SpheroidChip: Patterned Agarose Microwell Compartments Harboring HepG2 Spheroids are Compatible with Genotoxicity Testing
Three-dimensional tissue culture models are emerging as effective alternatives to animal testing. They are especially beneficial for liver toxicity studies, enabling hepatocytes to display improved levels of liver-specific functions. One common model is hepatocyte spheroids, which are spontaneously...
| Main Authors: | , , |
|---|---|
| Other Authors: | |
| Format: | Article |
| Language: | English |
| Published: |
American Chemical Society (ACS)
2020
|
| Online Access: | https://hdl.handle.net/1721.1/126762 |
| _version_ | 1826210335800950784 |
|---|---|
| author | Chao, Christy Ngo, Le P Engelward, Bevin P |
| author2 | Massachusetts Institute of Technology. Department of Chemical Engineering |
| author_facet | Massachusetts Institute of Technology. Department of Chemical Engineering Chao, Christy Ngo, Le P Engelward, Bevin P |
| author_sort | Chao, Christy |
| collection | MIT |
| description | Three-dimensional tissue culture models are emerging as effective alternatives to animal testing. They are especially beneficial for liver toxicity studies, enabling hepatocytes to display improved levels of liver-specific functions. One common model is hepatocyte spheroids, which are spontaneously formed cell aggregates. Techniques for spheroid formation include the use of ultralow attachment plates and the hanging drop method, both of which are technically challenging and relatively low throughput. Here, we describe a simple-to-use platform that improves spheroid production and is compatible with genotoxicity testing by the comet assay. To achieve this, we created a chip containing a microwell array where dozens of spheroids are contained within a single well of a 96-well plate. The microwells are made from agarose, a nontoxic material suitable for cell growth and spheroid formation. HepG2 cells loaded into customizable microwells formed spheroids through agarose-assisted aggregation within one to two days. In addition, the agarose matrix allows the same platform to be used in DNA damage analysis. Specifically, the comet assay enables quantification of DNA breaks based on the increased migration of damaged DNA through agarose during electrophoresis. Here, we developed a modified comet assay and show that intact HepG2 spheroids cultured in microwells can be electrophoresed to reveal the extent of DNA damage following exposure to inflammatory chemicals (H2O2 and SIN-1). With this SpheroidChip analysis method, we detected a dose-dependent increase in DNA damage and observed rapid repair of H2O2-induced DNA damage. In summary, we utilized an agarose microarray to condense what had required an entire 96-well plate into a single well, enabling analysis techniques that were cumbersome or impossible under conditions of a single spheroid per well of a 96-well plate. |
| first_indexed | 2024-09-23T14:48:10Z |
| format | Article |
| id | mit-1721.1/126762 |
| institution | Massachusetts Institute of Technology |
| language | English |
| last_indexed | 2024-09-23T14:48:10Z |
| publishDate | 2020 |
| publisher | American Chemical Society (ACS) |
| record_format | dspace |
| spelling | mit-1721.1/1267622022-10-01T22:34:35Z SpheroidChip: Patterned Agarose Microwell Compartments Harboring HepG2 Spheroids are Compatible with Genotoxicity Testing Chao, Christy Ngo, Le P Engelward, Bevin P Massachusetts Institute of Technology. Department of Chemical Engineering Massachusetts Institute of Technology. Department of Biological Engineering Three-dimensional tissue culture models are emerging as effective alternatives to animal testing. They are especially beneficial for liver toxicity studies, enabling hepatocytes to display improved levels of liver-specific functions. One common model is hepatocyte spheroids, which are spontaneously formed cell aggregates. Techniques for spheroid formation include the use of ultralow attachment plates and the hanging drop method, both of which are technically challenging and relatively low throughput. Here, we describe a simple-to-use platform that improves spheroid production and is compatible with genotoxicity testing by the comet assay. To achieve this, we created a chip containing a microwell array where dozens of spheroids are contained within a single well of a 96-well plate. The microwells are made from agarose, a nontoxic material suitable for cell growth and spheroid formation. HepG2 cells loaded into customizable microwells formed spheroids through agarose-assisted aggregation within one to two days. In addition, the agarose matrix allows the same platform to be used in DNA damage analysis. Specifically, the comet assay enables quantification of DNA breaks based on the increased migration of damaged DNA through agarose during electrophoresis. Here, we developed a modified comet assay and show that intact HepG2 spheroids cultured in microwells can be electrophoresed to reveal the extent of DNA damage following exposure to inflammatory chemicals (H2O2 and SIN-1). With this SpheroidChip analysis method, we detected a dose-dependent increase in DNA damage and observed rapid repair of H2O2-induced DNA damage. In summary, we utilized an agarose microarray to condense what had required an entire 96-well plate into a single well, enabling analysis techniques that were cumbersome or impossible under conditions of a single spheroid per well of a 96-well plate. National Institute of Environmental Health Sciences (Grants T32-ES007020, P42-ES027707, P30-ES002109) 2020-08-24T18:16:04Z 2020-08-24T18:16:04Z 2020-03 2019-12 2020-08-21T11:46:12Z Article http://purl.org/eprint/type/JournalArticle 2373-9878 https://hdl.handle.net/1721.1/126762 Chao, Christy et al. "SpheroidChip: Patterned Agarose Microwell Compartments Harboring HepG2 Spheroids are Compatible with Genotoxicity Testing." ACS Biomaterials Science & Engineering 6, 4 (March 2020): 2427–2439 © 2020 American Chemical Society en http://dx.doi.org/10.1021/acsbiomaterials.9b01951 ACS Biomaterials Science & Engineering Creative Commons Attribution-Noncommercial-Share Alike http://creativecommons.org/licenses/by-nc-sa/4.0/ application/pdf American Chemical Society (ACS) Kathleen L Reposa |
| spellingShingle | Chao, Christy Ngo, Le P Engelward, Bevin P SpheroidChip: Patterned Agarose Microwell Compartments Harboring HepG2 Spheroids are Compatible with Genotoxicity Testing |
| title | SpheroidChip: Patterned Agarose Microwell Compartments Harboring HepG2 Spheroids are Compatible with Genotoxicity Testing |
| title_full | SpheroidChip: Patterned Agarose Microwell Compartments Harboring HepG2 Spheroids are Compatible with Genotoxicity Testing |
| title_fullStr | SpheroidChip: Patterned Agarose Microwell Compartments Harboring HepG2 Spheroids are Compatible with Genotoxicity Testing |
| title_full_unstemmed | SpheroidChip: Patterned Agarose Microwell Compartments Harboring HepG2 Spheroids are Compatible with Genotoxicity Testing |
| title_short | SpheroidChip: Patterned Agarose Microwell Compartments Harboring HepG2 Spheroids are Compatible with Genotoxicity Testing |
| title_sort | spheroidchip patterned agarose microwell compartments harboring hepg2 spheroids are compatible with genotoxicity testing |
| url | https://hdl.handle.net/1721.1/126762 |
| work_keys_str_mv | AT chaochristy spheroidchippatternedagarosemicrowellcompartmentsharboringhepg2spheroidsarecompatiblewithgenotoxicitytesting AT ngolep spheroidchippatternedagarosemicrowellcompartmentsharboringhepg2spheroidsarecompatiblewithgenotoxicitytesting AT engelwardbevinp spheroidchippatternedagarosemicrowellcompartmentsharboringhepg2spheroidsarecompatiblewithgenotoxicitytesting |