Figure 3. Lineage tracing models show that transient Pkm2 expression increase CM cell division and suppress postnatal CM cell-cycle arrest.

A. Experimental timeline used for cardiac lineage tracing in R26^mTmG mice B. Representative images of transfected CMs and their progeny (GFP⁺, α-Actinin⁺) 28 days post MI. C. Quantification of GFP⁺ CMs 3 or 28 days post MI (n=3). D. Transfection efficiency (%GFP⁺) of CMs or non-CMs 28 days post MI (n=3). E. Representative images of GFP⁺ CMs, pH3⁺ or Ki67⁺ 28 days post MI. F&G. Quantification of GFP⁺ pH3⁺ (F. n=3) or GFP+ Ki67+ CMs (G. n=3) 28 days post transfection with _CMSLuc or _CMSPkm2 with _CMSCre modRNA in MI model. H-J. Ratio of heart to body weight (H. n=3), relative cross-sectional area of GFP⁺ CMs (I. n=3) and number of nuclei in GFP⁺ CMs (J. n=3) in hearts 28 days post MI and delivery of _CMSPkm2 in cardiac lineage tracing model using R26^mTmG mice. K. Experimental timeline to trace proliferating CMs using MADM mice L. Representative images of single-color- (Green (eGFP⁺/DsRed⁻), Red (eGFP⁻/DsRed⁺)) or double-color- (Yellow (eGFP⁺/DsRed⁺) labeled CMs 14 days post MI and injection of _CMSLuc or cmsPkm2 with _CMSCre modRNA in a MADM MI mouse model. M. Quantification of single-color CMs (Red or Green) amongst total labeled CMs 14 days post MI (n=5). N. Distribution of single-color CMs in the heart 14 days post MI (n=3, infarct, border or remote area). O. Representative images of isolated CMs from MADM mice 14 days post MI and modRNA injection. P. Quantification of isolated single-color CMs 14 days post MI (n=3). Q&R. Representative image (Q) and quantification (R, n=3, mono, bi or multi) of single-color CM nucleation 14 days post MI. Unpaired two-tailed t-test or Two-way ANOVA to analyze the data in C&D. ****, P<0.0001, ***, P<0.001, **, P<0.01, *, P<0.05, N.S, Not Significant. Scale bar = 50μm (B&L) or 5μm to E&O.