamamoto Y, Hirose H, Saito I, et al. Adiponectin, an adipocyte-derived protein, predicts future insulin resistance:two-year follow-up study in Japanese population. J Clin Endocrinol Metab.2004,89(1):87-90.
39. Hojlund K, Frystyk J, Levin K, et al. Reduced plasma adiponectin concentrations may contribute to impaired insulin activation of glycogen synthase in skeletal muscle of patients with type 2 diabetes. Diabetologia.2006,49(6):1283-1291.
40.张宁,陈子江,石玉华,等.血清脂联素水平测定在多囊卵巢综合征中的意义.实用妇产科杂志.2007,23(3)：154-156.
41. Ducluzeau, PH, Cousin, P, Malvoisin, E, et al. Glucose-to-insulin ratio rather than sex hormone-binding globulin and adiponectin levels is the best predictor of insulin resistance in nonobese women with polycystic ovary syndrome. Journal of Clinical Endocrinology & Metabolism.2003,88(8):3626-3631.
42. Spranger J, Mohlig M, Wegewitz U, et al. Adiponectin is independently associated with insulin sensitivity in women with polycystic ovary syndrome. Clinical Endocrinology.2004,61(6):738-746.
43. Aroda V, Ciaraldi, TP, Chang, SA, et al. Circulating and cellular adiponectin in polycystic ovary syndrome:Relationship to glucose tolerance and insulin action. Fertility & Sterility.2008,89(5):1200-1208.
44. Jensterle M, Weber M, Pfeifer M, et al. Assessment of insulin resistance in young women with polycystic ovary syndrome. International Journal of Gynecology and Obstetrics.2008,102(2):137-140.
45. Ehrmann DA. Medical progress:Polycystic ovary syndrome. New England Journal of Medicine.2005,24; 352(12):1223-1236.
46. Escobar-Morreale HF, Villuendas G, Botella-Carretero JI, et al. Adiponectin and resistin in PCOS:A clinical, biochemical and molecular genetic study. Human Reproduction.2006,21(9):2257-2265.
47. Glintborg D, Andersen M, Hagen C, et al. Evaluation of metabolic risk markers in polycystic ovary syndrome (PCOS). Adiponectin, ghrelin, leptin and body composition in hirsute PCOS patients and controls. European Journal of Endocrinology.2006,155(2):337-345.
48. Ardawi MS, Rouzi AA, Ardawi M SM,& Rouzi AA. Plasma adiponectin and insulin resistance in women with polycystic ovary syndrome. Fertility & Sterility. 2005,83(6):1708-1716.
49. Carmina E, Orio F, Palomba S, et al. Evidence for altered adipocyte function in polycystic ovary syndrome. European Journal of Endocrinology.2005,152(2): 389-394.
50. Orio F, Palomba S, Cascella T, et al. Adiponectin levels in women with polycystic ovary syndrome. Journal of Clinical Endocrinology & Metabolism.2003,88(6): 2619-2623.
51. Panidis D, Kourtis A, Farmakiotis D, et al. Serum adiponectin levels in women with polycystic ovary syndrome. Human Reproduction.2003,18(9):1790-1796.
52. Toulis KA, Goulis DG, Farmakiotis D, et al. Adiponectin levels in women with polycystic ovary syndrome:a systematic review and a meta-analysis. Hum Reprod Update.2009,15(3):297-307.
53. Xu A, Chan KW, Hoo RL, et al. Testosterone selectively reduces the high molecular weight form of adiponectin by inhibiting its secretion from adipocytes. J Biol Chem.2005,280(18):18073-18080.
54. Bik W, Baranowska-Bik A, Wolinska-Witort E, et al. The relationship between metabolic status and levels of adiponectin and ghrelin in lean women with polycystic ovary syndrome. Gynecol Endocrinol.2007,23(6):325-331.
55. Carmina E, Bucchieri S, Mansueto P, et al. Circulating levels of adipose products and differences in fat distribution in the ovulatory and anovulatory phenotypes of polycystic ovary syndrome. Fertil Steril.2009,91(4 Suppl):1332-1335.
