Skip to main content

Advertisement

SpringerLink
Log in

Probenecid Protects Against Transient Focal Cerebral Ischemic Injury by Inhibiting HMGB1 Release and Attenuating AQP4 Expression in Mice

  • Original Paper
  • Published:
Neurochemical Research Aims and scope Submit manuscript

Abstract

Stroke results in inflammation, brain edema, and neuronal death. However, effective neuroprotectants are not available. Recent studies have shown that high mobility group box-1 (HMGB1), a proinflammatory cytokine, contributes to ischemic brain injury. Aquaporin 4 (AQP4), a water channel protein, is considered to play a pivotal role in ischemia-induced brain edema. More recently, studies have shown that pannexin 1 channels are involved in cerebral ischemic injury and the cellular inflammatory response. Here, we examined whether the pannexin 1 channel inhibitor probenecid could reduce focal ischemic brain injury by inhibiting cerebral inflammation and edema. Transient focal ischemia was induced in C57BL/6J mice by middle cerebral artery occlusion (MCAO) for 1 h. Infarct volume, neurological score and cerebral water content were evaluated 48 h after MCAO. Immunostaining, western blot analysis and ELISA were used to assess the effects of probenecid on the cellular inflammatory response, HMGB1 release and AQP4 expression. Administration of probenecid reduced infarct size, decreased cerebral water content, inhibited neuronal death, and reduced inflammation in the brain 48 h after stroke. In addition, HMGB1 release from neurons was significantly diminished and serum HMGB1 levels were substantially reduced following probenecid treatment. Moreover, AQP4 protein expression was downregulated in the cortical penumbra following post-stroke treatment with probenecid. These results suggest that probenecid, a powerful pannexin 1 channel inhibitor, protects against ischemic brain injury by inhibiting cerebral inflammation and edema.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

Similar content being viewed by others

References

  1. Adams HP Jr, del Zoppo G, Alberts MJ, Bhatt DL, Brass L, Furlan A, Grubb RL, Higashida RT, Jauch EC, Kidwell C, Lyden PD, Morgenstern LB, Qureshi AI, Rosenwasser RH, Scott PA, Wijdicks EF (2007) Guidelines for the early management of adults with ischemic stroke: a guideline from the American Heart Association/American Stroke Association Stroke Council, Clinical Cardiology Council, Cardiovascular Radiology and Intervention Council, and the Atherosclerotic Peripheral Vascular Disease and Quality of Care Outcomes in Research Interdisciplinary Working Groups: the American Academy of neurology affirms the value of this guideline as an educational tool for neurologists. Stroke 38:1655–1711

    Article  PubMed  Google Scholar 

  2. Sattar N, Murray HM, Welsh P, Blauw GJ, Buckley BM, Cobbe S, de Craen AJ, Lowe GD, Jukema JW, Macfarlane PW, Murphy MB, Stott DJ, Westendorp RG, Shepherd J, Ford I, Packard CJ (2009) Are markers of inflammation more strongly associated with risk for fatal than for nonfatal vascular events? PLoS Med 6:e1000099

    Article  PubMed Central  PubMed  Google Scholar 

  3. Yue R, Yuan X, Liu X, Zhang J, Jiang P, He C, Shan L, Yu Y, Zhang W (2012) Cynandione A mitigates ischemic injuries in rats with cerebral ischemia. J Neurochem 121:451–464

    Article  CAS  PubMed  Google Scholar 

  4. Xiong X, White RE, Xu L, Yang L, Sun X, Zou B, Pascual C, Sakurai T, Giffard RG, Xie XS (2013) Mitigation of murine focal cerebral ischemia by the hypocretin/orexin system is associated with reduced inflammation. Stroke 44:764–770

