罗格列酮对体外培养的PCOS患者卵巢黄素化颗粒细胞胰岛素抵抗的影响

目的:探讨罗格列酮改善PCOS卵巢局部胰岛素抵抗的作用。方法:收集行IVF-ET治疗的11例PCOS患者(PCOS组)和15例排卵正常的输卵管性不孕患者(对照组)促排卵后黄素化颗粒细胞行体外培养,分别用不同浓度罗格列酮(0nmol/L、1nmol/L、10nmol/L、100nmol/L、1000nmol/L、10000nmol/L)处理细胞48h,采用RT-PCR和Westernblotting分别检测卵巢黄素化颗粒细胞胰岛素受体底物(IRS)-1和IRS-2mRNA的表达及蛋白含量。结果:①基础状态下,PCOS组IRS-1mRNA表达及蛋白含量较对照组显著增加;IRS-2mRNA表达及蛋白含量较对照组显著降低(P<0.05);②不同浓度罗格列酮作用后,PCOS组IRS-1mRNA及蛋白表达显著降低,IRS-2mRNA及蛋白表达显著增加,并呈剂量依赖性,而正常对照组无变化。结论:PCOS患者卵巢局部存在胰岛素抵抗,其原因可能与IRS-1和IRS-2mRNA及蛋白表达异常有关;罗格列酮可通过提高黄素化颗粒细胞IRS-2的表达,降低IRS-1的表达,改善PCOS患者卵巢局部胰岛素抵抗。     

PCOS患者卵巢胰岛素抵抗的发生机理探讨

目的:探讨胰岛素抵抗/高胰岛素血症在PCOS生殖功能障碍中的作用。方法:收集行IVF-ET治疗的11例PCOS患者(PCOS组)和15例排卵正常的输卵管性不孕患者(对照组)促排卵后卵巢黄素化颗粒细胞,行体外培养,分别用不同浓度胰岛素处理细胞48h,采用RT-PCR和Westernblot检测黄素化颗粒细胞胰岛素受体底物(IRS)-1和IRS-2mRNA及蛋白的表达。采用放射免疫法检测血清性激素及空腹血清胰岛素(FIN)水平;采用葡萄糖氧化酶法测定空腹血糖(FPG)水平;计算胰岛素抵抗指数(HOMA-IR)。结果:①PCOS患者血清LH、LH/FSH、T、FIN及HOMA-IR均明显高于对照组(P<0.05);②0mU/ml胰岛素时,PCOS组黄素化颗粒细胞IRS-1mRNA及蛋白的表达水平明显高于对照组(P<0.05),而IRS-2mRNA及蛋白的表达水平较对照组明显降低(P<0.05);③10mU/ml胰岛素对二组患者黄素化颗粒细胞IRS-1和IRS-2mRNA及蛋白的表达均无影响(P>0.05);④100mU/ml或1000mU/ml胰岛素作用后,二组患者IRS-1mRNA及蛋白的表达水平均明显增高(P<0.05),而IRS-2mRNA及蛋白的表达水平均明显降低(P<0.05)。结论:PCOS患者胰岛素抵抗/高胰岛素血症通过提高卵巢颗粒细胞IRS-1的表达,降低IRS-2的表达参与患者卵巢生殖功能障碍的发生。     

PCOS患者脂肪组织IRS-1、IRS-2蛋白表达及其酪氨酸磷酸化的研究

目的:研究多囊卵巢综合征(polycystic ovary syndrome,PCOS)患者脂肪组织胰岛素受体底物-1(insulin receptor substrate-1,IRS-1)及胰岛素受体底物-2(insulin recep-tor substrate-2,IRS-2)蛋白的表达及其酪氨酸磷酸化,探讨PCOS产生胰岛素抵抗(insulinresistance,IR)的分子机制。方法:用放射免疫法检测各实验组血清促黄体生成素(luteini-zing hormone,LH)、促卵泡激素(follicle stimulating hormone,FSH)、睾酮(testosterone,T)、血清空腹胰岛素(fasting insulin,FIN)的水平;用葡萄糖氧化酶法测定血浆空腹血糖(fast-ing plasma glucose,FPG);用HOMA(homeostasis model assessment,HOMA)模型计算胰岛素抵抗指数(HOMA-IR);用Western blot检测IRS-1及IRS-2蛋白的表达,应用免疫沉淀及增强化学发光法检测二者的酪氨酸磷酸化程度。结果:(1)PCOS IR组患者血清LH、LH/FSH、T、FIN及HOMA-IR均显著高于PCOS非IR组与对照组(均P(0.05);(2)PCOSIR组与PCOS非IR组及对照组相比,IRS-1蛋白表达量明显下降(P(0.05),IRS-2蛋白表达差异无统计学意义(P>0.05);(3)PCOS IR组IRS-1及IRS-2蛋白质酪氨酸磷酸化程度显著降低(均P(0.05)。结论:PCOS患者脂肪组织IRS-1蛋白表达下降,IRS-1及IRS-2蛋白质酪氨酸磷酸化程度降低,由此导致的受体后信号转导障碍可能是产生胰岛素抵抗的机制之一。     

