STOX1在子痫前期滋养细胞炎症反应中的作用研究

目的 探讨STOX1(storkhead box1)基因在人绒毛膜癌细胞系JEG3细胞模拟的滋养细胞炎症反应中的作用。方法 采用人绒毛膜癌细胞系JEG3细胞株模拟滋养细胞,构建过表达/敲除STOX1基因稳转细胞系模型,实验组及对照组加入0.05 mmol/L阿司匹林,采用Transwell测定细胞迁移能力,并采用流式细胞仪检测各组细胞凋亡,实时定量PCR和蛋白质免疫印迹法检测细胞相关炎性、缺氧、凋亡因子的表达、生物学行为、分子生物学的m RNA及蛋白的相对表达量。结果 过表达STOX1滋养细胞组中缺氧诱导因子α（HIF-1α）、内皮型一氧化氮合酶（e NOS）、诱导型一氧化氮合酶（i NOS）B淋巴细胞瘤-2(Bcl-2)均显著降低（P <0.01）;同时,炎性反应相关因子核因子κB(NF-κB)、肿瘤坏死因子α（TNF-α）、白介素8(IL-8)及凋亡相关因子、半胱氨酸蛋白酶-3(Caspase-3)在基因及蛋白水平表达均明显升高（P <0.05）。敲除STOX1后HIF-1α（P <0.01）、e NOS(P <0.05)、i NOS(P <0.001...     

PTEN在子痫前期胎盘滋养细胞浸润不足中的作用及机制研究

目的:检测子痫前期(PE)患者胎盘组织中PTEN表达,探讨PTEN对滋养细胞迁移功能的调控及其可能机制。方法:收集重度PE患者和正常孕妇的胎盘组织各20例,实时定量PCR和Western blot法检测胎盘组织中PTEN表达;将PTEN过表达质粒(pcDNA3.1-HA-PTEN)或特异性siRNA(siPTEN)瞬时转染至HTR-8/SVneo细胞,同时转染pcDNA3.1或siCTL作为对照。采用划痕实验评估细胞迁移能力,Western blot法检测p-Akt(Thr308)、p-Akt(Ser473)、Akt、MMP-2和MMP-9表达。结果:重度PE患者胎盘组织中PTEN mRNA和蛋白表达明显高于正常孕妇,差异均有统计学意义(P0.01)。与转染pcDNA3.1组比较,pcDNA3.1-HA-PTEN瞬时转染HTR-8/SVneo细胞48h后,细胞迁移率明显降低[(26.42±6.68)%vs(52.92±6.71)%,P0.01],p-Akt(Thr308)和p-Akt(Ser473)表达显著下调,Akt表达无明显改变,MMP-2和MMP-9蛋白表达下降;与转染siCTL组比较,siPTEN瞬时转染HTR-8/SVneo细胞48h后,细胞迁移率则明显升高[(50.39±6.84)%vs(30.04±5.40)%,P0.01],p-Akt(Thr308)和p-Akt(Ser473)的表达明显上调,Akt表达无明显改变,MMP-2和MMP-9蛋白表达升高。结论:重度PE患者胎盘组织中PTEN表达明显升高,PTEN可通过下调Akt活性降低MMP-2和MMP-9表达,抑制滋养细胞的迁移。提示PTEN在PE的发病中发挥了一定的作用。     

