本文關(guān)鍵詞： ADAR1基因 子宮內(nèi)膜異位癥(EMs) 子宮腺肌癥(AM)　出處：《鄭州大學(xué)》2017年碩士論文　論文類型：學(xué)位論文

【摘要】：子宮內(nèi)膜異位癥(endometriosis,EMs)簡(jiǎn)稱內(nèi)異癥,是一種很常見的婦科疾病,以子宮內(nèi)膜樣組織生長(zhǎng)在子宮以外為特征,如卵巢,輸卵管,盆腔和腹腔,其中以異位于卵巢最常見。子宮腺肌病(adenomyosis,AM)是指具有生長(zhǎng)功能的子宮內(nèi)膜腺體和間質(zhì)侵入子宮肌層,以月經(jīng)量過多,經(jīng)期延長(zhǎng)及繼發(fā)性痛經(jīng)為主要臨床表現(xiàn)的常見婦科疾病。AM被認(rèn)為是子宮內(nèi)膜異位癥的一種,其發(fā)病機(jī)制存在一定的相似性。EMs和AM被描述為非腫瘤性的疾病,但其在生物學(xué)行為上卻呈現(xiàn)出與惡性腫瘤一致的特性,如可異常增殖、侵潤(rùn)甚至侵襲。近年的研究還表明,細(xì)胞免疫和體液免疫功能失調(diào)均可能與Ems和AM的發(fā)生發(fā)展有關(guān)。國內(nèi)外有關(guān)ADAR1與子宮內(nèi)膜異位癥和子宮腺肌病發(fā)病關(guān)系的研究未見報(bào)道。ADAR1是一種RNA編輯酶,RNA編輯是指DNA轉(zhuǎn)錄成RNA后對(duì)RNA的修飾和加工,可能使基因在DNA水平的表達(dá)與其在蛋白水平的表達(dá)完全不同,從而改變了遺傳信息[2-6]。研究發(fā)現(xiàn)ADAR1在細(xì)胞增殖[7]、分化[8]及腫瘤發(fā)展[9]中起了十分重要的作用。此外,ADAR1與炎癥和免疫也關(guān)系密切[10]。本實(shí)驗(yàn)采用實(shí)時(shí)熒光定量PCR,Western blotting及免疫組織化學(xué)等方法檢測(cè)ADAR1在內(nèi)異癥和腺肌病患者在位內(nèi)膜與異位內(nèi)膜及腺肌瘤組織中的表達(dá),進(jìn)一步探討兩種疾病發(fā)病機(jī)制以期對(duì)臨床診治提供新的思路。目的研究ADAR1在子宮內(nèi)膜異位癥和子宮腺肌病中的表達(dá)差異,探討ADAR1與子宮內(nèi)膜異位癥和子宮腺肌病發(fā)病的關(guān)系及意義。資料與方法該研究經(jīng)鄭州大學(xué)第三附屬醫(yī)院即河南省婦幼保健院倫理委員會(huì)批準(zhǔn),所有患者均簽署知情同意書。1研究對(duì)象選取2015年7月至2016年10月在河南省鄭州大學(xué)第三附屬醫(yī)院(即河南省婦幼保健院)婦科行手術(shù)治療的EMs患者31例,年齡29~44歲,手術(shù)證實(shí)為子宮內(nèi)膜異位癥III型或IV型(按美國生殖醫(yī)學(xué)協(xié)會(huì)定義),對(duì)其中有生育要求的25例患者同時(shí)行宮腔鏡檢查,診刮得到子宮內(nèi)膜組織(實(shí)驗(yàn)組EU組),術(shù)中取異位內(nèi)膜組織(實(shí)驗(yàn)組EC組)。選同期在河南省鄭州大學(xué)第三附屬醫(yī)院就診行子宮全切的子宮腺肌病患者30例,年齡38~48,子宮全切后立即取患者在位子宮內(nèi)膜組織(實(shí)驗(yàn)組AM1),及腺肌瘤組織(實(shí)驗(yàn)組AM2);對(duì)照組為同期子宮肌瘤行子宮全切的患者30例,年齡34~49歲,子宮全切后立即取在位子宮內(nèi)膜組織(對(duì)照組C)。上述患者月經(jīng)規(guī)律,26~32天;術(shù)前六個(gè)月無激素及內(nèi)分泌疾病用藥史,無妊娠、哺乳,術(shù)前無絕經(jīng)期癥狀。以上標(biāo)本均在子宮內(nèi)膜增殖期獲取,且全部經(jīng)病理證實(shí)。2標(biāo)本處理在無菌狀態(tài)下獲取的子宮內(nèi)膜組織或肌瘤組織,一半立即放入液氮,后轉(zhuǎn)入-80℃保存用于實(shí)時(shí)熒光定量PCR和蛋白印記實(shí)驗(yàn),另一半放入10%中性福爾馬林溶液固定后石蠟包埋,用于組織學(xué)檢查。3試驗(yàn)方法2.1實(shí)時(shí)熒光定量PCR技術(shù)(qRT-PCR)檢測(cè)各組中ADAR1mRNA的表達(dá)。2.2 Western blotting半定量檢測(cè)各組ADAR1蛋白表達(dá)水平。2.3免疫組化對(duì)各組ADAR1蛋白表達(dá)進(jìn)行定位及半定量分析。4統(tǒng)計(jì)學(xué)分析實(shí)驗(yàn)數(shù)據(jù)以均數(shù)±標(biāo)準(zhǔn)差(x±s)表示,采用SPSS 21.0軟件進(jìn)行統(tǒng)計(jì)分析,組間兩兩比較用t檢驗(yàn),滿足正態(tài)性和方差齊性的多組比較采用單因素方差分析,若不滿足則采用K個(gè)獨(dú)立樣本檢驗(yàn)或Kruskal-Wallis秩和檢驗(yàn)。檢驗(yàn)水準(zhǔn)α=0.05。兩兩比較采用Bonferroni法,以Pα為差異有統(tǒng)計(jì)學(xué)意義。結(jié)果1各組年齡比較子宮內(nèi)膜異位癥組(36.4±3.8),子宮腺肌病組(39.13±5.61)和對(duì)照組(40.3±3.4),組間無統(tǒng)計(jì)學(xué)差異(P0.05)。2 ADAR1mRNA在子宮內(nèi)膜異位癥和子宮腺肌病中的表達(dá)實(shí)時(shí)熒光定量PCR結(jié)果顯示ADAR1 P150mRNA在EC(2.33±0.28)組和EU(2.18±0.36)組的表達(dá)明顯高于C(1.68±0.43)組(P0.05),但EU組和EC組ADAR1 P150 mRNA的表達(dá)差別無統(tǒng)計(jì)學(xué)意義,且ADAR1 P110 mRNA在子宮內(nèi)膜異位癥EU組(1.01±0.21),EC組(1.01±0.15)和C組(0.99±0.11)的表達(dá)差別無統(tǒng)計(jì)學(xué)意義(P0.05)。與EMs組結(jié)果相似,實(shí)時(shí)熒光定量PCR結(jié)果顯示ADAR1 P150 mRNA的表達(dá)在AM1組(2.09±0.26)和AM2組(2.09±0.25),均高于對(duì)照組(1.68±0.43)差異有統(tǒng)計(jì)學(xué)意義(P0.05);而ADAR1 P110 mRNA的表達(dá)在AM1組(0.97±0.21),AM2組(0.87±0.17),C組(0.99±0.11)差異無統(tǒng)計(jì)學(xué)意義(P0.05)。