本文選題：基底剛度 + 肌球蛋白Ⅱ��； 參考：《太原理工大學(xué)》2017年碩士論文

【摘要】：細(xì)胞外基質(zhì)是一個(gè)復(fù)雜的大分子網(wǎng)絡(luò),具有獨(dú)特的物理特性、生物化學(xué)特性和力學(xué)特性,這些性質(zhì)能夠直接或間接地影響細(xì)胞的生物學(xué)行為,成為正常組織發(fā)育必不可少的因素。當(dāng)細(xì)胞外基質(zhì)的代謝發(fā)生異常時(shí),細(xì)胞外基質(zhì)的組織結(jié)構(gòu)和力學(xué)特性發(fā)生改變,同時(shí)伴隨一些疾病的發(fā)生。例如組織的纖維化、癌細(xì)胞的增殖、遷移、上皮間質(zhì)轉(zhuǎn)換等。由此可見(jiàn)細(xì)胞所處的微環(huán)境剛度在癌癥的產(chǎn)生、遷移和惡化過(guò)程中起重要作用,研究癌細(xì)胞如何感知周?chē)M織剛度以及對(duì)硬度的響應(yīng)規(guī)律,能夠?yàn)閼?yīng)對(duì)癌癥的產(chǎn)生機(jī)制提供有價(jià)值的參考,并協(xié)助改進(jìn)治療方案。本文主要研究不同剛度的基底對(duì)于細(xì)胞生物力學(xué)特性的影響。采用具有良好生物兼容性的水溶性聚合物——聚丙烯酰胺水凝膠,作為模擬細(xì)胞外環(huán)境的培養(yǎng)基底。通過(guò)調(diào)節(jié)丙烯酰胺和甲叉雙丙烯酰胺的比例,制備不同剛度的彈性基底。把處于對(duì)數(shù)生長(zhǎng)期的宮頸癌細(xì)胞接種到1KPa、10KPa、150KPa的聚丙烯酰胺水凝膠基底上,進(jìn)行24-48小時(shí)的培養(yǎng),對(duì)培養(yǎng)后的宮頸癌細(xì)胞進(jìn)行以下實(shí)驗(yàn),觀(guān)測(cè)基底硬度對(duì)細(xì)胞生物力學(xué)行為特性的影響:(1)測(cè)量細(xì)胞骨架中分子馬達(dá)蛋白——肌球蛋白Ⅱ的分布與運(yùn)動(dòng)速率。將宮頸癌細(xì)胞穩(wěn)定轉(zhuǎn)染熒光肌球蛋白Ⅱ,利用激光共聚焦顯微鏡測(cè)定三種剛度基底上生長(zhǎng)的宮頸癌細(xì)胞熒光肌球蛋白Ⅱ沿細(xì)胞長(zhǎng)軸的熒光分布;運(yùn)用熒光漂白恢復(fù)技術(shù),測(cè)定熒光肌球蛋白Ⅱ在細(xì)胞末端的恢復(fù)速率。(2)測(cè)量肌動(dòng)蛋白纖維在整個(gè)細(xì)胞內(nèi)的分布情況。使用Phalloidin-Tetramethylrhodamine B Isothiocyanate對(duì)宮頸癌細(xì)胞肌動(dòng)蛋白纖維進(jìn)行熒光染色,測(cè)定三種剛度上培養(yǎng)的宮頸癌細(xì)胞肌動(dòng)蛋白纖維的分布。(3)采用微管吸吮技術(shù),結(jié)合半無(wú)限體模型,測(cè)定單個(gè)宮頸癌細(xì)胞在鋪展?fàn)顟B(tài)下的細(xì)胞彈性模量。結(jié)果顯示,不同硬度基底上生長(zhǎng)的宮頸癌細(xì)胞呈現(xiàn)出肌球蛋白Ⅱ和肌動(dòng)蛋白纖維的分布差異。在軟基底上生長(zhǎng)的宮頸癌細(xì)胞,細(xì)胞邊緣肌球蛋白Ⅱ和肌動(dòng)蛋白纖維分布較少,肌球蛋白Ⅱ的運(yùn)動(dòng)速率較慢,細(xì)胞彈性模量降低;在較硬基底上生長(zhǎng)的宮頸癌細(xì)胞,肌球蛋白Ⅱ和肌動(dòng)蛋白蛋白纖維呈現(xiàn)出明顯的細(xì)胞邊緣聚集狀態(tài),細(xì)胞彈性模量也明顯高于在較軟兩組基底上生長(zhǎng)的細(xì)胞。實(shí)驗(yàn)表明,基底硬度會(huì)明顯改變肌球蛋白Ⅱ、肌動(dòng)蛋白等重要的細(xì)胞骨架蛋白區(qū)域性作用的效果,對(duì)細(xì)胞的彈性模量造成影響。影響癌細(xì)胞在不同微環(huán)境下的細(xì)胞狀態(tài)。
[Abstract]:Extracellular matrix (ECM) is a complex network of macromolecules with unique physical, biochemical and mechanical properties that directly or indirectly affect the biological behavior of cells.To become an essential factor in the development of normal tissues.When the metabolism of extracellular matrix is abnormal, the tissue structure and mechanical properties of extracellular matrix change, and some diseases occur.Such as tissue fibrosis, cancer cell proliferation, migration, epithelial interstitial transition and so on.It can be seen that the microenvironmental stiffness of the cell plays an important role in the generation, migration and deterioration of cancer, and studies how cancer cells perceive the stiffness of the surrounding tissue and its response to hardness.It can provide valuable reference for the mechanism of cancer response and help to improve the treatment plan.In this paper, the effects of different stiffness substrates on the biomechanical properties of cells are studied.Polyacrylamide hydrogel, a water-soluble polymer with good biocompatibility, was used as the culture substrate to simulate the extracellular environment.The elastic substrates with different stiffness were prepared by adjusting the ratio of acrylamide to methacrylate.The cervical cancer cells in logarithmic growth phase were inoculated on the polyacrylamide hydrogel substrate of 1 KPA 10 KPA and 150 KPA, and cultured for 24-48 hours. The following experiments were carried out on the cultured cervical cancer cells.The effect of substrate hardness on the biomechanical behavior of cells was observed. (1) the distribution and movement rate of molecular motor protein-myosin 鈪，

本文編號(hào)：1756164