Neurochemical Research

, Volume 35, Issue 8, pp 1147–1155

Spatio-Temporal Distribution of Inflammatory Reaction and Expression of TLR2/4 Signaling Pathway in Rat Brain Following Permanent Focal Cerebral Ischemia

  • Xian-kun Tu
  • Wei-zhong Yang
  • Song-sheng Shi
  • Chun-hua Wang
  • Guo-liang Zhang
  • Tian-rui Ni
  • Chun-mei Chen
  • Rui Wang
  • Jian-wen Jia
  • Qi-min Song
Original Paper


Toll-like receptors (TLRs) are considered to mediate the inflammatory reaction, which are involved in the pathophysiological processes of cerebral ischemia injury. To elucidate the possible role of inflammatory reaction and TLR2/4 signaling pathway in cerebral ischemia, in the present study, we explored the spatio-temporal distribution of inflammatory reaction, and further investigated the time-course expression of TLR2/4 and the downstream effector molecules after focal cerebral ischemia in rats. Sprague–Dawley rats underwent permanent middle cerebral artery occlusion (pMCAO) for 6, 12, 24, 48 and 72 h. Neurological deficit, cerebral infarction and neutrophil infiltration were measured at different time points following pMCAO. Expression of TLR2/4 were examined by immunohistochemistry, reverse transcription-polymerase chain reaction (RT-PCR) and western blot. Nuclear factor-kappaB (NF-κB) and cyclooxygenase-2 (COX-2) were determined by western blot. Serum content of tumor necrosis factor-α (TNF-α) was detected by enzyme-linked immunosorbent assay (ELISA). Experimental results showed that pMCAO caused an increase of neutrophil infiltration in infarcted brain tissue, with a peaked activity at 24 h of ischemia. The inflammatory molecules including TLR2, TLR4, NF-κB, COX-2 and TNF-α were significantly increased after pMCAO, especially during 12–24 h of ischemia, which were correlated with pMCAO-induced brain injury and cerebral inflammation. Our studies suggested that TLR2/4 signaling pathway likely aggravated ischemic brain injury through mediating the inflammatory reaction. TLR2/4 signaling pathway may be a promising therapeutic target for cerebral ischemia injury.


Cerebral ischemia Cyclooxygenase-2 Inflammation Nuclear factor-kappaB Toll-like receptors Tumor necrosis factor-α Stroke 


