Dottorato in MALATTIE INFETTIVE, MICROBIOLOGIA E SANITA' PUBBLICA

Titolo della tesi: Cytokine storm in COVID-19-related deaths: from SARS-CoV-2 RNA detection to immunohistochemical quantification of proinflammatory cytokines IL-1β, IL-6, IL-15 and TNF-α

Background: Current evidence suggests that a cytokine storm is the major cause of ARDS and multiple organ failure, and is consistently linked with fatal outcomes. Also SARS-CoV-2 infection is characterized, in the early stages, by initial local inflammatory response to the virus infiltrates and progressive parenchymal lung damage, while a systemic inflammation damage can occur when the host inflammatory response continues to amplify. Aims: The aim of this study is to clarify the correlation between SARS-CoV-2 infection and the inflammatory response, to evaluate the tissue localization of the virus in various organs and to investigate the expression of cytokines/chemokines (IL-1β, IL-6, IL-10, IL-15, TNF-α, MCP-1), leukocyte markers (CD4, CD8, CD20, CD45) and fibrinogen, in an attempt to verify and define the role of cytokines in COVID-19-related deaths. Methods: In all the selected cases SARS-CoV-2 RT-PCR was performed on swabs collected from the upper (naso-oropharynx) and/or the lower respiratory (trachea and primary bronchi). A total of 60 SARS-CoV-2 positive subjects and 20 controls were included. Histological (hematoxylin–eosin staining) and immunohistochemical semi-quantitative analysis were performed on the multiorgan samples taken during autopsy. Tissue localization of SARS-CoV-2 was detected using antibodies against the nucleoprotein and the spike protein. The presence of viral particles was assessed using electron microscopy. Results: Immunohistochemical staining showed that endothelial cells expressed IL-1β in lung samples obtained from COVID-19 group (p < 0.001). Similarly, alveolar capillary endothelial cells showed strong and diffuse immunoreactivity for IL-6 and IL-15 in the COVID-19 group (p < 0.001). TNF-α showed a higher immunoreactivity and CD8+ T cells where more numerous in the lung samples obtained from COVID-19 group (p < 0.01). Electron microscopy revealed viral particles within the cells of the lungs, trachea, kidney, and large intestine. Discussion: Our study highlighted the morphological impact of the cytokine storm triggered by SARS-CoV-2 infection and the potent inflammatory response involved in the pathogenesis of COVID-19. We have experimentally confirmed that proinflammatory cytokines, such as IL-1β, IL-6, IL-15, and TNF-α, are produced at sites of tissue inflammation, determining a specific immune response. However, further autopsy studies are needed to expand this evidence and highlight the pathophysiology of SARS-CoV-2 infection.