本文選題：子癇前期 + 自噬　； 參考：《青島大學(xué)》2017年博士論文

【摘要】：子癇前期是妊娠期婦女特有的常見病癥,是妊娠期高血壓疾病最嚴重的階段,是妊娠期高血壓疾病所致母兒死亡的最主要原因。發(fā)展中國家,因子癇前期而死亡的孕婦占20%~80%,發(fā)達國家,子癇前期的孕婦圍生期死亡率是未患該病的5倍。為了探討子癇前期的病因和發(fā)病機制,廣大學(xué)者從流行病學(xué)角度和免疫學(xué)等方面都進行了廣泛的研究,目前認為該疾病是母體、胎盤、胎兒等眾多因素共同作用的結(jié)果,主要病因包括子宮螺旋小動脈重鑄不足,炎癥免疫過度激活,血管內(nèi)皮細胞受損,遺傳因素,營養(yǎng)缺乏和胰島素抵抗。但疾病的具體發(fā)病機制至今仍未完全闡明,目前普遍認為的該疾病病理生理學(xué)發(fā)生發(fā)展的兩個主要階段,即胎盤異常階段及母體異常階段,胎盤異常階段的核心問題是滋養(yǎng)細胞缺血、缺氧造成的氧化應(yīng)激反應(yīng),而母體異常階段主要是由胎盤局部炎性反應(yīng)誘發(fā)的母體全身炎性反應(yīng)。盡管子癇前期是全球多發(fā)疾病,其臨床表現(xiàn)具有多樣化,但到目前仍沒有有效的預(yù)測或預(yù)防性治療措施。臨床通用的監(jiān)測手段僅為孕期新發(fā)而孕后自愈性的高血壓及尿蛋白的出現(xiàn),但始終缺乏明確診斷的特異性指標。目前期待治療措施如降壓、擴容、皮質(zhì)激素的使用等處理療效甚微,且理論依據(jù)不足,明確有效的治療方法是終止妊娠,從而帶來早產(chǎn)等一系列不良后果;與此同時,目前關(guān)于子癇前期的治療中還未曾有關(guān)于抗炎治療的報道,因此我們認為大量研究工作應(yīng)從氧化應(yīng)激反應(yīng)、炎性反應(yīng)等各方面同時入手來探討子癇前期的發(fā)病機制并為子癇前期的臨床治療提供新的科研思路。自噬是細胞在自噬相關(guān)基因(Atg)的調(diào)控下由溶酶體參與的一種分解代謝機制,以消化自身受損或功能異常的細胞成分,在細胞增殖、分化過程中維持細胞結(jié)構(gòu)、代謝和功能的平衡,在饑餓或低代謝、缺氧、藥物影響的情況下自噬是保證細胞的存活的重要機制。自噬作為抵御病理狀態(tài)的代償機制,其調(diào)控機制的失調(diào)與感染、腫瘤、神經(jīng)退行性變、心臟疾病等多種疾病的發(fā)生、發(fā)展有關(guān),并逐漸成為多種疾病領(lǐng)域的研究熱點。自噬的發(fā)生依賴Atg的啟動以及一系列自噬相關(guān)蛋白的共同參與,自噬基因的研究最初是在酵母菌中進行的,其中,Beclin 1是哺乳動物體內(nèi)酵母Atg6的基因同源物,通過II型磷脂酰肌醇-3激酶(PI3K)/Beclin 1途徑可激活自噬,Beclin 1通過介導(dǎo)其與自噬相關(guān)蛋白的交互作用來完成自噬體的雙層膜結(jié)構(gòu),對于自噬的發(fā)生發(fā)揮關(guān)鍵性的作用。另外,Beclin 1可與凋亡調(diào)節(jié)基因Bcl-2產(chǎn)物的蛋白家族相結(jié)合,在細胞自噬與凋亡的過程中起到橋梁作用。LC3是哺乳動物體內(nèi)酵母Atg8的基因同源物,LC3在細胞中以LC3 I與LC3 II兩種形式存在,LC3的C末端被Atg4基因剪切后形成胞質(zhì)型LC3(LC3 I),LC3 I經(jīng)泛素樣加工修飾與磷脂酰乙醇胺結(jié)合形成LC3 II并表達在自噬體膜上,LC3 II特異性的結(jié)合在自噬體膜上,LC3 II最終被降解后可重新回到細胞質(zhì)中。因此,LC3 I向LC3 II的轉(zhuǎn)變是自噬發(fā)生的重要標志,自噬相關(guān)蛋白Beclin 1及LC3的表達是自噬研究的常用檢測指標。子癇前期所具有的缺氧、氧化應(yīng)激及炎癥反應(yīng)等諸多因素都與自噬有著密切關(guān)系,據(jù)此推測子癇前期的疾病狀態(tài)可能存在自噬水平的異常。目前關(guān)于自噬與子癇前期發(fā)病的關(guān)系研究較少且沒有達成統(tǒng)一的共識。氧化低密度脂蛋白(ox LDL)的生成系低密度脂蛋白(LDL)中大量不飽和脂肪酸在各種氧化條件下生成一種脂類自由基,從而介導(dǎo)產(chǎn)生更多的過氧化脂質(zhì)引起鏈式反應(yīng),形成多種活性醛,與低密度脂蛋白中的載脂蛋白B結(jié)合,產(chǎn)生新的抗原決定簇,形成氧化低密度脂蛋白。氧化低密度脂蛋白及血凝集素樣氧化低密度脂蛋白受體1即LOX-1共同作用參與了子癇前期的病理生理變化。較早的研究表明子癇前期患者血漿中氧化低密度脂蛋白水平升高,可能通過引起內(nèi)皮損傷參與子癇前期的發(fā)病。我們之前的研究也表明子癇前期患者血漿中氧化低密度脂蛋白水平升高,且重度子癇前期患者高于輕度子癇前期患者,并通過上調(diào)胎盤組織中的LOX-1表達水平參與子癇前期的發(fā)病及疾病的進展。通過對滋養(yǎng)細胞的研究發(fā)現(xiàn),LOX-1可能通過影響滋養(yǎng)細胞凋亡而參與子癇前期的發(fā)病。我們的研究則表明LOX-1可能通過其介導(dǎo)的氧化應(yīng)激反應(yīng)影響滋養(yǎng)細胞血管因子的表達而參與子癇前期的病理生理變化。近期的研究認為氧化低密度脂蛋白與多種細胞自噬有密切關(guān)系,其在不同細胞類型中發(fā)揮的作用有所不同,可呈濃度依賴的上調(diào)臍靜脈內(nèi)皮細胞的自噬水平;而在巨噬細胞及平滑肌細胞中卻能抑制自噬的發(fā)生,但是關(guān)于氧化低密度脂蛋白對于滋養(yǎng)細胞自噬影響的研究卻不多見。研究目的:本研究旨在探討子癇前期患者氧化低密度脂蛋白及胎盤滋養(yǎng)細胞自噬水平的變化以及氧化低密度脂蛋白對滋養(yǎng)細胞自噬及炎癥反應(yīng)的影響,進一步說明自噬對滋養(yǎng)細胞炎性反應(yīng)的保護作用,為子癇前期的病理生理學(xué)研究及疾病的治療提供新的理論基礎(chǔ)。研究方法:1、選擇2014年6月至2015年1月在青島大學(xué)附屬醫(yī)院產(chǎn)科住院且經(jīng)剖宮產(chǎn)分娩的子癇前期孕婦26例為子癇前期組(PE組),選取同期正常晚期妊娠婦女20例為正常孕婦組(NP組)。兩組孕婦均于入院后空腹抽取肘靜脈血5ml,分離血清,內(nèi)置于-70℃冰箱保存。兩組孕婦均經(jīng)剖宮產(chǎn)終止妊娠,在娩出胎盤后的30分鐘內(nèi)在無菌條件下獲取胎盤標本,避開胎盤母體面的壞死及鈣化區(qū),采集兩塊大小約1 cm×1 cm×1 cm的胎盤組織,經(jīng)滅菌生理鹽水反復(fù)漂凈殘余的血液后,將其中的一塊放于1.5 ml的無菌離心管中,15分鐘內(nèi)置于-70℃冰箱保存,以待RNA和蛋白提取,另一塊甲醛固定備用。2、采用ELISA法檢測兩組孕婦血清中ox LDL的水平,以及炎性因子TNF-α和IL-6水平的變化。采用免疫組化法對胎盤組織中TNF-α和IL-6的表達進行定位測定;采用Western Blot法檢測各組胎盤組織中TNF-α和IL-6蛋白的表達。3、采用免疫組化法對胎盤組織中自噬相關(guān)蛋白(Beclin 1和LC3)進行定位測定;采用real-time PCR法對胎盤組織中Beclin 1和LC3的m RNA進行測定;采用Western Blot法檢測各組胎盤組織中Beclin 1和LC3蛋白的表達。