Abstract
Hepatitis C virus (HCV) remains a global public health challenge with an estimated 71 million people chronically infected, with surges in new cases and no effective vaccine. New methods are needed to study the human immune response to HCV since in vivo animal models are limited and in vitro cancer cell models often show dysregulated immune and proliferative responses. Here, we developed a CD8+ T cell and adult stem cell liver organoid system using a microfluidic chip to coculture 3D human liver organoids embedded in extracellular matrix with HLA-matched primary human T cells in suspension. We then employed automated phase contrast and immunofluorescence imaging to monitor T cell invasion and morphological changes in the liver organoids. This microfluidic coculture system supports targeted killing of liver organoids when pulsed with a peptide specific for HCV non-structural protein 3 (NS3) (KLVALGINAV) in the presence of patient-derived CD8+ T cells specific for KLVALGINAV. This demonstrates the novel potential of the coculture system to molecularly study adaptive immune responses to HCV in an in vitro setting using primary human cells.
Highlights
Hepatitis C virus (HCV) is a positive-sense single-stranded RNA virus that targets hepatocytes, usually establishes chronic infection and, if untreated, can progress to cirrhosis and hepatocellular carcinoma [1]
We report the establishment of a tractable coculture system of liver organoids and antigen-specific T cells, which we envision could be applied towards identifying immunogenic HCV epitopes that lead to T cell-mediated killing of virally infected liver cells
We report here that organoid medium is suitable for coculture of the two cell populations in a (a) media + T cells liver organoid and T cell coculture royalsocietypublishing.org/journal/rsob Open Biol. 12: 210320 peptide-pulsed organoids in basement membrane extract (BME)
Summary
Hepatitis C virus (HCV) is a positive-sense single-stranded RNA virus that targets hepatocytes, usually establishes chronic infection and, if untreated, can progress to cirrhosis and hepatocellular carcinoma [1]. Despite the recent introduction of direct-acting antivirals against HCV, the global disease burden remains high, related to lack of access to treatment, expense of drugs and the possibility of reinfection after successful treatment is completed [4]. Despite a sustained virologic response (SVR), the risk of hepatocellular carcinoma in patients treated when advanced fibrosis was present remains higher in previously HCV-infected individuals compared to those who have never been infected [5]. This underscores the need for an effective vaccine, the development of which has been hampered by the high mutation rate of the virus and the lack of broadly neutralizing antibody induction after natural infection or vaccination strategies [6]
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