| Preeclampsia is a disease which occures only in pregnancy. Epidemiological survey of its incidence in China is about 9.4% -10.4%, and foreign reports are 7% -12%. Although a lot of researches were done on the pathogenesis of PE, the mechanism remains largely unknown. And it cause maternal and child morbidity and mortality.Annexins is a very ancient and widespread protein, which is a calcium-dependent manner to acidic phospholipid binding proteins. A large number of studies have shown that annexins play a role in a variety of physiological processes, including endocytosis, exocytosis, cells and extracellular matrix interactions, formation of ion channels. Some diseases is associated with the expression and the activity of annexins. In the annexin family, annexin A2 and annexin A5 are mainly related to the coagulation and fibrinolysis systerm. For a long time, the study shows that annexin A2 and annexin A5 are closely related to the adverse pregnancy outcomes, such as recurrent miscarriage, antiphospholipid syndrome. But the relationship of PE and annexins are rare. Part one The growth and apoptosis of trophoblast cells in hypoxia and pathological test.Objective: To investigate the growth and apoptosis of trophoblast cells in hypoxia.Methods: Detect the growth of trophoblast cells in hypoxia in vitro. Trophoblast apoptosis in hypoxic in vivo were detceted by flow cytometry. Also placental pathology was detected by immunohistochemistry.Results:1 Trophoblast cells in hypoxia in vitro was found that with prolonged hypoxia, trophoblast growth restriction, decreased protein levels.2 In hypoxia, the apoptosis of trophoblast cell, especially early apoptosis increased: early-onset mild preeclampsia group was 3.21%, early-onset severe preeclampsia group was 5.32%, late-onset mild preeclampsia group was 2.43%, late-onset severe preeclampsia group was 4.28%, and the healthy pregnant group was 1.05%.3 Compared with normal pregnancy, the formation of microthrombi can be seen in placenta of preeclampsia with disease severity.Conclusion: In hypoxia, the JAR cell growth was restriction in vitro, the apoptosis of trophoblastic in vivo (placental trophoblast cells) was significantly increased, especially the early apoptosis. And microthrombosis was found in the placenta of PE. Part two The expression of annexin A5 in patients of preeclampsiaObjective: To investigate the annexinA5 in maternal blood and placental tissue and its relationship with the pathogenesis of preeclampsia.Methods: 120 preeclampsia and 30 matched normal pregnant controls were included in the study. Expression of annexin A5 mRNA in placental tissues was analyzed using quantitative real time RT-PCR. Annexin A5 protein in placentas as well as blood was determined by western blot analysis and immunohistochemistry.Results: The expression levels of annexin A5 mRNA were significantly decreased in preeclampsia compared to normal pregnancies. The levels of annexin A5 protein in placentas were also significantly reduced in patients with preeclampsia, compared with healthy pregnant women.1 Annexin A5 mainly expressed in the surface of trophoblast cell. Compared with normal pregnancy, the expression of annexin A5 in placental trophoblast of preeclampsia cell surface is reduced.2 The expression level of annexin A5 in placental tissue and peripheral blood. Placental tissue and peripheral blood levels of annexin A5: early-onset mild preeclampsia group are 0.54±0.12 and 0.62±0.17, early-onset severe preeclampsia group are 0.47±0.15 and 0.56±0.24, late-onset mild preeclampsia group are 0.74±0.23 and 1.08±0.32, late-onset severe preeclampsia group are 0.68±0.28 and 0.72±0.21, and healthy pregnant group are 1.73±0.35 and 1.55±0.27.3 The mRNA levels of annexin A5 by RT-PCR in placental tissue: early-onset mild preeclampsia group is 25.0±3.04, early-onset severe preeclampsia group is 24.8±2.96, late-onset mild preeclampsia group is 25.4±3.92, late-onset severe preeclampsia group is 25.1±2.74, and healthy pregnancy group is 30.6±2.68.Conclusion: Annexin A5 mainly expresses on the surface of trophoblast cell. Compared with normal pregnancy, annexin A5 in placental of preeclampsia was decreased, which affect placental blood coagulation - fibrinolytic system. It may play an important role