In:
PLOS ONE, Public Library of Science (PLoS), Vol. 18, No. 2 ( 2023-2-6), p. e0280406-
Abstract:
Recent advances in human induced pluripotent stem cell (hiPSC)-derived cardiac microtissues provide a unique opportunity for cardiotoxic assessment of pharmaceutical and environmental compounds. Here, we developed a series of automated data processing algorithms to assess changes in action potential (AP) properties for cardiotoxicity testing in 3D engineered cardiac microtissues generated from hiPSC-derived cardiomyocytes (hiPSC-CMs). Purified hiPSC-CMs were mixed with 5–25% human cardiac fibroblasts (hCFs) under scaffold-free conditions and allowed to self-assemble into 3D spherical microtissues in 35-microwell agarose gels. Optical mapping was performed to quantify electrophysiological changes. To increase throughput, AP traces from 4x4 cardiac microtissues were simultaneously acquired with a voltage sensitive dye and a CMOS camera. Individual microtissues showing APs were identified using automated thresholding after Fourier transforming traces. An asymmetric least squares method was used to correct non-uniform background and baseline drift, and the fluorescence was normalized (ΔF/F 0 ). Bilateral filtering was applied to preserve the sharpness of the AP upstroke. AP shape changes under selective ion channel block were characterized using AP metrics including stimulation delay, rise time of AP upstroke, APD 30 , APD 50 , APD 80 , APD mxr (maximum rate change of repolarization), and AP triangulation (APD tri = APD mxr −APD 50 ). We also characterized changes in AP metrics under various ion channel block conditions with multi-class logistic regression and feature extraction using principal component analysis of human AP computer simulations. Simulation results were validated experimentally with selective pharmacological ion channel blockers. In conclusion, this simple and robust automated data analysis pipeline for evaluating key AP metrics provides an excellent in vitro cardiotoxicity testing platform for a wide range of environmental and pharmaceutical compounds.
Type of Medium:
Online Resource
ISSN:
1932-6203
DOI:
10.1371/journal.pone.0280406
DOI:
10.1371/journal.pone.0280406.g001
DOI:
10.1371/journal.pone.0280406.g002
DOI:
10.1371/journal.pone.0280406.g003
DOI:
10.1371/journal.pone.0280406.g004
DOI:
10.1371/journal.pone.0280406.g005
DOI:
10.1371/journal.pone.0280406.g006
DOI:
10.1371/journal.pone.0280406.g007
DOI:
10.1371/journal.pone.0280406.g008
DOI:
10.1371/journal.pone.0280406.t001
DOI:
10.1371/journal.pone.0280406.t002
DOI:
10.1371/journal.pone.0280406.t003
DOI:
10.1371/journal.pone.0280406.t004
DOI:
10.1371/journal.pone.0280406.s001
DOI:
10.1371/journal.pone.0280406.s002
DOI:
10.1371/journal.pone.0280406.s003
DOI:
10.1371/journal.pone.0280406.r001
DOI:
10.1371/journal.pone.0280406.r002
DOI:
10.1371/journal.pone.0280406.r003
DOI:
10.1371/journal.pone.0280406.r004
Language:
English
Publisher:
Public Library of Science (PLoS)
Publication Date:
2023
detail.hit.zdb_id:
2267670-3
Bookmarklink