糖耐量试验正常的多囊卵巢综合征患者子宫内膜胰岛素及其受体表达的研究

目的 :了解糖耐量正常的多囊卵巢综合征 (PCOS)患者子宫内膜局部胰岛素水平及其受体表达的情况 ,以了解它们在多囊卵巢综合征发病中的作用。方法 :采用免疫组化方法对口服葡萄糖耐量试验 (OGTT)正常的PCOS患者子宫内膜石蜡标本进行研究。结果 :PCOS的子宫内膜大部分腺体发育均达到增殖中晚期的发育程度 ,其局部胰岛素水平及其受体表达均显著高于增殖早期对照组 (P <0 .0 1) ;其胰岛素水平与增殖中晚期对照组差异无显著性 (P >0 .0 5) ;PCOS组胰岛素受体表达则低于增殖晚期组 ,差异有显著性 (P <0 .0 5) ;增殖晚期对照组与增殖早中期对照组相比胰岛素及其受体差异有显著性(P <0 .0 1)。结论 :糖耐量试验正常的PCOS患者子宫内膜可能依然存在胰岛素抵抗现象 ,这种子宫内膜的胰岛素抵抗可能影响子宫内膜发育 ,因此 ,可能是PCOS妊娠率低及受孕后流产率高的原因之一。     

多囊卵巢综合征子宫内膜局部胰岛素抵抗的研究

目的探讨多囊卵巢综合征(PCOS)患者子宫内膜是否存在局部胰岛素抵抗(IR)。方法分离28例PCOS患者(PCOS组)和32例不孕症患者(对照组)的子宫内膜上皮和间质细胞并体外培养;采用葡萄糖氧化酶法测定PCOS子宫内膜细胞对葡萄糖的摄取量;应用免疫组化法检测胰岛素(Ins)、胰岛素受体(InsR)和葡萄糖转运蛋白4(GLUT4)在PCOS患者子宫内膜的表达。结果 PCOS组子宫内膜细胞葡萄糖摄取量与对照组比较,差异无统计学意义(P0.05);加入1×10-7mmol/L胰岛素后,PCOS组葡萄糖摄取量低于对照组(P0.05);PCOS肥胖及IR组葡萄糖摄取量低于非肥胖及非IR组(P0.05)。PCOS组子宫内膜Ins表达与对照组比较,差异无统计学意义(P0.05),但InsR及GLUT4表达低于对照组(P0.05);PCOS肥胖和IR组子宫内膜InsR及GLUT4表达低于非肥胖和非IR组,但差异无统计学意义(P0.05)。结论 P-COS患者子宫内膜存在局部IR,肥胖及全身IR加重子宫内膜局部IR。     

多囊卵巢综合征患者胰岛素、胰岛素样生长因子及其受体的表达

目的: 探讨胰岛素(Ins)、胰岛素样生长因子(IGFs)及其受体在多囊卵巢综合征(PCOS)发展中的作用。方法:应用放射免疫方法检测病例组22例PCOS患者和对照组14例卵巢冠囊肿患者空腹血清Ins、IGF-Ⅰ、睾酮(T)和卵泡液内IGF-Ⅱ水平;应用免疫组化法检测卵泡组织中胰岛素受体(Ir)、IGF-Ⅰ受体(IGF-Ⅰr)的表达及分布情况。结果:①病例组空腹血清 Ins(22.51±10.23 mIU/mL)及IGF-Ⅰ(115.55±35.06 ng/mL)均明显高于对照组(分别为10.95±2.82 mIU/mL、57.96±37.46 ng/mL),P<0.001;②病例组空腹血糖/空腹胰岛素(G/I)比值(4.56±1.42)和卵泡液IGF-Ⅱ(0.133±0.02 ng/mL)均低于对照组(分别为9.36±2.85、0.200±0.04 ng/mL),P<0.001;③病例组内T与Ins(r=0.636,P<0.001)、 T与G/I(r=-0.558,P<0.01)呈明显直线相关;④卵泡组织石蜡切片显示,Ir定位于细胞膜,在卵泡膜细胞、颗粒细胞和间质细胞均有表达,尤其在血管内皮细胞表达更丰富,病例组和对照组间Ir表达强度差别无显著性;⑤各组切片均未见IGF-Ⅰr表达。结论:Ins、IGFs可能通过Ir参与促进PCOS患者的雄激素分泌及卵泡成熟障碍,这为PCOS的临床治疗提供了新的启示。     

