axoCells^TM atrial cardiomyocytes

Phenotypic characterization: ICC

axoCells atrial cardiomyocytes express the cardiac- and atrial-specific markers troponin T, atrial myosin light chain 2 (MLC2a) and atrial natriuretic peptide (ANP).

Immunocytochemistry data of cardiac- and atrial-specific proteins. Troponin T staining (red) confirmed the presence of cardiac myocytes, ANP is specifically secreted by atrial myocytes upon atrial stretching and MLC2a facilitates cardiac contractility. The nuclear marker DAPI was used as a counterstain.

Functional relevance: patch clamp

Here we show the functional validation of our axoCells atrial cardiomyocytes against 3 major drug classes: Lidocaine (an Na+ channel blocker), Nifedipine (a Ca2+ channel blocker) and E-4031 (a hERG channel blocker), measured by patch clamp, which is the “gold standard” electrophysiology method. There is expected response to the major drug classes, validating these cells for use in in vitro cardiac models.

axoCells human iPSC-Derived Atrial Cardiomyocytes express functional core cardiac ion channels, INa, ICa,L and IKr. Work done in collaboration with Metrion Biosciences.

A) Lidocaine (INa), Nifedipine (ICa,L), and E-4031 (IKr) were used to characterize the activity of core cardiac currents during iPSC-derived atrial cardiomyocyte action potentials. Each trace shows representative spontaneous action potentials under control conditions (grey) and in the presence of 100 μM Lidocaine (green), 100 nM Nifedipine (blue), and 100 nM E-4031 (red). Early afterdepolarizations (EADS) were observed after application of the hERG blocker E-4031 (arrow), indicative of arrhythmic activity.

B) Average effect of Lidocaine, Nifedipine, and E-4031 on atrial action potential parameters (% of control) (N=4).

Functional relevance: compound testing on patch clamp

Here we show the functional validation of our axoCells atrial cardiomyocytes against 2 atrial-specific compounds, 4-AP and carbachol, measured by patch clamp which is the “gold standard” electrophysiology method. There is expected response to these compounds, validating these cells for use in in vitro cardiac models.

Atrial cardiomyocyte phenotype was confirmed by the modulation of spontaneous action potentials using selective pharmacological compounds. Work done in collaboration with Metrion Biosciences.

Representative spontaneous action potentials are shown under control conditions (grey) and after the application of 50 μM 4-AP (A, blue) or 1 μM Carbachol (B, orange). The bar graphs show the average effect (% of control) on action potential parameters (N=5). (A) A low concentration of 4-aminopyridine (4-AP) selectively inhibits the IKur current to broaden action potential duration and slow spontaneous firing. (B) Carbachol was used to activate the IKACh current, resulting in action potential shortening and profound slowing of firing frequency.

Functional relevance: contractility

We tested the functional validation of our axoCells atrial cardiomyocytes on the InnoVitro Flexcyte 96 which measures contractility. We found expected responses to several compounds, demonstrating functional relevance, and clear chamber-specific responses between our axoCells atrial and ventricular cardiomyocytes.

Here you can see the effect of carbachol on mean beat rate, showing distinct chamber specificity and expected responses (carbachol is an activator of I(kAch) in atrial cells specifically).

The effect of carbachol, an activator of IKAch in atrial cardiomyocytes, on the contractility of axoCells atrial (left) and ventricular (right) hiPSC-CMs, as measured on the Flexcyte 96.

The effect of carbachol, an activator of IKAch in atrial cardiomyocytes, on the mean beat rate of axoCells atrial (left) and ventricular (right) hiPSC-CMs, as measured on the Flexcyte 96.

Functional relevance: MEA

We’ve extensively characterized our axoCells atrial cardiomyocytes using our industry-leading Axion Maestro Pro MEA system, which we also use for our axoServices electrophysiology assays.

Here you can see multiple parameters measured via the MEA system which demonstrate the functional relevance and chamber specificity of our atrial and ventricular cardiomyocytes.

Here you can see the field action potential (fAP, top) and contractility waveforms (middle) of our atrial and ventricular cardiomyocytes measured on the Axion Maestro Pro MEA system.

In the bottom graph, you can also see a comparison of response to dofetilide (a hERG blocker) of our axoCells atrial and ventricular cardiomyocytes on the Maestro Pro measuring the local extracellular action potential (LEAP), which enables electrophysiological recordings that are more similar to patch clamp.