Role of the Jak/STAT pathway in the regulation of interleukin-8 transcription by oxidized phospholipids in vitro and in atherosclerosis in vivo.
Oxidized 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphorylcholine (Ox-PAPC) and its component phospholipid, 1-palmitoyl-2-epoxyisoprostane-sn-glycero-3-phosphorylcholine, induce endothelial cells (EC) to synthesize chemotactic factors, such as interleukin 8 (IL-8). Previously, we demonstrated a role for c-Src kinase activation in Ox-PAPC-induced IL-8 transcription. In this study, we have examined the mechanism regulating IL-8 transcription by Ox-PAPC downstream of c-Src. Our findings demonstrate an important role for JAK2 in the regulation of IL-8 transcription by Ox-PAPC. Treatment of human aortic EC with Ox-PAPC and 1-palmitoyl-2-epoxyisoprostane-sn-glycero-3-phosphorylcholine induced a rapid yet sustained activation of JAK2; activation of JAK2 by Ox-PAPC was dependent on c-Src kinase activity. Furthermore, pretreatment with selective JAK2 inhibitors significantly reduced Ox-PAPC-induced IL-8 transcription. In previous studies, we also demonstrated activation of STAT3 by Ox-PAPC. Here we provide evidence that STAT3 activation by Ox-PAPC is dependent on JAK2 activation and that STAT3 activation regulates IL-8 transcription by Ox-PAPC in human EC. Transfection with small interfering RNA against STAT3 significantly reduced Ox-PAPC-induced IL-8 transcription. Using chromatin immunoprecipitation assays, we demonstrated binding of activated STAT3 to the sequence flanking the consensus gamma-interferon activation sequence (GAS) in the IL-8 promoter; site-directed mutagenesis of GAS inhibited IL-8 transcription by Ox-PAPC. Finally, these studies demonstrate a role for STAT3 activation in atherosclerosis in vivo. We found increased staining for activated STAT3 in the inflammatory regions of human atherosclerotic lesions and reduced fatty streak formation in EC-specific STAT3 knock-out mice on the atherogenic diet. Taken together, these data demonstrate an important role for the JAK2/STAT3 pathway in Ox-PAPC-induced IL-8 transcription in vitro and in atherosclerosis in vivo.
Gharavi
Nima M
NM
Division of Cardiology, Department of Medicine, Department of Pathology, Molecular Biology Institute, UCLA, Los Angeles, California 90095, USA.
Alva
Jackelyn A
JA
Mouillesseaux
Kevin P
KP
Lai
Chi
C
Yeh
Michael
M
Yeung
Winnie
W
Johnson
Jaclyn
J
Szeto
Wan Lam
WL
Hong
Longsheng
L
Fishbein
Michael
M
Wei
Lai
L
Pfeffer
Lawrence M
LM
Berliner
Judith A
JA
D000818:Animals; D001011:Aorta / Q000166:cytology; D050197:Atherosclerosis / Q000378:metabolism*; D002196:Capillaries / Q000166:cytology; D002478:Cells, Cultured; D047369:Chromatin Immunoprecipitation; D042783:Endothelial Cells / Q000187:drug effects / Q000378:metabolism*; D004730:Endothelium, Vascular / Q000166:cytology; D004789:Enzyme Activation / Q000187:drug effects; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D007150:Immunohistochemistry; D066298:In Vitro Techniques; D016209:Interleukin-8 / Q000096:biosynthesis* / Q000235:genetics; D053614:Janus Kinase 2 / Q000378:metabolism*; D051379:Mice; D018345:Mice, Knockout; D008954:Models, Biological; D010084:Oxidation-Reduction; D010713:Phosphatidylcholines / Q000378:metabolism* / Q000494:pharmacology / Q000502:physiology; D010766:Phosphorylation; D010957:Plasmids; D034741:RNA, Small Interfering / Q000378:metabolism; D050796:STAT3 Transcription Factor / Q000378:metabolism*; D013997:Time Factors; D015534:Trans-Activators / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects; D014162:Transfection
D016209:Interleukin-8; D010713:Phosphatidylcholines; D034741:RNA, Small Interfering; D050796:STAT3 Transcription Factor; C494086:STAT3 protein, human; C494087:Stat3 protein, mouse; D015534:Trans-Activators; C472349:oxidized-L-alpha-1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphorylcholine; D053614:Janus Kinase 2