in the occurrence and the development of preeclampsia. Part three The expression of annexin A2 in patients of preeclampsiaObjective: To investigate the annexinA2 in maternal blood and placental tissue and its relationship with the pathogenesis of preeclampsia.Methods: 120 preeclampsia and 30 matched normal pregnant controls were included in the study. Expression of annexin A2 mRNA in placental tissues was analyzed using quantitative real time RT-PCR. Annexin A2 protein in placentas as well as blood was determined by western blot analysis and immunohistochemistry.Results: The expression levels of annexin A2 mRNA were significantly decreased in preeclampsia compared to normal pregnancies. The levels of annexin A2 protein in placentas were also significantly reduced in patients with preeclampsia, compared with healthy pregnant women.1 Annexin A2 mainly expressed in the surface of trophoblast cell. Compared with normal pregnancy, the expression of annexin A2 in placental trophoblast of preeclampsia cell surface is reduced.2 The expression level of annexin A2 in placental tissue and peripheral blood. Placental tissue and peripheral blood levels of annexin A2: early-onset mild preeclampsia group were 0.77±0.15 and 0.63±0.09, early-onset severe preeclampsia group were 0.76±0.19 and 0.66±0.13, late-onset mild preeclampsia group were 0.74±0.23 and 0.71±0.23, late-onset severe preeclampsia group were 0.81±0.35 and 0.59±0.14, and healthy pregnant group were 1.82±0.18 and 0.50±0.04. Compared with normal pregnancy, annexin A2 in preeclampsia is significantly reduced.3 The expression of annexin A2 mRNA by RT-PCR in placental tissue: early-onset mild preeclampsia group 27.94±1.49, early-onset severe preeclampsia group was 27.23±1.26, late-onset mild preeclampsia group was 28.27±0.85, late-onset severe preeclampsia group was 28.54±0.65, and healthy pregnancy group was 32.34±0.71.Conclusion: Annexin A2 mainly expresses on the surface of trophoblast cell. Compared with normal pregnancy, annexin A2 in placental of preeclampsia was decreased. It plays an important role in the occurrence and the development of preeclampsia. Part four The purification of annexin A5 and its anticoagulantion in vitroObjective: To investigate the anticoagulation of annexin A5 in vivo and in vitro.Methods: Placental tissues of healthy pregnancy women were collected for purification. The anticoagulation of annexin A5 was measured in vitro with different concentrations of annexin A5.Results:1 When the concentration of annexin A5≥30μg/ml, APTT gradually extended. There are significant differences between the groups.2 Compared with the normal group, the coagulation of peripheral blood was no significant difference.Conclusion: Annexin A5 anticoagulant activity is closely related with its concentration. When the concentration of annexin A5≥30μg/ml, APTT extended with the increasing concentration of annexin A5. However, the APTT in peripheral blood of patients with preeclampsia did not change. Part five Antibody detection in preeclampsiaObjective: To detect related antibodies in maternal blood of the preeclampsia and analyze the ralationship between of antibody and pathogenesis of preeclampsia.Methods: The level of anti-annexin A5, anti-annexin A2, anti-β2GPI and ACL in maternal blood of preeclampsia and normal pregnancy were dectected by ELISA.Results:1 Compared with normal pregnancy, levels of anti-annexin A5 in the peripheral blood of patients with preeclampsia is increased. But there was no significant difference between them (P=0.83).2 Compared with normal pregnancy, levels of anti-annexin A2 in the peripheral blood of patients with preeclampsia are increased, and it has a significant difference between them (P=0.04).3 Compared with normal pregnancy, levels of anti-β2GPI in the peripheral blood of patients with preeclampsia are increased, and it has a significant difference between them (P=0.01).4 Compared with normal pregnancy, ACL-positive rate in preeclampsia was increased.Conclusion:The antibody in peripheral blood, which binding to antigen antibody decreased the level of annexin A5, annexin A2 expression. And they influcene the coagulation and fibrinolysis system, which creats hypercoagulation of PE.
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