CB₂Receptor Agonist Promote Cardiac Stem/Progenitor Cells Endogenous Regeneration In A Mouse Model Of Myocardial Infarction

BACKGROUNDCoronary heart disease become extremely common and seriously harm to people’shealth in China. Traditional treatment for coronary artery disease includes calciumblockers, β2recepter blockers，thrombolysis, percutaneous coronary intervention, hearttransplantation and so on.Though medication therapy and revascularization couldimprove the symptoms of coronary heart disease patients to some extent, end-stageheart failure caused by myocardial necrosis is still a serious problem. The prevalenceof heart transplantation becomes difficult due to fewer sources of donor heart andexpensive surgery. In past decade, stem cell therapy for coronary heart diseasebecomes a hot topic. Although a large number of basic and clinical researches onexogenous stem cell transplantation made huge progress, complex in vitroamplification procedures and low cell survival rate of recipients after transplantationcould not be avoided. Recent study showed the presence of cardiac stem/progenitorcells which is capable of supporting the endogenous cardiac regeneration. But numbers of cardiac stem/progenitor cells are small and their activation and proliferation areaffected by many factors. Primary cannabinoid receptors include receptor typeⅠ andtype Ⅱ.Cannabinoid receptor Ⅱ agonists have been proven not only to beanti-inflammatory, anti-oxidative stress and to Anti-apoptotic, but also to promote theproliferation of heart cells. Moreover, cannabinoid receptor Ⅱagonists have beenreported to mediate hematopoietic stem/progenitor cell mobilization and migration, aswell as to promote neural stem/progenitor cell proliferation in vitro and in vivo.Therefore, we hypothesized that cannabinoid receptor Ⅱ agonists can activate cardiacstem/progenitor cells and promote their proliferation, which plays an important rolein endogenous cardiac regeneration.ObjectiveThis study aimed to investigate whether the CB2receptor activation-inducedcardiac stem/progenitor cell proliferation could promote endogenous cardiacregeneration and to explore the underlying mechanism.Methods and ResultsC57BL/6male mice (8-week-old, weighing20to25g) were randomly allocatedinto four groups:(1) Sham group (Sham),(2) MI+PBS group (MI),(3) MI+CB2agonist AM1241group(MI+AM),(4) MI+CB2agonist AM1241+Wortmannin group(MI+AM+W). Mice myocardial infarction (MI) was induced by ligation of the leftanterior descending artery. Echocardiography was performed to evaluate leftventricular ejection fraction (LVEF). Masson’s trichrome staining was performed toobserved myocardial fibrosis level4weeks after myocardial infarction. The number ofcardiac stem/progenitor cells were observed with immunostaining for cardiac stem/progenitor cells surface marker c-kit and Sca-1in peri-infarction rigion at day7postMI. Real-time PCR analysis was applied to examine the expression of c-kit, Sca-1andMDR1of border zone of infracted myocardium at day7and14post MI. MDA, TNF-αand IL-6levels in plasma was measured by ELISA. Moreover, expression of p-Akt, HO-1and Nrf-2protein were detected by Western blot. Echocardiography revealedthat AM1241ameliorated LVEF compared with MI Group at day7(43.42%±3.737%vs20.39%±4.961%, p<0.01) and day14(49.17%±3.567%vs22.15%±6.913%, p<0.01)post MI. Also, AM1241ameliorated myocardial fibrosis level compared with MIGroup at week4（28.9%±1.14%vs.39.29%±1.58%, p<0.05）.Moreover, bothimmunofluorescence staining and RT-PCR demonstrated that administration ofAM1241significantly increased expression of c-kit and Sca-1in border zone ofinfracted myocardium (p<0.05). In addition, MDA, TNF-α and IL-6levels in plasmacan be reduced by AM1241administration after myocardial infarction（p<0.05）.Western blot suggested that AM1241could significantly increase cytoplasmicexpression of p-Akt and HO-1(p <0.05),along with the increasing nucleus expressionof Nrf-2（p<0.05）.Furthermore, the protective effect of AM1241on infarctedmyocardium was inhibited by PI3K inhibitor wortmannin.ConclusionsCB2receptor agonists can improve cardiac function and myocardial remodeling aswell as promote endogenous myocardial regeneration after myocardial in-farctionthrough modulating PI3K/Akt/Nrf-2signal pathway.