PTEN在子痫前期胎盘滋养细胞浸润不足中的作用及机制研究26

目的:检测子痫前期(PE)患者胎盘组织中PTEN表达,探讨PTEN对滋养细胞迁移功能的调控及其可能机制.方法:收集重度PE患者和正常孕妇的胎盘组织各20例,实时定量PCR和Western blot法检测胎盘组织中PTEN表达;将PTEN过表达质粒(pcDNA3.1-HA-PTEN)或特异性siRNA(siPTEN)瞬时转染至HTR-8/SVneo细胞,同时转染pcDNA3.1或siCTL作为对照.采用划痕实验评估细胞迁移能力,Western blot法检测p-Akt(Thr308)、p-Akt(Ser473)、Akt、MMP-2和MMP-9表达.结果:重度PE患者胎盘组织中PTEN mRNA和蛋白表达明显高于正常孕妇,差异均有统计学意义(P<0.01).与转染pcDNA3.1组比较,pcDNA3.1-HA-PTEN瞬时转染HTR-8/SVneo细胞48 h后,细胞迁移率明显降低[(26.42±6.68)%vs(52.92±6.71)%,P<0.01],p-Akt(Thr308)和p-Akt(Ser473)表达显著下调,Akt表达无明显改变,MMP-2和MMP-9蛋白表达下降;与转染siCTL组比较,siPTEN瞬时转染HTR-8/SVneo细胞48 h后,细胞迁移率则明显升高[(50.39±6.84)%vs(30.04±5.40)%,P<0.01],p-Akt(Thr308)和p-Akt(Ser473)的表达明显上调,Akt表达无明显改变,MMP-2和MMP-9蛋白表达升高.结论:重度PE患者胎盘组织中PTEN表达明显升高,PTEN可通过下调Akt活性降低MMP-2和MMP-9表达,抑制滋养细胞的迁移.提示PTEN在PE的发病中发挥了一定的作用.     

子痫前期滋养细胞功能障碍病因学研究进展

子痫前期(preeclapmpsia,PE)是围生期母婴死亡的主要原因之一,发病率为1%～5%,其病理生理改变发生在妊娠早期[1]。但在妊娠20周后发病,主要表现为高血压、蛋白尿、水肿,病情严重时会出现全身抽搐及多脏器功能损伤。子痫前期的病情变化复杂且很难预知,通常以终止妊娠控制病情发展,其病因和发病机制目前尚不明确。但是,从子痫前期的临床转归来看,患者娩出胎盘后该病相关的     

胎盘滋养细胞离子通道及其与子痫前期关系的研究进展

胎盘构成了母胎间物质交换的主要屏障。研究发现,在人类胎盘有氯离子通道、钙离子通道、钠离子通道和钾离子通道等的存在,这些通道与胎盘生理功能的调控密切相关,但其机制尚不清楚。目前离子通道功能改变与子痫前期关系的研究正在逐步开展。作者简要总结了胎盘离子通道研究的新进展,并讨论了离子通道与子痫前期可能的关系。     

葡萄糖调节蛋白78在子痫前期胎盘滋养细胞中的表达及意义

目的:探讨葡萄糖调节蛋白78( GRP78)在子痫前期孕妇胎盘组织中表达及其与子痫前期发病机制的关系.方法:选择2009年1 ～12月在青岛市市立医院及青岛市第八人民医院住院的子痫前期孕妇120例,其中早发型轻度子痫前期孕妇30例(早发轻度组),晚发型轻度子痫前期孕妇30例(晚发轻度组),早发型重度子痫前期孕妇30例(早发重度组),晚发型重度子痫前期孕妇30例(晚发重度组);另选同期健康孕妇30例为健康孕妇组.采用半定量逆转录-聚合酶链反应(RT-PCR)检测胎盘组织中GRP78mRNA相对表达量;采用免疫组织化学方法、West-ern-Blotting检测胎盘组织GRP78蛋白的表达.结果:①子痫前期各组胎盘组织中GRP78 mRNA表达水平均高于健康孕妇组,差异有统计学意义(P＜0.05);但子痫前期各组间分别比较,差异无统计学意义(P＞0.05).②子痫前期各组和健康孕妇组胎盘组织中都有GRP78蛋白表达,主要表达于细胞质,呈棕黄色染色.Western-Blotting检测结果显示,子痫前期各组胎盘组织中GRP78蛋白表达水平均高于健康孕妇组,差异有统计学意义(P＜0.05);但子痫前期各组间分别比较,差异无统计学意义(P＞0.05).结论:GRP78在子痫前期胎盘组织中表达升高,GRP78的表达水平变化可能与子痫前期发病有关.     

