Fig. 1: CCNA2 expression induces cytokinesis in adult human cardiomyocytes in vitro.
From: Cyclin A2 induces cytokinesis in human adult cardiomyocytes and drives reprogramming in mice
A Clinical-use grade replication-deficient human adenovirus type 5 (Ad5) was used for this study. The vector was designed with cardiac Troponin T (cTnT) promoter to express human cyclin A2 (hCCNA2) specifically in cardiomyocytes. B The cTnT-hCCNA2 adenovirus was used to transduce cultured adult human cardiomyocytes to induce the expression of CCNA2. Significantly higher CCNA2 expression was observed in cells transduced with cTnT-hCCNA2 compared to the control virus. C–H From Movies S1 and S2: still images from representative time-lapse epifluorescence microscopy of cultured cardiomyocytes isolated from adult humans (C) 55-year-old and (F) 41-year-old. Please note that still images of 55 and 41-years-old have scale bars of 50 µm and 100 µm as it is shown in the Movies S1 and S2 respectively. Cells were transduced with cTnT-hCCNA2, cTnT-H2B-GFP, cTnT-eGFP (to label cardiomyocytes), and CMV-α-actinin-m-Cherry adenoviruses prior to the beginning of the time-lapse imaging. C At time 0 h, a cell (yellow) expressing both cTnT (green) and α-actinin (red) was followed for 70 h via time-lapse microscopy. The green channel was closed, and cells were only followed through the red channel to avoid UV photo-toxicity. The observed human cardiomyocytes show the 1st cell division at 50 h of imaging, and one of the daughter cells subsequently undergoes division at 70 h of imaging. D, G At 70/72 h, a daughter cell is partially magnified with a grayscale version to demonstrate the presence of an intact sarcomere structure. E, H 1 day after live imaging ended, these wells were fixed with subsequent labeling of nuclei with DAPI. The green fluorescence of the original cTnT-eGFP and cTnT-H2B-GFP transduction is visible, further confirming that these cells are cardiomyocytes. I The cytokinetic events were counted in the control and test samples from the 21-, 41-, and 55-year-old subjects. A significantly higher rate of cytokinesis was observed in the test samples from the 41-year-old (p = 0.001) and 55-year-old (p < 0.0001) compared to the control. On the other hand, the control and test samples from the 21-year-old did not exhibit a significant difference in cytokinetic events (p = 0.47). Data are represented as mean ± s.e.m.
