Thymosin beta4 is an essential paracrine factor of embryonic endothelial progenitor cell-mediated cardioprotection.
BACKGROUND
Prolonged myocardial ischemia results in cardiomyocyte loss despite successful revascularization. We have reported that retrograde application of embryonic endothelial progenitor cells (eEPCs) provides rapid paracrine protection against ischemia-reperfusion injury. Here, we investigated the role of thymosin beta4 (Tbeta4) as a mediator of eEPC-mediated cardioprotection.


RESULTS
In vitro, neonatal rat cardiomyocytes were subjected to hypoxia-reoxygenation in the absence or presence of eEPCs with or without Tbeta4 short hairpin RNA (shRNA) transfection. In vivo, pigs (n=9 per group) underwent percutaneous left anterior descending artery occlusion for 60 minutes on day 1. After 55 minutes of ischemia, control eEPCs (5x10(6) cells) or cells transfected with Tbeta4 shRNA when indicated or 15 mg Tbeta4 alone were retroinfused into the anterior interventricular vein. Segmental endocardial shortening in the infarct zone at 150-bpm atrial pacing, infarct size (triphenyl tetrazolium chloride viability and methylene blue exclusion), and inflammatory cell influx (myeloperoxidase activity) were determined 24 hours later. Survival of neonatal rat cardiomyocytes increased from 32+/-4% to 90+/-2% after eEPC application, an effect sensitive to shRNA transfection compared with Tbeta4 (45+/-7%). In vivo, infarct size decreased with eEPC application (38+/-4% versus 54+/-4% of area at risk; P<0.01), an effect abolished by Tbeta4 shRNA (62+/-3%). Segmental subendocardial shortening improved after eEPC treatment (22+/-3% versus -3+/-4% of control area) unless Tbeta4 shRNA was transfected (-6+/-4%). Retroinfusion of Tbeta4 mimicked eEPC application (infarct size, 37+/-3%; segmental endocardial shortening, 34+/-7%). Myeloperoxidase activity (3323+/-388 U/mg in controls) was decreased by eEPCs (1996+/-546 U/mg) or Tbeta4 alone (1455+/-197 U/mg) but not Tbeta4 shRNA-treated eEPCs (5449+/-829 U/mg).


CONCLUSIONS
Our findings show that short-term cardioprotection derived by regional application of eEPCs can be attributed, at least in part, to Tbeta4.
Hinkel
Rabea
R
Medizinische Klinik I, University Clinic Grosshadern, Munich, Germany.
El-Aouni
Chiraz
C
Olson
Tonia
T
Horstkotte
Jan
J
Mayer
Stefan
S
Müller
Sebastian
S
Willhauck
Michael
M
Spitzweg
Christine
C
Gildehaus
Franz-Josef
FJ
Münzing
Wolfgang
W
Hannappel
Ewald
E
Bock-Marquette
Ildiko
I
DiMaio
J Michael
JM
Hatzopoulos
Antonis K
AK
Boekstegers
Peter
P
Kupatt
Christian
C
D000818:Animals; D002448:Cell Adhesion / Q000276:immunology; D002470:Cell Survival / Q000502:physiology; D002478:Cells, Cultured; D053595:Embryonic Stem Cells / Q000637:transplantation; D042783:Endothelial Cells / Q000166:cytology / Q000502:physiology; D015316:Genetic Therapy / Q000379:methods; D007962:Leukocytes / Q000166:cytology; D051379:Mice; D008822:Mice, Transgenic; D017202:Myocardial Ischemia / Q000378:metabolism / Q000503:physiopathology / Q000628:therapy; D015428:Myocardial Reperfusion Injury / Q000378:metabolism / Q000503:physiopathology* / Q000628:therapy*; D032383:Myocytes, Cardiac / Q000166:cytology* / Q000502:physiology*; D019899:Paracrine Communication / Q000502:physiology; D051381:Rats; D033581:Stem Cell Transplantation; D013552:Swine; D013947:Thymosin / Q000235:genetics* / Q000378:metabolism; D014162:Transfection; D000099201:Thymosin beta(4)
D013947:Thymosin; D000099201:Thymosin beta(4)