4、購置JEG-3細胞系并進行傳代培養(yǎng),經(jīng)不同濃度ox LDL(25,50,100,150mg/l)以及不同作用時間(6,12,24,48小時)處理后收集細胞,通過免疫熒光法檢測不同濃度及不同作用時間下ox LDL對滋養(yǎng)細胞自噬相關(guān)蛋白LC3的影響。5、采用JEG-3細胞系,利用不同濃度ox LDL(25,50,100,150mg/l)作用6小時進行研究,通過real-time PCR法檢測經(jīng)ox LDL處理前后滋養(yǎng)細胞中炎性因子(TNF-α和IL-6)m RNA表達的變化。6、采用JEG-3細胞系,將其分成兩組,一組利用濃度為100mg/l的ox LDL作用6小時后進行實驗,另一組經(jīng)雷帕霉素100n M預(yù)處理預(yù)處理1小時后,再利用濃度為100mg/l的ox LDL作用6小時為研究條件,通過real-time PCR法對滋養(yǎng)細胞中炎性因子(TNF-α和IL-6)的m RNA進行測定;采用Western Blot法檢測滋養(yǎng)細胞中自噬相關(guān)蛋白Beclin 1及LC3的蛋白表達水平的變化。7、統(tǒng)計學(xué)方法應(yīng)用SPSS 17.0進行數(shù)據(jù)處理,所有測定結(jié)果以_x±s表示,兩組間比較采用t檢驗,P㩳0.05為差別有統(tǒng)計學(xué)意義。結(jié)果:1、通過ELISA法檢測結(jié)果提示子癇前期組患者血清中ox LDL、TNF-α及IL-6水平較正常對照組明顯升高,差異有統(tǒng)計學(xué)意義(p0.05)。免疫組化結(jié)果提示TNF-α及IL-6在子癇前期組及正常對照組胎盤滋養(yǎng)細胞細胞中均有表達,呈棕色陽性染色,同時,子癇前期組胎盤中染色較深且陽性細胞數(shù)目較正常對照組增多。Western Blot法檢測TNF-α及IL-6蛋白表達結(jié)果提示子癇前期組胎盤組織中炎性因子的表達較正常對照組升高,差異有統(tǒng)計學(xué)意義(p0.05)。2、免疫組化結(jié)果顯示Beclin 1在滋養(yǎng)細胞中呈高于背景的淡棕色陽性染色,主要表達于細胞滋養(yǎng)細胞;LC3高于背景的淡棕色染色主要表達于細胞滋養(yǎng)細胞,在合體滋養(yǎng)細胞中也有表達。在正常對照組(NP組)中Beclin 1及LC3蛋白的陽性細胞數(shù)量明顯增多且染色增強。real-time PCR法檢測結(jié)果顯示正常對照組胎盤組織中Beclin 1及LC3 m RNA的表達水平明顯高于子癇前期組,差異有統(tǒng)計學(xué)意義(p0.05)。Western Blot法檢測Beclin 1及LC3蛋白表達結(jié)果提示正常對照組胎盤組織中自噬相關(guān)蛋白的表達較子癇前期組升高,差異有統(tǒng)計學(xué)意義(p0.05)。3、在不同濃度ox LDL(25,50,100,150mg/l)以及不同作用時間(6,12,24,48小時)下培養(yǎng)滋養(yǎng)細胞細胞,利用免疫熒光法均不能在細胞內(nèi)觀察到胞漿內(nèi)紅染的LC3點狀聚集。當細胞由雷帕霉素100n M預(yù)處理1h后,再經(jīng)ox LDL(100mg/l)刺激的細胞在不同作用時間下均可見到胞漿內(nèi)紅染的LC3點狀聚集。經(jīng)雷帕霉素預(yù)處理后再加入ox LDL培養(yǎng)細胞后,經(jīng)觀察得ox LDL作用6h及12h后細胞形態(tài)仍是完整的,但作用24小時及48小時即可出現(xiàn)細胞核的腫脹、變形;因此,作用6小時為后續(xù)其他實驗條件。4、通過real-time PCR法檢測TNF-α和IL-6 m RNA的表達發(fā)現(xiàn)經(jīng)50mg/l及100mg/l ox LDL處理后滋養(yǎng)細胞中炎性因子的表達均高于空白對照組,差異有統(tǒng)計學(xué)意義(p0.05)。同時,經(jīng)100mg/l ox LDL培養(yǎng)后滋養(yǎng)細胞中m TOR的表達較空白對照組無明顯變化,因此選定100mg/l ox LDL為后續(xù)實驗條件。5、細胞經(jīng)雷帕霉素100n M預(yù)處理1小時后,再經(jīng)100mg/l ox LDL培養(yǎng)6小時,Western Blot法檢測得Beclin 1及LC3蛋白表達明顯高于未經(jīng)雷帕霉素處理組,差異有統(tǒng)計意義(p0.05);同時real-time PCR法檢測TNF-α和IL-6m RNA結(jié)果提示炎性因子的表達較未經(jīng)雷帕霉素處理組明顯降低,差異有統(tǒng)計意義(p0.05)。結(jié)論及意義:1、子癇前期患者氧化低密度脂蛋白及炎性因子(TNF-α、IL-6)表達水平的升高參與了子癇前期疾病的病理生理過程。2、氧化低密度脂蛋白可引起滋養(yǎng)細胞炎性反應(yīng),釋放炎性因子;但ox LDL自身不能引起滋養(yǎng)細胞自噬。3、在自噬誘導(dǎo)劑的作用下,增強滋養(yǎng)細胞自噬后可減弱氧化低密度脂蛋白對滋養(yǎng)細胞引起的炎性反應(yīng),降低其炎性因子的表達。氧化低密度脂蛋白及炎性因子在子癇前期患者血清中顯著升高,細胞實驗證實,氧化低密度脂蛋白可誘導(dǎo)滋養(yǎng)細胞炎性因子的表達,相同實驗條件下不能誘導(dǎo)滋養(yǎng)細胞自噬的發(fā)生,也就是說當氧化低密度脂蛋白通過炎癥因素參與子癇前期發(fā)病的同時抑制了滋養(yǎng)細胞自噬的能力,因此認為氧化低密度脂蛋白在子癇前期疾病發(fā)生的氧化應(yīng)激反應(yīng)及炎性反應(yīng)中起到橋梁作用。同時,自噬對氧化低密度脂蛋白誘導(dǎo)滋養(yǎng)細胞炎性反應(yīng)具有保護作用,子癇前期胎盤滋養(yǎng)細胞自噬能力的減低不足以保護由氧化低密度脂蛋白引起的炎性反應(yīng);因此本研究在氧化低密度脂蛋白及自噬參與子癇前期發(fā)病的機制探討中均具有重要意義;并為子癇前期疾病的治療提供了新的理論基礎(chǔ)。
[Abstract]:Preeclampsia is a common common disease of women in pregnancy. It is the most serious stage of hypertensive disorder in pregnancy. It is the most important cause of maternal mortality in pregnancy induced hypertension. In developing countries, pregnant women who died during preeclampsia and preeclampsia are 20%~80%, and the perinatal mortality of pregnant women in the pre eclampsia period is 5 times as high as that of the disease. In order to explore the etiology and pathogenesis of preeclampsia, many scholars have conducted extensive research on epidemiology and immunology. It is believed that the disease is the result of