Journal: iScience

Article Title: Phenotype specific nuclear lamina remodeling in hiPSC derived cardiomyocytes bearing TNNT2 sarcomeric variants

doi: 10.1016/j.isci.2025.113901

Figure Lengend Snippet: Cardiac differentiation schematic and maturation of hiPSC-CMs show striations and cardiac markers (A) Schematic of fibroblast collection and cardiac differentiation processes: From two distinct donors, fibroblasts were reprogrammed into hiPSC and validated by the presence of pluripotency markers. Knock-in of TNNT2 variants was performed, and cells were differentiated into hiPSC-CMs. Finally, the distinct cell lines were treated with hormonal treatment (T3 - triiodothyronine and DEXA - dexamethasone) for enhanced maturation before experiments. (B) Representative image of maturation in hiPSC-CM cell lines. In the top image (WT cell line), fluorescently labeled phalloidin for F-actin (red) displays frequently found striations of mature cardiomyocytes. In the bottom image: fluorescently labeled antibody for TNNT2 (purple), a marker for protein Troponin T encoded by this gene, which is commonly present in mature cardiomyocytes. Yellow arrows point to visible striation patterns. Scale bars = 10 μm. (C) Representative fluorescence image of hiPSC-CMs culture. In the top image (WT cell line): fluorescently labeled antibody for TNNT2 marks Troponin T (purple), fluorescently labeled phalloidin marks F-actin in the cytoskeleton (red), fluorescently labeled antibody for MLC2V marks ventricular specific myosin light chain 2 (green), and DAPI marks the nucleus (blue). In the bottom image (WT cell line), fluorescently labeled phalloidin marks F-actin in the cytoskeleton (red), fluorescently labeled antibody for MLC7 marks atrial specific myosin light chain 7 (green), and DAPI marks the nucleus (blue). Scale bars = 30 μm. (D) Graph representation of the relative percentual of hiPSC-CMs culture, which had positive ventricular marker – ventricular specific myosin light chain 2 ( MLC2V ), and positive atrial marker – myosin light chain 7 ( MLC7 ) fluorescence. Statistical analysis was conducted using a t test to evaluate differences in cardiac chamber markers, with results indicating a statistically significant difference ( p < 0.0001). Data are represented as mean ± SD with N = 6.

Article Snippet: TNNT2 Antibody , DSHB , Cat #RV-C2; RRID: AB_2240831.

Techniques: Knock-In, Labeling, Marker, Fluorescence

Journal: iScience

Article Title: Phenotype specific nuclear lamina remodeling in hiPSC derived cardiomyocytes bearing TNNT2 sarcomeric variants

doi: 10.1016/j.isci.2025.113901

Figure Lengend Snippet: RNA sequencing analysis shows regulated genes in each TNNT2 variant, and overlapping regulated genes in HCM and DCM (A) Heatmap of DEGs in I79N, R141W, and respective isogenic control. (B) Heatmap of DEGs in R92Q, R134G, and respective isogenic control. (C) Regulated pathways of the I79N variant associated with the HCM phenotype compared to respective isogenic control, upregulated pathways are shown in orange, and downregulated pathways in blue. (D) Regulated pathways of the R141W variant associated with the DCM phenotype compared to respective isogenic control, upregulated pathways are shown in orange, and downregulated pathways in blue. (E) Regulated pathways of the R92Q variant associated with the HCM phenotype compared to the respective isogenic control, upregulated pathways are shown in orange, and downregulated pathways in blue. (F) Regulated pathways of the R134G variant associated with the DCM phenotype compared to respective isogenic control, upregulated pathways are shown in orange, and downregulated pathways in blue. (G) Venn diagrams show overlapping DEGs in HCM variants R92Q (yellow) and I79N (green), with 78 up-regulated genes and 161 down-regulated genes in common. (H) Venn diagrams show overlapping DEGs in DCM variants R134G (red) and R141W (purple), with 43 up-regulated and 179 down-regulated similar genes.

Article Snippet: TNNT2 Antibody , DSHB , Cat #RV-C2; RRID: AB_2240831.

Techniques: RNA Sequencing, Variant Assay, Control