發(fā)布時(shí)間：2018-03-19 13:44

  本文選題：SHP-2　切入點(diǎn)：miR-100　出處：《安徽醫(yī)科大學(xué)》2015年碩士論文　論文類型：學(xué)位論文

【摘要】：腫瘤是嚴(yán)重威脅人類健康的重大疾病,不僅發(fā)病率和死亡率逐年上升,近年來(lái)更呈現(xiàn)出高發(fā)和早發(fā)的趨勢(shì)。而腫瘤的發(fā)生是多因素作用,經(jīng)過(guò)多階段的長(zhǎng)期的復(fù)雜過(guò)程,受環(huán)境因素與基因因素的共同調(diào)控。因此,研究?jī)烧吖餐饔孟履[瘤發(fā)生發(fā)展的分子機(jī)制對(duì)解決腫瘤的高發(fā)和早發(fā)問(wèn)題至關(guān)重要。SHP-2是由PTPN11編碼的非受體型酪氨酸磷酸酶,參與了包括P13K.MAPK. JAK/STAT等多種信號(hào)通路的活化。最初在努南綜合征中發(fā)現(xiàn)其突變體的存在,后來(lái)在多種血液病及腫瘤中也發(fā)現(xiàn)了這一突變體的存在,并且這一突變體對(duì)腫瘤的發(fā)生發(fā)展有著至關(guān)重要的作用。SHP-2最常見(jiàn)的激活突變體是D61G/D61Y,這也是我們一直致力于研究的一個(gè)突變體。砷主要存在于水源和土壤中,是國(guó)際公認(rèn)的的化學(xué)致癌物,與皮膚癌、肺癌等多種腫瘤的發(fā)生發(fā)展密切相關(guān)。我們實(shí)驗(yàn)室前期的研究過(guò)程中發(fā)現(xiàn)激活突變SHP-2酪氨酸磷酸酶促進(jìn)砷誘導(dǎo)的MEFs惡性轉(zhuǎn)化。在對(duì)其機(jī)制進(jìn)行初步探究的過(guò)程中發(fā)現(xiàn)：As203急性處理MEF細(xì)胞48h后,miR-100在As203誘導(dǎo)前后的細(xì)胞或SHP2激活突變與否的細(xì)胞出現(xiàn)減少,且受三氧化二砷和SHP2激活突變影響變化一致。MiR-100是小的非編碼RNA,已經(jīng)被報(bào)道在不同腫瘤中通過(guò)靶向不同的靶蛋白而發(fā)揮著致癌基因或是抑癌基因的作用。包括乳腺癌、肺癌、宮頸癌、淋巴瘤在內(nèi)的多種腫瘤的發(fā)生發(fā)展都與miR-100的異常表達(dá)有關(guān)。但是關(guān)于環(huán)境污染與基因突變共同作用下腫瘤的發(fā)生過(guò)程中miR-100的作用機(jī)制并不明確。我們?cè)O(shè)想miR-100可能參與了激活突變SHP-2酪氨酸磷酸酶促進(jìn)砷誘導(dǎo)的MEFs惡性轉(zhuǎn)化。為了驗(yàn)證這一假說(shuō),首先,我們采用RT-PCR技術(shù)檢測(cè)了SHP-2+/+、SHP-2D61G/+以及其對(duì)應(yīng)的As203慢性處理40代的SHP-2+/+-40、SHP-2D61G/+-40細(xì)胞中miR-100的表達(dá)水平,結(jié)果發(fā)現(xiàn)miR-100在四種細(xì)胞中的表達(dá)逐漸下降,提示miR-100參與了MEF細(xì)胞的惡性轉(zhuǎn)化過(guò)程。接下來(lái),為了明確miR-100在其中發(fā)揮的效應(yīng),本實(shí)驗(yàn)通過(guò)轉(zhuǎn)染miR-100inhibitor或miR-100 mimics改變相應(yīng)的MEF細(xì)胞中miR-100的表達(dá)水平,結(jié)果發(fā)現(xiàn)miR-100可以減少細(xì)胞的克隆形成能力和非錨定生長(zhǎng)能力以及體外遷移和增殖能力。雙熒光素酶報(bào)告基因?qū)嶒?yàn)發(fā)現(xiàn)IGF1Rβ和nTOR是miR-100靶基因,WB實(shí)驗(yàn)表明IGF1Rβ在MEFs細(xì)胞中的表達(dá)隨著As2O3誘導(dǎo)SHP-2D61G/+激活突變表達(dá)逐漸升高。最后,采用攜帶有目的基因的慢病毒載體感染細(xì)胞建立穩(wěn)轉(zhuǎn)細(xì)胞系,細(xì)胞學(xué)實(shí)驗(yàn)得到與上述同樣的結(jié)果。以上結(jié)果表明：miR-100參與激活突變SHP-2酪氨酸磷酸酶促進(jìn)砷誘導(dǎo)的MEFs惡性轉(zhuǎn)化過(guò)程,并在其中發(fā)揮抑癌基因的作用。
[Abstract]:Cancer is a serious disease that threatens human health seriously. Not only the incidence and mortality rate increase year by year, but also the tendency of high incidence and early onset in recent years. Therefore, it is very important to study the molecular mechanism of tumorigenesis and development under the interaction of environmental and genetic factors. SHP-2 is a non-recipient tyrosine phosphatase encoded by PTPN11. Involved in the activation of multiple signaling pathways, including P13K.MAPK. JAK/STAT. The mutant was initially found in Nunan syndrome and later found in various hematological diseases and tumors. The most common active mutants of SHP-2 are D61G / D61Y, which is one of the mutants that we have been working on. Arsenic is mainly found in water and soil. Is an internationally recognized chemical carcinogen, associated with skin cancer, SHP-2 tyrosine phosphatase is found to promote arsenic induced malignant transformation of MEFs. In the course of preliminary study on the mechanism, we found that the activation mutation of SHP-2 tyrosine phosphatase promotes arsenic induced malignant transformation of MEFs. It was found that after 48 hours of acute treatment of MEF cells with 1: As203, the cells of miR-100 before and after As203 induction or those with or without SHP2 activation mutation were decreased. MiR-100, a small noncoding RNAs, has been reported to play the role of oncogene or tumor suppressor gene by targeting different target proteins in different tumors, including breast cancer and lung cancer. Cervical cancer, The occurrence and development of various tumors, including lymphoma, are related to the abnormal expression of miR-100. However, the mechanism of miR-100 in tumorigenesis under the action of environmental pollution and gene mutation is not clear. We assume that miR-100 may play an important role in the development of tumor. Participate in the activation of mutated SHP-2 tyrosine phosphatase to promote arsenic induced malignant transformation of MEFs. Firstly, we detected the expression of miR-100 in the SHP-2 / -40 SHP-2D61G / -40 cells treated with SHP-2 / HP-2D61G/ and its corresponding As203 chronically by RT-PCR technique. The results showed that the expression of miR-100 in four kinds of cells decreased gradually. The results suggest that miR-100 is involved in the malignant transformation of MEF cells. In order to understand the effect of miR-100, the expression of miR-100 in MEF cells was altered by transfection of miR-100inhibitor or miR-100 mimics. The results showed that miR-100 could reduce the ability of cell clone formation, unanchored growth, migration and proliferation in vitro. Double luciferase reporter gene assay showed that IGF1R 尾 and nTOR were miR-100 target genes in MEFs cells. The expression of SHP-2D61G/ activation mutation increased with the induction of As2O3. Finally, Stable transformed cell lines were established by infection with lentivirus vector carrying the target gene, and the same results were obtained by cytological experiments. The above results indicated that: miR-100 was involved in the activation of mutated SHP-2 tyrosine phosphatase to promote arsenic induced malignant transformation of MEFs. And play the role of tumor suppressor gene.
【學(xué)位授予單位】：安徽醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2015
【分類號(hào)】：R730.2

