發(fā)布時(shí)間：2018-02-02 15:31

  本文關(guān)鍵詞： P21 亞細(xì)胞定位 TGF-β1 凋亡 人支氣管上皮細(xì)胞　出處：《南昌大學(xué)》2013年碩士論文　論文類型：學(xué)位論文

【摘要】：背景： P21蛋白是一種廣泛細(xì)胞周期抑制蛋白，能夠調(diào)節(jié)細(xì)胞周期，隨著研究的深入，發(fā)現(xiàn)其還可以參與細(xì)胞衰老、凋亡、分化等。有研究證實(shí)其胞漿胞核定位與細(xì)胞凋亡關(guān)系密切，在胞漿和胞核中的表達(dá)有不一樣的功能，作用途徑也不一樣。在胞核中過(guò)度表達(dá)能夠促進(jìn)細(xì)胞凋亡，而在胞漿中過(guò)度表達(dá)抑制細(xì)胞凋亡。 人支氣管上皮細(xì)胞是氣道的第一道屏障，其能夠?qū)獾榔鸨Ｗo(hù)作用，同時(shí)能夠分泌一些保護(hù)性細(xì)胞因子，其凋亡與抗凋亡機(jī)制不平衡參與許多肺部疾病發(fā)生和發(fā)展，如COPD、哮喘、肺纖維化、急性肺損傷等。 TGF-β1是一種多功能的細(xì)胞因子，參與細(xì)胞生長(zhǎng)、分化、細(xì)胞周期、凋亡等。文獻(xiàn)報(bào)道能刺激肺上皮細(xì)胞凋亡，同時(shí)P21蛋白表達(dá)增加，P21蛋白表達(dá)增高又有保護(hù)TGF-β1導(dǎo)致的肺上皮細(xì)胞的凋亡，但鮮有研究TGF-β1與p21亞細(xì)胞定位的關(guān)系，本研究探討TGF-β1誘導(dǎo)細(xì)胞凋亡是否與p21亞細(xì)胞定位的關(guān)系。 目的： 研究P21蛋白胞漿、胞核亞細(xì)胞定位差異在TGF-β1誘導(dǎo)人支氣管上皮細(xì)胞系16HBE凋亡中的關(guān)系。方法： 用TGF-β1刺激16HBE細(xì)胞，建立細(xì)胞凋亡模型，再分別提取胞漿胞核蛋白，用western blot半定量的方法研究P21蛋白在細(xì)胞漿和細(xì)胞核中亞細(xì)胞分布情況。 結(jié)果： 1、TGF-β1在不同濃度下（0.3ng/ml,1ng/ml,3ng/ml,10ng/ml,33ng/ml）、在刺激時(shí)間為（12h,24h,48h）時(shí)能誘導(dǎo)16HBE細(xì)胞凋亡，在濃度≤10ng/ml、時(shí)間≤24h時(shí)，隨著濃度增大和時(shí)間延長(zhǎng)16HBE細(xì)胞凋亡增高，，而在刺激時(shí)間48h或TGF-β1濃度33ng/ml時(shí)晚期凋亡和死亡細(xì)胞明顯增多。 2、在TGF-β1濃度為(3ng/ml、10ng/ml)，刺激24小時(shí)后p21蛋白胞漿胞核中表達(dá)均比無(wú)TGF-β1組明顯增高（p0.05）；而TGF-β1在不同濃度（3ng/ml,10ng/ml）刺激下，胞核p21蛋白表達(dá)無(wú)差異（p0.05）；在TGF-β1同一濃度刺激下細(xì)胞漿和細(xì)胞核內(nèi)p21蛋白表達(dá)有明顯差異（P0.05），主要在胞漿中表達(dá)，并且隨著16HBE細(xì)胞凋亡的增加胞漿p21蛋白表達(dá)下降（p0.05），這與P21蛋白在細(xì)胞漿發(fā)揮抗凋亡的作用下降有關(guān)。 結(jié)論： TGF-β1在誘導(dǎo)人支氣管上皮細(xì)胞系16HBE凋亡中與P21蛋白的胞漿、胞核定位有關(guān)。
[Abstract]:Background: P21 protein is a kind of extensive cell cycle inhibitor protein, which can regulate cell cycle. With the development of research, it has been found that P21 protein can also participate in cell senescence and apoptosis. Differentiation and so on. Some studies have confirmed that its cytoplasmic nuclear localization is closely related to apoptosis, and its expression in cytoplasm and nucleus has different functions and different pathways. Overexpression in nucleus can promote cell apoptosis. Overexpression in cytoplasm inhibits apoptosis. Human bronchial epithelial cells are the first barrier of the airway, which can protect the airway and secrete some protective cytokines. The imbalance between apoptosis and antiapoptotic mechanism is involved in the occurrence and development of many pulmonary diseases, such as COPD, asthma, pulmonary fibrosis, acute lung injury and so on. TGF- 尾 1 is a multifunctional cytokine involved in cell growth, differentiation, cell cycle and apoptosis. The high expression of P21 protein can protect lung epithelial cells from apoptosis induced by TGF- 尾 1, but there is little research on the relationship between TGF- 尾 1 and p21 subcellular localization. The aim of this study was to investigate the relationship between apoptosis induced by TGF- 尾 1 and p21 subcellular localization. Objective: To study the relationship between cytoplasmic and nuclear subcellular localization of P21 protein in human bronchial epithelial cell line 16HBE induced by TGF- 尾 1. Methods: TGF- 尾 1 was used to stimulate 16HBE cells to establish apoptosis model and extract cytoplasmic nuclear protein. The distribution of P21 protein in cytoplasm and nucleus was studied by western blot semi-quantitative method. Results: 1TGF- 尾 _ 1 at different concentrations of 0.3ng / ml ~ (-1) ng / ml ~ (-1) ~ 3ng / ml ~ (-1) ~ (10) ng / ml ~ (10) ng / ml ~ (-1) ~ (33) ng / ml ~ (-1), and the time of stimulation is 12 h. When the concentration was 鈮

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