Journal: Annals of Biomedical Engineering

Article Title: A Study of Gene Expression, Structure, and Contractility of iPSC-Derived Cardiac Myocytes from a Family with Heart Disease due to LMNA Mutation

doi: 10.1007/s10439-021-02850-8

Figure Lengend Snippet: iPSC-derived cardiomyocytes. ( a ) Confocal images of cardiac tissues for one of the individuals. Stained for actin fibrils (phalloidin: green), nuclei (DAPI: blue), and sarcomeric z -lines ( α -actinin: red). Scale bar 20 μ m. ( b ) Single cell RNA sequencing of PA1 and CA1 cell lines represented with a heat map for the top 90 genes shows a varied expression profile for the two cells lines. ( c ) Feature maps for the two cells lines show mostly matching clusters for both cell lines, and the two cardiac-specific markers indicate that 99% of the cells are considered cardiomyocytes.

Article Snippet: Usually, on the 16th day of differentiation, purified cultures were dissociated with STEMdiffTM Cardiomyocyte Dissociation Kit (StemCell Technologies, Cat# 05025).

Techniques: Derivative Assay, Staining, RNA Sequencing, Expressing

Journal: Annals of Biomedical Engineering

Article Title: A Study of Gene Expression, Structure, and Contractility of iPSC-Derived Cardiac Myocytes from a Family with Heart Disease due to LMNA Mutation

doi: 10.1007/s10439-021-02850-8

Figure Lengend Snippet: Nuclei analysis. ( a ) Nuclei related gene expressions from single cell RNA-sequencing of two cell lines—PA1 (cell number = 13,028) and CA1 (cell number = 12,591). ( b ) Example images of normal and defective nuclei, stained for DAPI, for a patient and control cell line. Scale bar 5 μ m. ( c ) Quantitative analysis of nuclear morphology for all five cell lines (error bars are standard deviations; black horizontal lines indicate significance of p < 0.05). Unless otherwise indicated sample sizes ( n ) is the number of nuclei analyzed for each condition: (i) percent dysmorphic nuclei grouped by mutation status—controls (CTRL) and patients (PT) (sample sizes, n based on the number of coverslips). (ii) Mean negative curvature, (iii) relative concavity, (iv) maximum negative curvature, and (v) area for all controls and for each individual patient line. (vi) Comparison of dysmorphic nuclei percentage for both cardiomyocytes and fibroblasts in all individual cell lines (sample sizes, n based on the number of coverslips). (vii) The increase in dysmorphic nuclei percentage from fibroblasts to cardiomyocytes for each individual [errors were propagated from part (vi) and smaller sample size was used for each comparison].

Article Snippet: Usually, on the 16th day of differentiation, purified cultures were dissociated with STEMdiffTM Cardiomyocyte Dissociation Kit (StemCell Technologies, Cat# 05025).

Techniques: RNA Sequencing, Staining, Control, Mutagenesis, Comparison

Journal: Annals of Biomedical Engineering

Article Title: A Study of Gene Expression, Structure, and Contractility of iPSC-Derived Cardiac Myocytes from a Family with Heart Disease due to LMNA Mutation

doi: 10.1007/s10439-021-02850-8

Figure Lengend Snippet: Cardiomyocyte cytoskeleton. ( a ) Expression levels for cytoskeleton related genes that were differentially expressed in CA1 (cell number = 12,591) and PA1 (cell number = 13,028) lines. ( b ) Example analysis of actin orientation (left image, green arrows) and sarcomere identification [right image sarcomeres identified with purple (longer) and blue (shorter)]. ( c ) The density of sarcomeres averaged for three control lines (CTRL) and two patient lines (PT). ( d ) Quantification of myofibril architecture (error bars are standard deviations; black horizontal lines indicate significance of p < 0.05): (i) average sarcomere length for each individual cell line; orientational order parameter (OOP) for the (ii) sarcomeres and (iii) actin. Sample sizes ( n ) are based on the number of coverslips analyzed for each condition.

Article Snippet: Usually, on the 16th day of differentiation, purified cultures were dissociated with STEMdiffTM Cardiomyocyte Dissociation Kit (StemCell Technologies, Cat# 05025).

Techniques: Expressing, Control

Journal: International Journal of Molecular Sciences

Article Title: A Possible Role of Tetrodotoxin-Sensitive Na + Channels for Oxidation-Induced Late Na + Currents in Cardiomyocytes

doi: 10.3390/ijms25126596

Figure Lengend Snippet: Distinct effects on Na + currents in mouse cardiomyocytes and hiPSC-CMs: ( A , B ). Activation of Na + currents in cardiomyocytes ( A ) or hiPSC-CMs ( B ) at 0.1 Hz and a holding potential of −120 mV. While peak current amplitudes declined, non-inactivating Na + currents emerged upon application of 500 µM ChT. ( C , D ). Development of Na + current peak amplitudes following application of control solution or 500 µM ChT on mouse cardiomyocytes ( C ) hiPSC-CMs ( D ). Current amplitudes were normalized to the amplitude of the first current evoked prior application of ChT. ( E , F ). Current densities of late Na + currents following application of control solution or 500 µM ChT on cardiomyocytes ( E ) or hiPSC-CMs ( F ). The amplitudes of late currents were determined between 40 and 60 ms.

