P21蛋白亚细胞定位在TGF-β1诱导人支气管上皮细胞凋亡中的研究

发布时间：2018-02-02 15:31

  本文关键词： P21 亚细胞定位 TGF-β1 凋亡 人支气管上皮细胞　出处：《南昌大学》2013年硕士论文　论文类型：学位论文

【摘要】：背景： P21蛋白是一种广泛细胞周期抑制蛋白，能够调节细胞周期，随着研究的深入，发现其还可以参与细胞衰老、凋亡、分化等。有研究证实其胞浆胞核定位与细胞凋亡关系密切，在胞浆和胞核中的表达有不一样的功能，作用途径也不一样。在胞核中过度表达能够促进细胞凋亡，而在胞浆中过度表达抑制细胞凋亡。 人支气管上皮细胞是气道的第一道屏障，其能够对气道起保护作用，同时能够分泌一些保护性细胞因子，其凋亡与抗凋亡机制不平衡参与许多肺部疾病发生和发展，如COPD、哮喘、肺纤维化、急性肺损伤等。 TGF-β1是一种多功能的细胞因子，参与细胞生长、分化、细胞周期、凋亡等。文献报道能刺激肺上皮细胞凋亡，同时P21蛋白表达增加，P21蛋白表达增高又有保护TGF-β1导致的肺上皮细胞的凋亡，但鲜有研究TGF-β1与p21亚细胞定位的关系，本研究探讨TGF-β1诱导细胞凋亡是否与p21亚细胞定位的关系。 目的： 研究P21蛋白胞浆、胞核亚细胞定位差异在TGF-β1诱导人支气管上皮细胞系16HBE凋亡中的关系。方法： 用TGF-β1刺激16HBE细胞，建立细胞凋亡模型，再分别提取胞浆胞核蛋白，用western blot半定量的方法研究P21蛋白在细胞浆和细胞核中亚细胞分布情况。 结果： 1、TGF-β1在不同浓度下（0.3ng/ml,1ng/ml,3ng/ml,10ng/ml,33ng/ml）、在刺激时间为（12h,24h,48h）时能诱导16HBE细胞凋亡，在浓度≤10ng/ml、时间≤24h时，随着浓度增大和时间延长16HBE细胞凋亡增高，，而在刺激时间48h或TGF-β1浓度33ng/ml时晚期凋亡和死亡细胞明显增多。 2、在TGF-β1浓度为(3ng/ml、10ng/ml)，刺激24小时后p21蛋白胞浆胞核中表达均比无TGF-β1组明显增高（p0.05）；而TGF-β1在不同浓度（3ng/ml,10ng/ml）刺激下，胞核p21蛋白表达无差异（p0.05）；在TGF-β1同一浓度刺激下细胞浆和细胞核内p21蛋白表达有明显差异（P0.05），主要在胞浆中表达，并且随着16HBE细胞凋亡的增加胞浆p21蛋白表达下降（p0.05），这与P21蛋白在细胞浆发挥抗凋亡的作用下降有关。 结论： TGF-β1在诱导人支气管上皮细胞系16HBE凋亡中与P21蛋白的胞浆、胞核定位有关。
[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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