Yu Haitao, Jiang Wei, Du Hong, Xing Yuan, Bai Guangzhen, Zhang Ye, Li Yu, Jiang Hong, Zhang Ying, Wang Jiuping, Wang Pingzhong, Bai Xuefan
Department of Infectious Diseases, Tangdu Hospital, Fourth Military Medical University, Xi'an, Shaanxi Province, China.
Department of Physiology, Fourth Military Medical University, Xi'an, Shaanxi Province, China.
PLoS One. 2014 Apr 8;9(4):e93810. doi: 10.1371/journal.pone.0093810. eCollection 2014.
Hantaan virus (HTNV) infection causes a severe form of HFRS(hemorrhagic fever with renal syndrome)in Asia. Although HTNV has been isolated for nearly forty years, the pathogenesis of HFRS is still unknown, and little is known regarding the signaling pathway that is activated by the virus.
METHODOLOGY/PRINCIPAL FINDINGS: Cardamonin was selected as a NF-κB inhibitor, and indirect immunofluorescence assays were used to detect the effect of cardamonin on HTNV-infected HUVECs. The effect of cardamonin on the HTNV-induced phosphorylation of Akt and DNA-binding activity of NF-κB were determined using Western blot analysis and electrophoretic mobility shift assays (EMSAs), respectively. Then, flow cytometric and quantitative real-time PCR analyses were performed to quantify the expression levels of the adhesion molecules ICAM-1 and VCAM-1, and the concentrations of IL-6, IL-8, and CCL5 in HUVEC supernatants were examined using ELISA. The results showed that cardamonin did not effect the proliferation of HUVECs or the replication of HTNV in HUVECs. Instead, cardamonin inhibited the phosphorylation of Akt and nuclear transduction of NF-κB and further reduced the expression of the adhesion molecules ICAM-1 and VCAM-1 in HTNV-infected HUVECs. Cardamonin also inhibited the secretion of IL-6 and CCL5, but not IL-8.
CONCLUSION/SIGNIFICANCE: HTNV replication may not be dependent upon the ability of the virus to activate NF-κB in HUVECs. The Akt/NF-κB pathways may be involved in the pathogenesis of HFRS; therefore, cardamonin may serve as a potential beneficial agent for HFRS therapy.
汉坦病毒（HTNV）感染在亚洲引发严重形式的肾综合征出血热（HFRS）。尽管HTNV已被分离近四十年，但HFRS的发病机制仍不清楚，关于该病毒激活的信号通路也知之甚少。
方法/主要发现：选择小豆蔻明作为核因子κB（NF-κB）抑制剂，采用间接免疫荧光试验检测小豆蔻明对HTNV感染的人脐静脉内皮细胞（HUVECs）的影响。分别使用蛋白质免疫印迹分析和电泳迁移率变动分析（EMSA）确定小豆蔻明对HTNV诱导的蛋白激酶B（Akt）磷酸化和NF-κB DNA结合活性的影响。然后，进行流式细胞术和定量实时聚合酶链反应分析以量化黏附分子细胞间黏附分子-1（ICAM-1）和血管细胞黏附分子-1（VCAM-1）的表达水平，并使用酶联免疫吸附测定（ELISA）检测HUVECs上清液中白细胞介素-6（IL-6）、白细胞介素-8（IL-8）和趋化因子配体5（CCL5）的浓度。结果表明，小豆蔻明不影响HUVECs的增殖或HTNV在HUVECs中的复制。相反，小豆蔻明抑制Akt的磷酸化和NF-κB的核转导，并进一步降低HTNV感染的HUVECs中黏附分子ICAM-1和VCAM-1的表达。小豆蔻明还抑制IL-6和CCL-５的分泌，但不抑制IL-8的分泌。
结论/意义：HTNV复制可能不依赖于该病毒激活HUVECs中NF-κB的能力。Akt/NF-κB信号通路可能参与HFRS的发病机制；因此，小豆蔻明可能是HFRS治疗的一种潜在有益药物。
