Figure S1 . Scale bar = 800 μm. (B) Representative spontaneous CaT traces indicate the functional effects of Cx43 gene therapy over 10-second recordings. (C) Aligned single beat traces indicate the increase of calcium rise kinetics induced by Cx43 gene therapy. (D) Bipolar patient monolayers harboring the CACNA1C SNP conduction velocity could be increased using Cx43 gene therapy. Conduction velocity increased significantly in a gene therapy dose-dependent manner. One-way analysis of variance, multiple comparisons to 0 moi data, ∗∗∗indicates significant difference from control, 0 moi data, p = .0004; ∗∗∗∗ p < .0001. (E) Cx43 gene therapy increased the rate of calcium rise/release in a dose-dependent manner. One-way analysis of variance, multiple comparisons to 0 moi data, ∗∗∗∗ p < .0001. (F) Western blot analysis for CaV1.2 protein expression indicated low baseline expression in the CACNA1C SNP cells, and this expression was restored with Cx43 gene therapy. (G) Quantification of Cav1.2 protein expression as a function of Cx43 gene therapy dose indicates significant elevation of L-type calcium channel expression. (H) Cx43 gene therapy reduced the calcium triangulation index significantly, unpaired t test values indicated; p < .05 indicates significant difference, n = 7 to 9 monolayers. (I) Cx43 gene therapy reduced the effect of thioridazine (1 μM) to increase CaTD 80, an in vitro surrogate marker for the QT interval of the electrocardiogram. ∗unpaired t test within each group, p < .05, n = 7 to 8 monolayers. CaTD, calcium transient duration; moi, multiplicity of infection; RFP, red fluorescent protein; SNP, single nucleotide polymorphism. " width="100%" height="100%">
Journal: Biological Psychiatry Global Open Science
Article Title: Bipolar Patient–Specific In Vitro Diagnostic Test Reveals Underlying Cardiac Arrhythmia Phenotype Caused by Calcium Channel Genetic Risk Factor
doi: 10.1016/j.bpsgos.2024.100296
Figure Lengend Snippet: Cx43 gene therapy increases cardiac conduction velocity in bipolar patient cardiomyocyte monolayers expressing the CACNA1C SNP. (A) Cx43 gene therapy effectiveness was monitored via live-cell time-lapse imaging using RFP as a marker. The number of RFP-positive cells increased over time and was used as a surrogate marker for Cx43 protein expression. Gene therapy was tested using multiple concentrations of virus ranging from 0 to 10 moi as indicated in Figure S1 . Scale bar = 800 μm. (B) Representative spontaneous CaT traces indicate the functional effects of Cx43 gene therapy over 10-second recordings. (C) Aligned single beat traces indicate the increase of calcium rise kinetics induced by Cx43 gene therapy. (D) Bipolar patient monolayers harboring the CACNA1C SNP conduction velocity could be increased using Cx43 gene therapy. Conduction velocity increased significantly in a gene therapy dose-dependent manner. One-way analysis of variance, multiple comparisons to 0 moi data, ∗∗∗indicates significant difference from control, 0 moi data, p = .0004; ∗∗∗∗ p < .0001. (E) Cx43 gene therapy increased the rate of calcium rise/release in a dose-dependent manner. One-way analysis of variance, multiple comparisons to 0 moi data, ∗∗∗∗ p < .0001. (F) Western blot analysis for CaV1.2 protein expression indicated low baseline expression in the CACNA1C SNP cells, and this expression was restored with Cx43 gene therapy. (G) Quantification of Cav1.2 protein expression as a function of Cx43 gene therapy dose indicates significant elevation of L-type calcium channel expression. (H) Cx43 gene therapy reduced the calcium triangulation index significantly, unpaired t test values indicated; p < .05 indicates significant difference, n = 7 to 9 monolayers. (I) Cx43 gene therapy reduced the effect of thioridazine (1 μM) to increase CaTD 80, an in vitro surrogate marker for the QT interval of the electrocardiogram. ∗unpaired t test within each group, p < .05, n = 7 to 8 monolayers. CaTD, calcium transient duration; moi, multiplicity of infection; RFP, red fluorescent protein; SNP, single nucleotide polymorphism.
Article Snippet: Primary antibodies were used to detect specific expression of connexin 43 (Cx43) (anti-Cx43 antibody, rabbit polyclonal, Alomone Labs) and the L-type calcium channel (CaV1.2, rabbit polyclonal, Alomone Labs).
Techniques: Expressing, Imaging, Marker, Virus, Functional Assay, Control, Western Blot, In Vitro, Infection
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Figure S1 . Scale bar = 800 μm. (B) Representative spontaneous CaT traces indicate the functional effects of Cx43 gene therapy over 10-second recordings. (C) Aligned single beat traces indicate the increase of calcium rise kinetics induced by Cx43 gene therapy. (D) Bipolar patient monolayers harboring the CACNA1C SNP conduction velocity could be increased using Cx43 gene therapy. Conduction velocity increased significantly in a gene therapy dose-dependent manner. One-way analysis of variance, multiple comparisons to 0 moi data, ∗∗∗indicates significant difference from control, 0 moi data, p = .0004; ∗∗∗∗ p < .0001. (E) Cx43 gene therapy increased the rate of calcium rise/release in a dose-dependent manner. One-way analysis of variance, multiple comparisons to 0 moi data, ∗∗∗∗ p < .0001. (F) Western blot analysis for