孕妇血浆中sCD40及其配体与子痫前期发病及肾功能损害的关系

目的 探讨孕妇血浆中可溶性白细胞分化抗原(sCD40)和sCD40配体(sCD40L)水平变化与子痫前期发病及肾功能损害的关系.方法 选择2008年8月-2010年6月在青岛大学医学院附属医院产科分娩的轻度子痫前期孕妇28例(轻度子痫前期组),重度子痫前期孕妇35例(重度子痫前期组);另选同期妊娠结局良好的健康孕妇30例为对照组.比较3组孕妇分娩孕周及血压变化、血小板计数并检测其血常规、C反应蛋白(CRP)、尿常规、24h尿蛋白定量,以及血清尿酸(UA)、肌酐(Cr)、尿素氮(BUN)等生化指标.采用ELISA法检测3组孕妇血浆中sCD40和sCD40L的水平,并对血浆sCD40和sCD40L的水平与各临床指标的相关性进行分析.结果 (1)血常规及l临床指标:重度子痫前期组和轻度子痫前期组孕妇血浆CRP水平(分别为10.8及7.1 mg/L)均明显高于对照组(3.3 mg/L),前后两者分别比较,差异均有统计学意义(P＜0.05);重度子痫前期组高于轻度子痫前期组,两组比较,差异也有统计学意义(P＜0.05).重度子痫前期组孕妇分娩孕周(32.5周)明显低于轻度子痫前期组(37.2周)和对照组(38.6周),分别与对照组比较,差异均有统计学意义(P＜0.01);轻度子痫前期组与对照组比较,差异无统计学意义(P＞0.05).重度子痫前期组孕妇血小板计数(132×109/L)明显低于轻度子痫前期组(212×109/L)和对照组(216×109/L),分别比较,差异有统计学意义(P＜0.01);轻度子痫前期组与对照组比较,差异无统计学意义(P＞0.05).3组孕妇之间血红蛋白水平及白细胞数分别比较,差异均无统计学意义(P＞0.05).(2)血浆sCD40及sCD40L水平:重度子痫前期组、轻度子痫前期组和对照组孕妇血浆sCD40水平分别为133.6、126.5和90.7 ng/L,sCD40L水平分别为12.5、10.4和4.4 ng/L,24h尿蛋白定量分别为4.5、0.8 g和0,UA水平分别为486、289和162 μmol/L,重度子痫前期组以上各指标均明显高于轻度子痫前期组和对照组,差异均有统计学意义(P＜0.01);轻度子痫前期组也明显高于对照组,差异也有统计学意义(P＜0.01).重度子痫前期组孕妇血浆Cr(89 μmol/L)、BUN(5.32 mmol/L)水平高于轻度子痫前期组(分别为66μmol/L及4.49mmol/L)和对照组(分别为57 μmol/L及3.32 mmol/L),分别比较,差异均有统计学意义(P＜0.05);轻度子痫前期组与对照组比较,差异无统计学意义(P＞0.05).(3)相关性分析:轻、重度子痫前期组孕妇血浆sCD40水平与24 h尿蛋白定量呈正相关(r=0.434,P＜0.05),与UA、CRP呈明显正相关(r=0.536、0.528,P＜0.01),与收缩压、舒张压、分娩孕周、Cr、BUN、血小板计数无明显相关(r分别为0.135、0.183、-0.133、0.190、0.167、-0.221,P均＞0.05).轻、重度子痫前期组孕妇血浆sCD40L水平与24 h尿蛋白定量、UA、CRP均呈明显正相关(r分别为0.591、0.445、0.539,P均＜0.01),与收缩压、舒张压、分娩孕周、Cr、BUN、血小板计数无明显相关(r分别为0.178、0.212、-0.292、0.144、0.135、-0.273,P均＞0.05).轻、重度子痫前期组孕妇血浆sCD40L水平与sCD40呈明显正相关(r均为0.707,P＜0.01).对照组孕妇血浆sCD40、sCD40L水平与各项临床指标均无相关性(P＞0.05).结论 子痫前期孕妇血浆中sCD40和sCD40L水平明显升高,可能参与了子痫前期的发病,并导致肾功能损害.sCD40和sCD40L水平变化也与子痫前期的严重程度相关.

