The identification and modulation of proliferating cells is critical for stem cell research and regenerative medicine. However, current approaches to quantify cell proliferation are imprecise in particular in tissue types undergoing variations of the cell cycle, namely acytokinetic mitosis and endoreduplication. The only definitive proof for cell division in cardiac muscle is the observation of a contractile ring or a midbody prior to abscission. To visualize these cell cycle specific events we developed a new in vivo proliferation marker that indicates M-phase in a high spatial and temporal resolution. This was achieved by fusing eGFP to the scaffolding protein anillin. Transgenic murine embryonic stem cell (mESC) lines were generated that stably expressed the eGFP-anillin fusion protein under control of the ubiquitous chicken b-actin promoter with CMV enhancer (CAG). The fusion protein allowed the accurate and easy determination of proliferating cell populations in undifferentiated and differentiated mESCs as well as monitoring of sub-phases of the M-phase. Transduction with a CAG-eGFP-anillin lentivirus enabled us to easily measure cell-cycle kinetics of human induced pluripotent stem cells (hiPS) and human ESCs via time lapse.

The CAG-eGFP-anillin transgenic mouse line allowed live tracking of dividing cardiomyocytes in the embryonic heart and determination of cell cycle kinetics. During development of the forebrain eGFP-anillin specifically marked proliferating progenitor cells of the neuroepithelium. EGFP-anillin mice also enabled direct analysis of cell-cycle activity in the adult heart post-injury, indicating endoreduplication but no division of cardiomyocytes.

This approach will enable us to address open questions concerning the proliferation of heart muscle as well as other tissue types during development and upon injury.