摘要
目的:探讨Ripa裂解液蛋白提取法和Invent离心管柱蛋白提取法对慢性应激模型大鼠海马组织PI3K/Akt/mTOR信号通路相关蛋白表达量的影响。方法:空白组和模型组大鼠各3只,取双侧海马组织,每个样品分别采用Ripa裂解液、Invent离心管柱蛋白提取法进行蛋白提取,分为空白+Invent组、空白+Ripa组、模型+Invent组、模型+Ripa组。采用Western Blot法检测PI3K/Akt/mTOR信号通路相关蛋白表达情况,析因设计方法分析各因素的效果。结果:大鼠海马PI3K、p-Akt/Akt、p-mTOR/mTOR表达均受组别和提取方法的影响,组别和提取方法存在交互作用,运用Invent离心管柱蛋白提取法提取蛋白,模型组的PI3K、p-Akt/Akt、p-mTOR/mTOR表达较空白组减少,差异有统计学意义(P<0.05)。结论:慢性应激抑制大鼠海马PI3K/Akt/mTOR信号通路, Invent离心管柱蛋白提取法操作便捷,蛋白提取率高,尤其对信号通路中磷酸化蛋白的提取具有明显的优势。
Objective:To investigate the effect of Ripa lysate protein extraction and Invent centrifugal column protein extraction on the expression of mTOR signaling pathway related proteins in hippocampus of chronic stress model rats. Methods: Bilateral hippocampus was taken out from three rats in the blank group and the model group. Each sample was extracted with Ripa lysate and Invent centrifuge column protein extraction method, and divided into blank+Invent group, blank+Ripa group, model+Invent group, model+Ripa group. Western blot was used to detect the expressions of mTOR signaling pathway-related proteins. Effects of various factors were analyzed by factorial design. Results:The expressions of PI3 K, p-Akt/Akt and p-mTOR/mTOR in rats were affected by modeling and extraction methods, and there were interactions between modeling and extraction methods. When the protein was extracted by Invent centrifugal column protein extraction, the expressions of PI3 K, p-Akt/Akt and p-mTOR/mTOR in the model group were significantly lower than those in the blank group(P<0.05). Conclusions:Chronic stress inhibits the PI3 K/Akt/mTOR signaling pathway in hippocampal of rats. The Invent centrifuge column protein extraction method is easy to operate and has a high protein extraction rate, especially for the extraction of phosphorylated proteins in the signaling pathway.
作者
倪慧
张铭珈
敖海清
朱艳芳
徐志伟
NI Hui;ZHANG Mingjia;AO Haiqing;ZHU Yanfang;XU Zhiwei(Guangzhou University of Chinese Medicine,Guangzhou 510006,Guangdong,China)
出处
《中华中医药学刊》
CAS
北大核心
2019年第7期1669-1672,共4页
Chinese Archives of Traditional Chinese Medicine
基金
国家自然科学基金项目(81673838)
