Institute of Military Veterinary Medicine, Academy of Military Medical Sciences, Changchun, 130122, People's Republic of China.
Academician Workstation of Jilin Province, Changchun University of Chinese Medicine, Changchun, 130117, People's Republic of China.
Virol J. 2021 Feb 27;18(1):46. doi: 10.1186/s12985-021-01515-1.
Coronavirus disease 2019 (COVID-19) is caused by SARS-CoV-2 and broke out as a global pandemic in late 2019. The acidic pH environment of endosomes is believed to be essential for SARS-CoV-2 to be able to enter cells and begin replication. However, the clinical use of endosomal acidification inhibitors, typically chloroquine, has been controversial with this respect.
In this study, RT-qPCR method was used to detect the SARS-CoV-2N gene to evaluate viral replication. The CCK-8 assay was also used to evaluate the cytotoxic effect of SARS-CoV-2. In situ hybridization was used to examine the distribution of the SARS-CoV-2 gene in lung tissues. Hematoxylin and eosin staining was also used to evaluate virus-associated pathological changes in lung tissues.
In this study, analysis showed that endosomal acidification inhibitors, including chloroquine, bafilomycin A1 and NHCL, significantly reduced the viral yields of SARS-CoV-2 in Vero E6, Huh-7 and 293T-ACE2 cells. Chloroquine and bafilomycin A1 also improved the viability and proliferation of Vero E6 cells after SARS-CoV-2 infection. Moreover, in the hACE2 transgenic mice model of SARS-CoV-2 infection, chloroquine and bafilomycin A1 reduced viral replication in lung tissues and alleviated viral pneumonia with reduced inflammatory exudation and infiltration in peribronchiolar and perivascular tissues, as well as improved structures of alveolar septum and pulmonary alveoli.
Our research investigated the antiviral effects of endosomal acidification inhibitors against SARS-CoV-2 in several infection models and provides an experimental basis for further mechanistic studies and drug development.
2019 年末，新型冠状病毒病（COVID-19）由严重急性呼吸综合征冠状病毒 2（SARS-CoV-2）引发，迅速在全球蔓延。内体的酸性 pH 环境被认为是 SARS-CoV-2 进入细胞并开始复制的必要条件。然而，内体酸化抑制剂（如氯喹）在临床上的应用存在争议。
本研究采用 RT-qPCR 方法检测 SARS-CoV-2N 基因以评估病毒复制情况，采用 CCK-8 法检测 SARS-CoV-2 对细胞的细胞毒性作用，采用原位杂交法检测 SARS-CoV-2 基因在肺组织中的分布，采用苏木精-伊红（H&E）染色法评估病毒感染引起的肺组织病理变化。
本研究结果表明，内体酸化抑制剂，包括氯喹、巴弗洛霉素 A1 和 NHCL，可显著降低 SARS-CoV-2 在 Vero E6、Huh-7 和 293T-ACE2 细胞中的病毒产量。氯喹和巴弗洛霉素 A1 还可提高 SARS-CoV-2 感染后 Vero E6 细胞的活力和增殖能力。此外，在 hACE2 转基因 SARS-CoV-2 感染小鼠模型中，氯喹和巴弗洛霉素 A1 可降低肺组织中的病毒复制，减轻病毒性肺炎，减少细支气管和血管周围组织的炎症渗出和浸润，并改善肺泡间隔和肺泡结构。
本研究在多种感染模型中研究了内体酸化抑制剂对 SARS-CoV-2 的抗病毒作用，为进一步的机制研究和药物开发提供了实验依据。