Abstract：

Objective To investigate the variance levels of plasma soluble leukocyte differentiation antigens CD40 (sCD40) and soluble CD40 ligand (sCD40L) in preeclamptic patients with renal damage and its relationship. Methods A total of 63 pregnant women attended the Department of Obstetrics, Affiliated Hospital of Qingdao University Medical College between August 2008 and June 2010. In the present study included 28 pregnant women with mild preeclampsia and 35 patients with severe preeclampsia. Thirty matched normotensive pregnant women were enrolled in the study as the control group. Expression of sCD40 and sCD40L were determined by ELISA. At the same time, the blood routine, C reaction protein ( CRP),urine routine, 24 hours urine protein excretion, and serum uric acid (UA), creatinine (Cr), blood urea nitrogen (BUN) were measured. The correlation analysis was performed between the sCD40/sCD40L and the blood biochemical indexes in 3 groups. Results ( 1 ) The median levels of CRP in severe preeclampsia (10. 8 mg/L)and mild preeclampsia group(7. I mg/L)are significantly higher than that of control group (3. 3 mg/L,P ＜ 0. 05 ); The level of CRP in severe preeclampsia group was also higher than that of mild preeclampsia group ( P ＜ 0. 05 ). The median gestational age at delivery in severe preeclampsia ( 32. 5 weeks)was significantly less than that of mild preeclampsia group ( 37. 2 weeks) and normal group ( 38. 6 weeks,P ＜ 0. 05). However no significant differences were observed between mild preeclampsia group and normal group ( P ＞0. 05 ). The platelet count in severe preeclampsia ( 132 × 109/L) was significantly less than those of mild preeclampsia group (212 × 109/L) and normal group ( 216 × 109/L, P ＜ 0. 01 ), but no significant differences were observed in blood platelet amount between mild preeclampsia group and normal group ( P ＞0. 05 ). There was no significant difference in hemoglobin level and white blood cell in three groups ( P ＞0. 05). (2) The sCD40 plasma concentration in severe, mild preeclampsia and normal group was 133.6,126. 5 and 90. 7 ng/L, respectively. The sCD40 L plasma concentrations were 12. 5, 10. 4 and 4. 4 ng/L respectively in the 3 groups. 24 hours urinary protein quantitative was 4. 5 g/d,0. 8 g/d and 0 in the 3 groups respectively. And the UA level was 486 μ mol/L,289 μmol/L and 162 μmol/L. In the above three groups,the monitoring indicators were significantly higher in women with severe preeclampsia group compared with mild preeclampsia and control groups (P ＜ 0. 01 ), and there were also higher in mild preeclampsia group than that in control groups ( P ＜ 0. 01 ). The level of plasma Cr ( 89 μmol/L) and BUN ( 5. 32 mmol/L) in severe preeclampsia group were higher than those of mild preeclampsia group (66 μmol/L and 4. 49mmol/L) and control group ( 57 μmol/L and 3.32 mmol/L, P ＜ 0. 05 ). There was no significant difference between mild preeclampsia group and normal group (P ＞ 0. 05 ). (3) The correlation analysis indicated that the level of sCD40 has a positive correlation with 24 hours urinary protein quantitative( r = 0. 434, P ＜ 0. 05 ),also significant positive correlation( r =0. 536,0. 528 ,P ＜ 0. 01 ) between the level of sCD40 and UA or CRP in women with preeclampsia. There was no significant correlation between the level of sCD40 and systolic blood pressure, diastolic blood pressure, delivery gestational age, Cr, BUN, and platelet count(r =0. 135,0. 183, -0. 133,0. 190,0. 167, -0. 221 ,all P ＞0. 05 ). There were positive correlation between the level of sCD40L and 24 hours urine protein excretion, either UA or CRP( r =0. 591,0. 445,0. 539 ,all P ＜0. 01 ). No significant correlation was found between sCD40 L and systolic blood pressure, diastolic blood pressure,delivery gestational age, Cr, BUN, and platelet count( r =0. 178,0. 212, -0. 292,0. 144,0. 135, -0. 273,all P ＞0. 05). There was significant positive correlation between plasma sCD40 and sCD40L ( r =0. 707 ,P ＜0. 01 ). There was no relationship between the level of sCD40, sCD40L and the blood biochemical indexes in normotensive pregnant women ( P ＞ 0. 05 ). Conclusions The plasma concentrations of sCD40 and sCD40 L are significantly higher in pregnant women with preeclampsia compared with the control, which may be involved in the development of preeclampsia and contribute to the kidney damage. The variance levels of sCD40 and sCD40L may be also related to the severity of preeclampsia.     