    Article  CAS  PubMed  Google Scholar 

  5. Zhang J, Takahashi HK, Liu K, Wake H, Liu R, Maruo T, Date I, Yoshino T, Ohtsuka A, Mori S, Nishibori M (2011) Anti-high mobility group box-1 monoclonal antibody protects the blood-brain barrier from ischemia-induced disruption in rats. Stroke 42:1420–1428

    Article  CAS  PubMed  Google Scholar 

  6. Liu K, Mori S, Takahashi HK, Tomono Y, Wake H, Kanke T, Sato Y, Hiraga N, Adachi N, Yoshino T, Nishibori M (2007) Anti-high mobility group box 1 monoclonal antibody ameliorates brain infarction induced by transient ischemia in rats. FASEB J 21:3904–3916

    Article  CAS  PubMed  Google Scholar 

  7. Kim JB, Lim CM, Yu YM, Lee JK (2008) Induction and subcellular localization of high-mobility group box-1 (HMGB1) in the postischemic rat brain. J Neurosci Res 86:1125–1131

    Article  CAS  PubMed  Google Scholar 

  8. Muhammad S, Barakat W, Stoyanov S, Murikinati S, Yang H, Tracey KJ, Bendszus M, Rossetti G, Nawroth PP, Bierhaus A, Schwaninger M (2008) The HMGB1 receptor RAGE mediates ischemic brain damage. J Neurosci 28:12023–12031

    Article  CAS  PubMed  Google Scholar 

  9. Aghayev K, Bal E, Rahimli T, Mut M, Balci S, Vrionis F, Akalan N (2012) Aquaporin-4 expression is not elevated in mild hydrocephalus. Acta Neurochir (Wien) 154:753–759 discussion 759

    Article  Google Scholar 

  10. Iacovetta C, Rudloff E, Kirby R (2012) The role of aquaporin 4 in the brain. Vet Clin Pathol 41:32–44

    PubMed  Google Scholar 

  11. Manley GT, Fujimura M, Ma T, Noshita N, Filiz F, Bollen AW, Chan P, Verkman AS (2000) Aquaporin-4 deletion in mice reduces brain edema after acute water intoxication and ischemic stroke. Nat Med 6:159–163

    Article  CAS  PubMed  Google Scholar 

  12. Hiroaki Y, Tani K, Kamegawa A, Gyobu N, Nishikawa K, Suzuki H, Walz T, Sasaki S, Mitsuoka K, Kimura K, Mizoguchi A, Fujiyoshi Y (2006) Implications of the aquaporin-4 structure on array formation and cell adhesion. J Mol Biol 355:628–639

    Article  CAS  PubMed  Google Scholar 

  13. Saadoun S, Papadopoulos MC, Watanabe H, Yan D, Manley GT, Verkman AS (2005) Involvement of aquaporin-4 in astroglial cell migration and glial scar formation. J Cell Sci 118:5691–5698

    Article  CAS  PubMed  Google Scholar 

  14. Fukuda AM, Badaut J (2012) Aquaporin 4: a player in cerebral edema and neuroinflammation. J Neuroinflammation 9:279

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  15. Baranova A, Ivanov D, Petrash N, Pestova A, Skoblov M, Kelmanson I, Shagin D, Nazarenko S, Geraymovych E, Litvin O, Tiunova A, Born TL, Usman N, Staroverov D, Lukyanov S, Panchin Y (2004) The mammalian pannexin family is homologous to the invertebrate innexin gap junction proteins. Genomics 83:706–716

    Article  CAS  PubMed  Google Scholar 

  16. Qu Y, Misaghi S, Newton K, Gilmour LL, Louie S, Cupp JE, Dubyak GR, Hackos D, Dixit VM (2011) Pannexin-1 is required for ATP release during apoptosis but not for inflammasome activation. J Immunol 186:6553–6561

    Article  CAS  PubMed  Google Scholar 

  17. Pui K, Gow PJ, Dalbeth N (2013) Efficacy and tolerability of probenecid as urate-lowering therapy in gout; clinical experience in high-prevalence population. J Rheumatol 40:872–876