56. Stumvoll M, Tschritter O, Fritsche A, et al. Association of the T-G polymorphism in adiponectin (exon 2) with obesity and insulin sensitivity:interaction with family history of type 2 diabetes. Diabetes.2002,51(1):37-41.
57. Hara K, Boutin P, Mori Y, et al. Genetic variation in the gene encoding adiponectin is associated with an increased risk of type 2 diabetes in the Japanese population. Diabetes.2002,51(2):536-540.
58. Fumeron F, Aubert R, Siddiq A, et al. Adiponectin gene polymorphisms and adiponectin levels are independently associated with the development of hyperglycemia during a 3-year period:the epidemiologic data on the insulin resistance syndrome prospective study. Diabetes.2004,53(4):1150-1157.
59. Zacharova J, Chiasson JL, Laakso M, et al. The common polymorphisms (single nucleotide polymorphism [SNP]+45 and SNP +276) of the adiponectin gene predict the conversion from impaired glucose tolerance to type 2 diabetes:the STOP-NIDDM trial. Diabetes.2005,54(3):893-899.
60. Jang Y, Lee JH, Kim OY, et al. The SNP276G>T polymorphism in the adiponectin (ACDC) gene is more strongly associated with insulin resistance and cardiovascular disease risk than SNP45T>G in nonobese/nondiabetic Korean men independent of abdominal adiposity and circulating plasma adiponectin. Metabolism.2006,55(1):59-66.
61. Qj L, Doria A, Manson JE, et al. Adiponectin genetic variability, plasma adiponectin, and cardiovascular risk in patients with type 2 diabetes. Diabetes. 2006,55(5):1512-1516.
62. Panidis D, Kourtis A, Kukuvitis A, et al. Association of the T45G polymorphism in exon 2 of the adiponectin gene with polycystic ovary syndrome:role of Delta4-androstenedione. Hum Reprod.2004,19(8):1728-1733.
63. Xita N, Georgiou I, Chatzikyriakidou A, et al. Effect of adiponectin gene polymorphisms on circulating adiponectin and insulin resistance indexes in women with polycystic ovary syndrome. Clin Chem.2005,51(2):416-423.
64. Haap M, Machicao F, Stefan N, et al. Genetic determinants of insulin action in polycystic ovary syndrome. Exp Clin Endocrinol Diabetes.2005,113(5): 275-281.
65. Heinonen S, Korhonen S, Helisalmi S, et al. Associations between two single nucleotide polymorphisms in the adiponectin gene and polycystic ovary syndrome. Gynecol Endocrinol.2005,21(3):165-169.
66. Ranjzad F, Mahmoudi T, Irani Shemirani A, et al. A common variant in the adiponectin gene and polycystic ovary syndrome risk. Mol Biol Rep,2012,39(3): 2313-2319.
67. Yoshihara K, Yahata T, Kashima K, et al. Association of single nucleotide polymorphisms in adiponectin and its receptor genes with polycystic ovary syndrome. J Reprod Med.2009,54(11/12):669-674.
68. Ning Zhang, Yu-Hua Shi, Cui-Fang Hao, et al. Association of +45G15G(T/G) and +276(G/T) polymorphisms in the ADIPOQ gene with polycystic ovary syndrome among Han Chinese women. European Journal of Endocrinology.2008,158(2): 255-260.
69. Li L, Yun JH, Lee JH, et al. Association study of +45G15G(T/G) and +276(G/T) polymorphisms in the adiponectin gene in patients with polycystic ovary syndrome. Int J Mol Med.2011,27(2):283-287.
70. Demirci H, Yilmaz M, Ergun MA, et al. Frequency of adiponectin gene polymorphisms in polycystic ovary syndrome and the association with serum adiponectin, androgen levels, insulin resistance and clinical parameters. Gynecol Endocrinol.2010,26(5):348-355.
71. Campos DB, Palin MF, Bordignon V, et al. The beneficial adipokines in reproduction and fertility. Int J Obes.2008,32(2):223-231.
72. Ledoux S, Campos DB, Lopes FL, et al. Adiponectin induces periovulatory changes in ovarian follicular cells. Endocrinology.2006,147(11):5178-5186.