many factors such as mother, placenta, fetus and so on. The main causes include the insufficiency of the uterine spiral arterioles, the excessive activation of inflammatory immunity, and the intravascular injection. The damage of skin cells, genetic factors, nutritional deficiency and insulin resistance. But the specific pathogenesis of the disease has still not been fully elucidated. At present, the two main stages of pathophysiology of the disease are the placental abnormal stage and the maternal abnormal stage. The core of the placental abnormal stage is the ischemia of trophoblast and hypoxia. The oxidative stress reaction is caused by the maternal systemic inflammatory response, which is mainly caused by the local inflammatory response of the placenta. Although preeclampsia is a global multiple disease, its clinical manifestations are diverse, but there is still no effective prediction or preventive treatment at present. The self healing of hypertension and the appearance of urine protein after pregnancy, but there is always a lack of specific diagnostic criteria. At present, the treatment measures, such as decompression, dilatation, and the use of corticosteroids, are very small, and the theoretical basis is insufficient. The clear and effective treatment is to terminate pregnancy and bring a series of adverse consequences, such as premature delivery. At present, there is no report about the treatment of anti - inflammation in preeclampsia, so we think a lot of research should start with the oxidative stress reaction, inflammatory response and other aspects to explore the pathogenesis of preeclampsia and provide new scientific research ideas for the clinical treatment of preeclampsia. Autophagy is autophagy related to autophagy. A mechanism of catabolism involved in the lysosome under the regulation of the gene (Atg) to digest the cellular components of their own damage or function and maintain the balance of cell structure, metabolism and function during cell proliferation and differentiation. Autophagy is an important mechanism for the survival of the cells under the condition of starvation or low metabolism, anoxia, and drug effects. As a compensatory mechanism for resisting pathological state, the disorder of its regulatory mechanism is related to the development of a variety of diseases such as infection, tumor, neurodegenerative change, heart disease, development, and gradually become a hot spot in many fields of disease. The occurrence of autophagy depends on the initiation of Atg and the involvement of a series of autophagy related proteins. The study was originally carried out in yeast, in which Beclin 1 is a gene homologue of yeast Atg6 in mammals, which activates autophagy through the II type phosphatidylinositol -3 kinase (PI3K) /Beclin 1 pathway. Beclin 1 mediates its interaction with autophagy related proteins to complete the bilayer membrane structure of autophagosol, and plays a role in autophagy. In addition, Beclin 1 can be combined with the protein family of the Bcl-2 product of apoptosis regulating gene, which plays a bridge role in the process of autophagy and apoptosis,.LC3 is the gene homologue of yeast Atg8 in mammalian body, LC3 is stored in the two forms of LC3 I and LC3 II, and the C end of