The TLR9 ligand, CpG-ODN, induces protection against cerebral ischemia/reperfusion injury via activation of PI3K/Akt signaling.
BACKGROUND
Toll-like receptors (TLRs) have been shown to be involved in cerebral ischemia/reperfusion (I/R) injury. TLR9 is located in intracellular compartments and recognizes CpG-DNA. This study examined the effect of CpG-ODN on cerebral I/R injury.


RESULTS
C57BL/6 mice were treated with CpG-ODN by i.p. injection 1 hour before the mice were subjected to cerebral ischemia (60 minutes) followed by reperfusion (24 hours). Scrambled-ODN served as control-ODN. Untreated mice, subjected to cerebral I/R, served as I/R control. The effect of inhibitory CpG-ODN (iCpG-ODN) on cerebral I/R injury was also examined. In addition, we examined the therapeutic effect of CpG-ODN on cerebral I/R injury by administration of CpG-ODN 15 minutes after cerebral ischemia. CpG-ODN administration significantly decreased cerebral I/R-induced infarct volume by 69.7% (6.4±1.80% vs 21.0±2.85%, P<0.05), improved neurological scores, and increased survival rate, when compared with the untreated I/R group. Therapeutic administration of CpG-ODN also significantly reduced infarct volume by 44.7% (12.6±2.03% vs 22.8±2.54%, P<0.05) compared with untreated I/R mice. Neither control-ODN, nor iCpG-ODN altered I/R-induced cerebral injury or neurological deficits. Nissl staining showed that CpG-ODN treatment preserved neuronal morphology in the ischemic hippocampus. Immunoblot showed that CpG-ODN administration increased Bcl-2 levels by 41% and attenuated I/R-increased levels of Bax and caspase-3 activity in ischemic brain tissues. Importantly, CpG-ODN treatment induced Akt and GSK-3β phosphorylation in brain tissue and cultured microglial cells. PI3K inhibition with LY294002 abolished CpG-ODN-induced protection.


CONCLUSIONS
CpG-ODN significantly reduces cerebral I/R injury via a PI3K/Akt-dependent mechanism. Our data also indicate that CpG-ODN may be useful in the therapy of cerebral I/R injury.
Department of Surgery, East Tennessee State University, Johnson City, 37614, TN.
Chen
C
Lu
Tuanzhu
T
Ha
Xiaohui
X
Wang
Li
L
Liu
Xia
X
Zhang
Erinmarie Olson
EO
Kimbrough
Zhanxin
Z
Sha
Meijian
M
Guan
John
J
Schweitzer
John
J
Kalbfleisch
David
D
Williams
Chuanfu
C
Li
D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D001921:Brain / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology / Q000503:physiopathology; D002478:Cells, Cultured; D002544:Cerebral Infarction / Q000378:metabolism / Q000473:pathology / Q000503:physiopathology / Q000517:prevention & control*; D004195:Disease Models, Animal; D004789:Enzyme Activation; D008024:Ligands; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D017628:Microglia / Q000187:drug effects / Q000201:enzymology / Q000473:pathology; D009474:Neurons / Q000187:drug effects / Q000201:enzymology / Q000473:pathology; D018696:Neuroprotective Agents / Q000494:pharmacology*; D009838:Oligodeoxyribonucleotides / Q000494:pharmacology*; D058539:Phosphatidylinositol 3-Kinase / Q000378:metabolism*; D010766:Phosphorylation; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D015427:Reperfusion Injury / Q000378:metabolism / Q000473:pathology / Q000503:physiopathology / Q000517:prevention & control*; D015398:Signal Transduction / Q000187:drug effects; D013997:Time Factors; D051217:Toll-Like Receptor 9 / Q000819:agonists* / Q000378:metabolism
C408982:CPG-oligonucleotide; D008024:Ligands; D018696:Neuroprotective Agents; D009838:Oligodeoxyribonucleotides; C495370:Tlr9 protein, mouse; D051217:Toll-Like Receptor 9; D058539:Phosphatidylinositol 3-Kinase; D051057:Proto-Oncogene Proteins c-akt
CpG‐ODN; apoptosis; cerebral ischemia/reperfusion injury; toll‐like receptors