A 3D microfluidic liver model for high throughput compound toxicity screening in the OrganoPlate®
| Autor: | Mark T. Miedel, Anthony Saleh, Lawrence Vernetti, Richard DeBiasio, Alaa Alsebahi, Kristin M. Bircsak, Ryan Reddinger, Albert Gough, Tongying Shun |
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| Rok vydání: | 2020 |
| Předmět: |
0301 basic medicine
Cell Survival High-throughput screening Induced Pluripotent Stem Cells Microfluidics Cell Culture Techniques Toxicology 03 medical and health sciences Troglitazone 0302 clinical medicine Toxicity Tests medicine Cytochrome P-450 CYP3A Humans Viability assay Induced pluripotent stem cell Dose-Response Relationship Drug Chemistry Hepatotoxin Albumin High-Throughput Screening Assays 030104 developmental biology medicine.anatomical_structure Biochemistry Liver Hepatocyte Liquid handling robot Hepatocytes 030217 neurology & neurosurgery medicine.drug |
| Zdroj: | Toxicology. 450 |
| ISSN: | 1879-3185 |
| Popis: | We report the development, automation and validation of a 3D, microfluidic liver-on-a-chip for high throughput hepatotoxicity screening, the OrganoPlate LiverTox™. The model is comprised of aggregates of induced pluripotent stem cell (iPSC)-derived hepatocytes (iHep) seeded in an extracellular matrix in the organ channel and co-cultured with endothelial cells and THP-1 monoblasts differentiated to macrophages seeded in the vascular channel of the 96 well Mimetas OrganoPlate 2-lane. A key component of high throughput screening is automation and we report a protocol to seed, dose, collect and replenish media and add assay reagents in the OrganoPlate 2-lane using a standard laboratory liquid handling robot. A combination of secretome measurements and image-based analysis was used to demonstrate stable 15 day cell viability, albumin and urea secretion. Over the same time-period, CYP3A4 activity increased and alpha-fetoprotein secretion decreased suggesting further maturation of the iHeps. Troglitazone, a clinical hepatotoxin, was chosen as a control compound for validation studies. Albumin, urea, hepatocyte nuclear size and viability staining provided Robust Z’factors > 0.2 in plates treated 72 h with 180 μM troglitazone compared with a vehicle control. The viability assay provided the most robust statistic for a Robust Z’ factor = 0.6. A small library of 159 compounds with known liver effects was added to the OrganoPlate LiverTox model for 72 h at 50 μM and the Toxicological Prioritization scores were calculated. A follow up dose-response evaluation of select hits revealed the albumin assay to be the most sensitive in calculating TC50 values. This platform provides a robust, novel model which can be used for high throughput hepatotoxicity screening. |
| Databáze: | OpenAIRE |
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