LC3 is formed after the Atg4 gene is cut. LC3 (LC3 I), LC3 I is modified with phosphatidyl ethanolamine to form LC3 II and is expressed on the autophagosome membrane, and LC3 II specificity combines on the autophagic membrane. LC3 II is finally degraded to the cytoplasm. Therefore, the transformation of LC3 II is an important sign of autophagy, autophagy associated protein 1 The expression of C3 is a common detection index for autophagy. Many factors such as hypoxia, oxidative stress and inflammatory response in preeclampsia are closely related to autophagy. According to this, it is suggested that the state of autophagy may exist in preeclampsia. The relationship between autophagy and preeclampsia is less and less than that of the preeclampsia. A unified consensus. The generation of oxidized low density lipoprotein (ox LDL) generation of low density lipoprotein (LDL) produces a fat free radical under various oxidation conditions, which mediates more lipid peroxide induced chain reaction, forms a variety of active aldehydes, and combines with apolipoprotein B in low density lipoprotein, and produces a combination of apolipoprotein B. New antigenic determinants are produced to form oxidized low density lipoprotein. Oxidative low density lipoprotein and blood lectin like oxidized low density lipoprotein receptor 1, LOX-1, are involved in the pathophysiological changes in preeclampsia. Skin injury is involved in the onset of preeclampsia. Our previous study also showed that the plasma levels of oxidized low density lipoprotein in preeclampsia patients were elevated, and severe preeclampsia patients were higher than those with mild preeclampsia, and were involved in the pathogenesis of preeclampsia and the progress of the disease by raising the level of LOX-1 in placenta tissue. Cell culture studies have found that LOX-1 may participate in the onset of preeclampsia by affecting the apoptosis of trophoblastic cells. Our study suggests that LOX-1 may participate in the pathophysiological changes in the preeclampsia through its mediated oxidative stress response and its involvement in the expression of trophoblast vascular factors. Many cells are closely related to autophagy, and they play a different role in different cell types. They can increase the autophagy level of umbilical vein endothelial cells in a concentration dependent manner, but inhibit the occurrence of autophagy in macrophages and smooth muscle cells, but studies on the effect of oxidized low density lipoprotein on the autophagy of trophoblastic cells The purpose of this study is to investigate the changes in the level of autophagy of oxidized low density lipoprotein and placental trophoblast in preeclampsia and the effect of oxidized low density lipoprotein on the autophagy and inflammation of trophoblastic cells, and further explain the protective effect of autophagy on the inflammatory response to trophoblast and the pathophysiology of preeclampsia. The study and the treatment of disease provide a new theoretical basis. 