Hepatic ACE2 expression is upregulated in early NAFLD and in patients who died with severe COVID-19: a histological analysis

Non-alcoholic fatty liver disease (NAFLD) has been identified as a predictor of liver injury in patients hospitalised with coronavirus disease 2019 (COVID-19) and there are emerging reports of COVID-19-associated cholangiopathies, highlighting the requirement for further study of the principal mechanisms underlying liver injury. Our study aimed to (i) study hepatic expression of the key severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) entry point, angiotensin converting enzyme 2 (ACE2), across the NAFLD spectrum to determine the time course and cellular source(s); (ii) determine whether the level of hepatic ACE2 expression in patients who died with severe COVID-19 correlated with the presence of virus in liver tissue. We carried out a histological analysis of human liver tissue sections across the NAFLD spectrum (from isolated steatosis, through to non-alcoholic steatohepatitis (NASH) +/- fibrosis, to cirrhosis) and from patients who died with severe COVID-19 and had varying degrees of pre-existing chronic liver disease. ACE2 immunostaining was quantified using machine learning-based pixel classifiers and picrosirius red-stained sections were analysed for fibrosis content and lipid droplet accumulation. Hepatic ACE2 protein expression determined by immunohistochemistry is upregulated in early stage NAFLD, maximally in NASH samples compared with healthy control livers. Hepatic ACE2 protein levels did not correlate with SARS-CoV-2 RNA transcripts in liver tissue, although, ACE2 immunostaining was more marked in patients who died with severe COVID-19 compared to healthy controls. Greater hepatocyte lipid droplet accumulation was associated with a positive hepatic SARS-CoV-2 PCR result. In addition, ACE2 and SARS-CoV-2 spike protein expression was colocalised in cholangiocytes. The increase in hepatic ACE2 protein levels in patients with NASH and more severe COVID-19 is consistent with reports in in vitro and in vivo models of NAFLD of the beneficial anti-inflammatory effects of the counter-regulatory renin angiotensin system (Song et al., 2020). In conjunction with previous findings showing that cellular lipid accumulation can augment SARS-CoV-2 replication (Dias et al., 2020), our results suggest that hepatic lipid droplet accumulation may be a risk factor for direct viral-mediated liver damage in severe COVID-19. Moreover, colocalization of ACE2 and viral spike protein in cholangiocytes suggests that direct viral-mediated damage could play a role in COVID-19-associated cholangiopathies.