Differential gene expressions of various interferon, antiviral response genes as well as tissue factor (F3) and TMPRSS genes in primary airway epithelial cells infected with SARS-CoV-2

Clinical data show SARS-CoV-2 infection causes activation of host inflammatory antiviral response, including the type I interferon pathway and the expression of many inflammatory cytokines, as well as the infiltration of neutrophils and macrophages. It has been hypothesized that the overt host inflammatory response is at least partially responsible for COVID-19 associated diffuse alveolar damage (DAD). Our research focuses on one defining feature of DAD, hyaline membrane formation which is characterized by fibrin deposition in lung airspace. With primary normal human bronchial/tracheal epithelial (NHBE) cells and human small airway epithelial cells (HSAEC) as in vitro models, we have evidence of turbidity assay, confocal microscopy, and scanning electron microscopy (SEM) proving that infection with field isolates of SARS-CoV-2 variants and SARS-CoV-2 spike pseudotyped lentiviruses (pSARS2) induces fibrin clot formation in the NHBE and HSAEC cells. We have also observed that the infection-induced fibrin formation in NHBE and HSAEC cells requires thrombin but is independent of tissue factor and other classical plasma coagulation factors. In addition, we have validated involvement of type II transmembrane serine proteases (TMPRSS) in the activation of thrombin, the last enzyme in the coagulation cascade. Therefore, we performed RNA sequencing analyses to infected NHBE and HSAEC cells and further investigated how SARS-CoV-2 variants and pSARS2 regulate type I interferon response, antiviral innate response, and expression of TMPRSS genes. By doing this, we found both pSARS2 and the field delta variant viruses upregulated type I interferon and innate antiviral responses in the infected NHBE and HSAEC cells compared to their uninfected controls. However, we did not observe significant upregulation in the tissue factor transcription in both pSARS2 and the delta variant infected samples. To investigate how viral infection regulates type I interferon response, antiviral innate response, and expression of TMPRSS genes in lung epithelial cells, we cultured two types of lung epithelial cells, NHBE and HSAEC in vitro and uninfected or infected these cells. We then performed gene expression profiling analysis using data obtained from RNA-seq of these 2 different cells either uninfected or infected with SARS-CoV-2 delta variant or SARS-CoV-2 spike pseudotyped lentiviruses. Comparative gene expression profiling analysis of RNA-seq data for infected NHBE and HSAEC cells and their uninfected counterparts. Each condition contains duplicate samples .