| Erythropoietin (EPO), well known for its role in stimulation oferythropoiesis, has recently been shown to have a dramatic protective effect inanimal models of myocardial ischemia/reperfusion (I/R) injury mainly byreducing apoptosis, other studies show that EPO and subsequent EPO-Rsignaling provides cellular protection by inhibiting apoptosis involving specificprotein kinase cascades, including the stress-responsive Janus-associatedkinase-2 (Jak2), PI3K/Akt, and ras-MAP kinase pathways. However, it has notbeen elucidated entirely which transcription factor plays an important role in theEPO's cardioprotection.Transcription factors such as GATA-4, GATA-6, NKX2.5, MEF2, FOG-2are involved in modulating cardiomyocyte development and apoptosis. GATA-4is a member of the GATA family of zinc finger transcription factors, whichplays important roles in transducing nuclear events that modulate cell lineagedifferentiation during development and hypertrophy of adult cardiac myocytes.GATA-4 is zinc finger transcription factor that is expressed in the developingheart and GATA-4 continues expression in the adult cardiac myocytes. Besidesregulating cell growth and differentiation, GATA factors also play importantroles in controlling cell survival by modulating apoptosis. The present study was to investigate the anti-apoptotic properties of EPO administered at the timeof reperfusion after myocardial infarct and whether transcription factor GATA-4is involved. In order to investigate whether EPO protects cardiomyocyte fromI/R injury via GATA-4, we replicated in vivo and in vitro models of I/R, A/R toobserve the influence of blocking GATA-4 by transfection of dnGATA4 on EPOeffect.In experiments in vivo, male mice were treated with EPO at the time ofreperfusion, following 45 min of left anterior descending coronary arteryligation and hearts were harvested 4 hours later. EPO not only reducedmyocardial IA/RA, IA/LV by 43.40%, 47.77% respectively, decreased 43.14%of apoptosis and 68.15% of necrosis in the myocardial area at risk, but inhibitedcardiac arrhythmia, improved cardiac function expressed as the left ventricularejection fraction (EF)increased by 12.8% and fractional shortening (ES)increased by 25.10%, compared with I/R. In order to determine whetherGATA-4 is involved in the myocyte protection, further study was done. Ourresults indicate phosphorylated GATA-4, total GATA-4 were increased by41.04% 29.50% respectively, GATA-4 mRNA was increased by 110.27%compared with I/R (p＜0.05). In addition, Akt phosphorylation and bcl-2 werealso increased by 31.82% and 178.7% respectively. Our in vivo results indicatethat EPO remarkbly reduced infarct size, apoptosis and necrosis, improvedcardiac function and increased phosphorylated-Akt and GATA-4, total GATA-4and bcl-2 expression (p＜0.05). The results suggest transcription factor GATA-4 is closely associated with myocyte protection of EPO, accompanied with Aktactivation and bcl-2 increasement.To eliminate the influence of other factors, A/R model was used. EPO(2u/1×10~6 cells) was administered at the time of reoxygenation after anoxia.EPO inhibited A/R-induced cardiomyocyte apoptosis measured by Tunel andflow cytometry. Our results by Western blot indicate the protein expressions ofGATA-4 phosphorylation and bcl-2 were increased by 151.71%, 119.40%respectively compared with anoxia/reoxygenation. In order to determineGATA-4 played an important role in the myocyte protection of EPO, dnGATA4was transfected into cultured neonatal Sprague-Dawley rat myocytes, EPOprotection against I/R-induced cardiomyocyte apoptosis and the influence ofEPO on GATA-4 were abolished.In summary, our results invo and invitro demonstrate EPO, administeredat the time of reperfusion after ischemia, significantly protects cardiomyocytesfrom I/R or A/R injury, GATA-4 plays an important role in the anti-apoptosis ofEPO and the cardioprotection of EPO may effect by an Akt-GATA-4-bcl-2dependent pathway. |
PDF Full Text Request