High-throughput bioengineering of homogenous and functional human-induced pluripotent stem cells-derived liver organoids via micropatterning technique

Abstract

Human pluripotent stem cell-derived liver organoids are emerging as more human-relevant in vitro models for studying liver diseases and hepatotoxicity than traditional hepatocyte cultures and animal models. The generation of liver organoids is based on the Matrigel dome method. However, the organoids constructed by this method display significant heterogeneity in their morphology, size, and maturity. Additionally, the formed organoid is randomly encapsulated in the Matrigel dome, which is not convenient for in situ staining and imaging. Here, we demonstrate an approach to generate a novel type of liver organoids via micropatterning technique. This approach enables the reproducible and high-throughput formation of bioengineered fetal liver organoids with uniform morphology and deterministic size and location in a multiwell plate. The liver organoids constructed by this technique closely recapitulate some critical features of human liver development at the fetal stage, including fetal liver-specific gene and protein expression, glycogen storage, lipid accumulation, and protein secretion. Additionally, the organoids allow whole-mount in-situ staining and imaging. Overall, this new type of liver organoids is compatible with the pharmaceutical industry’s widely-used preclinical drug discovery tools and will facilitate liver drug screening and hepatotoxic assessment.

Toward reproducible tumor organoid culture: focusing on primary liver cancer

“…Similarly, Xu et al. ( 76 ) designed a micropatterning technique that can be used to generate liver organoids with a uniform deterministic size in a multi-well plate in a reproducible and high-throughput manner. Tienderen et al.…”

The native liver as inspiration to create superior in vitro hepatic models

“…Since organoids are currently being cultured using the ECM-dome method, there is only little control on the microenvironment, distribution, reproducibility, as well as organoid size, density and mass. To overcome these problems, Xu et al 200 developed a PEG-based micropatterned cell-adhesion substrate to culture individual liver organoids in a controlled way in order to promote the application of organoids for high-throughput DILI testing resulting in an in vitro model representative for the fetal liver. Jiang et al 201 developed another micropatterned organoid model using microfabricated hexagonal closely packed cavity arrays, with a single organoid in each cavity.…”

Reproducible generation of human liver organoids (HLOs) on a pillar plate platformviamicroarray 3D bioprinting

“…Additionally, a micropatterned cell-adhesion substrate (MPCS) in a 24-well plate was introduced by oxygen plasma treatment for high-throughput generation of functional HLOs 17 . Despite improved organoid availability, these organoid culture systems are not easily amenable to HTS of compounds due to the need for isolation and encapsulation of HLOs in biomimetic hydrogels, imposing low throughput for downstream analyses.…”