Understanding cell type responses to SARS-CoV-2 coronavirus spike proteins

Abstract

The angiotensin converting enzyme 2 (ACE2) receptor is important for the regulation of blood pressure and as an entry receptor for the SARS-CoV-2 coronavirus. ACE2 degrades the vasoconstrictive Ang II protein to assist in the maintenance of fluid homeostasis. As the entry receptor for SARS-CoV-2, ACE2 expression can predict susceptibility to infection alongside host priming enzymes including TMPRSS2. Whilst SARS-CoV-2 is largely associated with lung pathology, extrapulmonary organs including the heart, kidney and small intestine are also implicated in COVID-19. Disruption to ACE2 levels during COVID-19 is associated with exacerbated immune responses leading to the development of cytokine storms and severe disease states. In this thesis, ACE2 expression has been assessed in keratinocyte, placental and cervical carcinoma cell lines. Furthermore, the regulation of ACE2 by cytokines was also investigated. The administration of recombinant spike proteins did not alter ACE2 expression or localisation in keratinocytes, although
ACE2 downregulation was indicated within placental cells, demonstrating potential cell line specific responses to SARS-CoV-2 protein. Assays were also devised to investigate downstream responses to spike proteins in HaCaT, BeWo and HeLa cell lines. The gamma, kappa and omicron spike protein
variants were unable to affect the gross expression of the ACE2 receptor or cytokine responses. This thesis assesses the suitability of ACE2 antibodies for protein expression and immunofluorescence analysis and provides a basis for future investigation of SARS-CoV-2 spike responses within
susceptible cell lines.