Mechanism of NLRP3 activation by human adenovirus 7 infection
10.3760/cma.j.cn112866-20191122-00186
- VernacularTitle:人腺病毒7型感染激活NLRP3炎症小体的机制研究
- Author:
Wei WANG
1
;
Yali DUAN
;
Meng ZHANG
;
Xiangpeng CHEN
;
Lili XU
;
Yun ZHU
;
Zhengde XIE
Author Information
1. 国家儿童医学中心 首都医科大学附属北京儿童医院 北京市儿科研究所 感染与病毒研究室 儿科重大疾病研究教育部重点实验室 儿童呼吸道感染性疾病研究北京重点实验室 中国医学科学院儿童危重感染诊治创新单元，北京 100045
- Keywords:
Human adenovirus type 7;
NLRP3 inflammasome;
Interleukin-1 β;
Toll-like receptor4;
Cathepsin B
- From:
Chinese Journal of Experimental and Clinical Virology
2020;34(2):113-121
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To investigate the mechanism of activation of NOD-like receptors protein (NLRP)3 in the process of human adenovirus type 7 (HAdV-7) infection.Methods:THP-1 cells were treated with adenosine triphosphate (ATP), an NLRP3 inhibitor MCC950, and a caspase-1 inhibitor Ac-YVDK-cmk, ammonium pyrrolidinedithiocarbamate (APDC), N-acetyl-L-cysteine (NAC), cathepsin B-selective inhibitor CA-074-me, glibenclamide and potassium chloride (KCl) respectively. THP-1 cells without infection were used as the control group and those infected with HAdV-7 were used as the experimental group. The expression of NLRP3, caspase-1 and IL-1β in the supernatant was detected by enzyme-linked immunosorbent assay (ELISA), Q-PCR and western blot(WB).Results:Immunofluorescence detection showed that HAdV-7 infected THP-1, while WB showed that HAdV-7 infection could induce the pro-IL-1β protein expression in cells, and caspase-1 cleavage induce IL-1β maturation and release to the outside of cells. The expression of IL-1β in the supernatant of cells was significantly increased as detected by ELISA (MOI=0.5, P=0.0008). After inhibition of caspase-1 expression, the expression of IL-1 β in the supernatant was significantly inhibited ( P=0.0025). The expression of pro-IL-1β and NLRP3 mRNA in HAdV-7 infected cells was up-regulated (NLRP3: P=0.0004; pro-IL-1β: P=0.0007), and WB showed that HAdV-7 could up regulate the expression of pro-IL-1β and NLRP3 protein in the cells. After THP-1 was treated with NLRP3 specific inhibitor MCC950 the expression of IL-1β in the supernatant of HAdV-7 infected cells were significantly inhibited ( P=0.002) 7). Toll-like receptor (TLR)2, TLR4 and TLR7/9 signal transduction inhibitors were used respectively to inhibit HAdV-7 infection of THP-1. The result showed that TLR4 signal transduction was inhibited, and the expression level of IL-1 β in cell supernatant was significantly decreased ( P=0.0122). Using reactive oxygen species (ROS) inhibitor, cathepsin B inhibitor, or K + outflow inhibitor to inhibit the infection of THP-1 differentiated macrophages by HAdV-7, the result showed that when cathepsin B was inhibited ( P=0.0292), or K + outflow was inhibited (KCl, P=0.0022; glibenclamide, P=0.0275), the expression level of IL-1 β in the supernatant was significantly decreased. Conclusions:HAdV-7 infection of THP-1 cells can activate NLRP3 inflammasome. The first signal of NLRP3 activation is transmitted by TLR4, and the second signal is mainly activated by K + outflow and cathepsin B release.