    Article  CAS  PubMed  Google Scholar 

  18. Mason RM (1954) Studies on the effect of probenecid (benemid) in gout. Ann Rheum Dis 13:120–130

    Article  CAS  PubMed  Google Scholar 

  19. Cunningham RF, Israili ZH, Dayton PG (1981) Clinical pharmacokinetics of probenecid. Clin Pharmacokinet 6:135–151

    Article  CAS  PubMed  Google Scholar 

  20. Suadicani SO, Brosnan CF, Scemes E (2006) P2X7 receptors mediate ATP release and amplification of astrocytic intercellular Ca2+ signaling. J Neurosci 26:1378–1385

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  21. Silverman W, Locovei S, Dahl G (2008) Probenecid, a gout remedy, inhibits pannexin 1 channels. Am J Physiol Cell Physiol 295:C761–C767

    Article  CAS  PubMed  Google Scholar 

  22. Bruzzone R, Hormuzdi SG, Barbe MT, Herb A, Monyer H (2003) Pannexins, a family of gap junction proteins expressed in brain. Proc Natl Acad Sci USA 100:13644–13649

    Article  CAS  PubMed  Google Scholar 

  23. Vanden Abeele F, Bidaux G, Gordienko D, Beck B, Panchin YV, Baranova AV, Ivanov DV, Skryma R, Prevarskaya N (2006) Functional implications of calcium permeability of the channel formed by pannexin 1. J Cell Biol 174:535–546

    Article  CAS  PubMed  Google Scholar 

  24. Peters MA, Teramoto T, White JQ, Iwasaki K, Jorgensen EM (2007) A calcium wave mediated by gap junctions coordinates a rhythmic behavior in C. elegans. Curr Biol 17:1601–1608

    Article  CAS  PubMed  Google Scholar 

  25. Locovei S, Bao L, Dahl G (2006) Pannexin 1 in erythrocytes: function without a gap. Proc Natl Acad Sci USA 103:7655–7659

    Article  CAS  PubMed  Google Scholar 

  26. Jiang H, Zhu AG, Mamczur M, Falck JR, Lerea KM, McGiff JC (2007) Stimulation of rat erythrocyte P2X7 receptor induces the release of epoxyeicosatrienoic acids. Br J Pharmacol 151:1033–1040

    Article  CAS  PubMed  Google Scholar 

  27. Silverman WR, de Rivero Vaccari JP, Locovei S, Qiu F, Carlsson SK, Scemes E, Keane RW, Dahl G (2009) The pannexin 1 channel activates the inflammasome in neurons and astrocytes. J Biol Chem 284:18143–18151

    Article  CAS  PubMed  Google Scholar 

  28. Kanneganti TD, Lamkanfi M, Kim YG, Chen G, Park JH, Franchi L, Vandenabeele P, Nunez G (2007) Pannexin-1-mediated recognition of bacterial molecules activates the cryopyrin inflammasome independent of Toll-like receptor signaling. Immunity 26:433–443

    Article  CAS  PubMed  Google Scholar 

  29. Bargiotas P, Krenz A, Hormuzdi SG, Ridder DA, Herb A, Barakat W, Penuela S, von Engelhardt J, Monyer H, Schwaninger M (2011) Pannexins in ischemia-induced neurodegeneration. Proc Natl Acad Sci USA 108:20772–20777

    Article  CAS  PubMed  Google Scholar 

  30. Chekeni FB, Elliott MR, Sandilos JK, Walk SF, Kinchen JM, Lazarowski ER, Armstrong AJ, Penuela S, Laird DW, Salvesen GS, Isakson BE, Bayliss DA, Ravichandran KS (2010) Pannexin 1 channels mediate ‘find-me’ signal release and membrane permeability during apoptosis. Nature 467:863–867

