解剖学报

解剖学报 ›› 2014, Vol. 45 ›› Issue (5): 646-651.doi: 10.3969/j.issn.0529-1356.2014.05.011

• 细胞和分子生物学 • 上一篇    下一篇

微管相关蛋白1轻链3及Beclin-1在硅沉着病大鼠肺泡巨噬细胞中的表达

张娟1 赵曼曼1,4 李冉1 朱会兴1 田艳霞1 崔建忠5 高俊玲1,2,3*   

  1. 1. 河北联合大学基础医学院组织学胚胎学系，河北 唐山 063000; 2. 河北省煤矿卫生与安全实验室，河北 唐山 063000; 3. 唐山市慢性病基础与临床实验室，河北 唐山 063000; 4. 河北联合大学冀唐学院，河北 唐山 063000; 5. 唐山市工人医院神经外科，河北 唐山 063000
     
  • 收稿日期:2014-01-13 修回日期:2014-03-19 出版日期:2014-10-06 发布日期:2013-10-06
  • 通讯作者: 高俊玲 E-mail:junlinggao@163.com
  • 基金资助:

    河北省科技支撑重点专项;河北省重点职业病防治技术研究

Expression of microtubule-associated protein 1 light chain 3 and Beclin-1 in alveolar macrophages of silicosis rats

ZHANG Juan1 ZHAO Man-man 1,4 LI Ran1 ZHU Hui-xing1 TIAN Yan-xia1 CUI Jian-zhong5 GAO Jun-ling 1,2,3*   

  1. 1. Department of Histology and Embryology, School of Basic Medical Sciences, Hebei United University, Hebei Tangshan 063000, China;2. Key Laboratory of Occupational Health and Safety for Coal Industry, Hebei Tangshan 063000, China; 3. Chronic Disease Basic and Clinical Laboratory, Hebei Tangshan 063000, China; 4. Jitang College, Hebei United University, Hebei Tangshan 063000, China;5. Department of Neurosurgery, Tangshan Gongren Hospital, Hebei Tangshan 063000, China )
     
  • Received:2014-01-13 Revised:2014-03-19 Online:2014-10-06 Published:2013-10-06
  • Contact: GAO Jun-ling E-mail:junlinggao@163.com

摘要：

目的 观察自噬相关蛋白——微管相关蛋白1轻链3（LC-3）、Beclin-1在硅沉着病大鼠肺泡巨噬细胞中的表达，从细胞自噬角度探讨硅沉着病形成的分子机制。方法 50只成年雄性SD大鼠随机分为2组（n=25）：对照组和硅沉着病模型组，每组再分5个时相组。采用非暴露气管灌注二氧化硅（SiO2）粉尘混悬液法（50g/L）建立大鼠硅沉着病模型。分别于造模后1、3、7、14及28 d分批处死5只大鼠，进行原位支气管肺泡灌洗，获取肺泡巨噬细胞，进行培养纯化和富集后用于后续研究。HE染色及透射电子显微镜观察肺泡巨噬细胞形态学改变；免疫细胞化学法检测LC-3、Beclin-1的表达及分布；免疫印迹法检测LC-3、Beclin-1的蛋白表达量。
结果 与对照组相比模型组肺泡巨噬细胞体积较大，胞质丰富，部分细胞内可见硅沉着吞噬颗粒，电镜下可见自噬体形成；模型组LC-3、Beclin-1在各时间点的表达较对照组均增多（P<0.05），1 d即开始增多，随着时间的延长表达逐渐增多，至14 d时达高峰（P<0.05），28d时回落，但仍高于对照组的表达。
结论 在硅沉着病大鼠肺泡巨噬细胞中有自噬的激活，肺泡巨噬细胞自噬参与了大鼠硅沉着病的病理进程。

关键词: 硅沉着病, 肺泡巨噬细胞, 自噬, 微管相关蛋白1轻链3, Beclin-1, 免疫印迹法, 透射电子显微术, 大鼠

Abstract:

Objective To observe expression of autophagy-related proteins LC-3 and Beclin-1 in alveolar macrophages in the silicosis model of rats, and to investigate the molecular mechanisms of silicosis formation from cells autophagy perspective. Methods Fifty adult male SD rats were randomly divided into control and model groups, 25 rats for each group. The silicosis model was made by one-time infusion of silica dust suspension through the trachea without exposed. The rats were sacrificed on the 1st, 3rd, 7th, 14th or 28th day after the modeling. Alveolar macrophages were obtained from bronchoalveolar lavage and used for subsequent research after culture and purification. Morphological characteristics of alveolar macrophages were observed by HE staining and transmission electron microscope; The expression of LC-3 and Beclin-1 was detected by means of immunocytochemistry and Western blotting in each group.  Results Compared with the control group, alveolar macrophages of the model group had larger volume and abundant cytoplasm, the phagocytic silica dust particles were observed in some cells, and autophagosomes were detected by transmission electron microscope. The expressions of LC-3 and Beclin-1 were increased at all time points in the model group(P<0.05). Both LC-3 and Beclin-1 began to increase at the 1st day. As the extension of time the expression gradually enhanced, peaked at the 14th day(P<0.05), and decreased at the 28th day, but higher than the basal expression. Conclusion Autophagy is activated in alveolar macrophages of the silicosis model, and alveolar macrophages autophagy is involved in the pathological process of silicosis in the rat.
 

Key words: Alveolar macrophage, Autophagy, Microtubule-associated protein1 light chain 3, Beclin-1, Western blotting, Transmission electron microscopy, Rat