子痫前期胎盘中生长阻滞和DNA损伤45α基因及p38丝裂原活化蛋白激酶的表达

目的 探讨子痫前期胎盘组织中生长阻滞和DNA损伤45α(growth arrest and DNA damage-inducible 45 alpha,Gadd45α)基因和p38丝裂原活化蛋白激酶(p38 mitogen-activated protein kinase,p38 MAPK)信号分子的表达,及其与血清中可溶性血管内皮生长因子受体-1(soluble vascular endothelial growth factor receptor-1,sFlt-1)和可溶性endoglin(soluble endoglin,sEng)的相关性分析.方法 选择2009年9月至2010年3月在重庆医科大学附属第一医院住院分娩的孕妇54例为研究对象,按病情分为子痫前期轻度组20例、子痫前期重度组16例,以足月择期剖宫产孕妇18例为对照组.采用免疫组织化学SP法检测Gadd45α及磷酸化p38 MAPK(phospho-p38 MAPK,p-p38 MAPK)蛋白在各组胎盘组织中的定位;实时荧光定量PCR检测各组胎盘组织中Gadd45α mRNA水平;Western印迹法检测各组胎盘组织中Gadd45α蛋白的表达水平、p38 MAPK及p-p38 MAPK的蛋白表达差异;双抗体夹心酶联免疫吸附法检测各组孕妇血清样本中sFlt-1及sEng的含量,并进行单因素方差及LSD-t检验分析.结果 (1)免疫组织化学检测Gadd45α与p-p38 MAPK蛋白均定位于胎盘滋养层细胞胞质及胞核、血管内皮细胞胞核及少量间质细胞胞核.(2)子痫前期重度及轻度组Gadd45α mRNA相对表达水平(3.33±0.13和2.10±0.11)较正常对照组(1.01±0.18)明显上调,且子痫前期重度组高于轻度组,两两比较差异均有统计学意义(P＜0.05).(3)Western 印迹法检测正常对照组、子痫前期轻度组及重度组患者胎盘组织中Gadd45α蛋白表达水平分别为0.22±0.11、0.65±0.15、1.34±0.17;p-p38 MAPK蛋白的表达水平分别为0.32±0.08、0.72±0.12、1.45±0.21,两两比较差异均有统计学意义(P＜0.05);p38 MAPK蛋白水平组间比较差异无统计学意义(P＞0.05).(4)子痫前期轻度组、重度组孕妇血清sFlt-1、sEng水平明显高于正常对照组,且子痫前期重度组高于轻度组,两两比较差异均有统计学意义(P＜0.05).(5)各组Gadd45α蛋白水平与孕妇血清中的sFlt-1、sEng含量呈正相关(r分别为0.88和0.87,P均＜0.05).结论 Gadd45α在子痫前期患者胎盘组织中的表达明显升高,其可能通过调控p38 MAPK信号转导通路,诱使循环中的sFlt-1、sEng释放增加,从而加重胎盘血管重铸障碍和抑制滋养细胞浸润,参与子痫前期的发病.

Abstract：

Objective To evaluate the expression of Gadd45α and p38 MAPK in placentas and the correlations of Gadd45α protein and serum soluble vascular endothelial growth factor receptor-1 (sFlt-1) and soluble endoglin (sEng) in preeclampsia(PE). Methods Fifty-four pregnant women who delivered from September 2009 to March 2010 in the First Affiliated Hospital of Chongqing Medical University were chosen as the subjects. They were classified into mild preeclampsia group (n=20),severe preeclampsia group (n=16) and the control group (normal pregnant women underwent elective cesarean sections at term without labor and perinatal complications, n=18). Western blot and immunohistochemistry were employed to determine the expression and localization of Gadd45α and p-p38 MAPK protein respectively. Gadd45α mRNA level was determined by quantitative real-time PCR. The levels of seum sFlt-1 and sEng were measured by enzyme-linked immunosorbent assay (ELISA). One-way ANOVA and LSD-t test were applied for statistical analysis. Results (1)Immunohistochemistry identified that the positive stained cells were mostly located in trophoblast cells in normotensive placentas, whereas in preeclamptic placentas Gadd45α protein and p-p38 MAPK protein were detected in trophoblast and endothelial cells, as well as a few stromal cells at increased levels.(2)The mRNA levels of Gadd45α was significantly elevated in mild and severe preeclampsia groups compared to the control group (2.10±0.11 and 3.33±0.13 vs 1.01±0.18, P＜0.05), and Gadd45α mRNA level in severe group was significantly higher than in mild group (P＜0.05).(3)The data of Western blot revealed that the Gadd45α protein levels in each group were 0.22±0.11, 0.65±0.15 and 1.34±0.17, respectively, with significant differences between each group(P＜0.05). The p-p38 MAPK protein levels in each group were 0.32±0.08, 0.72±0.12 and 1.45±0.21, respectively, with significant differences between each group (P＜0.05). p38 MAPK protein levels in the total groups showed no difference(P＞0.05).(4)Compared with the control group, sFlt-1 and sEng concentrations in maternal circulation were significantly increasing in mild and severe preeclampsia groups, and concentrations in severe group were significantly higher than those in mild group (P＜0.05).(5) There were positive correlations between Gadd45α protein levels and the concentrations of serum sFlt-1 and sEng in each group( r=0.88 and 0.87, respectively all P＜0.05). Conclusions Upregulation of Gadd45α in preeclampsia placentas may play an important role in the pathogenesis of preeclampsia. It may induce the increased maternal serum levels of sFlt-l and sEng by activating p38 MAPK signaling pathway, leading to deficient cytotrophoblastic invasion and abnormal placental vascular reconstruction during pregnancy.     

