RhoA/Rock2 And Preeclampsia

PartⅠThe expression of RhoA and Rock2 in placenta of preeclampsia and normal pregnancyBackgound: Preeclampsia is a pregnancy-specific syndrome in late trimester of pregnancy. It threatens maternal and fetal health. The etiology and pathogenesy have not yet elucidated. However, The fact that the disease fully recover after delivery indicates placental mainly effect in etiology. RhoA is an important member of Rho family in small GTPases. Rho associated coiled coil forming protein kinase(Rock)belongs to Serine/Threonine kinases family. It is downstream effector of Rho that studied most distinctly at present. It involves in cytolergy in many ways, and plays key roles in essential aspects such as migration, proliferation, survival, et al. We suppose RhoA/Rock2 may be involved in develpoment of preeclampsia.Objection:To determine the localization and expression of RhoA and Rock2 in placenta of normal pregnancy and preeeclampsia. To evaluate the role of RhoA/Rock2 in preeclampsia.Methods: Fifteen 1st trimester placental specimens between 8 and 10 weeks'gestation, twenty 3rd trimester placentas between 38 and 40 weeks'gestation and forty preeclampsia plcentas were collected following termination of pregnancy. Immunohistochemistry was employed to locate the expression of RhoA and Rock2 in three group placenta. RhoA and Rock2 mRNA were detected by reverse transcript polymerase chain reaction. The quantity of the expression of RhoA and Rock2 protein was determined by Western blot analysis. Results: The expression of RhoA was more prominent in cytoplasm of trophoblast cells. The expression of Rock2 was in cytoplasm and nucleus of trophoblast cells. In early pregnancy, there was seldom staining of RhoA and Rock2 in other position besides cytotrophoblast. The level of of RhoA and Rock2 protein in early pregnancy were higher than those from normal 3rd trimester pregnancies (P﹤0.05). In the 3rd trimester placenta, there was a little staining in stromal. RhoA and Rock2 mRNA and protein in mild preeclampsia group and severe preeclampsia group were significantly higher than those from normal pregnancies(P<0.05). The severe preeclampsia group was higher than those from mild preeclampsia group, the difference was not significance(P>0.05).Conclusion: The expression change of RhoA and Rock2 during placentation may be involved in invasion by trophoblast, hypoxia reaction, vasomotor function, et al. RhoA and Rock2 may participate in normal placentation and a series of adaptive changes. The abnormal expression may be associated with pathological change of preeclampsia such as placenta shallow invasion, arteriospasm, endothelial dysfunction.PartⅡThe expression of RhoA and Rock2 in human umbilical artery smooth muscle cells of preeclampsia and normal pregnancyBackgound:The successful pregnancy dependes on the function and adaptability of cardiovascular system . As all know, preeclampsia is a syndrome charactered by peripheral vascular smooth muscle spasm and increased blood pressure. More and more researches discovered RhoA/Rock pathway playing an important role in various disorders development especially in cardiovascular system, such as hypertention, atherosclerosis and coronary artery spasm. Despite the pathogenesy of preeclampsia is different from essential hypertension, the basal pathological changes like vasospam, increased peripheral vascular resistance are similarity. The expression of RhoA/Rock2 in vascular of preeclampsia has not been evaluated yet.Objection: To determine the expression of RhoA/Rock2 protein and mRNA in human umbilical artery smooth muscle cells of normal pregnancy and preeeclampsia. To evaluate the role of RhoA/Rock2 in vascular lesion of preeclampsia.Methods: Four ortho-arteriotony 3st trimester umbilical core specimens and six preeclampsia umbilical core were collected following termination of pregnancy. Primary umbilical artery vascular smooth muscle cells were cultured. RhoA and Rock2 mRNA were detected by reverse transcript polymerase chain reaction. The quantity expression of RhoA and Rock2 protein was determined by Western blot analysis.Results: Primary umbilical artery vascular smooth muscle cells were charactered by typical"peak and valley". Immunocytochemical stain by antibody againstα-actin display silkiness and fog staining, parallelled to cell long axis. The purity of cells exceeded 98%. RhoA and Rock2 mRNA and protein in preeclampsia group were significantly higher than those from normal pregnancies(P﹤0.05).Conclusion: The upregulated expression of RhoA and Rock2 in umbilical artery vascular smooth muscle cells from preeclampsia may be associated with vascular lesion. Inhibiting RhoA /Rock2 pathway may improve the vascular lesion of preeclampsia. PartⅢThe RhoA / Rock pathway mediates hypoxia reaction in human umbilical vein endothelial cellsBackgound: The theory of endothelial dysfunction is one of the pathogenesis of preeclampsia. It is thought to be pathological foundation of impaired multisystem and carrier that pathological changes transport from placenta to all over the body. Hypoxia is one of the reason of endothelial dysfunction. HIF-1αis to mediates cellular adaptations by facilitating increased oxygen delivery, decreasing oxygen consumption and/or adapting cellular metabolic activities. HIF-1αhas been known to regulate the transcription of more than twenty genes in reponse to hypoxia, including endothelin-1, leptin, TNF-αand IL-1 et al. The abnormal expression of these genes may be associated with preeclampsia by influencing vascular tone and the function of trophoblast and endothelial cell. Little is known about the regulatory mechanisms of HIF-1αexpression in cells. Resent researches discover RhoA/Rock pathway involved in various aspects of cell biology. Whether the pathway mediates hypoxia reaction in human umbilical vein endothelial cells should be evaluated.Objection: To investigate the expression of RhoA,HIF-1αand NO in conditions of low oxygen. To evaluate the role of RhoA/Rock in regulation hypoxia reaction in human umbilical vein endothelial cells .Methods: To induce umbilical vein endothelial cell line ECV304 hypoxia by chemical method. To investigate the expression of RhoA and HIF-1αby reverse transcript polymerase chain reaction and Western blot analysis. The expression of NO in supernate fluid was investigated by nitrate reductase method. To evaluate the varied expression when cell line was given Rock inhibitor.Results: In conditions of low oxygen, gene transcripts and protein of RhoA and HIF-1α were found to up regulate significantly (P﹤0.05), the expression of NO was significantly lower(P﹤0.05). In uniform conditions of low oxygen but preincubated by Rock inhibitor, gene transcripts and protein of RhoA and HIF-1α, and the expression of NO were found no variation comparising with normoxia condition (P>0.05).Conclusion: Hypoxia regulated the expression of HIF-1αand NO though activating RhoA/Rock passway. Inhibiting this passway may reverse hypoxia reaction. This suggested Rock inhibitor may down regulate the expression of several genes in reponse to hypoxia induced by placental shallow invasion in preeclampsia. This pathway is considered a potential target for preeclampsia.