子痫前期患者胎盘组织中Rho激酶Ⅱ表达与滋养细胞凋亡的研究

目的探讨Rho激酶Ⅱ(Rho-associatedProteinKinaseⅡ,ROCKⅡ)在正常妊娠和子痫前期患者胎盘的分布和表达及与滋养细胞凋亡的关系。方法采用免疫组织化学法检测30例子痫前期(轻、重度)孕妇和15例正常妊娠妇女胎盘中ROCKⅡ的分布和表达,采用TUNEL法检测三组胎盘滋养细胞的凋亡情况。结果ROCKⅡ在正常妊娠和子痫前期患者胎盘的滋养细胞、内皮细胞、基质细胞中均有表达,以滋养细胞表达为主。子痫前期重度和轻度组以及正常妊娠组ROCKⅡ的免疫组化积分分别为(4.53±0.58)、(3.18±0.59)和(2.06±0.63),P<0.01。正常妊娠和子痫前期患者胎盘的滋养细胞、内皮细胞、基质细胞均存在凋亡,以滋养细胞凋亡为主。重度子痫前期组胎盘滋养细胞凋亡(0.28±0.03)%显著高于子痫前期轻度组(0.22±0.04)%,子痫前期轻度组显著高于正常妊娠组(0.16±0.05)%(P<0.01)。滋养细胞ROCKⅡ表达和滋养细胞凋亡具有相关性(r=0.96,P<0.01)。结论子痫前期患者胎盘组织中Rho激酶Ⅱ的高表达可能与其滋养细胞凋亡增加有关。     

sFlt-1及缺氧因素导致子痫前期孕鼠模型滋养细胞凋亡的分析

目的:探讨可溶性血管内皮因子受体-1(sFlt-1)及缺氧因素对子痫前期(PE)孕鼠动物模型滋养细胞凋亡的作用。方法:将120只孕鼠随机分为对照组(A组)和PE模型组(B、C、D组)。建立以L-NAME、PS/PC、STBM全面的复合PE和体外sFlt-1浓度孕鼠模型。妊娠10天开始、B、D组孕鼠腹腔注射sFlt-1,A、C组腹腔注射0.9%生理盐水,均持续13天。妊娠第22天测定孕鼠血压及蛋白尿。断鼠尾取静脉血,ELISA法检测各组孕鼠血浆sFlt-1水平。剖宫产取各组孕鼠胎盘绒毛组织清洗后行原代滋养细胞组织皿培养。A、B、C、D组滋养细胞分别于正常氧压、50ng/ml sFlt-1 2ml+正常氧压、低氧和50ng/ml sFlt-1 2ml+低氧培养箱中培养。48h后,收集培养的原代滋养细胞。流式细胞技术检测细胞调亡;Western blot法检测凋亡相关蛋白Bax、Bcl-2及Caspase-3表达。结果:对照组与PE模型组孕鼠的血浆sFlt-1水平、收缩压、舒张压及尿蛋白指标比较,差异有统计学意义(P0.01);PE模型组的胎盘滋养细胞凋亡率与对照组(3.95%)比较,差异有统计学意义(P0.01),PE模型组间两两比较,差异有统计学意义(P0.05)。与对照组比较,PE模型组胎盘组织中的促凋亡蛋白Bax明显上调,抑凋亡蛋白Bcl-2明显下调,凋亡执行蛋白caspase-3表达上调;PE模型组两两比较,差异均有统计学意义(P0.05)。结论:sFlt-1及缺氧因素与PE滋养细胞过度凋亡发生密切相关,两者促进PE的发生发展。     

子痫前期中lncRNA对滋养细胞的作用及机制研究

子痫前期是妊娠期高血压疾病的一种类型,病因及发病机制至今尚未阐明,其中滋养细胞行为异常起关键作用。长链非编码RNA(lncRNA)是指长度超过200个核苷酸(nt)非编码蛋白质的RNA转录本。随着研究的进展,人们发现lncRNA在不同水平对基因表达进行调控,影响滋养细胞增殖、分化、迁移、侵袭、凋亡及管型形成,导致血管重塑受损,从而引发子痫前期。本文旨在介绍子痫前期发病中lncRNA对滋养细胞的作用及其调控机制。     

合体滋养细胞微绒毛膜与子痫前期

子痫前期是严重危害母儿健康的产科常见病，是导致孕产妇和围生儿发病率和死亡率升高的主要原因之一，其病因至今未明。近年来合体滋养细胞微绒毛膜（STBM）与子痫前期发病密切相关得到重视，合体滋养细胞微绒毛膜是胎盘合体滋养细胞碎片脱落进入母体血循环后形成的，可经多种途径损害母体血管内皮细胞，影响母体自身免疫系统，从而在子痫前期的发病中起重要作用。就合体滋养细胞微绒毛膜与子痫前期的关系综述。     

