本文選題：消癥方 + 子宮腺肌病�。� 參考：《廣州中醫(yī)藥大學(xué)》2017年碩士論文

【摘要】：目的:通過(guò)采用膠原酶消化法處理子宮腺肌病在位及異位病灶子宮內(nèi)膜組織,體外分離培養(yǎng)腺肌病在位及異位子宮內(nèi)膜間質(zhì)細(xì)胞,觀察腺肌病內(nèi)膜間質(zhì)細(xì)胞的生長(zhǎng)增殖曲線,通過(guò)將藥物直接作用于細(xì)胞,采用MTT法檢測(cè)藥物對(duì)細(xì)胞的增殖抑制作用,顯微鏡下觀察凋亡細(xì)胞形態(tài),采用流式細(xì)胞術(shù)檢測(cè)細(xì)胞凋亡及細(xì)胞周期,觀察消癥方對(duì)腺肌病在位及異位子宮內(nèi)膜間質(zhì)細(xì)胞增殖和凋亡的影響,闡釋消癥方治療子宮腺肌病的作用機(jī)理,為臨床治療子宮腺肌病提供實(shí)驗(yàn)論證和理論基礎(chǔ),為中醫(yī)采用消ve化瘀法治療子宮腺肌病賦予新的內(nèi)涵,為子宮腺肌病的保守治療提供新的思路。方法:1.本實(shí)驗(yàn)研究選擇2014年12月—2015年12月在廣東省中醫(yī)院大德路總院婦科行全子宮切除術(shù),且術(shù)后病理診斷為子宮腺肌病的患者6例,子宮全切后,無(wú)菌操作分別取在位子宮內(nèi)膜、異位病灶內(nèi)膜組織,采用膠原酶消化法體外分離培養(yǎng)子宮腺肌病在位及異位內(nèi)膜間質(zhì)細(xì)胞,并采用免疫組織化學(xué)方法對(duì)細(xì)胞進(jìn)行純度鑒定。2.采用水提法制備含藥培養(yǎng)液,并將實(shí)驗(yàn)分為5組:消癥方高劑量組、消癥方中劑量組、消癥方低劑量組、孕三烯酮組以及空白對(duì)照組。通過(guò)將藥物直接作用于子宮腺肌病在位及異位內(nèi)膜間質(zhì)細(xì)胞24h,采用MTT法觀察消癥方含藥培養(yǎng)液對(duì)腺肌病細(xì)胞的增殖抑制作用,采用蘇木素-伊紅染色(HE染色)方法顯微鏡下觀察消癥方直接作用于細(xì)胞之后的凋亡細(xì)胞形態(tài);同時(shí)利用流式細(xì)胞儀檢測(cè)藥物對(duì)細(xì)胞凋亡、細(xì)胞周期的作用。成果:1.本實(shí)驗(yàn)研究通過(guò)采用Ⅰ型膠原酶消化、篩網(wǎng)過(guò)濾等方法原代培養(yǎng)子宮腺肌病在位及異位子宮內(nèi)膜間質(zhì)細(xì)胞,取得標(biāo)本共6例,最終培養(yǎng)成功5例。并通過(guò)選用對(duì)腺肌病間質(zhì)細(xì)胞具有特異性的波形蛋白,對(duì)原代培養(yǎng)的腺肌病在位及異位內(nèi)膜細(xì)胞進(jìn)行免疫組織化學(xué)方法鑒定,由細(xì)胞鑒定可知,原代培養(yǎng)出的子宮腺肌病在位及異位內(nèi)膜細(xì)胞中內(nèi)膜間質(zhì)細(xì)胞的純度均高達(dá)99%以上。2.消癥方及孕三烯酮組均可抑制子宮腺肌病在位及異位內(nèi)膜間質(zhì)細(xì)胞的增殖,且消癥方在一定范圍內(nèi)能呈劑量依賴性的抑制腺肌病細(xì)胞增殖。3.消癥方組及孕三烯酮組顯微鏡下均可觀察到:部分細(xì)胞變得固縮,細(xì)胞質(zhì)變得致密,細(xì)胞核染色質(zhì)濃集,為凋亡細(xì)胞的形態(tài)學(xué)特征。4.消癥方對(duì)子宮腺肌病在位及異位內(nèi)膜間質(zhì)細(xì)胞的凋亡的流式細(xì)胞術(shù)檢測(cè):藥物直接作用于腺肌病細(xì)胞24h后,與空白對(duì)照組相比,消癥方高、中、低劑量組及孕三烯酮組均能顯著促進(jìn)在位及異位內(nèi)膜間質(zhì)細(xì)胞凋亡(P0.05),且消癥方組隨著藥物濃度的升高,凋亡率也隨之增加。5.消癥方對(duì)子宮腺肌病在位及異位內(nèi)膜間質(zhì)細(xì)胞的細(xì)胞周期的流式細(xì)胞術(shù)檢測(cè):消癥方高、中、低劑量組及孕三烯酮組均可引起腺肌病在位及異位內(nèi)膜間質(zhì)細(xì)胞發(fā)生S期阻滯,且消癥方組阻滯細(xì)胞周期呈一定的濃度依賴性。結(jié)論:1.消癥方能有效抑制子宮腺肌病在位及異位內(nèi)膜間質(zhì)細(xì)胞的增殖,對(duì)子宮腺肌病在位及異位內(nèi)膜間質(zhì)細(xì)胞的生長(zhǎng)有促進(jìn)其凋亡的作用,并且能通過(guò)將子宮腺肌病在位及異位內(nèi)膜間質(zhì)細(xì)胞阻斷于S期以阻斷周期進(jìn)程。2.消癥方治療子宮腺肌病的作用可能與其通過(guò)抑制子宮腺肌病在位及異位內(nèi)膜間質(zhì)細(xì)胞增殖、促進(jìn)細(xì)胞凋亡、阻斷周期進(jìn)程有關(guān),從細(xì)胞生物學(xué)方面解釋了消癥方限制子宮腺肌病發(fā)生和發(fā)展的過(guò)程,為消癥方臨床治療子宮腺肌病提供了實(shí)驗(yàn)論證及理論基礎(chǔ),為臨床對(duì)子宮腺肌病的治療提供了新的思路。
[Abstract]:Objective: to treat endometriosis and ectopic endometrium of adenomyosis by collagenase digestion, in vitro and in vitro culture of adenomyosis in vitro and ectopic endometrial stromal cells, and observe the growth and proliferation of endometrial stromal cells in adenomyosis. By direct action of drugs to cells, the proliferation of cells was detected by MTT method. The morphology of apoptotic cells was observed under microscope, apoptosis and cell cycle were detected by flow cytometry. The effect of Xiaozheng prescription on the proliferation and apoptosis of endometrial stromal cells in adenomyosis and ectopic endometrium was observed. The mechanism of Xiaozheng prescription in the treatment of adenomyosis was explained, and the experimental demonstration of the clinical treatment of adenomyosis and the clinical treatment of adenomyosis were provided. The theory foundation provides new connotation for the treatment of adenomyosis by eliminating ve and removing stasis in traditional Chinese medicine. It provides a new idea for the conservative treatment of uterine adenomyosis. Methods: 1. experimental studies were selected from December 2014 to December 2015 in the Department of gynecology in the General Hospital of Guangdong Province Traditional Chinese Medical Hospital, and the pathological diagnosis of uterine adenomyosis after operation was made. In 6 patients, after total hysterectomy, the eutopic endometrium, endometrium and ectopic endometrium were isolated and cultured by collagenase digestion. The purity of the cells was identified by immunohistochemical method and.2. was