吡咯列酮对PCOS大鼠下丘脑、卵巢中瘦素受体表达的影响及其意义

目的:研究吡咯列酮对PCOS大鼠下丘脑、卵巢中瘦素受体(Lep-R)表达的影响及其意义。方法:将30只雌性SD大鼠分为3组:正常对照组、PCOS组、吡咯列酮干预组。放射免疫法测定各组大鼠血清T、LH、空腹血清血糖(fasting serum glucose,FSG)、胰岛素(insulin,Ins)、瘦素(leptin,Lep)水平,通过免疫组织化学、实时荧光相对定量PCR(Real-time RT-PCR)方法检测各组大鼠下丘脑、卵巢中Lep-R的相对表达。结果:经吡咯列酮干预后,PCOS大鼠血清T、LH、Ins、Lep水平明显降低,下丘脑Lep-R表达增强,而卵巢中Lep-R表达减弱。结论:下丘脑Lep-R表达的下调可能是PCOS大鼠瘦素抵抗的原因之一;吡咯列酮可以改善PCOS大鼠内分泌紊乱。     

PCOS是卵巢对胰岛素超敏的综合征

多囊卵巢综合征(PCOS)以胰岛素抵抗或高胰岛素血症导致高雄激素血症为主要发病机制。卵巢内胰岛素调节雄激素合成的信号传导途径亢进,即卵巢局部组织对胰岛素作用的敏感性升高。胰岛素可以刺激正常妇女和PCOS妇女卵巢产生雄激素,但在体外PCOS妇女卵巢细胞对胰岛素刺激雄激素合成的反应性更高。多囊卵巢对促性腺激素的刺激敏感性也升高,即卵巢的反应性增强。部分发展为PCOS的妇女是由于卵巢内雄激素合成通路选择性的和特异性地对胰岛素敏感性增强引起的。因此,卵巢内部雄激素合成对胰岛素信号通路的过度敏感是一部分多囊卵巢妇女发展为PCOS的病因。     

曲格列酮在治疗与胰岛素抵抗相关的PCOS对改善内分泌及排卵功能的疗效

多囊卵巢综合征(PCOS)是育龄妇女最常见的内分泌疾病,在育龄妇女中约占6%,其特征为慢性无排卵,LH水平升高,高雄激素血症。最近的研究提示胰岛素抵抗在PODS发病机制中起关键性作用。曲格列酮(Troglitazone)是一种改善胰岛素敏感性的新药。     

多囊卵巢综合征胰岛素抵抗与卵巢局部IGF-I系统的相关性研究

目的探讨卵巢局部胰岛素样生长因子-I(IGF-I)系统在PCOS胰岛素抵抗发病机制中的作用。方法选择30例PCOS胰岛素抵抗患者为研究组,30例输卵管性不孕患者为对照组。测定并比较两组的血清、小卵泡液中IGF-I、胰岛素样生长因子结合球蛋白-1(IGFBP-1)及各项性激素、糖代谢及卵巢超声指标,分析其与各项指标之间的相关性。结果研究组小卵泡液IGF-I高于对照组和其血清水平(P0.01),血清和小卵泡液的IGFBP-1低于对照组(P0.05,P0.01),小卵泡液IGFBP-1水平低于血清(P0.05)。研究组小卵泡液IGF-I与总睾酮(T0)、雌二醇(E2)及卵巢体积(OV)、卵巢总面积(TA)、卵泡数(FN)、空腹胰岛素(FI)和口服葡萄糖后2h胰岛素(2h胰岛素)呈正相关(r分别为0.94、0.51、0.52、0.49、0.65、0.76和0.58,P值均0.05);血清IGF-I与胰岛素敏感指数(ISI)、体重指数(BMI)和腰臀比(WHR)呈正相关(r分别为0.47、0.61和0.58,P0.05),与定量胰岛素敏感指数(QUICKI)呈负相关(r=-0.34,P0.05);研究组小卵泡液及血清IGFBP-1与FINS、2h胰岛素呈负相关(r=-0.48,P0.001;r=-0.39,P0.05;r=0.54,P0.05;r=-0.52,P0.05)。结论 PCOS患者的外周胰岛素抵抗可能通过影响卵巢局部IGF-I系统,刺激卵巢组织产生大量雄激素,导致排卵障碍。     

