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A non-invasive platform for functional characterization of stem-cell-derived cardiomyocytes with applications in cardiotoxicity testing.

Maddah M, Heidmann JD, Mandegar MA, Walker CD, Bolouki S, Conklin BR, Loewke KE - Stem Cell Reports (2015)

Bottom Line: We present a non-invasive method to characterize the function of pluripotent stem-cell-derived cardiomyocytes based on video microscopy and image analysis.The platform, called Pulse, generates automated measurements of beating frequency, beat duration, amplitude, and beat-to-beat variation based on motion analysis of phase-contrast images captured at a fast frame rate.Using Pulse, we demonstrate recapitulation of drug effects in stem-cell-derived cardiomyocytes without the use of exogenous labels and show that our platform can be used for high-throughput cardiotoxicity drug screening and studying physiologically relevant phenotypes.

View Article: PubMed Central - PubMed

Affiliation: Cellogy, Inc., Palo Alto, CA 94301, USA. Electronic address: mmaddah@alum.mit.edu.

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Ca2+ Imaging Experiments on GCaMP Cells(A) Response of the beating signal extracted by Pulse to different doses of blebbistatin. The amplitude of the biomechanical beating signal decreases as the dose increases.(B) Comparison of the relative amplitude of the biomechanical beating signal before blebbistatin is applied, after 30 min of applying this compound, and after a media change to wash out blebbistatin. Each drug concentration mean value is based on technical replicates of four wells with four recordings per well.(C) No change is seen in the Ca+2 signal.(D) Beating signal extracted by Pulse and Ca2+ imaging with and without quinidine. Both methods capture an increase in the duration of the beats. See also Figure S1 for description of genomic modification of the GCaMP cell line used in these experiments.
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fig4: Ca2+ Imaging Experiments on GCaMP Cells(A) Response of the beating signal extracted by Pulse to different doses of blebbistatin. The amplitude of the biomechanical beating signal decreases as the dose increases.(B) Comparison of the relative amplitude of the biomechanical beating signal before blebbistatin is applied, after 30 min of applying this compound, and after a media change to wash out blebbistatin. Each drug concentration mean value is based on technical replicates of four wells with four recordings per well.(C) No change is seen in the Ca+2 signal.(D) Beating signal extracted by Pulse and Ca2+ imaging with and without quinidine. Both methods capture an increase in the duration of the beats. See also Figure S1 for description of genomic modification of the GCaMP cell line used in these experiments.

Mentions: Figure 4A shows the beating signal extracted by Pulse 30 min after addition of different concentrations of blebbistatin. As the concentration of blebbistatin is increased, the relative amplitude of the beating signal decreases, serving as a functional readout for the expected reduction in contractility. Figure 4B shows a comparison of the amplitude of the beating signal before blebbistatin is added, 30 min after compound application, and after a subsequent media change. Note that the biomechanical beating of the cardiomyocytes is restored back to normal levels after blebbistatin is washed away. For a concentration of 1 μM or above, no biomechanical beating is observed. In contrast, fluorescence imaging of GCaMP continues to generate a calcium transient signal even at 5 μM, as seen in Figure 5C. These data demonstrate that Pulse is able to capture and quantify the effect of a myosin II blocker in inhibiting the biomechanical contraction of the cardiomyocytes, an effect that MEA or Ca+2 imaging systems do not capture (Abassi et al., 2012).


A non-invasive platform for functional characterization of stem-cell-derived cardiomyocytes with applications in cardiotoxicity testing.

Maddah M, Heidmann JD, Mandegar MA, Walker CD, Bolouki S, Conklin BR, Loewke KE - Stem Cell Reports (2015)

Ca2+ Imaging Experiments on GCaMP Cells(A) Response of the beating signal extracted by Pulse to different doses of blebbistatin. The amplitude of the biomechanical beating signal decreases as the dose increases.(B) Comparison of the relative amplitude of the biomechanical beating signal before blebbistatin is applied, after 30 min of applying this compound, and after a media change to wash out blebbistatin. Each drug concentration mean value is based on technical replicates of four wells with four recordings per well.(C) No change is seen in the Ca+2 signal.(D) Beating signal extracted by Pulse and Ca2+ imaging with and without quinidine. Both methods capture an increase in the duration of the beats. See also Figure S1 for description of genomic modification of the GCaMP cell line used in these experiments.
© Copyright Policy - CC BY-NC-ND
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4400609&req=5

fig4: Ca2+ Imaging Experiments on GCaMP Cells(A) Response of the beating signal extracted by Pulse to different doses of blebbistatin. The amplitude of the biomechanical beating signal decreases as the dose increases.(B) Comparison of the relative amplitude of the biomechanical beating signal before blebbistatin is applied, after 30 min of applying this compound, and after a media change to wash out blebbistatin. Each drug concentration mean value is based on technical replicates of four wells with four recordings per well.(C) No change is seen in the Ca+2 signal.(D) Beating signal extracted by Pulse and Ca2+ imaging with and without quinidine. Both methods capture an increase in the duration of the beats. See also Figure S1 for description of genomic modification of the GCaMP cell line used in these experiments.
Mentions: Figure 4A shows the beating signal extracted by Pulse 30 min after addition of different concentrations of blebbistatin. As the concentration of blebbistatin is increased, the relative amplitude of the beating signal decreases, serving as a functional readout for the expected reduction in contractility. Figure 4B shows a comparison of the amplitude of the beating signal before blebbistatin is added, 30 min after compound application, and after a subsequent media change. Note that the biomechanical beating of the cardiomyocytes is restored back to normal levels after blebbistatin is washed away. For a concentration of 1 μM or above, no biomechanical beating is observed. In contrast, fluorescence imaging of GCaMP continues to generate a calcium transient signal even at 5 μM, as seen in Figure 5C. These data demonstrate that Pulse is able to capture and quantify the effect of a myosin II blocker in inhibiting the biomechanical contraction of the cardiomyocytes, an effect that MEA or Ca+2 imaging systems do not capture (Abassi et al., 2012).

Bottom Line: We present a non-invasive method to characterize the function of pluripotent stem-cell-derived cardiomyocytes based on video microscopy and image analysis.The platform, called Pulse, generates automated measurements of beating frequency, beat duration, amplitude, and beat-to-beat variation based on motion analysis of phase-contrast images captured at a fast frame rate.Using Pulse, we demonstrate recapitulation of drug effects in stem-cell-derived cardiomyocytes without the use of exogenous labels and show that our platform can be used for high-throughput cardiotoxicity drug screening and studying physiologically relevant phenotypes.

View Article: PubMed Central - PubMed

Affiliation: Cellogy, Inc., Palo Alto, CA 94301, USA. Electronic address: mmaddah@alum.mit.edu.

Show MeSH
Related in: MedlinePlus