0202 : Adult cardiomyocytes proliferation blockage by nuclear ephrin-B1: involvement of the PI3Kγ pathway

The possibility to reboot the proliferation of adult resident cardiomyocytes (CMs) has emerged as a promising avenue in cardiac regenerative medicine. For that purpose, there is an urgent need for identifying the molecular mechanisms involved in the natural blockage of adult CMs proliferation. Recently, we recently demonstrated that ephrin-B1 expression in the CMs nuclei acts as a specific inhibitor of the adult CMs proliferation (unpublished data). Here, we have explored the specific role of the PI3Kγin the nuclearization of ephrin-B1. Fluorescent immunostaining showed a specific loss of nuclear ephrin-B1 in p110-γKO but not in KI mice, suggesting a specific role of the PKAanchoring function of the p110-γcatalytic subunit of PI3Kγin ephrin-B1 nuclearization. Interestingly, p110-γKO mice demonstrated a significant increase of mononucleated CMs (19,1 ± 1,1% vs 13,3 ± 1,0% in WT) correlated to a decrease in binucleated CMs (70,9 ± 1,9% vs 79,7 ±1,0% in WT) possibly indicative of a proliferative potential. In agreement with a proliferative potential, we measured a higher level of replicative-BrdU± CMs in KO PI3Kγmice (WT: 6,23±2,2% ; KO: 19,3±4,9%; KI: 3,0±0,8%), but also a higher CMs density in cardiac tissue from 8 month old mice (WT: 2633±79,4cm/mm 2 ; KO: 3208±155,6CM/mm 2 ; KI: 2659,7±99,3CM/mm 2 ) mosy likely reflecting CMs hyperplasia. To confirm the involvement of a cAMP/PKA pathway in ephrin-B1 nuclearization, we studied a neonatal cardiomyoblast cell line (H9C2) naturally expressing a nuclear ephrin-B1 pool. We showed that H89 treatment prevented ephrin-B1 nuclearization while forskolin promoted nuclearization of a recombinant ephrin-B1-GFP. These results demonstrated for the first time the specific role of the cAMP/ PKA/PI3Kγ pathway in ephrin-B1 nuclearization in the CM. In the future, the accurate characterization of this pathway could allow identifying new and more specific targets for heart regenerative medicine strategies.