【相似文獻(xiàn)】

相關(guān)期刊論文 前7條

1 胡波;Shp-2酪氨酸磷酸酶在細(xì)胞信號(hào)傳導(dǎo)的作用[J];實(shí)用醫(yī)藥雜志;2003年05期

2 王莉;楊艷麗;宋子博;廖世秀;黃飛飛;;酪氨酸磷酸酶PRL-3單抗的制備及特性鑒定[J];腫瘤防治研究;2009年11期

3 王小明，，肖殿模;蛋白酪氨酸磷酸酶在疾病發(fā)生中的意義[J];基礎(chǔ)醫(yī)學(xué)與臨床;1996年02期

4 郝冬梅;酪氨酸磷酸酶在腫瘤發(fā)生中的作用[J];國(guó)外醫(yī)學(xué).遺傳學(xué)分冊(cè);1995年02期

5 王鳳玫;SHP-2酪氨酸磷酸酶:信號(hào)轉(zhuǎn)導(dǎo)機(jī)制及生物學(xué)作用[J];醫(yī)學(xué)綜述;2003年05期

6 張梅;李菲菲;鄭紅;倪芳;瞿成奎;汪思應(yīng);;SHP-2酪氨酸磷酸酶在DNA損傷性放化療引發(fā)的小鼠骨髓毒中的作用[J];安徽醫(yī)科大學(xué)學(xué)報(bào);2008年05期

7 ;[J];;年期

相關(guān)會(huì)議論文 前3條

1 張宏利;朱達(dá);駱天紅;王琴琴;趙列賓;蘇莉珍;陸駱;戴蒙;李紀(jì)平;劉優(yōu)萍;楊鍵;劉峗;江凌;李果;羅敏;;酪氨酸磷酸酶-1B基因多態(tài)性與2型糖尿病和肥胖的相關(guān)性研究[A];全國(guó)首屆代謝綜合征的基礎(chǔ)與臨床專題學(xué)術(shù)會(huì)議論文匯編[C];2004年

2 曾王勇;汪迎華;吳季輝;戴懷恩;施蘊(yùn)渝;;蛋白質(zhì)雙功能酪氨酸磷酸酶VHR的結(jié)構(gòu)與功能[A];中國(guó)生物化學(xué)與分子生物學(xué)會(huì)第八屆會(huì)員代表大會(huì)暨全國(guó)學(xué)術(shù)會(huì)議論文摘要集[C];2001年

3 彭麗蓉;邢曉芳;王莉;吳健;孟麟;壽成超;;酪氨酸磷酸酶PRL-3在腫瘤組織表達(dá)的臨床意義及促腫瘤轉(zhuǎn)移的分子機(jī)制探討[A];中國(guó)病理生理學(xué)會(huì)受體、腫瘤和免疫專業(yè)委員會(huì)聯(lián)合學(xué)術(shù)會(huì)議論文匯編[C];2010年

相關(guān)重要報(bào)紙文章 前1條

1 黎彬；相洪琴;抗癌研究不斷求解[N];中國(guó)醫(yī)藥報(bào);2005年

相關(guān)碩士學(xué)位論文 前2條

1 朱其蘋;MiR-100參與激活突變SHP-2酪氨酸磷酸酶促進(jìn)砷誘導(dǎo)的MEFs惡性轉(zhuǎn)化[D];安徽醫(yī)科大學(xué);2015年

2 劉晶;淋巴特異的酪氨酸磷酸酶（Lyp）不同突變（S201F、R263Q和R266W）的生化特性與細(xì)胞功能研究[D];山東大學(xué);2012年

本文編號(hào)：1634527