Expression of insulin-receptor substrate-1 and -2 in ovaries from women with insulin resistance and from controls

OBJECTIVE: To evaluate the role of insulin-receptor substrate (IRS)-1 and -2 in ovary dysfunction in women with insulin resistance. DESIGN: Immunoblotting and immunohistochemical analyses of the localization and staining intensity of IRS-1 and IRS-2 in the ovaries of women with the polycystic ovary syndrome (PCOS) and gestational diabetes mellitus. SETTING: Department of Obstetrics and Gynecology, Turku University Central Hospital. PATIENT(S): Sections of ovary were obtained at the time of cesarean section from five volunteers without medical complications and three patients with gestational diabetes mellitus. Paraffin-embedded ovary sections were selected from those on file from the department of pathology; four were from women with a histologic diagnosis of PCOS and seven were from women with endometriosis (controls). INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Protein expression of IRS in human ovary samples. RESULT(S): Immunoblotting with specific monoclonal and polyclonal antibodies showed the presence of 165-kDa and 183-kDa proteins that corresponded to the size of IRS-1 and IRS-2, respectively, in normal pregnant ovaries and human cultured follicles. Immunohistochemical staining showed that positive IRS-2 expression in antral follicles was restricted to theca internal cells in ovulatory ovaries but was distributed widely in all compartments of follicles in different stages in polycystic ovaries. Compared with follicles at a similar stage of development in ovulatory ovaries, follicles in polycystic ovaries showed decreased staining for IRS-1 in granulosa cells but increased staining for IRS-2 in theca internal cells. These features of IRS-1 and -2 expression were also noted in preantral and atretic follicles from patients with gestational diabetes mellitus compared with those who had uncomplicated pregnancy. CONCLUSION(S): This study highlights a shift of the follicular insulin signal protein from IRS-1 to IRS-2 in insulin-resistant states and suggests an association between this change and ovarian abnormality in PCOS and gestational diabetes mellitus.     

Troglitazone: a possible modulator of ovarian steroidogenesis

OBJECTIVE: Troglitazone increases insulin sensitivity. When used to treat women with insulin-resistant polycystic ovary syndrome (PCOS), troglitazone lowers androgen concentrations and improves ovulatory, endocrine, and metabolic disturbances. However, it is not known whether these effects are due to increased peripheral insulin sensitivity only or to direct effects on steroidogenesis. To determine whether troglitazone has a direct effect on ovarian steroidogenesis, we studied the effect of troglitazone on androgen production in cultured rat theca interstitial cells (rTICs) and on progesterone production in cultured human granulosa lutein cells (hGLCs). METHODS: Primary cell cultures of rTICs were isolated from the ovaries of hypophysectomized immature Sprague-Dawley rats, and hGLCs was obtained from women who had in vitro fertilization-embryo transfer. Treatments included troglitazone, insulin, LH, hCG, and combinations of these agents. Media were collected and assayed for androsterone and progesterone. RESULTS: Troglitazone decreased both basal and insulin-, LH-, and hCG-stimulated androsterone production by TIC and progesterone production by hGLCs in a dose-dependent manner. CONCLUSION: We found a direct effect of troglitazone on the production of androsterone by rTICs and progesterone by hGLCs. These results suggest that the beneficial effects of troglitazone in PCOS may not be due solely to improvement of peripheral insulin resistance and hyperinsulinemia, but also to a possible direct effect on ovarian steroidogenesis.     

多囊卵巢综合征合并胰岛素抵抗患者脂肪组织胰岛素受体底物2蛋白的表达及酪氨酸磷酸化的研究

目的检测多囊卵巢综合征(PCOS)患者脂肪组织胰岛素受体底物(IRS)2蛋白表达及其酪氨酸磷酸化的程度,探讨发生PCOS合并胰岛素抵抗(IR)的机制。方法采用蛋白印迹(Westernblot)法,检测PCOS合并IR患者19例(PCOS合并IR组)、PCOS非IR患者17例(PCOS非IR组)及因单纯子宫肌瘤行子宫切除术的患者20例(对照组)的IRS2蛋白的表达;并采用免疫组化技术,检测IRS2在脂肪组织中的分布;采用免疫沉淀及增强化学发光法,检测IRS2的酪氨酸磷酸化程度。结果(1)PCOS合并IR组、PCOS非IR组及对照组IRS2蛋白表达分别为115±026、113±026及100±025,3组比较,差异无统计学意义(P>005)。(2)PCOS合并IR组、PCOS非IR组及对照组IRS2蛋白酪氨酸磷酸化程度分别为077±016、091±025及100±012,3组比较,差异有统计学意义(P<005);PCOS非IR组与对照组比较,差异无统计学意义(P>005)。结论PCOS合并IR患者脂肪组织的IRS2蛋白酪氨酸磷酸化程度的降低,可能是发生IR的机制之一。     

Immunohistochemical detection of plasminogen activator inhibitor-1 in polycystic ovaries.

