本文主要研究内容
作者周莹（2019）在《功能化类纸基的构建及其在细胞三维培养中的应用》一文中研究指出：细胞培养是体外研究细胞功能的重要技术手段。长期以来,基于培养皿的二维平面培养(2-dimensional culture,2D culture)在细胞生物学研究中得到了最为广泛的应用。然而,细胞在2D培养模式中的单层生长(monolayer)与细胞在体内环境中的生长方式的极大差异引起了研究者对体外细胞培养模式的思考。特别是在肿瘤药物的开发过程中,很多在体外细胞实验中展示良好抗肿瘤效果的药物,在体内实验中抗瘤效果却并不理想。三维细胞培养(3-dimensional culture,3D culture)是指在体外培养中通过使用特殊培养皿或支架材料,形成营养物质、代谢物和氧气的梯度,从而重现细胞体内环境,弥补单层培养缺陷的一种细胞培养模式。纸是人类最古老最重要的发明之一,而将纸基类材料作为一种新型的基底用于细胞培养则是古代智慧与现代科技的完美结合。首先,纸基中大量类似组织间隙的微小孔隙利于营养物质扩散,为细胞提供养分。其次,与基于聚二甲基硅氧烷(poly-dimethylsiloxane,PDMS)的细胞培养芯片相比,纸基的独特之处在于通过多层叠加可方便地实现3D结构组织/器官构建,而通过拆解多层纸基实现从3D至2D的结构转化,从而实现组织内部细胞形态和功能的观测分析。由于将纸类材料作为细胞培养的尝试刚刚开始,目前用于细胞培养的纸基材料主要是Whatman@#114号滤纸和Whatman@#105号擦镜纸,类型较为单一且同类比较下价格比较昂贵;有必要寻找适用于细胞培养的价廉物美的类纸基材料。此外,在实验中多需要将细胞先混合于水凝胶溶胶中再滴加到滤纸上。而且,在CiGiPs细胞培养模型中,纸质基底仅作为一种支架材料为水凝胶提供网络支撑,虽然解决了水凝胶塑形的问题,但仍缺乏相关的生物功能;同时,水凝胶现用现配的特性,降低了芯片制作使用的便捷性和普适性。基于对上述相关领域和关键技术现状与问题的分析,本论文首次探索了将中国传统书画用纸-宣纸用来探究细胞三维培养的潜力;将明胶溶液与聚丙烯无纺布采用冷冻干燥的方式制备即用型明胶@聚丙烯无纺布复合材料,用于细胞三维培养;进一步在即用型明胶@聚丙烯无纺布复合材料上有效负载细胞因子,构建具有生物活性的纸基,为细胞提供仿生三维生长环境和构建仿生微环境,为探究功能化类纸基为基材的仿生芯片奠定基础。本文主要研究内容如下:1、探索宣纸作为经济适用型细胞三维培养基材的潜力宣纸是我国所特有的书画用纸,具有柔韧、洁白、纹理清晰、润墨性好等特点,有“纸寿千年”的美誉。宣纸种类选择多,厚度可调控、价格相对低廉。本工作首先从市售种类的宣纸中选择了生宣、熟宣、半生半熟这三种宣纸作为研究对象。通过液体实验对宣纸的液体浸润性进行评估,结果显示生宣具备更加优异的液体浸润性,为细胞培养过程中细胞培养液的补给提供了条件。进一步将生宣与纸基材料(Whatman@#114号滤纸和Whatman@#105号擦镜纸)进行比较,研究发现宣纸的纤维素成分更加纯粹、纤维与纤维之间的小腔室有利于营养物质的运输和细胞之间的迁移。不同厚度的宣纸用于细胞培养。通过乳酸脱氢酶(Lactate Dehydrogenase,LDH)释放实验发现宣纸对细胞的生长无毒性影响;通过WST-1细胞增殖测试发现细胞可以在宣纸上正常生长,而且在80μm厚的宣纸中细胞增殖最高,展示了良好的细胞相容性。宣纸纤维为细胞黏附与生长提供3D网络结构。通过细胞染色可以观察到,细胞在宣纸网络上大量黏附,而且随着培养时间的延长,细胞呈现增殖生长的趋势且依附宣纸纤维走向生长。最后通过构建基于宣纸的细胞免疫分析平台测定了前列腺癌中O-GaINAc的表达。与传统细胞微孔板相比,宣纸的纤维结构所提供的更大的比表面积促进了细胞的黏附和增殖,增强了细胞免疫分析中信号的表达,提高了细胞免疫检测的灵敏度,展示了宣纸应用到细胞分析平台的潜力。2、即用型明胶@聚丙烯无纺布复合材料的构建及其在3D细胞培养中的应用明胶(gelatin)是一种从动物体内提取出的胶原蛋白,具有良好的生物相容性,并被用于包被细胞培养皿以促进细胞黏附。聚丙烯(polypropylene)是应用最为广泛的聚合物之一,具有机械强度强、易于加工等特点。本工作中,我们首次提出了即用型CiGiPs的概念。我们将明胶溶液滴加到以聚丙烯为原材料的无纺布(polypropylene nonwoven fabric)基底上,采用冷冻干燥的方式制备即用型明胶@聚丙烯无纺布复合材料。在即用型明胶@聚丙烯无纺布复合材料中,明胶溶液的引入在聚丙烯无纺布的纤维与纤维之间形成蜂窝状的小腔室,增加了复合材料的比表面积。材料的孔隙率、溶胀性质和刚性测定实验表明,明胶@聚丙烯无纺布复合材料孔隙率显著增加,良好的溶胀性质有利于营养物质的储存。明胶@聚丙烯无纺布复合材料的生物相容性实验证明,明胶@聚丙烯无纺布复合材料可以应用于细胞培养,细胞大量黏附并呈现增殖生长的趋势。为了突出所提出的复合材料即用型这一概念,我们对最长放置了90天的复合材料进行了性质测试,结果显示明胶@聚丙烯无纺布复合材料化学性质和溶胀性质都未发生改变,更为重要的是细胞相容性保持良好,展现了作为细胞3D培养基材的应用潜力。3、细胞因子修饰的明胶@聚丙烯无纺布复合材料细胞因子是由免疫细胞和基质细胞合成的低分子量蛋白质,其通过与特定的受体结合来调控、参与人体的各种生理活动。在本工作中,将血管内皮生长因子(Human vascular endothelial growth factor,hrVEGF)负载于明胶@聚丙烯无纺布复合材料,构建具有生物活性的类纸基材料,为血管内皮细胞提供仿生三维生长环境。首先,探究如何在明胶@聚丙烯无纺布复合材料上有效负载细胞因子。通过吸附法、EDC/NHS共价结合方法和混合物冷冻干燥法制备负载生长因子的明胶@无纺布复合材料。研究结果显示具有生物活性的明胶@无纺布材料有利于细胞的快速黏附,并促进血管内皮细胞增殖。此外,将负载细胞因子的明胶@聚丙烯复合材料多层叠加构建细胞迁移模型。结果显示细胞可以从种植层迁移到负载细胞因子的迁移层,为构建功能化类纸基为基材的仿生芯片奠定基础。