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  31. Gu L, Xiong X, Zhang H, Xu B, Steinberg GK, Zhao H (2012) Distinctive effects of T cell subsets in neuronal injury induced by cocultured splenocytes in vitro and by in vivo stroke in mice. Stroke 43:1941–1946

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  32. Joo SP, Xie W, Xiong X, Xu B, Zhao H (2013) Ischemic postconditioning protects against focal cerebral ischemia by inhibiting brain inflammation while attenuating peripheral lymphopenia in mice. Neuroscience 243:149–157

    Google Scholar 

  33. Xiong X, Barreto GE, Xu L, Ouyang YB, Xie X, Giffard RG (2011) Increased brain injury and worsened neurological outcome in interleukin-4 knockout mice after transient focal cerebral ischemia. Stroke 42:2026–2032

    Article  PubMed Central  PubMed  Google Scholar 

  34. Zheng YY, Lan YP, Tang HF, Zhu SM (2008) Propofol pretreatment attenuates aquaporin-4 over-expression and alleviates cerebral edema after transient focal brain ischemia reperfusion in rats. Anesth Analg 107:2009–2016

    Article  CAS  PubMed  Google Scholar 

  35. Zhao XC, Zhang LM, Tong DY, An P, Jiang C, Zhao P, Chen WM, Wang J (2013) Propofol increases expression of basic fibroblast growth factor after transient cerebral ischemia in rats. Neurochem Res 38:530–537

    Article  CAS  PubMed  Google Scholar 

  36. Shin YJ, Lee JH, Oh JH, Lee YJ (2013) Low-dose probenecid selectively inhibits urinary excretion of phenolsulfonphthalein in rats without affecting biliary excretion. J Appl Toxicol 33:511–515

    Article  CAS  PubMed  Google Scholar 

  37. Xiong X, Gu L, Zhang H, Xu B, Zhu S, Zhao H (2012) The protective effects of T cell deficiency against brain injury are ischemic model-dependent in rats. Neurochem Int 62:265–270

    Article  PubMed  Google Scholar 

  38. Gu L, Xiong X, Wei D, Gao X, Krams S, Zhao H (2013) T cells contribute to stroke-induced lymphopenia in rats. PLoS ONE 8:e59602

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  39. Zhu SM, Xiong XX, Zheng YY, Pan CF (2009) Propofol inhibits aquaporin 4 expression through a protein kinase C-dependent pathway in an astrocyte model of cerebral ischemia/reoxygenation. Anesth Analg 109:1493–1499

    Article  CAS  PubMed  Google Scholar 

  40. Pelegrin P, Surprenant A (2006) Pannexin-1 mediates large pore formation and interleukin-1beta release by the ATP-gated P2X7 receptor. EMBO J 25:5071–5082

    Article  CAS  PubMed  Google Scholar 

  41. Locovei S, Scemes E, Qiu F, Spray DC, Dahl G (2007) Pannexin1 is part of the pore forming unit of the P2X(7) receptor death complex. FEBS Lett 581:483–488

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  42. Scaffidi P, Misteli T, Bianchi ME (2002) Release of chromatin protein HMGB1 by necrotic cells triggers inflammation. Nature 418:191–195

    Article  CAS  PubMed  Google Scholar 

  43. Gardella S, Andrei C, Ferrera D, Lotti LV, Torrisi MR, Bianchi ME, Rubartelli A (2002) The nuclear protein HMGB1 is secreted by monocytes via a non-classical, vesicle-mediated secretory pathway. EMBO Rep 3:995–1001

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  44. Asavarut P, Zhao H, Gu J, Ma D (2013) The role of HMGB1 in inflammation-mediated organ injury. Acta Anaesthesiol Taiwan 51:28–33

    Article  PubMed  Google Scholar 

  45. Zhao H, Watts HR, Chong M, Huang H, Tralau-Stewart C, Maxwell PH, Maze M, George AJ, Ma D (2013) Xenon treatment protects against cold ischemia associated delayed graft function and prolongs graft survival in rats. Am J Transplant 13:2006–2018