1, 26 cases of preeclampsia (group PE) were selected from June 2014 to January 2015 in the obstetrics department of the Affiliated Hospital of Qiingdao University and cesarean section, and 20 cases of normal pregnant women were selected as normal pregnant women group (group NP). All the two groups of pregnant women were enrolled. The blood 5ml of the elbow vein was extracted on the empty stomach after the hospital, and the serum was separated and stored in the refrigerator of -70 C. The two groups of pregnant women were treated with cesarean section to terminate the pregnancy. The placenta samples were obtained under sterile conditions for 30 minutes after delivery of the placenta, avoiding the necrosis and calcification of the placental maternal surface, and collecting two pieces of placental tissue about 1 cm * 1 cm x 1 cm, and sterilized physiological salt After rinsing the residual blood repeatedly, one of them was placed in a sterile centrifuge tube of 1.5 ml. 15 minutes were stored in the refrigerator of -70 centigrade for RNA and protein extraction. Another block of formaldehyde was fixed to the.2. The level of ox LDL in the serum of two groups of pregnant women, and the changes of TNF- A and IL-6 levels of inflammatory factors were detected by ELISA. Immunohistochemistry was used. The expression of TNF- alpha and IL-6 in placenta tissue was determined by method. The expression of TNF- alpha and IL-6 protein in the placental tissues of each group was detected by Western Blot method. The localization of autophagy related proteins (Beclin 1 and LC3) in placenta tissue was detected by immunohistochemical method, and real-time PCR method was used to detect the Beclin 1 in placenta tissue. The expression of Beclin 1 and LC3 protein in the placental tissues of each group was detected by Western Blot, and JEG-3 cell lines were purchased and cultured. The cells were collected after different concentrations of ox LDL (25,50100150mg/l) and different action time (6,12,24,48 hours). The cells were detected by immunofluorescence and different concentrations and different action times were detected by immunofluorescence. The effect of ox LDL on the autophagy related protein LC3 of trophoblast.5, using JEG-3 cell line and using ox LDL (25,50100150mg/l) at different concentrations for 6 hours, and using real-time PCR method to detect the changes of inflammatory factors in the trophoblast cells before and after ox LDL, and divide them into two groups. A group of ox LDL with concentration of 100mg/l was used for 6 hours, and another group was pretreated by preconditioning with rapamycin 100N M for 1 hours, and then ox LDL with a concentration of 100mg/l was used for 6 hours as the study condition. The real-time PCR method was used to determine the inflammatory factors (TNF- alpha and IL-6) in trophoblast. The changes in the protein expression level of autophagy related protein Beclin 1 and LC3 in trophoblast were detected.7, the statistical method was used for data processing with SPSS 17, all the results were _x + s, the two groups were compared with t test and P 0.05 was statistically significant. Results: 1, the results of ELISA method were used to prompt the sera of the preeclampsia group. The level of ox LDL, TNF- alpha and IL-6 was significantly higher than that in the normal control group. The difference was statistically significant (P0.05). The immunohistochemical results suggested that TNF- alpha and IL-6 were expressed in the placental trophoblast cells of preeclampsia