Developing a 3D tri-culture cardiovascular in vitro model for assessing drug cardiotoxicity

Abstract:

Introduction

Cardiotoxicity is a leading cause of drug withdrawal from potential drug development. There is a need to improve current in vitro cardiotoxicity models to provide an earlier and more efficient assessment of a drug’s cardiotoxicity, minimising drug attrition in the late phases of development. Current in vitro assays use 2D monolayers of single cell types, and do not take into the account the effect of flow. Therefore, we are developing a 3D in vitro model, using a tri-culture of human heart cells and exposing them to dynamic culture.

 

Materials and Methods

Primary human coronary artery smooth muscle cells, endothelial cells and cardiomyocytes (Promocell, uk) were seeded (in an equal ratio of 1:1:1) at a seeding density of 3.33 x 103 cells/ml onto gelatin coated 13mm2 glass coverslips. A perfusion chamber (QV-500 Quasi-Vivo®, Kirkstall Ltd.) was used to expose cells to a flow rate of 50µl/min, for up to 7 days in culture. Cells were assessed for cell viability, using Live/dead assay and alamar blue (mono-cultures) and stained with cell tracker (tri-culture). Polyurethane 3D scaffolds were manufactured using a ProJet MJP 2500 Plus printer (R&D Systems). Scaffolds were characterised using scanning electron microscopy.

 

Results and Discussion

Cell viability of each cell type in mono-culture, under dynamic conditions, significantly increased compared to cells cultured under static conditions. Dynamic conditions significantly increased the number of human endothelial cells after 7 days in culture, compared to static conditions, 97.48± 6.85 live cells per mm2 compared to 62.32 ± 4.01 live cells per mm2. A tri-culture of cardiac cells (Cardiomyocytes, endothelial cells, smooth muscle cells) was established on glass coverslips (2D). Cell survival, of all three cell types, was demonstrated in 2D tri-culture and was applied to 3D conditions.

 

Conclusions

Overall, the effect of flow has been seen to amplify the response/sensitivity of the cells, bringing the culture environment closer to in vivo. Future work includes improving 3D tri-culture, as well as exposing the model to known cardiotoxicity causing drug Doxorubicin, to validate its efficiency.

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