Abstract
xi bao pei yang shi ti wai yan jiu xi bao gong neng de chong yao ji shu shou duan 。chang ji yi lai ,ji yu pei yang min de er wei ping mian pei yang (2-dimensional culture,2D culture)zai xi bao sheng wu xue yan jiu zhong de dao le zui wei an fan de ying yong 。ran er ,xi bao zai 2Dpei yang mo shi zhong de chan ceng sheng chang (monolayer)yu xi bao zai ti nei huan jing zhong de sheng chang fang shi de ji da cha yi yin qi le yan jiu zhe dui ti wai xi bao pei yang mo shi de sai kao 。te bie shi zai zhong liu yao wu de kai fa guo cheng zhong ,hen duo zai ti wai xi bao shi yan zhong zhan shi liang hao kang zhong liu xiao guo de yao wu ,zai ti nei shi yan zhong kang liu xiao guo que bing bu li xiang 。san wei xi bao pei yang (3-dimensional culture,3D culture)shi zhi zai ti wai pei yang zhong tong guo shi yong te shu pei yang min huo zhi jia cai liao ,xing cheng ying yang wu zhi 、dai xie wu he yang qi de ti du ,cong er chong xian xi bao ti nei huan jing ,mi bu chan ceng pei yang que xian de yi chong xi bao pei yang mo shi 。zhi shi ren lei zui gu lao zui chong yao de fa ming zhi yi ,er jiang zhi ji lei cai liao zuo wei yi chong xin xing de ji de yong yu xi bao pei yang ze shi gu dai zhi hui yu xian dai ke ji de wan mei jie ge 。shou xian ,zhi ji zhong da liang lei shi zu zhi jian xi de wei xiao kong xi li yu ying yang wu zhi kuo san ,wei xi bao di gong yang fen 。ji ci ,yu ji yu ju er jia ji gui yang wan (poly-dimethylsiloxane,PDMS)de xi bao pei yang xin pian xiang bi ,zhi ji de du te zhi chu zai yu tong guo duo ceng die jia ke fang bian de shi xian 3Djie gou zu zhi /qi guan gou jian ,er tong guo ca jie duo ceng zhi ji shi xian cong 3Dzhi 2Dde jie gou zhuai hua ,cong er shi xian zu zhi nei bu xi bao xing tai he gong neng de guan ce fen xi 。you yu jiang zhi lei cai liao zuo wei xi bao pei yang de chang shi gang gang kai shi ,mu qian yong yu xi bao pei yang de zhi ji cai liao zhu yao shi Whatman@#114hao lv zhi he Whatman@#105hao ca jing zhi ,lei xing jiao wei chan yi ju tong lei bi jiao xia jia ge bi jiao ang gui ;you bi yao xun zhao kuo yong yu xi bao pei yang de jia lian wu mei de lei zhi ji cai liao 。