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Multilayered Cell Sheets of Cardiac Reprogrammed Cells for the Evaluation of Drug Cytotoxicity
Sung Pil Kwon1, Seuk Young Song1, Jin Yoo1
1School of Chemical and Biological Engineering, Seoul National University, Seoul, 08826, Republic of Korea.
Background:
Various cell-culture systems have been used to evaluate drug toxicity in vitro. However, factors that affect cytotoxicity outcomes in drug toxicity evaluation systems remain elusive. In this study, we used multilayered sheets of cardiac-mimetic cells, which were reprogrammed from human fibroblasts, to investigate the effects of the layer number on drug cytotoxicity outcomes.
Methods:
Cell sheets of cardiac-mimetic cells were fabricated by reprogramming of human fibroblasts into cardiac-mimetic cells via coculture with cardiac cells and electric stimulation, as previously described. Double-layered cell sheets were prepared by stacking the cell sheets. The mono- and double-layered cell sheets were treated with 5-fluorouracil (5-FU), an anticancer drug, in vitro. Subsequently, apoptosis and lipid peroxidation were analyzed. Furthermore, effects of cardiac-mimetic cell density on cytotoxicity outcomes were evaluated by culturing cells in monolayer at various cell densities.
Results:
The double-layered cell sheets exhibited lower cytotoxicity in terms of apoptosis and lipid peroxidation than the mono-layered sheets at the same 5-FU dose. In addition, the double-layered cell sheets showed better preservation of mitochondrial function and plasma membrane integrity than the monolayer sheets. The lower cytotoxicity outcomes in the double-layered cell sheets may be due to the higher intercellular interactions, as the cytotoxicity of 5-FU decreased with cell density in monolayer cultures of cardiac-mimetic cells.
Conclusion:
The layer number of cardiac-mimetic cell sheets affects drug cytotoxicity outcomes in drug toxicity tests. The in vitro cellular configuration that more closely mimics the in vivo configuration in the evaluation systems seems to exhibit lower cytotoxicity in response to drug.
Insights
Multilayered cardiac-mimetic cell sheets show reduced drug cytotoxicity compared to single-layered sheets. This finding suggests that in vitro models mimicking in vivo cellular structures improve drug toxicity evaluations.
Area of Science:
- Biomedical Engineering
- Cell Biology
- Pharmacology
Background:
- In vitro cell-culture systems are crucial for evaluating drug toxicity, but factors influencing cytotoxicity outcomes are not fully understood.
- Existing models often lack the complex cellular architecture found in vivo, potentially affecting toxicity assessments.
- Cardiac-mimetic cells, reprogrammed from human fibroblasts, offer a promising model for studying drug effects.
Purpose of the Study:
- To investigate the impact of cell layer number on drug cytotoxicity outcomes using multilayered cardiac-mimetic cell sheets.
- To compare the cytotoxicity of a model drug (5-fluorouracil) in mono-layered versus double-layered cardiac-mimetic cell sheets.
- To explore the relationship between cell density, intercellular interactions, and drug cytotoxicity.
Main Methods:
- Cardiac-mimetic cell sheets were generated by reprogramming human fibroblasts.
- Mono-layered and double-layered cell sheets were created and treated with 5-fluorouracil (5-FU).
- Apoptosis, lipid peroxidation, mitochondrial function, and plasma membrane integrity were analyzed. Cell density effects were studied in monolayer cultures.
Main Results:
- Double-layered cell sheets demonstrated significantly lower cytotoxicity (reduced apoptosis and lipid peroxidation) than mono-layered sheets when exposed to the same 5-FU dose.
- The multilayered configuration better preserved mitochondrial function and plasma membrane integrity compared to the monolayer.
- Reduced cytotoxicity in multilayered sheets correlated with increased intercellular interactions, mirroring findings where higher cell density in monolayers decreased 5-FU toxicity.
Conclusions:
- The number of cell layers in cardiac-mimetic cell sheets critically influences drug cytotoxicity outcomes in in vitro toxicity testing.
- In vitro cellular configurations that better replicate in vivo tissue structures appear to exhibit reduced cytotoxicity.
- This study highlights the importance of mimicking in vivo cellular architecture for more accurate drug toxicity evaluations.
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