p38丝裂原活化蛋白激酶信号分子在子痫前期患者血管内皮细胞损伤中的作用

目的 探讨p38丝裂原活化蛋白激酶(MAPK)信号分子在子痫前期患者血管内皮细胞损伤中的作用.方法选择2009年9月至2010年3月在重庆医科大学附属第一医院住院分娩的足月产妇54例为研究对象,其中36例为子痫前期患者,按病情分为子痫前期轻度组(20例)、子痫前期重度组(16例);同期足月择期剖宫产孕妇为对照组(18例).采用CD34标记血管内皮细胞,计数各组胎盘微血管密度(MVD);SP法检测磷酸化的p38 MAPK(p-p38 MAPK)在胎盘组织中的定位;蛋白印迹法检测各组胎盘p38 MAPK及p-p38 MAPK的蛋白表达差异;双抗体夹心ELISA法检测各组血清中可溶性血管内皮生长因子受体1(sFlt-1)及可溶性内皮糖蛋白(soluble endoglin,sEng)的水平.结果 (1)MVD:对照组、子痫前期轻度组及子痫前期重度组患者胎盘MVD分别为103±3、81±5、63±4,各组间两两比较,差异均有统计学意义(P＜0.05).(2)p38 MAPK和p-p38 MAPK蛋白表达水平:对照组、子痫前期轻度组及子痫前期重度组患者胎盘组织p38 MAPK蛋白表达水平分别为0.84±0.05、0.90±0.14、0.86±0.18,各组间两两比较,差异均无统计学意义(P＞0.05);p-p38 MAPK蛋白的表达水平分别为0.13±0.05、0.59±0.12和1.16±0.18,各组间两两比较差异均有统计学意义(P＜0.05).(3)p-p38 MAPK蛋白的表达部位:p-p38 MAPK蛋白主要表达于胎盘滋养层细胞的胞质及胞核、血管内皮细胞及少量间质细胞的胞核.(4)sFlt-1及sEng的水平:①对照组孕妇血清sFlt-1、sEng水平分别为(5.2±0.3)、(10.9-±0.4)μg/L;子痫前期轻度组分别为(12.5±1.2)、(20.4±5.3)μg/L;子痫前期重度组分别为(19.3±3.0)、(29.5±3.7)μg/L;各组sFlt-1、sEng水平两两比较,差异均有统计学意义(P＜0.05);②p-p38 MAPK蛋白表达水平与血清sFlt-1、sEng水平的相关性:子痫前期重度组和子痫前期轻度组p-p38 MAPK蛋白表达水平与血清sFlt-1、sEng水平均呈明显正相关(r=0.68,P＜0.05;r=0.87,P＜0.05).结论 子痫前期患者胎盘组织中p38 MAPK信号分子的显著活化,使血清中sFlt-l、sEng水平升高,损伤胎盘血管内皮细胞,导致胎盘MVD降低;p38 MAPK信号分子可能是子痫前期患者血管内皮细胞损伤的关键信号传导通路之一.

Abstract：

Objective To study expression and activation of p38 mitogen activated protein kinase (MAPK) in vascular endothelial cells dysfunction in preeclampsia. Methods From Sept. 2009 to Mar.2010, 54 pregnant women underwent deliveries in the First Affiliated Hospital of Chongqing Medical University were enrolled in this study, including 20 patients in mild preeclampsia group, 16 patients in severe preeclampsia group and 18 women with term cesarean section without perinatal complications as control group. Placental endothelial cells were labeled by CD34 to assay microvessel density (MVD) of each group. Immunohistochemical SP and western blot were used to detect localization and expression of p-p38 MAPK protein, respectively. The levels of sera soluble fms-like tyrosine kinase-1 (sFlt-1)and soluble endoglin(sEng) were measured by ELISA. Results ①The MVD of placenta were 103 ± 3 in control group, 81 ±5 in mild preeclampsia group and 63±4 in severe group, respectively, which showed statistical difference among each group (P＜0.05).②The expression of p38 MAPK protein were 0.84±0.05 in control group,0.90±0.14 in mild group and 0. 86 ±0.18 in severe group, which did not reach remarkable difference among each group (P＞0.05). The expression of p-p38 MAPK protein were 0.13±0.05 in control group,0.59±0.12 in mild group and 1.16±0.18 in severe group, which show statistical difference among each group(P＜0.05).(3) The localization of p-p38 was in trophoblast, endothelial cells and a few (5.2±0.3)and(10.9±0.4)μg/L in control group,(12.5±1.2) and (20.4±5.3)μg/L in mild group and (19.3±3.0) and (29. 5 ±3.7) μg/L in severe group. When drawing paired comparison in those p-p38 MAPK protein levels and the concentrations of serum sFlt-1, sEng in preeclampsia groups (r=0.68,P＜0.05;r=0.87,P＜0.05). Conclusions The remarkable activation of the p38 MAPK in the placenta of patients with preeclampsia induced the increased levels of sFlt-1 and sEng in maternal serum, which confer the injury of vascular endothelial cells that caused the significant decline of MVD in placentas. p38 MAPK signaling might be one of the key pathways in vascular endothelial cell dysfunction in preeclampsia.     

