SARS-CoV-2 N Protein Triggers Acute Lung Injury via Modulating Macrophage Activation and Infiltration in in vitro and in vivo

Background: SARS-CoV-2-induced acute lung injury but its nucleocapsid (N) and/or Spike (S) protein involvements in the disease pathology remain elusive. Methods: In vitro, the cultured THP-1 macrophages were stimulated with alive SARS-CoV-2 virus at different loading dose, N protein or S protein with/without TICAM2-siRNA, TIRAP-siRNA or MyD88-siRNA. The TICAM2, TIRAP and MyD88 expression in the THP-1 cells after N protein stimulation were determined. In vivo, naive mice or mice with depletion macrophages were injected with N protein or dead SARS-CoV-2. The macrophages in the lung were analyzed with flow cytometry, and lung sections were stained with H&E or immunohistochemistry. Culture supernatants and serum were harvested for cytokines measurements with cytometric bead array. Results: Alive SARS-CoV-2 virus or N protein but not S protein induced high cytokine releases from macrophages in a time or virus loading dependent manner. MyD88 and TIRAP but not TICAM2 were highly involved in macrophage activation triggered by N protein whilst both inhibited with siRNA decreased inflammatory responses. Moreover, N protein and dead SARS-CoV-2 caused systemic inflammation, macrophage accumulation and acute lung injury in mice. Macrophage depletion in mice decreased cytokines in response to N protein. Conclusion: SARS-CoV-2 and its N protein but not S protein induced acute lung injury and systemic inflammation, which was closely related to macrophage activation, infiltration and release cytokines.