Adenosine attenuates reperfusion-induced apoptotic cell death by modulating expression of Bcl-2 and Bax proteins.
This study tests the hypothesis that infarct reduction with adenosine (Ado) is associated with inhibition of apoptotic cell death by modulating expression of anti-apoptotic Bcl-2 and pro-apoptotic Bax proteins and reducing neutrophil accumulation. In three groups of dogs, the left anterior descending coronary artery was occluded for 60 min and reperfused for 6 h. Either saline (Control, n=8), Ado (140 microg/kg/min, n=8) or CGS21680, an adenosine A2A receptor analogue, (0.2 microg/kg/min, n=7) were infused during the first 2 h of reperfusion. Myocardial apoptosis was detected by histological TUNEL staining and DNA laddering. Expression of Bcl-2 and Bax proteins was analyzed using Western blot assay. Neutrophil localization was detected by immunohistochemistry with monoclonal anti-neutrophil CD18 antibody. There was no group difference in collateral blood flow (colored microspheres) during ischemia. Intra-left atrial administration of Ado and CGS21680 significantly decreased infarct size from 26+/-2% in Control to 13+/-1%* and 16+/-3%*, respectively. TUNEL positive cells in the peri-necrotic zone of the ischemic myocardium were also significantly reduced from 16+/-2% in Control group to 9+/-1%* and 10+/-2%*, respectively, consistent with the absence of DNA laddering in these two groups. Densitometrically, Ado and CGS21680 at reperfusion significantly increased the expression (% of normal myocardium) of downregulated Bcl-2 from 45+/-6% in Control group to 78+/-12%* and 69+/-10%*, respectively, and attenuated expression of upregulated Bax from 198+/-16% in Control group to 148+/-10%* and 158+/-12%*, respectively. Furthermore, the number of positive CD18 cells (mm(2) myocardium), which was significantly correlated with TUNEL positive cells in peri-necrotic zone, was significantly reduced from 403+/-42 in Control group to 142+/-18* in Ado group and 153+/-20%* in CGS21680 group, respectively. In conclusion, the present study suggests that inhibition of apoptosis by Ado at reperfusion involves alterations in anti-apoptotic Bcl-2 and pro-apoptotic Bax proteins and neutrophil accumulation, primarily mediated by an adenosine A2A receptor. * P<0.05 v Control group.
Zhao
Z Q
ZQ
Department of Cardiothoracic Surgery, Emory University School of Medicine, Atlanta, GA 30365-2225, USA. zzhao@emory.edu
Budde
J M
JM
Morris
C
C
Wang
N P
NP
Velez
D A
DA
Muraki
S
S
Guyton
R A
RA
Vinten-Johansen
J
J
D000241:Adenosine / Q000008:administration & dosage / Q000031:analogs & derivatives* / Q000494:pharmacology / Q000627:therapeutic use*; D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D018821:CD18 Antigens / Q000032:analysis; D003326:Coronary Circulation / Q000187:drug effects; D053938:DNA Fragmentation; D004285:Dogs; D004353:Drug Evaluation, Preclinical; D005260:Female; D005786:Gene Expression Regulation / Q000187:drug effects*; D019254:Genes, bcl-2; D006439:Hemodynamics / Q000187:drug effects; D007269:Injections, Intra-Arterial; D008297:Male; D009203:Myocardial Infarction / Q000473:pathology; D017202:Myocardial Ischemia / Q000150:complications; D015428:Myocardial Reperfusion Injury / Q000235:genetics / Q000473:pathology / Q000517:prevention & control*; D009336:Necrosis; D009504:Neutrophils / Q000473:pathology; D010627:Phenethylamines / Q000494:pharmacology / Q000627:therapeutic use; D011518:Proto-Oncogene Proteins / Q000096:biosynthesis* / Q000235:genetics; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000096:biosynthesis*; D018047:Receptors, Purinergic P1 / Q000187:drug effects / Q000502:physiology; D051028:bcl-2-Associated X Protein
D018821:CD18 Antigens; D010627:Phenethylamines; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D018047:Receptors, Purinergic P1; D051028:bcl-2-Associated X Protein; C061282:2-(4-(2-carboxyethyl)phenethylamino)-5'-N-ethylcarboxamidoadenosine; D000241:Adenosine