HO在子痫前期患者胎盘组织和滋养细胞中的表达及青心酮的干预

目的探讨血红素氧合酶（heme oxygenase，HO）在子痫前期胎盘中的表达和作用机制，以及青心酮（DHAP）治疗子痫前期的理论依据。方法应用半定量反转录一聚合酶链反应法检测正常孕妇和妊娠期高血压疾病子痫前期患者胎盘组织以及青心酮干预的滋养细胞中HOmRNA表达水平。结果与正常孕妇组比较，子痫前期组胎盘中HO-1、2mRNA表达明显降低（P〈0．05，P〈0．01）；青心酮能上调滋养细胞中HO-1mRNA的表达水平（P〈0．05），并呈浓度依赖关系。结论子痫前期患者胎盘组织中HO-1、2表达降低，可能系子痫前期疾病发病的重要机制之一。青心酮能够上调滋养细胞中HO-1基因表达，其作用机制可能为通过改善HO—CO系统防治子痫前期。     

过表达SATB1保护人滋养细胞氧化应激损伤的机制研究

目的 探讨富集AT序列的特异性结合蛋白1（SATB1）参与人绒毛外滋养细胞株（HTR8/SVneo）氧化应激损伤的机制。方法 收集2013年9月至2014年9月在重庆医科大学附属第一医院产科行选择性剖宫产术的子痫前期孕妇（n=22）和正常足月妊娠行择期剖宫产术的孕妇的胎盘组织（n=22）。采用免疫组化和Western blotting（WB）检测正常晚孕期及子痫前期胎盘组织中SATB1的表达。构建SATB1过表达慢病毒载体（LV-SATB1）和阴性对照载体（LV-NC）。将滋养细胞分为正常培养对照组、缺氧/复氧（H/R）培养组、LV-SATB1+H/R培养组、LV-NC+H/R培养组。利用免疫荧光和WB法检测SATB1在各组细胞中的表达。采用流式细胞仪检测细胞凋亡指数及细胞内活性氧（reactive oxygen species, ROS）水平。利用Transwell小室模型检测细胞的体外侵袭率。结果 SATB1在子痫前期中的表达显著低于正常晚孕组（0.24±0.02 vs. 0.12±0.01, t=9.35,P<0.05）。转染LV-SATB1可显著提升H/R培养组的SATB1蛋白水平（0.92±0.17 vs. 0.58±0.15,P<0.01）。LV-SATB1可抑制H/R导致的细胞凋亡率上升及ROS聚集（P<0.01）。H/R组细胞侵袭能力低于正常培养组;上调SATB1的表达可提升H/R组细胞的体外侵袭能力（34.33±10.08 vs. 19.33±6.52,P<0.05）。结论    过表达SATB1可以逆转滋养细胞氧化应激损伤,其有望成为研究子痫前期发病机制的新靶点。     

QSOX1 regulates trophoblastic apoptosis in preeclampsia through hydrogen peroxide production

Objective: Oxidative stress plays a significant role in the pathogenesis of preeclampsia (PE), by inducing trophoblast cell death and consequent placental dysfunction. Quiescin sulfhydryl oxidase 1 (QSOX1) is upregulated in many types of cancer cells; it promotes disulfide bond formation as well as hydrogen peroxide (H₂O₂) production. The aims of present study are to investigate the expression pattern of QSOX1 in placentae of pregnancies complicated by PE and the role of QSOX1 in the regulation of trophoblastic function, thus providing in-depth understanding of the putative involvement of QSOX1 in the development of PE.

Methods: Human term placenta from normal pregnancies and from pregnancies complicated by PE was collected to measure QSOX1 expression and H₂O₂ levels. Down-regulation of QSOX1 in HTR-8/SVneo cells was achieved by siRNA interference. An in vitro cellular PE model was generated by hypoxic incubation. Protein expression levels were assessed by Western blotting, and H₂O₂ levels were determined in the cell culture medium as well as in the cell lysate. Trophoblast apoptosis was evaluated by TUNEL staining.

