Modeling the cardiac pathological traits would be of paramount utility to elucidate possible targets of new therapeutics for unmet pathology (e.g., Dilated Cardiomyopathy (DCM )). Traditional in vitro systems lack the complexity of human physiological conditions, resulting poorly reliable for tissue engineering studies. Organs-on-chip (OoC) have been shown to be a promising alternative, offering the ease to integrate stimulation and sensors . In previously developed beating heart-on-chip –, despite advanced tissue maturation through mechanical stimulation have been demonstrated, the coupling with electrical readout allowing for a broader electrophysiological recording space and high throughput analyses is still missing.
To overcome these issues, we developed µPEA, an electrode array system coupled with the beating-heart-on-chip  able to provide 3D microtissues with mechanical stimulation and electrical readouts, using noninvasive 2D electrodes. We validated µPEA-Heart-on-Chip by assessing electrophysiology of neonatal rat cells and we exploited it to study the effect of mechanical stimulation on human DCM models.
Electrodes were designed using CAD and developed in glass substrate using physical vapor deposition (Cr/Au, 100nm). Electrodes arrays consist of five recordings, two references and one ground electrodes for measuring field potential. Chips were connected to a platform interface with a multiplexing system followed by an amplifier coupled with a bandpass filter (0.5-100Hz).
Electrodes characterizations were done by measuring impedance spectroscopy and by filling the OoC with PBS. iPSC-CM were generated by following well-established differentiation protocols of iPSC derived from DCM patients . Neonatal rat cardiomyocytes, DCM iPSC-CMs, and isogenic controls were embedded in fibrin gel at 80-120 · 106 cells/mL and cultured for 5-10 days in static or mechanically active (i.e. 10% uniaxial strain at 1Hz) environment. After maturation, electrophysiological signals were measured from different electrodes.
The µPEA-Heart-on-Chip was successfully assembled by integrating and aligning the innovative patterned surface with the mechanically active OoC (the electrodes are positioned along the cardiac microtissue). Electrodes impedance measurements were 124.5 ± 3.1 KΩ at 1Khz, in line with literature values . Neonatal rat cardiomyocytes microtissues started to spontaneously beat after 4 days in culture. Field potentials were successfully measured through all five different electrodes and key parameters were evaluated. DCM iPSC-CM were efficiently differentiated in 2D, as evidenced by immunofluorescence staining of cardiac Troponin T. The DCM diseased model was established within the µPEA-Heart-on-Chip, with cells that organized and interconnected within the 3D environment. Electrophysiological and contractility changes of the DCM model in response to the mechanical stimulation is currently under evaluation.
In Conclusion, here we described the development of the µPEA-Heart-on-Chip capable to integrate a mechanical stimulation with an electrophysiological activity recording system. The platform represents an unprecedented tool to investigate electrical cardiac alterations in healthy or diseased model in response to mechanical stimulation.
1.B. J. Maron et al, Circulation, 2006.
2.G. A. Clarke et al, Sensors, 2021.
3.A. Marsano et al., Lab Chip, 2016.
4.R. Visone et al, APL Bioeng., 2018.
5.R. Visone et al., Biofabrication, 2021.
6.N. Sun et al., Sci. Transl. Med., 2012.
7. H. Cui et al., Biomed. Eng. Online, 2019.