Abstract 20983: Size Based Microfluidic Enrichment of Human Inducedpluripotent Cell-derived Cardiomyocytes
While human induced pluripotent stem cell (iPSC)-derived CMs offer a tantalizing cellular model for drug discovery and understanding mechanisms underlying heart disease, current protocols yield highly heterogeneous populations [1,2]. The heterogeneity in iPSC-CM cultures suggests variation in the maturation, which has been correlated with cell size and shape [3-5]. This study offers a novel approach to enrich structurally mature-like iPSC-CMs, which would also address current inaccuracies in the assessment of properties of iPSC-CM cultures due to the cell-to-cell variations. We have used inertial microfluidics and have shown the feasibility of sorting live iPSC-CMs based on size and shape.
Materials and Methods: Cells were differentiated using previously described methods and cultured on Matrigel-coated dishes. iPSC-CMs were then re-suspended in RPMI media at 2-5x105 cells/mL density. After passing through the microfluidic chip (Fig. 1A) at 60μL/min , sorted cells were either collected for RNA extraction or cultured on Matrigel for further analysis.
Results and Discussion: We see ~2.5-fold increase in the diameter of cells in inner (O3) versus outer outlets (O2, O4). The purity of cells with diameter (d) larger than 7μm increased ~2.6-fold compared to the inlet (Fig. 1B). Cell viability after sorting did not change significantly relative to unsorted cells. Sorted cells showed spontaneous beating at day 2 or 3. We assessed the expression of multiple genes for cells captured in different outlets (Figure.1C). Larger cells, captured in the O3, showed higher expression of maturity markers. We observed a ~2.8-fold increase in the MYH7/MYH6 expression ratio in O3 compared to O2, O4. We are currently assessing the contractility and myofibril structural properties of cells captured in different outlets. The presented method offers a simple and elegant solution for selecting populations of iPSC-CM that are more relevant for drug discovery and disease studies.
Author Disclosures: A. Ribeiro: None. L. Lin: None. A. Dainis: None. E. Ashley: None. B. Pruitt: None.
- © 2016 by American Heart Association, Inc.