and normal control groups, and the positive staining of the placenta in the placenta was deeper and the number of positive cells was more positive in the preeclampsia group. The expression of TNF- alpha and IL-6 protein detected by.Western Blot method in normal control group showed that the expression of inflammatory factors in placental tissue of preeclampsia group was higher than that of normal control group, and the difference was statistically significant (P0.05).2. The immunohistochemical results showed that Beclin 1 was a light brown positive staining in the trophoblastic cells which was higher than the background, mainly expressed in the cells. In the normal control group (group NP), the number of Beclin 1 and LC3 protein positive cells increased significantly and the staining enhanced.Real-time PCR assay showed the Beclin 1 and LC3 m RNA in the normal control group of the placental tissue. The expression level of the preeclampsia group was significantly higher than that in the preeclampsia group (P0.05) the expression of Beclin 1 and LC3 protein by.Western Blot method suggested that the expression of autophagy related protein in the normal control group was higher than the preeclampsia group, and the difference was statistically significant (P0.05).3, at different concentrations ox LDL (25,50100150mg/l) and not The cytotrophoblast cells were cultured at the same time of action (6,12,24,48 hours), and the cytoplasmic red dye LC3 spot aggregation could not be observed by immunofluorescence. When the cells were pretreated with rapamycin 100N M, the cells stimulated by ox LDL (100mg/l) could see the LC3 dot aggregation of erythrocyte in the cytoplasm at different time. After pretreated with rapamycin, after adding ox LDL to ox LDL cells, it was observed that the cell morphology was still intact after the action of 6h and 12h, but the cell nucleus could be swelled and deformed for 24 hours and 48 hours. Therefore, the action of 6 hours was the subsequent other experimental conditions.4, and the expression of TNF- A and IL-6 m was detected by real-time PCR method. The expression of inflammatory factors in the trophoblastic cells after 50mg/l and 100mg/l ox LDL was higher than that in the blank control group. The difference was statistically significant (P0.05). At the same time, the expression of M TOR in the trophoblastic cells after 100mg/l ox LDL was not significantly changed. Therefore, 100mg/l ox was selected as a follow-up experimental condition, and the cells were treated by rapamycin. The expression of Beclin 1 and LC3 protein detected by Western Blot method was significantly higher than that without rapamycin treatment group after 1 hours of pretreatment with 100mg/l ox LDL, and the difference was statistically significant (P0.05), while real-time PCR method was statistically significant (P0.05), while real-time PCR assay showed that the expression of the inflammatory factor was significantly lower than that of the non rapamycin treatment group. The difference was statistically significant (P0.05). Conclusion and significance: 1, the expression of oxidized low density lipoprotein and TNF- (IL-6) in preeclampsia patients
【學(xué)位授予單位】：青島大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2017
【分類號】：R714.244

