A novel and cost-effective method for high-throughput 3D culturing and rhythmic assessment of hiPSC-derived cardiomyocytes using retroreflective Janus microparticles

다국어 초록 (Multilingual Abstract)

Background Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) gain attention as a potent cell source in regenerative medicine and drug discovery. With the necessity of the demands for experimental models to create a more physiologically relevant model of the heart in vitro we herein investigate a 3D culturing platform and a method for assessing rhythm in hiPSC-CMs.
Methods The 3D cell culture PAMCELL™ plate is designed to enable cells to attach exclusively to adhesive patterned areas. These cell adhesive zones, named as micro-patterned pads, feature micron silica beads that are surfacemodified with the well-known arginyl-glycyl-aspartic acid (RGD) peptide. RGD binding to the surface of hiPSC-CMs facilitates cell–cell attachment and the formation of uniform-size spheroids, which is controlled by the diameter of the micro-patterned pads. The assessment and evaluation of 3D hiPSC-CMs beating pattern are carried out using reflective properties of retroreflective Janus micro-particle (RJP). These RJPs are modified with an antibody targeting the gap junction protein found on the surface of hiPSC-CM spheroids. The signal assessment system comprises a camera attached to an optical microscope and a white light source.
Results The 3D PAMCELL™ R100 culture plate efficiently generate approximately 350 uniform-sized hiPSC-CM spheroids in each well of a 96-well plate and supported a 20-day culture. Analysis of genes and protein expression levels reveal that iPSC-CM spheroids grown on PAMCELL™ R100 retain cardiac stem cell characteristics and functions, outperforming traditional 2D culture platform. Additionally, the RJPs enable monitoring and evaluation of in vitro beating properties of cardiomyocytes without using complex monitoring setup. The system demonstrates its capability to identify alteration in the rhythmic activity of cardiac cells when exposed to ion channel blockers, nifedipine and E4031.