ci wai ,zai shi yan zhong duo xu yao jiang xi bao xian hun ge yu shui ning jiao rong jiao zhong zai di jia dao lv zhi shang 。er ju ,zai CiGiPsxi bao pei yang mo xing zhong ,zhi zhi ji de jin zuo wei yi chong zhi jia cai liao wei shui ning jiao di gong wang lao zhi cheng ,sui ran jie jue le shui ning jiao su xing de wen ti ,dan reng que fa xiang guan de sheng wu gong neng ;tong shi ,shui ning jiao xian yong xian pei de te xing ,jiang di le xin pian zhi zuo shi yong de bian jie xing he pu kuo xing 。ji yu dui shang shu xiang guan ling yu he guan jian ji shu xian zhuang yu wen ti de fen xi ,ben lun wen shou ci tan suo le jiang zhong guo chuan tong shu hua yong zhi -xuan zhi yong lai tan jiu xi bao san wei pei yang de qian li ;jiang ming jiao rong ye yu ju bing xi mo fang bu cai yong leng dong gan zao de fang shi zhi bei ji yong xing ming jiao @ju bing xi mo fang bu fu ge cai liao ,yong yu xi bao san wei pei yang ;jin yi bu zai ji yong xing ming jiao @ju bing xi mo fang bu fu ge cai liao shang you xiao fu zai xi bao yin zi ,gou jian ju you sheng wu huo xing de zhi ji ,wei xi bao di gong fang sheng san wei sheng chang huan jing he gou jian fang sheng wei huan jing ,wei tan jiu gong neng hua lei zhi ji wei ji cai de fang sheng xin pian dian ding ji chu 。ben wen zhu yao yan jiu nei rong ru xia :1、tan suo xuan zhi zuo wei jing ji kuo yong xing xi bao san wei pei yang ji cai de qian li xuan zhi shi wo guo suo te you de shu hua yong zhi ,ju you rou ren 、jie bai 、wen li qing xi 、run mo xing hao deng te dian ,you “zhi shou qian nian ”de mei yu 。xuan zhi chong lei shua ze duo ,hou du ke diao kong 、jia ge xiang dui di lian 。ben gong zuo shou xian cong shi shou chong lei de xuan zhi zhong shua ze le sheng xuan 、shou xuan 、ban sheng ban shou zhe san chong xuan zhi zuo wei yan jiu dui xiang 。tong guo ye ti shi yan dui xuan zhi de ye ti jin run xing jin hang ping gu ,jie guo xian shi sheng xuan ju bei geng jia you yi de ye ti jin run xing ,wei xi bao pei yang guo cheng zhong xi bao pei yang ye de bu gei di gong le tiao jian 。jin yi bu jiang sheng xuan yu zhi ji cai liao (Whatman@#114hao lv zhi he Whatman@#105hao ca jing zhi )jin hang bi jiao ,yan jiu fa xian xuan zhi de qian wei su cheng fen geng jia chun cui 、qian wei yu qian wei zhi jian de xiao qiang shi you li yu ying yang wu zhi de yun shu he xi bao zhi jian de qian yi 。