Apoptosis in an Ovine Model of Intrauterine Growth Restriction (IUGR)

胎儿宫内生长受限(LUGR)是导致婴儿死亡率和发病率增加的重要原因。一个 在LUGR期间已经描述了许多胎盘异常，包括滋养层细胞的增加 细胞凋亡。滋养层细胞凋亡异常可能导致氧和营养交换受损，提示 细胞凋亡在LUGR发生发展中的作用为了更好地了解LUGR妊娠中的细胞凋亡，我们 将利用一种已建立的高温症绵羊LUGR模型，该模型在很大程度上模拟了人类疾病。 我们建议观察妊娠早期绵羊胎盘的细胞凋亡以及暴露于 热疗20天，胎盘生长高峰期55天，胎儿近足月80天 增长是最大的。我们的长期目标是：1)确定体内胎盘细胞凋亡效应，DNA 我们的高温模型中的降解、增殖率、端粒酶活性和细胞角蛋白18的裂解 诱导绵羊PI-IUGR，2)阐明抗凋亡分子bcl-2和XIAP的表达及 促凋亡分子Bax、caspase 3和caspase 9在PI-IUGR绵羊胎盘中的表达。要确定 HSP27和HIF-2a在高温诱导PI-IUGR细胞凋亡过程中的相关性及表达 3)确定高温、低氧或两者兼而有之对细胞凋亡、细胞角蛋白18裂解、 培养细胞端粒酶活性及bcl2、xIAP、bax、caspase3和9、HSP27、HIF-2a的表达 从正常绵羊胎盘中获得胎盘双核细胞(BNC)。为了实现这些目标，我们 将使用末端脱氧核苷酸转移酶介导的dUTP缺口末端标记来确定细胞凋亡 技术(TUNEL)和细胞角蛋白18裂解表达。此外，我们还将测定端粒酶活性 与我们使用端粒酶重复扩增协议(TRAP)技术的lUGR模型相关联。 为了进一步研究细胞凋亡，我们将测定Bcl2、Bax以及caspase3和9的蛋白激活水平 在高温处理的动物胎盘中，与对照组相比。此外，我们还将确定 高温对胎盘XIAP和HIF-2a表达的影响学习 体外细胞凋亡我们将确定高温和低氧对培养的胎盘BNCs的影响。

Intrauterine growth restriction (lUGR) is a significant cause of increased infant mortality and morbidity. A number of placental abnormalities have been described during lUGR including an increase in trophoblast apoptosis. Abnormal trophoblast apoptosis may lead to impaired oxygen and nutrient exchange, suggesting a role for apoptosis in the development of lUGR. To better understand apoptosis in lUGR pregnancies we will utilize an established hyperthermia sheep lUGR model which mimics the human disease considerably. We propose to look at apoptosis in the sheep placentae early in gestation and in sheep exposed to hyperthermia for 20 days, 55 days when placental growth is at its peak and 80 days near term when fetal growth is maximal. Our long-term goals are: 1) to determine the in-vivo placental apoptotic effects, DNA degradation, proliferation rate, telomerase activity and cytokeratin 18 cleavage in our model of hyperthermia induced PI-IUGR in the sheep, 2) to elucidate the expression anti-apoptotic molecules BCL-2 and XIAP and the pro-apoptotic molecules BAX, caspase 3 and caspase 9 in the sheep PI-IUGR placentae. To determine the correlation and expression of HSP27 and of HIF-2a during apoptosis in hyperthermia induced PI-IUGR, and 3) to determine the effect of hyperthermia, hypoxia or both on apoptosis, cytokeratin 18 cleavage, telomerase activity and expression of BCL-2, XIAP, BAX, caspase 3 and 9, HSP27 and HIF-2a in cultured placental binucleated cells (BNC) obtained from normal sheep placentomes. In order to accomplish these we will determine apoptosis using the terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling technique (TUNEL) and cytokeratin 18 cleavage expression. Also we will determine the telomerase activity associated with our model of lUGR using the Telomerase Repeat Amplification Protocol (TRAP) technique. To further study apoptosis, we will determine the protein levels of Bcl-2, Bax, and caspase 3 and 9 activation in placentae from animals treated with hyperthermia as compared to controls. Also, we will determine the effects of hyperthermia in the expression of XIAP and HIF-2a in the placentae from treated animals. To study apoptosis in-vitro we will determine the effects of hyperthermia and hypoxia on cultured placental BNCs.