73. Bersinger NA, Birkhauser MH, Wunder DM. Adiponectin as a marker of success in intracytoplasmic sperm injection/embryo transfer cycles. Gynecol Endocrinol. 2006,22(9):479-483.
74. Chabrolle C, Tosca L, Rame C, et al. Adiponectin increases insulin-like growth factor I induced progesterone and estradiol secretion in human granulose cells. Fertil Steril.2008,92(6):1988-1996.
75. Chabrolle C, Tosca L, Crochet S, et al. Expression of adiponectin and its receptors (AdipnRl and AdipoR2) in chicken ovary:potential role in ovarian steroidogenesis. Domest Anim Endocrinol.2007,33(4):480-487.
76. Chabrolle C, Tosca L, Dupont J. Regulation of adiponectin and its receptors in rat ovary by human chorionic gonadotrophin treatment and potential involvement of adiponectin in granulosa cell steroidogenesis. Reproduction.2007,133(4): 719-731.
77. Chappaz E, Albornoz MS, Campos D, et al. Adiponectin enhances in vitro development of swine embryos. Domest Anim Endocrinol.2008,35(2):198-207.
78. Tabandeh MR, Hosseini A, Sae b M, et al. Changes in the gene expression of adiponectin and adiponectin receptors (AdipoR1 and AdipoR2) in ovarian follicular cells of dairy cow at different stages of development. Theriogenology. 2010,73(5):659-669.
79. Maillard V, Uzbekova S, Guignot F, et al. Effect of adiponectin on bovine granulosa cell steroidogenesis, oocyte maturation and embryo development. Reproductive Biology and Endocrinology.2010,10; 8:23-37.
80. Lord E, Ledoux S, Murphy BD, et al. Expression of adiponectin and its receptors in swine. J Anim Sci.2005,83(3):565-578.
81. Lagaly DV, Aad PY, Grado-Ahuir JA, et al. Role of adiponectin in regulating ovarian theca and granulosa cell function. Molecular and Cellular Endocrinology. 2008,12; 284(1-2):38-45.
82. Chabrolle C, Jeanpierre E, Tosca L, et al. Effects of high levels of glucose on the steroidogenesis and the expression of adiponectin receptors in rat ovarian cells. Reproductive Biology and Endocrinology.2008,19; 6:11.
83. Chabrolle C, Tosca L, Rame C, et al. Adiponectin increase insulin-like growth factor 1 induced progesterone and estradiol secretion in human granulose cells. Fertil Steril.2009,92(6):1988-1996.
84. Pierre P, Froment P, Negre D, et al. Role of adiponectin receptors, AdipoRl and AdipoR2, in the steroidogenesis of the human granulosa tumor cell line, KGN. Hum Reprod.2009,24(11):2890-2901.
85. Kim ST, Marquard K, Stephens S, et al. Adiponectin and adiponectin receptors in the mouse preimplantation embryo and uterus. Human Reproduction.2011,26(1): 82-95.
86. Liu YH, Tsai EM, Chen YL, et al. Serum adiponectin levels increase after human chorionic gonadotropin treatment during in vitro fertilization. Gynecol Obstet Invest.2006,62(2):61-65.
87. Liu YH, Tsai EM, Wu LC, et al. Higher basal adiponectin levels are associated with better ovarian response to gonadotropin stimulation during in vitro fertilization. Gynecol Obstet Invest.2005,60(3):167-170.
88. Bersinger NA, Birkhauser MH, Wunder DM. Adiponection as a marker of success in intracytoplasmic sperm injection/embryo transfer cycles. Gynecol Endocrinol.2006,22(9):479-483.
89. Gutman G, Barak V, Maslovitz S, et al. Recombinant luteinizing hormone induces increased production of ovarian follicular adiponectin in vivo:implications for enhanced insulin sensitivity. Fertil Steril.2008,91(5):1837-1841.
90. Richards JS, Liu Z, Kawai T, et al. Adiponectin and its receptors modulate granulosa cell and cumulus cell functions, fertility, and early embryo development in the mouse and human. Fertil Steril.2012,98(2):471-479.