NGF和BDNF聯(lián)合誘導(dǎo)NSC分化及其在192-IgG-saporin致阿爾茨海默病模型鼠中的應(yīng)用.pdf

1、廣州醫(yī)科大學(xué)博士研究生學(xué)位論文NGF和BDNF聯(lián)合誘導(dǎo)NSC分化及其在192IgGsapin致阿爾茨海默病模型鼠中的應(yīng)用CombinedEffectofNGFBDNFontheNeuronalDifferentiationofNeuralStemCellonRatModelofAlzheimer’sDiseasewith192IgGsapin專(zhuān)業(yè)名稱(chēng)：稱(chēng)：神經(jīng)病學(xué)研究生：劉：劉菲菲學(xué)號(hào)：號(hào)：2011310218指導(dǎo)老師：龍師：龍大宏教授

2、教授本課題受廣東省自然科學(xué)基金資助項(xiàng)目(9151063101000016)和廣州市屬高?？蒲杏?jì)劃項(xiàng)目(2012C042)資助。廣州醫(yī)科大學(xué)廣州二0一四年五月3.2影響NSC分化的因素................................................................................253.3NSC體外培養(yǎng)的不足..................................

3、................................................263.4神經(jīng)球轉(zhuǎn)化為單層貼壁培養(yǎng)的優(yōu)點(diǎn)..........................................................264.結(jié)論........................................................................................

4、................................27第二部分NGF聯(lián)合BDNF對(duì)體外培養(yǎng)的神經(jīng)干細(xì)胞分化為神經(jīng)元的影響...........281材料和方法..............................................................................................................281.1動(dòng)物、主要試劑和儀器.........

5、.............................................................................281.2NSC的取材和培養(yǎng)..............................................................................................301.3單獨(dú)使用NGF或BDNF和兩個(gè)因子的聯(lián)合應(yīng)用........

6、....................................301.3.1NGF和BDNF濃度的確定......................................................................301.3.2因子的單獨(dú)使用和聯(lián)合應(yīng)用對(duì)NSC分化及相關(guān)分子改變的影響......301.4免疫細(xì)胞化學(xué)免疫熒光......................................

7、...............................................311.5QPCR步驟..........................................................................................................311.6總蛋白的提取和濃度測(cè)定........................................

8、..........................................321.6.1細(xì)胞裂解提取總蛋白...............................................................................321.6.2蛋白濃度測(cè)定......................................................................

9、.....................321.7蛋白免疫印跡......................................................................................................331.7.1WB試劑準(zhǔn)備.........................................................................

10、....................331.7.2WB分離膠和濃縮膠的制備.....................................................................341.7.3WB操作步驟.............................................................................................351.8統(tǒng)

11、計(jì)學(xué)分析..........................................................................................................362結(jié)果.........................................................................................................

12、................362.1NGF和BDNF誘導(dǎo)單層貼壁培養(yǎng)NSC分化....................................................362.2比較因子的單獨(dú)使用和聯(lián)合應(yīng)用對(duì)NSC分化的影響....................................382.3WB檢測(cè)PD98059對(duì)PERK的水平變化和神經(jīng)元分化比例的影響.............422.4QPCR檢測(cè)N

13、GF或(和)BDNF對(duì)bHLH家庭基因的表達(dá)變化的影響..........443討論.........................................................................................................................473.1NGF和BDNF對(duì)NSC分化為神經(jīng)元的疊加作用.............................

14、.......473.2ERK的磷酸化參與NGF和BDNF對(duì)NSC的疊加作用..........................483.3MASH1、NGN1和NeuroD的表達(dá)變化參與NGF和BDNF對(duì)NSC的疊加作用..............................................................................................................