Anovulation in women with polycystic ovary syndrome (PCOS) is incompletely understood. The concentration of the glycoprotein plasminogen activator inhibitor-1 (PAI-1) is raised in insulin resistance. This has been described in the granulosa and theca cell layers of the animal but not the human ovary. This study was performed to investigate the location of PAI-1 in the human ovary and investigate whether it may contribute to anovulation in PCOS. PAI-1 was localized immunohistochemically and quantitated using computer image analysis in 17 ovarian follicles from five women with a diagnosis of PCOS and compared with 15 follicles from six normal ovaries. PAI-1 was predominantly found in the granulosa and theca cells in both polycystic and normal ovaries. Image analysis did not reveal a difference in the PAI-1 signal from polycystic compared with normal ovaries. This study shows that PAI-1 plays a role in human ovulation, but its role in PCOS requires further research.     

Concentration of vascular endothelial growth factor released by cultured human luteinized granulosa cells is higher in women with polycystic ovaries than in women with normal ovaries

To test the hypothesis that increased serum levels of vascular endothelial growth factor (VEGF) in women with polycystic ovaries or the polycystic ovary syndrome (PCOS) result from excess release by ovarian granulosa cells.Prospective study.Academic research setting.Twenty women undergoing IVF treatment, of whom 10 had normal ovaries and 10 had polycystic ovaries.Human granulosa lutein cells were isolated from follicular fluid obtained on the day of oocyte retrieval. Release of VEGF was assessed after co-incubation of granulosa lutein cells with gonadotropins and insulin. Serum and follicular fluid concentrations of VEGF were measured.Release of VEGF from granulosa lutein cells and serum levels of VEGF.Incubation with human hCG, and luteinizing hormone increased release of VEGF into the culture medium. Insulin alone did not increase release of VEGF, but addition of insulin increased hCG-stimulated release of VEGF. Serum and follicular fluid VEGF concentrations and the amount VEGF released from granulosa lutein cells obtained from women with polycystic ovaries or PCOS and those who developed the ovarian hyperstimulation syndrome were greater than those from granulosa lutein cells obtained from women with normal ovaries and those who did not develop the ovarian hyperstimulation syndrome.The amount of VEGF released by granulosa lutein cells is gonadotropin dependent and is augmented by insulin. The increased circulating concentrations of VEGF in women with PCOS may not only be due to an increased number of actively secreting granulosa lutein cells but also due to increased secretory capacity of each granulosa cell.     

脂肪组织中胰岛素受体底物1蛋白的表达及酪氨酸磷酸化在多囊卵巢综合征发病中的作用

目的探讨多囊卵巢综合征(PCOS)患者脂肪组织胰岛素受体底物1(IRS1)的蛋白表达及酪氨酸磷酸化在PCOS发病中的作用。方法采用免疫沉淀法、Western印迹法和增强化学发光蛋白免疫印迹法及图像分析半定量检测24例PCOS患者[PCOS组,根据体重指数分为肥胖(体重指数≥24kg/m2)和非肥胖(体重指数<24kg/m2)者各12例]及同期因卵巢囊肿或输卵管阻塞行开腹手术的非PCOS患者24例(对照组,根据体重指数分为肥胖和非肥胖者各12例)的脂肪组织中IRS1的蛋白表达及酪氨酸磷酸化程度。结果PCOS组肥胖者IRS1的蛋白表达为(82±15)%,PCOS组非肥胖者为(79±18)%;对照组肥胖者为(75±19)%,对照组非肥胖者为(70±19)%,各组间比较,差异均无统计学意义(P>005)。脂肪组织中IRS1的酪氨酸磷酸化程度在PCOS组肥胖者为(52±23)%,对照组非肥胖者为(88±12)%,两者比较,差异有统计学意义(P<001);对照组肥胖者为(45±22)%,PCOS组非肥胖者为(70±25)%,与对照组非肥胖者比较,明显降低,差异均有统计学意义(P<001,P<005);PCOS组肥胖者和对照组肥胖者间以及PCOS组肥胖者和PCOS组非肥胖者间比较,差异均无统计学意义(P>005)。结论PCOS组脂肪组织IRS1的蛋白表达及酪氨酸磷酸化程度明显减弱,可能参与胰岛素受体后信号传导抑制,并与PCOS胰岛素抵抗的发生有