重度子痫前期患者血清作用下自然杀伤细胞对血管内皮细胞功能的影响

目的 研究在重度子痫前期患者血清作用下,自然杀伤(natural killer,NK)细胞对血管内皮细胞功能的影响.方法 2006年1月1日至2008年12月31日中国医科大学附属盛京医院产科重度子痫前期患者、正常妊娠妇女各15例,采用1∶1配比的病例一对照研究.20%重度子痫前期及正常妊娠者(对照组)血清孵育NK细胞20 h,加入人脐静脉血管内皮细胞培养体系中培养24 h.采用流式细胞仪进行凋亡检测,透射电镜下观察细胞形态,放射免疫方法检测NK细胞与人脐静脉血管内皮细胞培养上清液中内皮素-1水平.结果 重度子痫前期组早期凋亡细胞(Annexin V-FITC+/PI-)为(23.81±4.79)%,晚期凋亡细胞(Annexin V-FITC+/PI+)为(3.29±1.04)%,对照组分别为(16.59±5.13)%和(2.24±0.72)%,差异均有统计学意义(P＜0.01);组间坏死细胞(Annexin V-FITC-/PI+)差异无统计学意义.透射电镜重度子痫前期组可见核膜溶解、核碎裂等细胞凋亡表现,上清液中内皮素-1水平为(39.08±9.04) μg/ml,较正常妊娠组[(29.98±7.04) μg/ml]明显升高,差异有统计学意义(t=3.439,P=0.004).结论 重度子痫前期患者血清可活化NK细胞,导致血管内皮细胞凋亡增多,分泌缩血管活性物质——内皮素-1增多,进而导致高血压等临床症状.

Abstract：

Objective To investigate the effect of natural killer (NK) cells treated by serum of severe preeclampsia patient on the function of endothelial cells.Methods Fifteen patients with severe preeclampsia and 15 normal pregnant women from the Obstetrics department,Shengjing Hospital,China Medical University were admitted into this case-control study from January 1,2006 to December 31,2008.NK cells from healthy non-pregnant woman were incubated with 20% serum from severe preeclampsia patients or normal pregnant women for 20 hours.Then,human umbilical vein endothelial cells ( HUVEC) and serum-treated NK cells were co-incubated for 24 hours.Apoptosis of HUVEC was checked by flow cytometry and electronic microscope.Endothelin-1 (ET-1) levels in the supernatants of HUVEC and NK cells were examined by radioimmunologic method.Results In severe preeclampsia group,the percentage of early apoptosis cell (Annexin V-FITC+ /PI+ ) was (23.81±4.79)%,that of late apoptosis cell (AnnexinV-FITC+/PI+ ) was (3.29±1.04) %,while those were (16.59±5.13)% and (2.24±0.72)% respectively in normal pregnant group (P＜0.01).There was no significant difference in dead cells (Annexin V-FITC- /PI+ ).Under electronic microscope,typical morphologic changes of apoptosis were shown in severe preeclampsia group.Level of ET-1 in     

氧化低密度脂蛋白及其受体与子痫前期发病的关系

目的 探讨氧化低密度脂蛋白(oxLDL)及血凝集素样氧化低密度脂蛋白受体1(LOX-1)与子痫前期发病的关系.方法 选择2007年6月至2008年1月在青岛大学医学院附属医院产科住院分娩的子痫前期孕妇73例,其中轻度子痫前期孕妇35例(轻度子痫前期组),重度子痫前期孕妇38例(重度子痫前期组).选取同期正常晚期妊娠妇女45例为对照组.采用酶联免疫吸附试验检测各组孕妇血浆中oxLDL水平;采用RT-PCR、蛋白印迹法分别检测各组孕妇胎盘组织中LOX-1 mRNA及蛋白表达水平;采用RT-PCR检测各组孕妇胎盘组织中半胱氨酸天门冬氨酸蛋白酶-3(caspase-3)mRNA表达水平.结果 (1)轻度及重度子痫前期组孕妇血浆中oxLDL水平分别为(0.42±0.11)及(0.68±0.12)mg/L,明显高于对照组的(0.35±0.14)mg/L,差异有统计学意义(P<0.01);重度子痫前期组又明显高于轻度子痫前期组(P<0.01).(2)轻度及重度子痫前期组孕妇胎盘组织中LOX-1mRNA表达水平分别为0.70±0.10及0.84±0.08,明显高于对照组的0.58±0.11,差异有统计学意义(P<0.01);重度子痫前期组又明显高于轻度子痫前期组(P<0.01).(3)轻度及重度子痫前期组孕妇胎盘组织中LOX-1蛋白表达水平分别为0.79±0.15及0.90±0.12,明显高于对照组的0.68±0.11,差异有统计学意义(P<0.01);重度子痫前期组又明显高于轻度子痫前期组(P<0.01).(4)轻度及重度子痫前期组孕妇胎盘组织中caspase-3 mRNA表达水平分别为3.82±0.18及5.39±0.14,明显高于对照组的2.19±0.20,差异有统计学意义(P<0.01);重度子痫前期组又明显高于轻度子痫前期组(P<0.01).(5)轻度及重度子痫前期组孕妇血浆中oxLDL水平与胎盘组织中LOX-1 mRNA表达水平呈正相关(r=0.93,P<0.05);胎盘组织中LOX-1 mRNA表达与胎盘组织caspase-3 mRNA表达水平呈正相关(r=0.84,P<0.05).结论 子痫前期孕妇血浆中oxLDL水平升高,并上调胎盘组织中的LOX-1表达水平,从而参与子痫前期的病理生理过程.     

