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SpheroidChip: Patterned Agarose Microwell Compartments Harboring HepG2 Spheroids are Compatible with Genotoxicity
Christy Chao1, P Ngo Le2, Bevin P Engelward2
1Department of Chemical Engineering, Massachusetts Institute of Technology, 77 Massachusetts Avenue, Cambridge, MA, 02139, USA.
ACS Biomaterials Science & Engineering
|November 4, 2020
Summary
A novel agarose microwell platform simplifies the production of hepatocyte spheroids for liver toxicity testing. This SpheroidChip enables high-throughput genotoxicity analysis using the comet assay, improving upon traditional methods.
Area of Science:
- Biotechnology and Biomedical Engineering
- Toxicology and Pharmacology
- Cell Biology and Tissue Engineering
Background:
- Three-dimensional (3D) tissue culture models, particularly hepatocyte spheroids, offer improved liver-specific functions compared to traditional animal testing.
- Existing spheroid formation techniques (e.g., ultralow attachment plates, hanging drops) are often low-throughput and technically demanding.
- Genotoxicity testing, such as the comet assay, is crucial for assessing chemical-induced DNA damage but can be challenging with small spheroid models.
Purpose of the Study:
- To develop a user-friendly platform for enhanced spheroid production and high-throughput genotoxicity assessment.
- To integrate spheroid culture and comet assay analysis within a single microwell array system.
- To validate the platform's utility for detecting DNA damage and repair in liver cells.
Main Methods:
- Fabrication of a SpheroidChip featuring an agarose microwell array within a standard 96-well plate format.
- Formation of HepG2 cell spheroids via agarose-assisted aggregation within the microwells over 1-2 days.
- Adaptation and application of the comet assay directly on intact spheroids cultured in the microwells for DNA damage quantification.
Main Results:
- The SpheroidChip successfully produced numerous hepatocyte spheroids per well, significantly increasing production efficiency.
- The platform enabled direct electrophoretic analysis of DNA damage in HepG2 spheroids using a modified comet assay.
- A dose-dependent increase in DNA damage was observed following exposure to inflammatory chemicals (H₂O₂ and SIN-1), along with evidence of rapid H₂O₂-induced DNA damage repair.
Conclusions:
- The developed agarose microwell platform (SpheroidChip) streamlines spheroid generation and facilitates complex analyses like the comet assay.
- This integrated system condenses a 96-well plate's worth of analysis into a single well, overcoming limitations of previous methods.
- The SpheroidChip represents a significant advancement for high-throughput liver toxicity and genotoxicity screening, offering a more efficient alternative to animal testing.
