Generation of vascularized liver spheroid on-chip model for in-vitro infectious disease studies

Background

Our lab develops organs-on-chip (OOC) models, focusing on the lungs, liver and gut. We are creating vascularized tissues-on-chip (patent filed) that better mimic the tissue microenvironment, enabling interaction studies between the vascular systems and their surroundings. Several projects, either already running or planned, will directly benefit from the new approach that will be learned at InSphero. After investigating the transcriptional and morphological differences between uni- and bidirectional flows on the endothelium (manuscript in preparation), I plan to create vascularized liver tissue based on this study and the InSphero spheroids. This project, funded by the MCID (Multidisciplinary Center for Infectious Diseases), aims to mimic HBV and malaria infection and will lay the groundwork for a larger high-throughput study aimed at evaluating the efficacy of various therapeutic agents against both malaria and HBV. This will be realized with an automated microscopy system for High throughput screening (HTS) of OOC (project applied).

Aim

University of Bern -> Insphero AG

InSphero, the spheroid global market leader, specializes in liver spheroids models. I am aiming to acquire skills in 3D liver cell culture techniques, isolation of RNA from the microtissues, cell viability assay, cell cytotoxicity (LDH), multiplex Luminex assay, functional characterization of the cultures (albumin, urea, …), preparation of the samples (immunohistochemistry, histology techniques) for fluorescent and confocal microscopy including high content imagining.

In vitro liver models have improved significantly with developing liver spheroids and incorporating primary human cells. However, cultivating primary human hepatocytes faces several challenges, including short-term in vitro culture, low proliferation rate, and dedifferentiation tendency. Therefore, cancer cell lines, such as HepG2, were widely used in 2D and 3D models. The InSphero method, introduced over 10 years ago, effectively mimics the microarchitecture of the liver, generating reproducible, long-term liver spheroid co-cultures of primary cells. The technique offers high-throughput physiologically relevant microtissues to study disease models and host-pathogen interactions.

Our project aims to study HBV and malaria infection on a vascularized liver spheroid model. Pathogens interact with host cells via surface markers, making it essential to recreate the physiological microenvironment to preserve the cell niche. Primary cells undergo alteration in vitro, affecting the gene and protein expression profile, which can negatively impact studies of host-pathogen interactions. This 3D co-culture of vascularized hepatocytes with fibroblasts and endothelial cells provides an in vivo-like environment to maintain cell phenotype and obtain the required infection rate, which cannot be achieved with cancer cell lines cultured in 2D; in order to develop the infection models, it is crucial to adapt the established protocols for vascularization at InSphero, screen the culture for the expression level of the entrance markers of the infection particle and to introduce the spheroids to the microfluidics device on a specific timeline.