重度子痫前期患者血清作用下自然杀伤细胞对血管内皮细胞功能的影响

Objective To investigate the effect of natural killer (NK) cells treated by serum of severe preeclampsia patient on the function of endothelial cells.Methods Fifteen patients with severe preeclampsia and 15 normal pregnant women from the Obstetrics department,Shengjing Hospital,China Medical University were admitted into this case-control study from January 1,2006 to December 31,2008.NK cells from healthy non-pregnant woman were incubated with 20% serum from severe preeclampsia patients or normal pregnant women for 20 hours.Then,human umbilical vein endothelial cells ( HUVEC) and serum-treated NK cells were co-incubated for 24 hours.Apoptosis of HUVEC was checked by flow cytometry and electronic microscope.Endothelin-1 (ET-1) levels in the supernatants of HUVEC and NK cells were examined by radioimmunologic method.Results In severe preeclampsia group,the percentage of early apoptosis cell (Annexin V-FITC+ /PI+ ) was (23.81±4.79)%,that of late apoptosis cell (AnnexinV-FITC+/PI+ ) was (3.29±1.04) %,while those were (16.59±5.13)% and (2.24±0.72)% respectively in normal pregnant group (P＜0.01).There was no significant difference in dead cells (Annexin V-FITC- /PI+ ).Under electronic microscope,typical morphologic changes of apoptosis were shown in severe preeclampsia group.Level of ET-1 in     

重度子(癎)前期和正常妊娠胎盘中微小RNA-155及富含半胱氨酸蛋白61的差异表达

目的 探讨微小RNA-155(microRNA-155,miR-155)和富含半胱氨酸蛋白61(cysteine-rich 61,CYR61)在重度子(癎)前期和正常妊娠胎盘中差异表达的意义. 方法取18例重度子(癎)前期患者(孕36～40周)胎盘和同期分娩且孕周匹配的18例正常产妇胎盘,从mRNA水平检测miR-155和CYR61 mRNA的表达情况,并在蛋白水平检测CYR61的表达情况. 结果 与正常产妇胎盘相比,重度子(癎)前期组胎盘的miR-155表达水平明显增高(165.7±16.4和527.9±49.1,t=7.00,P＜0.01);而CYR61 mRNA表达水平及蛋白表达水平均降低(31.7±2.7和16.4±1.2,t=5.10,P＜0.01;36.4±1.5和19.7±1.2,t=36.26,P＜0.01).重度子(癎)前期和正常妊娠胎盘组织中的miR-155与CYR61 mRNA的检测结果的相关性分析显示,两组内两者表达均呈负相关(r=-0.52,P＜0.05;r=-0.57,P＜0.05). 结论重度子(癎)前期患者胎盘中miR-155的上调,可能与促血管生成因子-CYR61的表达下降有关.miR-155、CYR61可能共同参与了子(癎)前期胎盘血管缺陷的发生.     