Article Snippet: For patch clamp experiment, hiPSC-CMs aggregates after ~14 to 18 days of differentiation were dissociated with the STEMdiff Cardiomyocyte Dissociation Kit (StemCell Technologies, Cologne, Germany) for 3 min.

Techniques: Activation Assay, Control

Journal: International Journal of Molecular Sciences

Article Title: A Possible Role of Tetrodotoxin-Sensitive Na + Channels for Oxidation-Induced Late Na + Currents in Cardiomyocytes

doi: 10.3390/ijms25126596

Figure Lengend Snippet: Inhibition of ChT-induced late Na + currents by TTX: ( A , B ). Representative current traces displaying activation of Na + currents in mouse cardiomyocytes before ( A ) and after application of 500 µM ChT ( B ). Application of 300 nM TTX (red traces) resulted in partially reduced amplitudes of peak currents as well as late Na + currents. ( C ). Bar graphs displaying inhibition of peak currents as well as late Na + currents by TTX before and after application of ChT. ( D , E ). Representative current traces displaying activation of Na + currents in hiPSC-CMs before ( A ) and after application of 500 µM ChT ( B ). Application of 300 nM TTX (red traces) resulted in partially reduced amplitudes of peak currents as well as late Na + currents. ( F ). Bar graphs displaying inhibition of peak currents as well as late Na + currents by TTX before and after application of ChT. Data are presented as mean ± S.E.M. * = p < 0.05. n.s. = not significant.

Article Snippet: For patch clamp experiment, hiPSC-CMs aggregates after ~14 to 18 days of differentiation were dissociated with the STEMdiff Cardiomyocyte Dissociation Kit (StemCell Technologies, Cologne, Germany) for 3 min.

Techniques: Inhibition, Activation Assay

Journal: International Journal of Molecular Sciences

Article Title: A Possible Role of Tetrodotoxin-Sensitive Na + Channels for Oxidation-Induced Late Na + Currents in Cardiomyocytes

doi: 10.3390/ijms25126596

Figure Lengend Snippet: UVA-light induced effects on Nav1.5 and Nav1.3: ( A – D ). Current traces displaying activation of Na + currents in cardiomyocytes ( A ) or HEK 293 cells with Nav1.5 ( B ), Nav1.3 ( C ), or Nav1.2 ( D ). Non-inactivating Na + currents emerged upon application of UVA-light. The first and last current traces are marked as bold black lines. ( E ). Development of Na + current peak amplitudes following application of UVA-light. ( F ). Development of late Na + currents following application of UVA-light. In ( D , E ), current amplitudes were normalized to the amplitude of the first current evoked prior application of UVA-light. ( G , I ). Current–voltage dependencies of Nav1.5 ( G ) or Nav1.3 ( I ) in control solution and after application of UVA-light. ( H , J ). Steady-state fast inactivation of Nav1.5 ( H ) and Nav1.3 ( J ) in control solution or after treatment with UVA-light. Normalized amplitudes were plotted against the membrane potential and data were fitted with the Boltzmann equation. All data are presented as mean ± S.E.M.

Article Snippet: For patch clamp experiment, hiPSC-CMs aggregates after ~14 to 18 days of differentiation were dissociated with the STEMdiff Cardiomyocyte Dissociation Kit (StemCell Technologies, Cologne, Germany) for 3 min.

Techniques: Activation Assay, Control, Membrane

Journal: International Journal of Molecular Sciences

Article Title: A Possible Role of Tetrodotoxin-Sensitive Na + Channels for Oxidation-Induced Late Na + Currents in Cardiomyocytes

doi: 10.3390/ijms25126596

Figure Lengend Snippet: Different effects of AS on Nav1.5 and Nav1.3: ( A , B , G , H ). Voltage-dependent activation of Nav1.5 ( A , B ) or Nav1.3 ( G , H ) in control solution or after application of 500 µM AS. ( C , I ) Bar graphs displaying the average inhibition of Nav1.5 ( C ) and potentiation of Nav1.3 induced by 500 µM AS ( I ) in control solution and after application of AS. ( D , J ) Normalized current-voltage dependencies of Nav1.5 ( D ) and Nav1.3 ( J ) in control solution and after application of AS. ( E , K ) Steady-state fast inactivation of Nav1.5 ( E ) and Nav1.3 ( K ) in control solution or after treatment with 500 µM AS. Normalized amplitudes were plotted against the membrane potential and data were fitted with the Boltzmann equation. ( F , L ) Recovery from fast inactivation of Nav1.5 ( F ) and Nav1.3 ( L ) in control solution and after treatment with 500 µM AS. Normalized amplitudes were plotted against the corresponding interval duration. The lines are best fits of the normalized data calculated with an exponential potential. All data are presented as mean ± S.E.M. ( M , N ) Current traces displaying voltage-dependent activation Na + currents in cardiomyocytes before ( M ) and after application of 500 µM AS ( N ). ( O ) Current-voltage dependencies of Na + currents in cardiomyocytes examined in control solution and after treatment with AS.

Article Snippet: For patch clamp experiment, hiPSC-CMs aggregates after ~14 to 18 days of differentiation were dissociated with the STEMdiff Cardiomyocyte Dissociation Kit (StemCell Technologies, Cologne, Germany) for 3 min.

Techniques: Activation Assay, Control, Inhibition, Membrane