Results: QSOX1 was overexpressed in the PE placenta. Inhibition of QSOX1 expression in HTR-8/SVneo cells attenuated cell apoptosis and intracellular H₂O₂ levels. Hypoxia-induced QSOX1 expression in HTR-8/SVneo cells and led to apoptosis of HTR-8/SVneo cells, and knock-down of QSOX1 rescued hypoxia-induced trophoblast apoptosis.

Conclusions: Hypoxia-induced upregulation of QSOX1 and a consequent elevation in intracellular H₂O₂ increased apoptosis in placentae of pregnancies complicated by PE.     

Detection of circulating trophoblast particles in maternal blood using density gradient centrifugation in preeclampsia and in normotensive pregnancies

Objective: Preeclampsia (PE) is a frequent pregnancy-related disease and a major cause of maternal and fetal morbidity and mortality. Despite that, exact mechanisms of its pathophysiology remain largely unknown. In pregnancies complicated by PE, changes in the regulation of apoptosis seem to result in increased apoptotic shedding of trophoblast particles (TPs) into maternal circulation. Since the number of TP in peripheral blood is low, their detection necessitates pre-analytical enrichment. Methods: In this prospective multicenter pilot study we aimed to analyze TP in peripheral blood of 29 women with PE and of 13 unaffected controls using the OncoQuick®plus system for cell enrichment. Using immunocytochemistry, slides were evaluated microscopically for TP. Statistical analyses were performed using Welch’s t-test or Fisher’s exact test. Results: TP were detected in 10 (34.5%) women with PE and in two (15.4%) of unaffected controls. More than one TP were only found in PE. Comparing the mean counts of TP between groups, we detected significantly more TP in PE (p = 0.046). Conclusions: The OncoQuick®plus system can be applied to detect TP in both women with PE and in normotensive pregnancies. Longitudinal studies investigating the role of TP as a screening method for patients at risk for PE are warranted.     

The role of SATB1 in HTR8/SVneo cells and pathological mechanism of preeclampsia

Objective: Special AT-rich sequence binding protein 1 (SATB1) play potential roles in invasion and metastasis of tumor cells, and involves in human placental and fetal development. The objective of this study is to explore the role of SATB1 in migration and invasion of trophoblast and the potential mechanism.

Methods: Human placental tissues from first trimester, second trimester, term, and preeclampsia (PE) pregnancies were used to detect the expression and subcellular location of SATB1 and β-catenin. The human trophoblast cell line HTR8/SVneo, which was treated with hypoxia/re-oxygenation (H/R), lithium chloride (LiCl) or SATB1-siRNA to investigate the role of SATB1 and β-catenin signaling in human trophoblast function.

Results: We observed that SATB1 specifically localized within trophoblast cells of placenta tissues. Gradually reduced expression of SATB1 was observed during gestation, and lower expression were detected in placenta of PE compared with normal pregnancy. Moreover, the expression of SATB1 was decreased in H/R-treated HTR8/Svneo cells and villous explants. The Wnt/β-catenin signaling pathway interacted with SATB1 expression and H/R treatment resulted in Wnt pathway inhibition in trophoblast, while lithium chloride (LiCl) treatment enhanced H/R-exposed HTR8/SVneo migration and invasion. Knockdown of SATB1 significantly reduced the level of β-catenin and the migratory and invasive abilities of trophoblast.

Conclusions: Our data suggested that oxidative stress reduced SATB1 leading to inhibition of Wnt/β-catenin, and participate in the subdued migration and invasion of trophoblast, which indicated a potential pathological mechanism of PE.     

阿司匹林预防子痫前期的相关问题

子痫前期是严重威胁母胎安全的常见产科疾病,降低其发病率具有重要意义。近年研究表明,阿司匹林作用于血小板环氧化酶1(cyclooxygenase 1,COX1)等多个子痫前期发病的关键靶点,从而在预防疾病发生中发挥重要作用。掌握阿司匹林预防子痫前期的应用指征及时机,排除应用禁忌证后合理用药,可有效降低子痫前期的发病率,或减缓其病情严重程度。