【摘要】：研究背景：血管平滑肌細(xì)胞(vascular smooth muscle cells, VSMCs)的表型變化引起的VSMCs異常增殖是動(dòng)脈粥樣硬化(atherosclerosis,AS)、腦梗死等心腦血管疾病的重要病理基礎(chǔ),但其表型調(diào)節(jié)的機(jī)制仍未被完全闡釋清楚。miR-145是一種與心血管疾病密切相關(guān)的microRNA,主要在VSMCs中表達(dá),對(duì)VSMCs具有表型調(diào)節(jié)作用。CD40、CD40L在VSMCs中均有表達(dá),且CD40-CD40L系統(tǒng)激活后具有促炎及促VSMCs增殖作用,在AS的發(fā)生發(fā)展過(guò)程中起重要作用。生物信息學(xué)軟件分析CD40可能是miR-145重要的靶基因,miR-145可能通過(guò)對(duì)CD40的調(diào)節(jié)作用介導(dǎo)VSMCs的增殖及炎癥因子生成。 阿司匹林是臨床上用于治療腦梗死的常用藥物。文獻(xiàn)報(bào)道阿司匹林具有抗血栓及抗炎作用,還可以抑制VSMCs增殖。但是阿司匹林抑制VSMCs增殖的機(jī)制仍不清楚。阿司匹林抗血栓及抗炎作用與調(diào)節(jié)機(jī)體CD40-CD40L系統(tǒng)的活性有關(guān),但尚無(wú)阿司匹林是否能影響VSMCs中CD40的水平及其與miR-145關(guān)系的報(bào)道。 研究目的：1.探討miR-145是否通過(guò)對(duì)CD40的調(diào)節(jié)介導(dǎo)VSMCs的增殖及炎癥因子生成,為miR-145在心血管疾病中的研究提供參考；2.探討miR-145是否通過(guò)對(duì)CD40的調(diào)節(jié)參與阿司匹林的抗VSMCs增殖及抗炎作用,為闡述阿司匹林的新作用機(jī)制提供理論依據(jù)。 實(shí)驗(yàn)方法：1.miR-145通過(guò)調(diào)節(jié)CD40介導(dǎo)VSMCs的增殖及炎癥因子生成：培養(yǎng)人血管平滑肌細(xì)胞(human vascular smooth muscle cells,HAVSMCs),應(yīng)用TNF-α建立細(xì)胞增殖模型,運(yùn)用miR-145的模擬物轉(zhuǎn)染細(xì)胞,分組為：(1)空白對(duì)照組(Control)、(2)造模組(TNF-α)、(3)轉(zhuǎn)染miR-145模擬物對(duì)照造模組(miR-145mimic control+TNF-α)、(4)轉(zhuǎn)染miR-145模擬物造模組(miR-145mimic+TNF-α)。EdU檢測(cè)細(xì)胞增殖情況,Real-time PCR法檢測(cè)miR-145、CD40、VSMC分化標(biāo)志基因Calponin的mRNA表達(dá),Western-blot法檢測(cè)CD40的蛋白表達(dá),ELISA檢測(cè)細(xì)胞上清炎性因子IL-6的濃度；2.miR-145通過(guò)對(duì)CD40的調(diào)節(jié)參與阿司匹林的抗VSMCs增殖及抗炎作用：培養(yǎng)HAVSMCs,應(yīng)用TNF-α建立細(xì)胞增殖模型,進(jìn)行藥物研究。分組為：(1)溶媒對(duì)照組(DMSO)、(2)造模組(TNF-α)、(3)轉(zhuǎn)染miR-145抑制劑對(duì)照造模組(miR-145inhibitor control+TNF-α)、(4)阿司匹林預(yù)處理造模組(ASA+TNF-α)、(5)轉(zhuǎn)染miR-145抑制劑及給藥阿司匹林預(yù)處理造模組(miR-145inhibitor+ASA+TNF-α)。EdU檢測(cè)細(xì)胞增殖情況,Real-time PCR法檢測(cè)miR-145、CD40、VSMC分化標(biāo)志基因Calponin的mRNA表達(dá),Western-blot法檢測(cè)CD40的蛋白表達(dá),ELISA檢測(cè)細(xì)胞上清炎性因子IL-6的濃度。 實(shí)驗(yàn)結(jié)果：1.miR-145通過(guò)調(diào)節(jié)CD40介導(dǎo)VSMCs的增殖及炎癥因子生成：過(guò)表達(dá)miR-145能抑制TNF-α誘導(dǎo)的HAVSMCs增殖,逆轉(zhuǎn)TNF-α引起的平滑肌細(xì)胞分化標(biāo)志基因Calponin的下調(diào)及逆轉(zhuǎn)TNF-α引起的CD40的升高,可以降低VSMCs激活后釋放的IL-6水平。2.miR-145通過(guò)對(duì)CD40的調(diào)節(jié)參與阿司匹林的抗VSMCs增殖及抗炎作用：TNF-α引起VSMCs增殖,平滑肌細(xì)胞分化標(biāo)志基因Calponin下調(diào),miR-145水平下調(diào)及CD40水平的上調(diào),并伴有IL-6水平顯著升高；預(yù)先使用20μg/mL的阿司匹林處理VSMCs后可明顯抑制TNF-a誘導(dǎo)的VSMCs增殖并升高Calponin的表達(dá),同時(shí)阿司匹林還能夠升高miR-145的水平,下調(diào)CD40及IL-6的水平 結(jié)論：1.miR-145通過(guò)抑制CD40的表達(dá)起抗VSMCs增殖及抗VSMCs炎癥因子生成的作用；2.miR-145通過(guò)抑制HAVSMCs中CD40的表達(dá)參與阿司匹林的抗VSMCs增殖及抗炎作用。
[Abstract]:Background: The abnormal proliferation of VSMCs caused by the phenotypic change of vascular smooth muscle cells (VSMCs) is an important pathological basis for cardiovascular and cerebrovascular diseases such as atherosclerosis (AS) and cerebral infarction, but the mechanism of phenotypic regulation is not fully explained. MiR-145 is a microRNA which is closely related to cardiovascular diseases, is mainly expressed in VSMCs, and has a phenotypic regulation effect on VSMCs. CD40 and CD40L are expressed in VSMCs, and it plays an important role in the development of AS after activation of the CD40-CD40L system. The analysis of CD40 by bioinformatics software may be an important target gene for miR-145, and miR-145 may mediate the proliferation of VSMCs and the production of inflammatory factors through the regulation of CD40. Aspirin is a common drug used in the treatment of cerebral infarction It is reported that aspirin has antithrombotic and anti-inflammatory effects and can inhibit the proliferation of VSMCs. However, the mechanism of aspirin to inhibit the proliferation of VSMCs is still unclear. The anti-thrombotic and anti-inflammatory effects of aspirin are related to the regulation of the activity of CD40-CD40L system in the body, but no aspirin can affect the level of CD40 in the VSMCs and its relationship with the miR-145. A. The study 1. To investigate whether miR-145 is produced by regulating the proliferation and inflammatory factors of the VSMCs mediated by the regulation of CD40, and to provide reference for the study of miR-145 in the cardiovascular disease. 