3 ADAR1蛋白在子宮內(nèi)膜異位癥和子宮腺肌病中的表達(dá)Western Blotting的結(jié)果顯示ADAR1蛋白的表達(dá)在EU組(0.39±0.05)和EC(0.40±0.07)組的均明顯高于C組(0.104±0.04)(P0.05),且EU和EC兩組ADAR1蛋白的表達(dá)差異無統(tǒng)計(jì)學(xué)意義(P0.05)。ADAR1蛋白表達(dá)在AM1組(0.34±0.06)和AM2組(0.35±0.06)均高于C組(0.104±0.04),差異均有統(tǒng)計(jì)學(xué)意義(P0.05)。AM1組的ADAR1蛋白表達(dá)與AM2組相比,差異無統(tǒng)計(jì)學(xué)意義(P0.05)。4 ADAR1蛋白在子宮內(nèi)膜異位癥和子宮腺肌病中的定位與表達(dá)免疫組化的結(jié)果顯示ADAR1蛋白在內(nèi)異癥在位、異位內(nèi)膜,腺肌瘤及在位內(nèi)膜,對(duì)照組內(nèi)膜的細(xì)胞胞漿中均可見棕黃色陽性染色。子宮內(nèi)膜異位癥組中ADAR1在位內(nèi)膜(EU組),異位內(nèi)膜(EC組)和正常內(nèi)膜(C組)表達(dá)水平分別為(87.29±11.87),(96.65±13.31),(26.08±8.49),差異有統(tǒng)計(jì)學(xué)意義(p0.05),子宮腺肌病組ADAR1于在位內(nèi)膜(AM1),腺肌瘤(AM2)和正常內(nèi)膜C組的表達(dá)水平分別為(68.77±11.20),(67.48±11.44),(26.08±8.49),差異有統(tǒng)計(jì)學(xué)意義(p0.05)。結(jié)論ADAR1基因可能與子宮內(nèi)膜異位癥和子宮腺肌病的發(fā)病及發(fā)生發(fā)展有一定關(guān)系。為診治子宮內(nèi)膜異位癥和子宮腺肌病提供了新思路。
[Abstract]:Endometriosis (endometriosis, EMs) referred to as endometriosis is a common gynecological disease, with endometrial tissue outside the uterus growth characteristics, such as ovarian, tubal, pelvic and abdominal cavity, which is the most common ovarian adenomyosis (adenomyosis, AM) refers to the uterus the growth of endometrial glands and interstitial myometrium, with menorrhagia, menostaxis and.AM of common gynecological diseases of secondary dysmenorrhea is the main clinical manifestation is believed to be an endometriosis, its pathogenesis there are some similarities between.EMs and AM is described as non neoplastic the disease, but its biological behavior is consistent with the characteristic of malignant tumors, such as proliferation, invasion and invasion. In recent years, the study also showed that cellular immunity and humoral immunity disorders are likely to occurrence and development of Ems and AM are . the domestic and foreign related ADAR1 in endometriosis and adenomyosis pathogenesis has not been reported that.ADAR1 is a RNA editing enzyme, RNA editing is the modification and processing of the RNA transcription of DNA into RNA, may make the gene expression in the level of DNA expression and protein level in a completely different, so as to change the study found that the genetic information of [2-6]. ADAR1 in [7] cells, play a very important role in the differentiation of [8] and [9] in tumor development. In addition, ADAR1 with inflammation and immunity is also closely related to the experiment of [10]. by real-time quantitative PCR, Western blotting and immunohistochemical method to detect expression of ADAR1 in endometrium and endometriosis and adenomyosis in the reign of patients with endometriosis and adenomyosis, to further explore the pathogenesis of the two diseases in order to provide new ideas for the clinical diagnosis and treatment. Objective to study the ADAR1 in endometriosis and sub The difference in the expression of uterine adenomyosis, to explore the relationship between ADAR1 and endometriosis and adenomyosis and its significance. Materials and methods the study by the Third Affiliated Hospital of Zhengzhou