prepared by the method of immunohistochemistry. The 5 groups: the high dose group of Xiaozheng Fang, the dose group of Xiaozheng Fang, the low dose group of Xiaozheng Fang, the pregnant three ketone group and the blank control group. Through the direct action of the drug on the eutopic and ectopic endometrium interstitial cells 24h of the uterine adenomyosis, the inhibitory effect of the culture solution of Xiaozheng Fang on the proliferation of adenomyosis cells was observed by the method of MTT, and the hematoxylin eosin was used. The staining (HE staining) method was used to observe the morphology of the apoptotic cells directly acting on the cells after the elimination of the disease. At the same time, the effect of drugs on cell apoptosis and cell cycle was detected by flow cytometry. The results of the 1. experimental studies were the primary culture of adenomyosis in situ and ectopia by using type I collagenase digestion and sieve filtration. A total of 6 cases of endometrial stromal cells were obtained, and 5 cases were successfully cultured. By selecting the specific vimentin for the interstitial cells of adenomyosis, the primary cultured adenomyosis was identified by immunohistochemistry. The primary and ectopic endometriosis of the primary cultured adenomyosis was identified by cell identification. The purity of endometrial stromal cells in membrane cells is more than 99%.2. and three ketones can inhibit the proliferation of endometriosis and ectopic endometrium stromal cells, and the elimination side can be observed under a certain range of dose-dependent inhibition of adenomyosis cell proliferation.3. elimination group and pregnancy three ketone group under microscope. Some cells become compact, the cytoplasm becomes dense, the nucleus chromatin is dense, and the morphological feature of the apoptotic cells is the flow cytometry of the apoptosis of the endometriosis and ectopic endometrium cells of.4.. The drug is directly acted on the 24h of the adenomyosis cells and compared with the blank control group, the high, middle, and low dose groups are compared with the blank control group. And gestation three ketone group can significantly promote the apoptosis of eutopic and ectopic endometrium stromal cells (P0.05). With the increase of drug concentration, the rate of apoptosis also increases the flow cytometry of.5. Xiao Zheng on the cell cycle of endometriosis and ectopic endometrial stromal cells: high, middle, low dose, and three ketones. The group of adenomyosis can cause S phase block in the eutopic and ectopic endometrial stromal cells, and the cell cycle of the Xiaozheng group has a certain concentration dependence. Conclusion: the 1. side can effectively inhibit the proliferation of the eutopic and ectopic endometrium stromal cells of the uterus adenomyosis, and promote the growth of the adenomyosis in the position and the ectopic endometrium cells. The effect of apoptosis, and the effect of blocking the interstitial cells of endometriosis and ectopic endometrium by blocking the interstitial cells of the endometrium in the S stage to block the periodic process of.2. elimination in the treatment of adenomyosis may be related to the inhibition of the proliferation of endometriosis and ectopic endometrium cells, the promotion of cell apoptosis and the interruption of the cycle process, from cell biology. It explains the process of restricting the occurrence and development of adenomyosis by Xiaozheng party. It provides an experimental demonstration and theoretical basis for the clinical treatment of adenomyosis by Xiaozheng party, and provides a new way of thinking for the treatment of adenomyosis in clinical.

【學(xué)位授予單位】：廣州中醫(yī)藥大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：R271.9

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