2. To investigate whether miR-145 has been involved in the anti-VSMCs proliferation and anti-inflammatory effects of aspirin on the regulation of CD40, and to provide a basis for elucidating the new mechanism of aspirin. On the basis of the experimental method,1. The expression of miR-145 by regulating the proliferation of the CD40-mediated VSMCs and the production of the inflammatory factors: the human vascular smooth muscle cells (HAVSMCs) were cultured, the cell proliferation model was established by using the TNF-145, the cells were transfected with the mimetic of the miR-145, and the group was: (1) the blank control group (Con (l), (2) Construction module (TNF-1), (3) Transfection of miR-145 mimetic control (miR-145 mic control + TNF-1), (4) Transfection of miR-145 mimetic (miR-145 mic + T) The expression of the expression of the expression of the expression of the expression of the expression of calcin in the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the expression of the VSMCs, the application of TNF-1 to establish a cell proliferation model, The group was: (1) Vehicle control group (DMSO), (2) Construction module (TNF-1), (3) Transfection of miR-145 inhibitor control building module (miR-145 in control + TNF-1), (4) Aspirin pre-treatment module (ASA + T NF-1, (5) Transfection of miR-145 inhibitor and administration of aspirin pre-treatment module (miR-145 in hibitor + ASA + T The expression of the expression of the expression of calcin in the expression of miR-145, CD40, and VSMC was detected by the real-time PCR, and the expression of the protein of CD40 was detected by Western-blot. -6. Experimental results:1. miR-145 is generated by regulating the proliferation and the inflammatory factors of the CD40-mediated VSMCs: the overexpression of the miR-145 can inhibit the proliferation of the TNF-1-induced HAVSMCs, reverse the down-regulation of the differentiation marker gene Calcin of the smooth muscle cells induced by the TNF-1 and the reversal of the TNF-1 induced by the TNF-145. The increase of CD40 can reduce the level of IL-6 released after activation of VSMCs.2. miR-145 is involved in the anti-VSMCs proliferation and anti-inflammatory effects of aspirin on the regulation of CD40: TNF-1 causes the proliferation of VSMCs, the down-regulation of the differentiation marker gene of smooth muscle cells, the down-regulation of miR-145 and the up-regulation of the level of CD40, and is accompanied by an IL- 6. The level of VSMCs induced by TNF-a was significantly inhibited after treatment with 20. m u.g/ mL of aspirin, and the expression of Calcin was increased, while aspirin was able to increase the level of miR-145 and down-regulate CD40. and IL -6. Horizontal conclusion:1. miR-145 acts to inhibit the proliferation of the anti-VSMCs and the anti-VSMCs inflammatory factors by inhibiting the expression of CD40;2. miR-145 inhibits the anti-VSM of aspirin by inhibiting the expression of CD40 in the HAVSMCs
【學(xué)位授予單位】：中南大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2014
【分類號(hào)】：R969

