Abstract 1266: Rapid eEPC-Mediated Cardioprotection After Ischemia/Reperfusion: Role of Thymosinβ4
Prolonged myocardial ischemia results in myocardial dysfunction, even after successful revascularisation. We have reported that retrograde application of embryonic EPCs, derived from murine d 7.5 embryos (Tie-2+, c-Kit+, Sca-1+, flk-1 low, MHC-1-), provide rapid cardioprotection via soluble factors. Now, we investigated the role of Thymosin beta 4/10 (TB4, TB10) as mediators of the cardioprotective potential of eEPCs.
Methods: In vitro, neonatal rat cardiomyocytes (NRCMs) were subjected to hypoxia (4h)/reoxygenation (1h) in the absence or presence of eEPCs with or without TB siRNA transfection, which decreased of TB mRNA levels in RT-PCR. In vivo, pigs (n=6 per group) underwent percutaneous LAD occlusion for 60 min at day 1. After 55 min of ischemia either saline solution or eEPCs (5 * 106 DiI labeled cells) with or without TB4 siRNA were applied via selective pressure-regulated retroinfusion into the anterior interventricular vein. Infarct size (TTC-viability and Methylene-blue exclusion) was determined 24h later. Myeloperoxidase assay and TUNEL-staining were performed on tissue slices.
Results: In vitro, survival of NRCMs was increased from 32 ± 4 to 90 ± 2 % after eEPC application, an effect sensitive to siRNA transfection versus TB4 (45 ± 7%) and TB10(54 ± 7%). In vivo, numerous eEPCs were detected for 24 h in the ischemic area. Moreover, eEPCs significantly decreased the infarct size compared to control group (36±3% vs. 58±5% of AAR, p<0.05), whereas TB4-siRNA transfected eEPCs blunted this effect (48 ± 7% of AAR). Embryonic EPC application decreased TUNEL-positive cells compared to controls, an effect diminished by TB4 siRNA transfection of the eEPCs.
Conclusion: Our findings show that embryonic EPCs reduce ischemia-reperfusion injury early in the postischemic time course. In vitro assays and TB4 siRNA transfection of the eEPCs in vivo showed, that the cardioprotection mediated through the murine eEPCs is at least partially mediated by Thymosin beta 4 which is abundantly present in eEPCs.