bu tong hou du de xuan zhi yong yu xi bao pei yang 。tong guo ru suan tuo qing mei (Lactate Dehydrogenase,LDH)shi fang shi yan fa xian xuan zhi dui xi bao de sheng chang mo du xing ying xiang ;tong guo WST-1xi bao zeng shi ce shi fa xian xi bao ke yi zai xuan zhi shang zheng chang sheng chang ,er ju zai 80μmhou de xuan zhi zhong xi bao zeng shi zui gao ,zhan shi le liang hao de xi bao xiang rong xing 。xuan zhi qian wei wei xi bao nian fu yu sheng chang di gong 3Dwang lao jie gou 。tong guo xi bao ran se ke yi guan cha dao ,xi bao zai xuan zhi wang lao shang da liang nian fu ,er ju sui zhao pei yang shi jian de yan chang ,xi bao cheng xian zeng shi sheng chang de qu shi ju yi fu xuan zhi qian wei zou xiang sheng chang 。zui hou tong guo gou jian ji yu xuan zhi de xi bao mian yi fen xi ping tai ce ding le qian lie xian ai zhong O-GaINAcde biao da 。yu chuan tong xi bao wei kong ban xiang bi ,xuan zhi de qian wei jie gou suo di gong de geng da de bi biao mian ji cu jin le xi bao de nian fu he zeng shi ,zeng jiang le xi bao mian yi fen xi zhong xin hao de biao da ,di gao le xi bao mian yi jian ce de ling min du ,zhan shi le xuan zhi ying yong dao xi bao fen xi ping tai de qian li 。2、ji yong xing ming jiao @ju bing xi mo fang bu fu ge cai liao de gou jian ji ji zai 3Dxi bao pei yang zhong de ying yong ming jiao (gelatin)shi yi chong cong dong wu ti nei di qu chu de jiao yuan dan bai ,ju you liang hao de sheng wu xiang rong xing ,bing bei yong yu bao bei xi bao pei yang min yi cu jin xi bao nian fu 。ju bing xi (polypropylene)shi ying yong zui wei an fan de ju ge wu zhi yi ,ju you ji xie jiang du jiang 、yi yu jia gong deng te dian 。ben gong zuo zhong ,wo men shou ci di chu le ji yong xing CiGiPsde gai nian 。wo men jiang ming jiao rong ye di jia dao yi ju bing xi wei yuan cai liao de mo fang bu (polypropylene nonwoven fabric)ji de shang ,cai yong leng dong gan zao de fang shi zhi bei ji yong xing ming jiao @ju bing xi mo fang bu fu ge cai liao 。zai ji yong xing ming jiao @ju bing xi mo fang bu fu ge cai liao zhong ,ming jiao rong ye de yin ru zai ju bing xi mo fang bu de qian wei yu qian wei zhi jian xing cheng feng wo zhuang de xiao qiang shi ,zeng jia le fu ge cai liao de bi biao mian ji 。cai liao de kong xi lv 、rong zhang xing zhi he gang xing ce ding shi yan biao ming ,ming jiao @ju bing xi mo fang bu fu ge cai liao kong xi lv xian zhe zeng jia ,liang hao de rong zhang xing zhi you li yu ying yang wu zhi de chu cun 。