  1. 1.
    Jemal A, Ward E, Hao Y et al (2005) Trends in the leading causes of death in the United States, 1970–2002. JAMA 294:1255–1259CrossRefPubMedGoogle Scholar
  2. 2.
    Durukan A, Tatlisumak T (2007) Acute ischemic stroke: overview of major experimental rodent models, pathophysiology, and therapy of focal cerebral ischemia. Pharmacol Biochem Behav 87:179–197CrossRefPubMedGoogle Scholar
  3. 3.
    Danton GH, Dietrich WD (2003) Inflammatory mechanisms after ischemia and stroke. J Neuropathol Exp Neurol 62:127–136PubMedGoogle Scholar
  4. 4.
    Huang J, Upadhyay UM, Tamargo RJ (2006) Inflammation in stroke and focal cerebral ischemia. Surg Neurol 66:232–245CrossRefPubMedGoogle Scholar
  5. 5.
    Kaur C, Ling EA (2008) Blood brain barrier in hypoxic-ischemic conditions. Curr Neurovasc Res 5:71–81CrossRefPubMedGoogle Scholar
  6. 6.
    Hoffman WH, Stamatovic SM, Andjelkovic AV (2009) Inflammatory mediators and blood brain barrier disruption in fatal brain edema of diabetic ketoacidosis. Brain Res 1254:138–148CrossRefPubMedGoogle Scholar
  7. 7.
    Jordan J, Segura T, Brea D et al (2008) Inflammation as therapeutic objective in stroke. Curr Pharm Des 14:3549–3564CrossRefPubMedGoogle Scholar
  8. 8.
    Tu XK, Yang WZ, Shi SS et al (2009) 5-Lipoxygenase inhibitor zileuton attenuates ischemic brain damage: involvement of matrix metalloproteinase 9. Neurol Res 31:848–852CrossRefPubMedGoogle Scholar
  9. 9.
    Aderem A, Ulevitch RJ (2000) Toll-like receptors in the induction of the innate immune response. Nature 406:782–787CrossRefPubMedGoogle Scholar
  10. 10.
    Kaisho T, Akira S (2006) Toll-like receptor function and signaling. J Allergy Clin Immunol 117:979–987CrossRefPubMedGoogle Scholar
  11. 11.
    Meylan E, Tschopp J, Karin M (2006) Intracellular pattern recognition receptors in the host response. Nature 442:39–44CrossRefPubMedGoogle Scholar
  12. 12.
    Barton GM, Medzhitov R (2003) Toll-like receptor signaling pathways. Science 300:1524–1525CrossRefPubMedGoogle Scholar
  13. 13.
    Lehnardt S, Lehmann S, Kaul D et al (2007) Toll-like receptor 2 mediates CNS injury in focal cerebral ischemia. J Neuroimmunol 190:28–33CrossRefPubMedGoogle Scholar
  14. 14.
    Tang SC, Arumugam TV, Xu X et al (2007) Pivotal role for neuronal Toll-like receptors in ischemic brain injury and functional deficits. Proc Natl Acad Sci USA 104:13798–13803CrossRefPubMedGoogle Scholar
  15. 15.
    Seegers H, Grillon E, Trioullier Y et al (2000) Nuclear factor-kappa B activation in permanent intraluminal focal cerebral ischemia in the rat. Neurosci Lett 288:241–245CrossRefPubMedGoogle Scholar
  16. 16.
    Nijboer CH, Heijnen CJ, Groenendaal F et al (2008) Strong neuroprotection by inhibition of NF-kappaB after neonatal hypoxia-ischemia involves apoptotic mechanisms but is independent of cytokines. Stroke 39:2129–2137CrossRefPubMedGoogle Scholar
  17. 17.
    Kim YA, Lim SY, Rhee SH et al (2006) Resveratrol inhibits inducible nitric oxide synthase and cyclooxygenase-2 expression in beta-amyloid-treated C6 glioma cells. Int J Mol Med 17:1069–1075PubMedGoogle Scholar
  18. 18.
    Mattson MP (2005) NF-kappaB in the survival and plasticity of neurons. Neurochem Res 30:883–893CrossRefPubMedGoogle Scholar
  19. 19.
    Nogawa S, Zhang F, Ross ME et al (1997) Cyclo-oxygenase-2 gene expression in neurons contributes to ischemic brain damage. J Neurosci 17:2746–2755PubMedGoogle Scholar
  20. 20.
    Liu T, Clark RK, McDonnell PC et al (1994) Tumor necrosis factor-alpha expression in ischemic neurons. Stroke 25:1481–1488PubMedGoogle Scholar
  21. 21.
    Jiang C, Wang J, Li X et al (2009) Progesterone exerts neuroprotective effects by inhibiting inflammatory response after stroke. Inflamm Res 58:619–624CrossRefPubMedGoogle Scholar
  22. 22.
    Tu XK, Yang WZ, Shi SS et al (2009) Neuroprotective effect of baicalin in a rat model of permanent focal cerebral ischemia. Neurochem Res 34:1626–1634CrossRefPubMedGoogle Scholar
  23. 23.
    Bederson JB, Pitts LH, Tsuji M et al (1986) Rat middle cerebral artery occlusion: evaluation of the model and development of a neurologic examination. Stroke 17:472–476PubMedGoogle Scholar
  24. 24.
    Matsuo Y, Onodera H, Shiga Y et al (1994) Correlation between myeloperoxidase-quantified neutrophil accumulation and ischemic brain injury in the rat: effects of neutrophil depletion. Stroke 25:1469–1475PubMedGoogle Scholar
  25. 25.
    Gill MB, Perez-Polo JR (2008) Hypoxia ischemia-mediated cell death in neonatal rat brain. Neurochem Res 33:2379–2389CrossRefPubMedGoogle Scholar
  26. 26.
    Zhou Y, Wei EQ, Fang SH et al (2006) Spatio-temporal properties of 5-lipoxygenase expression and activation in the brain after focal cerebral ischemia in rats. Life Sci 79:1645–1656CrossRefPubMedGoogle Scholar
  27. 27.
    Fang SH, Zhou Y, Chu LS et al (2007) Spatio-temporal expression of cysteinyl leukotriene receptor-2 mRNA in rat brain after focal cerebral ischemia. Neurosci Lett 412:78–83CrossRefPubMedGoogle Scholar
  28. 28.
    Guo G, Bhat NR (2007) p38alpha MAP kinase mediates hypoxia-induced motor neuron cell death: a potential target of minocycline’s neuroprotective action. Neurochem Res 32:2160–2166CrossRefPubMedGoogle Scholar
  29. 29.
    Ziegler G, Harhausen D, Schepers C et al (2007) TLR2 has a detrimental role in mouse transient focal cerebral ischemia. Biochem Biophys Res Commun 359:574–579CrossRefPubMedGoogle Scholar
  30. 30.
    Sironi L, Banfi C, Brioschi M et al (2006) Activation of NF-kB and ERK1/2 after permanent focal ischemia is abolished by simvastatin treatment. Neurobiol Dis 22:445–451CrossRefPubMedGoogle Scholar
  31. 31.
    Caso JR, Pradillo JM, Hurtado O et al (2007) Toll-like receptor 4 is involved in brain damage and inflammation after experimental stroke. Circulation 115:1599–1608CrossRefPubMedGoogle Scholar
  32. 32.
    Cao CX, Yang QW, Lv FL et al (2007) Reduced cerebral ischemia-reperfusion injury in Toll-like receptor 4 deficient mice. Biochem Biophys Res Commun 353:509–514CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2010

Authors and Affiliations

  • Xian-kun Tu
    • 1
  • Wei-zhong Yang
    • 1
  • Song-sheng Shi
    • 1
  • Chun-hua Wang
    • 1
  • Guo-liang Zhang
    • 1
  • Tian-rui Ni
    • 1
  • Chun-mei Chen
    • 1
  • Rui Wang
    • 1
  • Jian-wen Jia
    • 1
  • Qi-min Song
    • 1
  1. 1.Department of NeurosurgeryThe Affiliated Union Hospital of Fujian Medical UniversityFuzhouChina

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