Signal transducer and activator of transcription 3alpha and specificity protein 1 interact to upregulate intercellular adhesion molecule-1 in ischemic-reperfused myocardium and vascular endothelium.
OBJECTIVE
Intercellular adhesion molecule-1 (ICAM-1) is upregulated rapidly on endothelial cells during ischemia-reperfusion (I-R) and mediates tissue leukocyte accumulation. The ICAM-1 proximal promoter contains a signal transducer and activator of transcription (Stat) binding motif (gamma-interferon activation site [GAS] sequence), which flanks a specificity protein 1 (Sp1) binding site. We examined the roles of Stat and Sp1 in the regulation of ICAM-1 after myocardial I-R.


RESULTS
Open-chest anesthetized rats underwent coronary artery occlusion for 35 minutes and reperfusion for 0 to 240 minutes. Stat became activated within 15 minutes after reperfusion, primarily in vascular endothelial cells; the activated Stat protein was identified as Stat3 (alpha-isoform). After phosphorylation on serine 727 (p-S727), Stat3alpha was found in association with the transcriptional regulator Sp1, and the complex bound to an ICAM-1-GAS probe. ICAM-1 expression increased after I-R and lagged shortly behind Stat3alpha activation. In cultured human umbilical vein endothelial (HUVE) cells, activation of Stat3alpha after hypoxia-reoxygenation (H-R) was dependent on the small GTPase Rac1. Transfection of a dominant-negative Stat3 (Y705F) adenovirus or a GAS decoy oligonucleotide reduced ICAM-1 mRNA expression after H-R. Using a reporter gene transfected into HUVE cells, mutation of the GAS element in the ICAM-1 promoter resulted in reduced transcriptional activity after H-R. Sp1 coimmunoprecipitated with p-S727 Stat3 during H-R, and Sp1 or Stat3alpha interfering RNA markedly reduced ICAM-1 mRNA expression.


CONCLUSIONS
The Sp1-Stat3 complex appears to play an important role in the upregulation of ICAM-1 transcription after reoxygenation or reperfusion.
Yang
Xiao Ping
XP
Cardiology Division, Halsted 500, Johns Hopkins Hospital, 600 N Wolfe St, Baltimore, MD 21287-5500. lbecker@mail.jhmi.edu.
Irani
Kaikobad
K
Mattagajasingh
Subhendra
S
Dipaula
Anthony
A
Khanday
Firdous
F
Ozaki
Michitaka
M
Fox-Talbot
Karen
K
Baldwin
William M
WM
Becker
Lewis C
LC
D000818:Animals; D002478:Cells, Cultured; D003331:Coronary Vessels / Q000166:cytology / Q000502:physiology; D004730:Endothelium, Vascular / Q000166:cytology / Q000502:physiology; D006801:Humans; D018799:Intercellular Adhesion Molecule-1 / Q000235:genetics*; D015850:Interleukin-6 / Q000494:pharmacology; D008297:Male; D015425:Myocardial Reperfusion; D015428:Myocardial Reperfusion Injury / Q000378:metabolism* / Q000503:physiopathology*; D010766:Phosphorylation / Q000187:drug effects; D011401:Promoter Regions, Genetic / Q000502:physiology; D034622:RNA Interference; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D011917:Rats, Inbred Lew; D050796:STAT3 Transcription Factor / Q000235:genetics / Q000378:metabolism*; D016329:Sp1 Transcription Factor / Q000378:metabolism*; D015533:Transcriptional Activation / Q000502:physiology; D014443:Tyrosine / Q000378:metabolism; D014471:Umbilical Veins / Q000166:cytology; D015854:Up-Regulation / Q000502:physiology; D020830:rac1 GTP-Binding Protein / Q000378:metabolism
D015850:Interleukin-6; D012333:RNA, Messenger; D050796:STAT3 Transcription Factor; D016329:Sp1 Transcription Factor; D018799:Intercellular Adhesion Molecule-1; D014443:Tyrosine; C499134:Rac1 protein, rat; D020830:rac1 GTP-Binding Protein