【相似文獻】

相關(guān)期刊論文 前10條

1 孟濤;李輝;劉彤;潘莉莉;;脂聯(lián)素及氧化修飾低密度脂蛋白在子癇前期中的測定及意義[J];中國優(yōu)生與遺傳雜志;2006年06期

2 桑翠琴;張震宇;王淑珍;劉浩;郭淑麗;;重度子癇前期/子癇合并急性呼吸窘迫綜合征的診斷與處理[J];中國醫(yī)刊;2006年10期

3 趙桂花;;重度子癇前期與子癇82例臨床分析[J];臨床醫(yī)藥實踐雜志;2007年01期

4 楊小頎;嚴妮子;;子癇前期與血鈣水平相關(guān)性研究[J];實用臨床醫(yī)學(xué);2007年04期

5 楊小頎;嚴妮子;;子癇前期與血鈣水平相關(guān)性研究[J];四川生理科學(xué)雜志;2007年01期

6 陳茜;王澤華;;子癇前期的預(yù)測及預(yù)防[J];華中醫(yī)學(xué)雜志;2007年03期

7 朱倩;;對38例子癇前期(重)、子癇治愈的體會[J];實用預(yù)防醫(yī)學(xué);2007年05期

8 梁英;周麗麗;甘娟;姜碧洋;楊承東;;重度子癇前期在產(chǎn)科ICU監(jiān)護救治的臨床分析[J];中國婦幼保健;2008年19期

9 詹衛(wèi)星;鄭九生;;血管緊張素轉(zhuǎn)換酶基因多態(tài)性與重度子癇前期及腎功能損害的關(guān)系[J];江西醫(yī)藥;2008年08期

10 蔡穎;尹國武;姜鋒;李怡;閔保華;王s，

本文編號：1971330