【相似文獻(xiàn)】

相關(guān)期刊論文 前4條

1 王穎懿;房林;沈磊;錢春妹;宋力雯;;miR-145、miR-375在乳腺浸潤(rùn)性導(dǎo)管癌中的表達(dá)及臨床意義[J];同濟(jì)大學(xué)學(xué)報(bào)(醫(yī)學(xué)版);2012年03期

2 張晴;吳正升;張瑰紅;丁向東;黃焰;楊楓;徐曉春;吳強(qiáng);;miR-145在乳腺癌中表達(dá)及意義[J];臨床與實(shí)驗(yàn)病理學(xué)雜志;2009年01期

3 郭廣秀;張功亮;陳青;;miR-145與腫瘤研究進(jìn)展[J];江西醫(yī)藥;2011年11期

4 董文;林天歆;黃健;廖蓓;許可慰;江春;張彩霞;劉皓;;miR-143和miR-145在膀胱癌中的表達(dá)及意義[J];中山大學(xué)學(xué)報(bào)(醫(yī)學(xué)科學(xué)版);2011年06期

相關(guān)會(huì)議論文 前2條

1 馮陽(yáng);曹建華;李新云;趙書紅;;miR-145在豬骨骼肌發(fā)育和成肌細(xì)胞分化不同時(shí)期的差異表達(dá)分析[A];中國(guó)遺傳學(xué)會(huì)模式生物與人類健康研討會(huì)會(huì)議論文集[C];2010年

2 馮陽(yáng);曹建華;李新云;趙書紅;;miR-145在豬骨骼肌發(fā)育和成肌細(xì)胞分化不同時(shí)期的差異表達(dá)分析[A];中國(guó)遺傳學(xué)會(huì)模式生物與人類健康研討會(huì)會(huì)議論文集[C];2010年

相關(guān)碩士學(xué)位論文 前4條

1 陳敏;miR-145介導(dǎo)阿司匹林抗血管平滑肌細(xì)胞增殖及抗炎作用的研究[D];中南大學(xué);2014年

2 鄧鑫;miR-145對(duì)前列腺癌PC-3細(xì)胞增殖和凋亡及其調(diào)控基因表達(dá)的影響[D];蘇州大學(xué);2014年

3 陳嘉佳;miR-145靶向FSCN1在不同生長(zhǎng)方式胃癌中的作用及機(jī)制[D];廈門大學(xué);2014年

4 張鴻博;miR-145對(duì)宮頸癌Hela細(xì)胞的增殖、凋亡及其對(duì)紫杉醇藥物敏感性的研究[D];吉林大學(xué);2014年

，

本文編號(hào)：2489981