巨噬细胞移动抑制因子与受体CD_(74)在胎盘组织中的表达及其与子痫前期发病的关系

目的 探讨巨噬细胞移动抑制因子(MIF)与受体CD_(74)在胎盘组织中的表达及其与子痫前期发病的关系.方法 选取2008年3-11月在青岛大学医学院附属医院产科住院分娩的子痫前期孕产妇69例,其中轻度子痫前期孕产妇33例(轻度子痫前期组),重度子痫前期孕产妇36例(重度子痫前期组).另选取同期正常妊娠晚期孕产妇43例为对照组.采用免疫比浊法榆测3组孕产妇血中C反应蛋白水平;采用免疫组化链酶菌抗生物素蛋白-过氧化物酶连接(SP)法榆测3组孕产妇胎枯组织中的MIF和CD_(74)的表达;采用逆转录(RT)PCR技术检测3组孕产妇胎盘组织中MIFmRNA和CD_(74)mRNA的表达.并对轻度及重度子痫前期组孕产妇血中C反应蛋白水平与胎盘组织中MIF mRNA及CD_(74) mRNA表达的相关性进行分析.结果 (1)3组孕产妇胎盘组织中均有MIF和CD_(74)的表达,并呈棕黄色染色,轻度及莆度子痫前期组孕产妇胎盘组织中MIF及CD_(74)的表达明显增强.(2)轻度子痫前期组孕产妇胎盘组织中MIF mRNA及CD_(74)mRNA表达水平分别为0.70±0.13及0.96±0.16;重度子痫前期组分别为0.88±0.12及1.08±0.15,两组比较,差异有统计学意义(P<0.01).对照组孕产妇胎盘组织中MIF mRNA及CD_(74)mRNA表达水平分别为0.67±0.11及0.83±0.14,明显低于轻度及重度子痫前期组,分别比较,差异均有统计学意义(P<0.01).(3)轻度及重度子痫前期组孕产妇血中C反应蛋白水平分别为(15.3±7.0)及(21.6±9.1)mg/L,两组比较,差异有统计学意义(P<0.01);对照组为(4.8±1.8)mg/L,明显低于轻度及重度子痫前期组,分别比较,差异均有统计学意义(P<0.01).(4)轻度及重度子痫前期组孕产妇血中C反应蛋白水平与胎盘组织中MIF mRNA表达呈正相关关系(r=0.67,P<0.01);与胎盘组织中CD_(74)mRNA表达呈正相关火系(r=0.83,P<0.01).胎盘组织中MIF mRNA表达与CD_(74)mRNA表达呈正相关关系(r=0.93,P<0.01).结论 子痫前期患者胎盘组织中的MIF及其受体CD_(74)的过度表达,可上调血中炎性标志物C反应蛋白的水平,引起血管内皮损伤,从而参与子痫前期的发病.     

子(癎)前期孕妇胎盘硫氧还蛋白系统的表达及其意义

目的 研究子(癎)前期孕妇胎盘硫氧还蛋白还原酶(thioredoxin reductase,TR)、硫氧还蛋白(thioredoxin,Trx)mRNA的表达情况和胎盘TR的活性,探讨Trx系统与子(癎)前期发病的关系.方法 用半定量逆转录-聚合酶链反应检测26例子(癎)前期孕妇(10例轻度子(癎)前期,16例重度子(癎)前期)和28例正常妊娠妇女(对照组)胎盘TR、Trx mRNA表达的情况.采用紫外分光光度计测定胎盘TR活性.结果 (1)子(癎)前期组胎盘TR mRNA(轻度:0.865±0.018,重度0.800±0.025)和Trx mRNA表达(轻度:0.873±0.008,重度0.818±0.025)均低于对照组(TR:0.893±0.023,Trx:0.918±0.023;P均＜0.05),重度子(癎)前期组的表达水平低于轻度子(癎)前期组(P均＜0.05);(2)子(癎)前期组胎盘TR活性分别低于对照组(轻度:17.732±1.653,重度:15.626±1.543,对照:19.770±2.432;P＜0.05),重度子(癎)前期组低于轻度子(癎)前期组(P＜0.05);(3)两组胎盘TR活性和TR mRNA表达呈正相关(r=0.612,P＜0.001).结论 Trx系统的表达异常可能与子(癎)前期的发病有关,并有重要作用.     

人类白细胞抗原G的表达与子痫前期发病的关系

目的 通过检测子痫前期孕妇相关组织中人类白细胞抗原G (HLA-G)的表达,探讨其与子痫前期发病的关系.方法 选择2009年3月至12月在陕西省妇幼保健院产科住院分娩的子痫前期孕妇30例,根据病情分为轻度子痫前期组8例,重度子痫前期组22例.选择同期健康孕妇30例为对照组.3组孕妇均以剖宫产方式分娩.采用ELISA法检测孕妇外周血、脐血及羊水中的可溶性HLA-G(sHLA-G)水平;采用蛋白印迹法检测胎盘、胎膜及脐带组织中HLA-G蛋白的表达.结果 (1)各组孕妇外周血、脐血、羊水中sHLA-G水平:轻、重度子痫前期组孕妇外周血sHLA-G水平分别为(50±14)、(30±6) μg/L,新生儿脐血中sHLA-G水平分别为(34±10)、( 26±8)μg/L,均明显低于对照组的(100±16)、(70±9) μg/L,分别与对照组比较,差异均有统计学意义(P＜0.01);重度子痫前期组与轻度子痫前期组比较,差异也有统计学意义(P＜0.01).重度子痫前期组新生儿脐血中sHLA-G水平虽低于轻度子痫前期组,但差异无统计学意义(P＞0.05).轻、重度子痫前期组孕妇羊水中sHLA-G水平分别为(26±7)、( 25±5) μg/L,均低于对照组的(27±6)μg/L,但分别与对照组比较,差异无统计学意义(P＞0.05);轻度子痫前期组与重度子痫前期组比较,差异也无统计学意义(P＞0.05).(2)各组孕妇胎盘、胎膜及脐带组织中的HLA-G蛋白表达:对照组胎盘组织中HLA-G蛋白表达水平为1.59±0.36,胎膜组织中表达水平为0.42±0.09,胎盘组织中的HLA-G蛋白表达水平明显高于胎膜组织,两者比较,差异有统计学意义(P＜0.05);对照组脐带组织中HLA-G蛋白表达水平为0.24±0.17,分别与胎盘及胎膜组织中的HLA-G蛋白表达水平比较,差异均有统计学意义(P＜0.01).轻度子痫前期组胎盘组织中HLA-G蛋白表达水平为0.78±0.21,重度子痫前期组为0.29±0.17,分别与对照组比较,差异均有统计学意义(P＜0.01).轻、重度子痫前期组孕妇胎膜及脐带组织中均未检测出HLA-G蛋白的表达.结论 与健康孕妇相比,HLA-G在子痫前期孕妇外周血、脐血及胎盘组织中呈明显低表达,HLA-G表达水平的异常可能与子痫前期的发病有关.     

