1.U0126 prevents ERK pathway phosphorylation and interleukin-1beta mRNA production after cerebral ischemia.
Zhi-qiu WANG ; Xian-cheng CHEN ; Guo-yuan YANG ; Liang-fu ZHOU
Chinese Medical Sciences Journal 2004;19(4):270-275
OBJECTIVETo study the role of extracellular signal-regulated kinase (ERK) in cerebral ischemia and the mechanism of protective effects of U0126 (1,4-diamino-2,3-dicyano-1,4-bis[2-aminophenylthio] butadiene) on ischemic brain.
METHODSMice underwent left middle cerebral artery occlusion (MCAO) by introducing a suture in the lumen. U0126 was injected intravenously through the internal jugular vein. The immuno-activity of phosphorylated ERK1/2 (pERK1/2), phosphorylated mitogen activated protein kinase kinase (pMEK), and phosphorylated Elk-1 (pElk-1) was assessed by Western blot analysis and immunohistochemistry. Interleukin (IL)-1beta mRNA level was measured by ribonuclease protection assay.
RESULTSPhosphorylated ERK1/2 in 2 hours MCAO mice was down-regulated after intravenous injection of U0126. The inhibition was dose dependent and treatment time related. pMEK and pElk-1 were also reduced in a similar fashion after U0126 treatment. IL-1beta mRNA increased after 1 and 2 hours of MCAO. After injection of U0126, it was down-regulated during 1 to 4 hours after MCAO.
CONCLUSIONIntravenous administration of the MEK inhibitor U0126 inhibits pMEK, pERK1/2, and pElk-1 up-regulation induced by cerebral ischemia. The protective effect of U0126 against ischemic injury is probably resulted from the reduction of IL-1beta mRNA via the inhibition of ERK pathway.
Animals ; Butadienes ; pharmacology ; DNA-Binding Proteins ; metabolism ; Enzyme Inhibitors ; pharmacology ; Infarction, Middle Cerebral Artery ; metabolism ; Interleukin-1 ; biosynthesis ; genetics ; Male ; Mice ; Mitogen-Activated Protein Kinase 1 ; metabolism ; Mitogen-Activated Protein Kinase Kinases ; antagonists & inhibitors ; metabolism ; Nitriles ; pharmacology ; Phosphorylation ; Proto-Oncogene Proteins ; metabolism ; RNA, Messenger ; biosynthesis ; genetics ; Signal Transduction ; Transcription Factors ; metabolism ; ets-Domain Protein Elk-1
2.The translation inhibitor anisomycin induces Elk-1-mediated transcriptional activation of egr-1 through multiple mitogen-activated protein kinase pathways.
Soon Young SHIN ; Joon Ho LEE ; Byung MIN ; Young Han LEE
Experimental & Molecular Medicine 2006;38(6):677-685
The early growth response-1 gene (egr-1) encodes a zinc-finger transcription factor Egr-1 and is rapidly inducible by a variety of extracellular stimuli. Anisomycin (ANX), a protein synthesis inhibitor, stimulates mitogen-activated protein kinase (MAPK) pathways and thereby causes a rapid induction of immediate-early response genes. We found that anisomycin treatment of U87MG glioma cells resulted in a marked, time-dependent increase in levels of Egr-1 protein. The results of Northern blot analysis and reporter gene assay of egr-1 gene promoter (Pegr-1) activity indicate that the ANX- induced increase in Egr-1 occurs at the transcriptional level. Deletion of the serum response element (SRE) in the 5'-flanking region of egr-1 gene abolished ANX-induced Pegr-1 activity. ANX induced the phosphorylation of the ERK1/2, JNK, and p38 MAPKs in a time-dependent manner and also induced transactivation of Gal4-Elk-1, suggesting that Elk-1 is involved in SRE-mediated egr-1 transcription. Transient transfection of dominant-negative constructs of MAPK pathways blocked ANX-induced Pegr-1 activity. Furthermore, pretreatment with specific MAPK pathway inhibitors, including the MEK inhibitor U0126, the JNK inhibitor SP600125, and the p38 kinase inhibitor SB202190, completely inhibited ANX-inducible expression of Egr-1. Taken together, these results suggest that all three MAPK pathways play a crucial role in ANX-induced transcriptional activation of Pegr-1 through SRE-mediated transactivation of Elk
p38 Mitogen-Activated Protein Kinases/genetics/metabolism
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ets-Domain Protein Elk-1/genetics/*metabolism
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Trans-Activation (Genetics)/*drug effects
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Serum Response Element
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Protein Kinase Inhibitors/pharmacology
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Protein Biosynthesis/*drug effects
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Promoter Regions (Genetics)/genetics
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*MAP Kinase Signaling System/drug effects
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JNK Mitogen-Activated Protein Kinases/genetics/metabolism
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Humans
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Extracellular Signal-Regulated MAP Kinases/genetics/metabolism
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Early Growth Response Protein 1/genetics/*metabolism
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Cell Line, Tumor
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Anisomycin/*pharmacology