ming jiao @ju bing xi mo fang bu fu ge cai liao de sheng wu xiang rong xing shi yan zheng ming ,ming jiao @ju bing xi mo fang bu fu ge cai liao ke yi ying yong yu xi bao pei yang ,xi bao da liang nian fu bing cheng xian zeng shi sheng chang de qu shi 。wei le tu chu suo di chu de fu ge cai liao ji yong xing zhe yi gai nian ,wo men dui zui chang fang zhi le 90tian de fu ge cai liao jin hang le xing zhi ce shi ,jie guo xian shi ming jiao @ju bing xi mo fang bu fu ge cai liao hua xue xing zhi he rong zhang xing zhi dou wei fa sheng gai bian ,geng wei chong yao de shi xi bao xiang rong xing bao chi liang hao ,zhan xian le zuo wei xi bao 3Dpei yang ji cai de ying yong qian li 。3、xi bao yin zi xiu shi de ming jiao @ju bing xi mo fang bu fu ge cai liao xi bao yin zi shi you mian yi xi bao he ji zhi xi bao ge cheng de di fen zi liang dan bai zhi ,ji tong guo yu te ding de shou ti jie ge lai diao kong 、can yu ren ti de ge chong sheng li huo dong 。zai ben gong zuo zhong ,jiang xie guan nei pi sheng chang yin zi (Human vascular endothelial growth factor,hrVEGF)fu zai yu ming jiao @ju bing xi mo fang bu fu ge cai liao ,gou jian ju you sheng wu huo xing de lei zhi ji cai liao ,wei xie guan nei pi xi bao di gong fang sheng san wei sheng chang huan jing 。shou xian ,tan jiu ru he zai ming jiao @ju bing xi mo fang bu fu ge cai liao shang you xiao fu zai xi bao yin zi 。tong guo xi fu fa 、EDC/NHSgong jia jie ge fang fa he hun ge wu leng dong gan zao fa zhi bei fu zai sheng chang yin zi de ming jiao @mo fang bu fu ge cai liao 。yan jiu jie guo xian shi ju you sheng wu huo xing de ming jiao @mo fang bu cai liao you li yu xi bao de kuai su nian fu ,bing cu jin xie guan nei pi xi bao zeng shi 。ci wai ,jiang fu zai xi bao yin zi de ming jiao @ju bing xi fu ge cai liao duo ceng die jia gou jian xi bao qian yi mo xing 。jie guo xian shi xi bao ke yi cong chong zhi ceng qian yi dao fu zai xi bao yin zi de qian yi ceng ,wei gou jian gong neng hua lei zhi ji wei ji cai de fang sheng xin pian dian ding ji chu 。
论文参考文献
论文详细介绍
论文作者分别是来自西南大学的周莹，发表于刊物西南大学2019-09-24论文，是一篇关于细胞培养论文,三维生长论文,类纸基论文,即用型论文,细胞迁移芯片论文,西南大学2019-09-24论文的文章。本文可供学术参考使用，各位学者可以免费参考阅读下载，文章观点不代表本站观点，资料来自西南大学2019-09-24论文网站，若本站收录的文献无意侵犯了您的著作版权，请联系我们删除。
标签：细胞培养论文; 三维生长论文; 类纸基论文; 即用型论文; 细胞迁移芯片论文; 西南大学2019-09-24论文;