子痫前期孕妇胎盘组织中生长阻滞和DNA损伤45α基因的表达变化及其临床意义

目的 通过检测子痫前期孕妇胎盘组织中生长阻滞和DNA损伤45α(Gadd45α)基因的定位及表达变化,探讨其与子痫前期发病的关系.方法 选择2009年9月至2010年3月在重庆医科大学附属第一医院住院分娩的36例子痫前期孕妇为研究对象,按病情分为子痫前期轻度组(20例),子痫前期重度组(16例),另以同期择期行足月剖宫产术的18例健康孕妇为对照组.3组孕妇分娩后取胎盘组织标本,采用免疫组化链霉菌抗生物素蛋白-过氧化物酶连接(SP)法检测各组孕妇胎盘组织中Gadd45α蛋白的定位及分布,采用逆转录(RT)-PCR技术检测各组孕妇胎盘组织中Gadd45α mRNA表达水平,采用蛋白印迹(western blot)技术检测各组孕妇胎盘组织中Gadd45α蛋白的表达水平.结果 (1)子痫前期轻度组及重度组胎盘组织中均有Gadd45α蛋白表达,主要定位于胎盘滋养层细胞的胞质及胞核中,血管内皮细胞及少量间质细胞胞核也可见阳性染色;对照组胎盘组织中阳性染色较弱,且仅见于胎盘滋养层细胞中,血管内皮细胞中未见阳性染色.(2)子痫前期重度组及轻度组胎盘组织中Gadd45αmRNA表达水平分别为0.75±0.07及0.44±0.13,明显高于对照组的0.18±0.04,分别比较,差异均有统计学意义(P＜0.05);子痫前期重度组Gadd45α mRNA表达水平与子痫前期轻度组比较,差异也有统计学意义(P＜0.05).(3)子痫前期重度组及轻度组胎盘组织中Gadd45α蛋白表达水平分别为1.34±0.17及0.65±0.15,两组比较,差异有统计学意义(P＜0.05);两组也明显高于对照组的0.22±0.11,分别比较,差异均有统计学意义(P＜0.05).结论 子痫前期患者胎盘组织中Gadd45α mRNA和蛋白表达水平均明显高于健康孕妇,并且随子痫前期患者病情加重其表达上调;Gadd45α定位在与子痫前期发生密切相关的胎盘滋养层细胞中,Gadd45α基因可能参与了子痫前期的发生与发展.     

子痫前期患者血浆对人脐静脉内皮细胞生长活性及Edg4表达的影响

目的:探讨子痫前期患者血浆对人脐静脉内皮细胞(HUVECs)生长活性的影响以及与溶血磷脂酸受体蛋白内皮分化基因-4(Edg4)的关系。方法:体外培养HU-VECs,取对数生长的细胞,分组实验:对照组,正常晚孕组,轻度子痫前期组,重度子痫前期组。用MTT比色法和流式细胞仪检测各组细胞增殖及凋亡情况,免疫组化链霉菌抗生物素蛋白-过氧化物酶(SP)法和逆转录聚合酶链反应(RT-PCR)检测各组细胞Edg4蛋白及mRNA表达水平。结果:(1)子痫前期组HUVECs增殖减少,细胞凋亡增加;各组间细胞增殖率及细胞凋亡率差异显著(P<0.05);(2)Edg4蛋白主要表达于HUVECs的细胞浆和细胞膜,子痫前期病情越重,其血浆诱导的HUVECs Edg4蛋白表达越强;(3)子痫前期组Edg4 mRNA表达明显升高,与对照组和正常晚孕组相比显著差异(P<0.05);重度子痫前期组Edg4 mRNA表达较轻度组显著升高(P<0.05)。结论:子痫前期患者血浆可诱导HUVECs Edg4蛋白表达增强,并抑制HUVECs细胞增殖,促进细胞凋亡,提示子痫前期患者溶血磷脂酸升高可能与血管内皮细胞功能异常有关。