Plk1 & Myocardial Regeneration Figure 5. Over expression of Plk1 has no effect on adult cardiac myocyte binucleation. Ventricular cardiac myocytes were isolated and infected with EGFP, WT or T210D adenovirus and incubated for 48 hours (P3 and P7) or 24 hours (adult) in serum free media or media supplemented with 5% foetal calf serum (FCS). Cells were fixed and assessed for binucleation. A) Cardiac myocyte binucleation increases with age. Over expression of Plk1 had no effect on binucleation at P3 (B), P7 (C) or adult (D) stages of development. E) Plk1 over expression does not increase cardiac myocyte cell number, as determined by the number of troponin I positive cardiac myocytes per image (cells were counted for three separate images for EGFP, WT and T210D-infected cells from three separate experiments). doi:10.1371/journal.pone.0006752.g005 required for S-phase progression and the associated centriole duplication process [52,56–59] and this most likely explains the presence of these two proteins in the adult cardiac myocyte. Plk3 was originally identified as an immediate-early response gene that was upregulated in NIH3T3 cells in response to FGF.[75] It has been reported that Plk3 has a role in G2 checkpoint signaling [76,77] and is activated following treatment of cells with 100 mM doxorubicin or H2O2.[54,78] We show here that at the protein level, the expression of Plk3 does not change during rat cardiac myocyte development, with levels persisting to PLoS ONE | www.plosone.org 8 August 2009 | Volume 4 | Issue 8 | e6752