UCSF

RNA viruses, including SARS-CoV-2, pose a dire threat to human health around the globe. Single-stranded RNA viruses target a wide range of organs, causing a diverse set of clinical symptoms. Here, Hepatitis C virus (HCV) in the Flaviviridae family, which targets the liver, and SARS-CoV-2 in the Coronaviridae family, which primarily targets lung cells, are studied in stem cell models. However, in vitro studies have been limited by the lack of robust laboratory model systems. The recent development of organoids and other stem cell-derived systems has enabled studies with tractable, biologically relevant, genetically diverse human systems to understand viral replication and innate immune response. Other in vitro models lack cellular polarity and have a dysregulated immune response while naturally susceptible in vivo models such as non-human primates are often not accessible for many laboratories.

SARS-CoV-2 is the human pathogenic coronavirus causing COVID-19. While the primary target for the virus is lung epithelial cells, symptoms can be found across multiple organ systems including the gut, heart, and brain. The need to study the pathogenesis of SARS-CoV-2 in different cell types became clear early in the COVID-19 pandemic. Here, we used iPSC-derived cardiomyocytes to uncover sarcomeric fragmentation as a potential mechanism for cardiac-related symptoms during or after COVID-19 infection. To understand the effects of SARS-CoV-2 and viral variants that arose during the pandemic, an adult stem cell-derived airway organoid model was used. We found that organoids naturally had low levels of ACE2 receptor expression and supported low levels of SARS-CoV-2 infection. Overexpression of ACE2 significantly increased the percentage of cells that got productively infected. Single cell RNA-sequencing showed that cells had high expression of interferon-stimulated genes as well as of interferon beta and lambda.

HCV infects hepatocytes and in 70% of cases leads to hepatocellular carcinoma if left untreated. While direct-acting antivirals (DAAs) have been a breakthrough in HCV treatment, they are expensive and do not prevent reinfection. Vaccination which stimulates T cell responses is essential for reducing HCV incidence, and an in vitro system which is susceptible to HCV and can interact with T cells is critically missing. Here, we develop a new system to coculture primary liver organoids derived from HCV+ patients with cytotoxic T cells recognizing a specific HCV epitope. Using quantitative time course microscopy, organoid viability is successfully tracked after peptide pulsing, with organoids expressing the HCV peptide dying at significantly higher rates than organoids without the peptide. Collectively, these studies take advantage of novel organoid technology to bring insight into the pathogenesis of two important viral infections: SARS-CoV-2 and HCV. We expect our studies to be impactful in the future by therapeutically addressing cardiac comorbidities in COVID-19 and finding vaccines for HCV.