Abnormality of proliferation and differentiation of the trophoblasts in preeclampsia -Correlation between the cell cycle and oxidative stress-

异常妊娠滋养细胞细胞周期相关蛋白的表达分析获得葡萄胎、早期正常妊娠、早孕和足月妊娠的胎盘绒毛组织，并提取蛋白质和常规石蜡包埋组织。在正常滋养层绒毛细胞和葡萄胎组织中，免疫印迹分析证实细胞周期相关蛋白如增殖细胞核抗原、细胞周期蛋白CDK2、p21、p27、p57和细胞周期蛋白E的表达。在葡萄胎和早孕滋养细胞中，PCNA、CDK2和Cyclin E蛋白均呈强阳性表达。随着妊娠晚期，这些蛋白的表达逐渐减弱。P21在早孕正常滋养细胞中高表达，在妊娠晚期恶化，在葡萄胎中的表达强于早孕滋养细胞。至于p57，它在…中表达葡萄胎多为正常胎盘，未见异常表现。P27在正常早孕滋养层细胞中表达增强，晚期表达减弱，在葡萄胎中的表达强于妊娠晚期滋养细胞。用免疫组织化学方法研究p21、p27、p57在葡萄胎中的定位，并与正常妊娠滋养细胞进行比较。分析氧化应激对妊娠滋养细胞组织的影响。取足月妊娠和早产时的胎盘，提取RNA、蛋白质，部分组织包埋于石蜡包埋。免疫组织化学染色和Western印迹分析检测氧化应激所产生的过氧化脂质分解产物4-羟基-2-壬烯醛(HNE)的积聚。在子痫前期患者的滋养层组织中，1-INE的蓄积显著高于无并发症的滋养层组织。同样，绒毛膜羊水滋养层组织中HNE的积聚也非常显著。我们在培养的滋养层组织中添加HNE，并通过RT-PCR和Western印迹分析检测COX-2的表达。COX-2在转录和翻译水平均有表达，且随时间和浓度的增加而增加。前列腺素的产生也以时间依赖的方式增加。这些结果表明，滋养层组织在氧化应激时产生过氧化脂质，分解代谢产物增多，激活前列腺素合成系统的级联反应。较少

Manifestation analysis of a cell cycle related proteins of trophoblasts in abnormal pregnancyWe obtained chorionic tissues of molar pregnancy, early normal pregnancy, preterm pregnancy andfull-term pregnancy with informed consents at delivery, and we extracted protein and embedded tissues in conventional paraffin. In normal trophoblastic villous cells of each trimester of pregnancy and hydatidiform mole, we confirmed manifestation of cell cycle related proteins such as PCNA, CDK2, p21, p27, p57 and Cyclin Eby Western blot analysis. PCNA, CDK2 and Cyclin Eproteins were strongly expressed in hydatidiform mole and early gestational trophoblasts at the same level. With the late gestation period, the expression of these proteins was diminished. As for p21 which we showed high manifestation in normal trophoblasts of early pregnancy, and deteriorated in the latter period, its stronger expression was found in hydatidiform mole than early pregnancy trophoblasts. As for p57 which is expressed in … More the normal placenta, the manifestation was not found in hydatidiform mole. As for p27 which manifestation enhanced in normal trophoblasts at the early trimester and getting weaker in the latter period, strong manifestation was found in hydatidiform mole than a late gestational trophoblasts. We investigated the localization of p21, p27, p57 in hydatidiform mole by immunohistochemical analysis and compared its localization with the normal gestational trophoblasts.Analysis of influence of oxidative stress in gestational trophoblastic tissueWe obtained placentae at the time of a full-term pregnancy and preterm labor and extracted RNA, protein, and encapsulated a part of these tissues in paraffin. Accumulation of 4hydroxy-2nonenal (HNE), the catabolite of peroxide lipid produced by oxidative stress was investigated by Immunohistochemical staining and Western blot analysis. As for the trophoblastic tissue obtained from the patient suffering from preeclampsia, accumulationof 1-INEwas remarkable in comparison with trophoblastic tissue without any complication. In the same manner, accumulation of HNE was significantly remarkable in trophoblastic tissue with chorioamnionits. We supplemented HNE in cultured trophoblastic tissue and investigated the expression of cyclooxygenase-2 (COX-2) by RT-PCR and Western blot analysis. COX-2 was expressed at transcriptional and translational level at increased for level at time and concentration dependent manner. Production of Prostaglandin also increased at the time dependent manner. These results indicated that trophoblastic tissue caught oxidative stress produced lipid peroxide catabolite increased and activate the cascade of prostaglandin synthetic system. Less