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皮层星形胶质细胞Toll样受体4在炎症和β淀粉样蛋白形成中的作用

龚长银, 周爱玲, 茅家慧, 胡亚娥, 耿劲松

南通大学医学院病理生理学系，南通 226001；南京医科大学附属常州市第二人民医院病理科，常州 213003；南通大学医学院循证医学中心，南通 226001

摘要

为了探讨星形胶质细胞在炎症和β淀粉样蛋白(amyloid β-protein, Aβ)形成中的作用和可能的分子机制，本研究通过LPS刺激体外培养大鼠皮层星形胶质细胞，首先采用实时PCR和Western blot方法分别检测Toll样受体4 (Toll-like receptor 4, TLR4)、肿瘤坏死因子α (tumor necrosis factor α, TNF-α)、白细胞介素-1β (interleukin 1β, IL-1β)、β淀粉样前体蛋白(β-amyloid precursor protein, β-APP)和β-位点APP剪切酶1 (β-site APP clearing enzyme 1, BACE1) mRNA及TLR4、NF-κB/P65蛋白水平，而后用免疫荧光法进一步证明NF-κB/P65的核易位，ELISA法测定培养上清中TNF-α、IL-1β和Aβ含量。结果显示，这些指标在LPS刺激后均不同程度地上调。然而，如果预先用TLR4抗体处理，与仅用LPS刺激组相比，LPS对NF-κB/P65核易位及培养上清中TNF-α、IL-1β和Aβ含量的刺激作用显著减弱或消失。结果表明，星形胶质细胞TLR4可能通过TLR4/NF-κB信号通路在炎症和Aβ的形成中发挥重要的作用。

关键词： 阿尔茨海默病; 星形胶质细胞; Toll样受体4; 炎症; β淀粉样蛋白

分类号：R363

The role of Toll-like receptor 4 on inflammation and Aβ formation in cortex astrocytes

GONG Chang-Yin, ZHOU Ai-Ling, MAO Jia-Hui, HU Ya-E, GENG Jin-Song

Department of Pathophysiology, Medical School of Nantong University, Nantong 226001, China； Department of Pathology, Affiliated Changzhou No. 2 People’s Hospital, Nanjing Medical University, Changzhou 213003, China； Evidence-based Medicine Center, Medical School of Nantong University, Nantong 226001, China

Abstract

To investigate the role and possible molecular mechanism of astrocytes in inflammation and amyloid β-protein (Aβ) formation, in this research, by using LPS to stimulate cultured rat astrocytes in vitro with or without anti-Toll-like receptor 4 (TLR4) antibody pretreatment, we first detected the TLR4, TNF-α, IL-1β, β-amyloid precursor protein (β-APP) and β-site APP clearing enzyme 1 (BACE1) mRNA with real-time PCR, and TLR4, NF-κB/P65 protein in cultured astrocytes by Western blot, and then further probed the translocation of NF-κB/P65 using immunofluorescence and the contents of TNF-α, IL-1β and Aβ in culture supernatant through ELISA. We found that all of these indexes increased at different degrees after LPS-stimulation. However, if pretreatment with anti- TLR4 antibody, such stimulating effects of LPS on the nuclear translocation of NF-κB/P65 and TNF-α, IL-1β, Aβ contents in astrocytic culture supernatant were reduced significantly or disappeared in comparison with the group with only LPS-administration. Our results suggest that TLR4 in astrocytes might play an important role in the inflammation and Aβ formation through the TLR4/NF-κB signaling pathway, thus providing new knowledge and understanding of the inflammatory hypothesis of AD pathogenesis.

Key words: Alzheimer’s disease; astrocytes; Toll-like receptor 4; inflammation; β-amyloid protein

收稿日期：2014-04-29　　录用日期：2014-07-02

通讯作者：周爱玲　　E-mail: alz@ntu.edu.cn

引用本文：

龚长银, 周爱玲, 茅家慧, 胡亚娥, 耿劲松. 皮层星形胶质细胞Toll样受体4在炎症和β淀粉样蛋白形成中的作用[J]. 生理学报 2014; 66 (6): 631-638.

GONG Chang-Yin, ZHOU Ai-Ling, MAO Jia-Hui, HU Ya-E, GENG Jin-Song. The role of Toll-like receptor 4 on inflammation and Aβ formation in cortex astrocytes. Acta Physiol Sin 2014; 66 (6): 631-638 (in Chinese with English abstract).