University, Henan Provincial Maternal and child health hospital ethics committee approval, all patients signed the informed consent of.1 from July 2015 to October 2016 in the Affiliated Hospital of Henan Province third Zhengzhou University (Henan Provincial Maternal and child health hospital) 31 EMs patients in the treatment of gynecological surgery patients, aged 29~44 years, surgery confirmed endometriosis III type or IV type (as defined by the American Society for Reproductive Medicine), there are also 25 cases of hysteroscopy for patients with fertility requirements of the endometrial curettage obtained the organization (group EU), and the ectopic endometrial tissue during the operation (group EC). Selected uterine gland in the Affiliated Hospital of Zhengzhou University of Henan province from third hysterectomy muscle In 30 cases, patients aged 38~48, immediately after hysterectomy patients in eutopic endometrium (experimental group, AM1) and adenomyoma tissues (experimental group AM2); 30 cases in the control group for the same period of uterine fibroids hysterectomy patients, aged 34~49 years old, hysterectomy immediately after taking eutopic endometrium (control group C). The patients with regular menstrual cycle, 26~32 days; six months before the operation without hormone and endocrine disease medication history, pregnancy, lactation, pre menopausal symptoms. All the specimens were obtained in the proliferative phase of endometrium, and all pathologically confirmed.2 specimens processed in aseptic conditions of endometrium tissue or leiomyoma, half immediately placed in liquid nitrogen, and then transferred to -80 DEG C for preservation of real-time fluorescence quantitative PCR and Western blot experiments, the other half in 10% formalin fixed paraffin embedded, 2.1 for real-time.3 test method to examine the histology Quantitative PCR Technology (qRT-PCR) to detect the ADAR1 protein expression detected by semi quantitative.2.2 Western blotting groups ADAR1mRNA.2.3 expression level in each group by immunohistochemistry ADAR1 protein expression and localization analysis of.4 semi quantitative statistical analysis the experimental data to mean + standard deviation (x + s), were analyzed by SPSS 21 software. Between the 22 groups were compared with t test, meet the normality and homogeneity of variance were analyzed by single factor analysis of variance, if not satisfied with independent samples K test or Kruskal-Wallis rank sum test. The test level of alpha =0.05. 22 compared with Bonferroni method, using P alpha values. There was significant difference between the 1 the age group of endometriosis group (36.4 + 3.8), adenomyosis group (39.13 + 5.61) and control group (40.3 + 3.4), there was no significant difference between groups (P0.05).2 ADAR1mRNA in endometriosis and uterus 鑵鴻倢鐥呬腑鐨勮〃杈懼疄鏃惰崸鍏夊畾閲廝CR緇撴灉鏄劇ずADAR1 P150mRNA鍦‥C(2.33鹵0.28)緇勫拰EU(2.18鹵0.36)緇勭殑琛ㄨ揪鏄庢樉楂樹簬C(1.68鹵0.43)緇，

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