    Article  CAS  PubMed  Google Scholar 

  46. Qiu J, Nishimura M, Wang Y, Sims JR, Qiu S, Savitz SI, Salomone S, Moskowitz MA (2008) Early release of HMGB-1 from neurons after the onset of brain ischemia. J Cereb Blood Flow Metab 28:927–938

    Article  CAS  PubMed  Google Scholar 

  47. Fujita M, Tsuruta R, Kaneko T, Otsuka Y, Kutsuna S, Izumi T, Aoki T, Shitara M, Kasaoka S, Maruyama I, Yuasa M, Maekawa T (2010) Hyperoxia suppresses excessive superoxide anion radical generation in blood, oxidative stress, early inflammation, and endothelial injury in forebrain ischemia/reperfusion rats: laboratory study. Shock 34:299–305

    Article  CAS  PubMed  Google Scholar 

  48. Hayakawa K, Mishima K, Nozako M, Hazekawa M, Mishima S, Fujioka M, Orito K, Egashira N, Iwasaki K, Fujiwara M (2008) Delayed treatment with minocycline ameliorates neurologic impairment through activated microglia expressing a high-mobility group box 1-inhibiting mechanism. Stroke 39:951–958

    Article  CAS  PubMed  Google Scholar 

  49. Papadopoulos MC, Verkman AS (2008) Potential utility of aquaporin modulators for therapy of brain disorders. Prog Brain Res 170:589–601

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  50. Satin J (2013) The long and short of PKC modulation of the L-type calcium channel. Channels (Austin) 7:57–58

    Article  Google Scholar 

  51. MacVicar BA, Thompson RJ (2010) Non-junction functions of pannexin-1 channels. Trends Neurosci 33:93–102

    Article  CAS  PubMed  Google Scholar 

  52. Thompson RJ, Zhou N, MacVicar BA (2006) Ischemia opens neuronal gap junction hemichannels. Science 312:924–927

    Article  CAS  PubMed  Google Scholar 

  53. Anderson TR, Jarvis CR, Biedermann AJ, Molnar C, Andrew RD (2005) Blocking the anoxic depolarization protects without functional compromise following simulated stroke in cortical brain slices. J Neurophysiol 93:963–979

    Article  CAS  PubMed  Google Scholar 

  54. Bargiotas P, Monyer H, Schwaninger M (2009) Hemichannels in cerebral ischemia. Curr Mol Med 9:186–194

    Article  CAS  PubMed  Google Scholar 

  55. Dahl G, Keane RW (2012) Pannexin: from discovery to bedside in 11 ± 4 years? Brain Res 1487:150–159

    Article  CAS  PubMed Central  PubMed  Google Scholar 

Download references

Acknowledgments

This work was supported by the National Natural Science Foundation of China (Grant Nos. 81271274 and 81301019).

Conflict of interest

The authors have no conflict of interest.

Author information

Authors and Affiliations

  1. Department of Anesthesia, the First Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou, Zhejiang, China

    Xiao-Xing Xiong, Xian-Hui Kang, Yue-Ying Zheng & Sheng-Mei Zhu

  2. Department of Basic Medicine, Hangzhou Normal University, Hangzhou, Zhejiang, China

    Li-Juan Gu

  3. Department of Neurosurgery, the First Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou, Zhejiang, China

    Jian Shen

  4. Department of Biology, Johns Hopkins University, Baltimore, MD, USA

    Si-biao Yue

Authors
  1. Xiao-Xing Xiong
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Li-Juan Gu
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Jian Shen
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Xian-Hui Kang
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Yue-Ying Zheng
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Si-biao Yue
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Sheng-Mei Zhu
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding authors

Correspondence to Si-biao Yue or Sheng-Mei Zhu.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Xiong, XX., Gu, LJ., Shen, J. et al. Probenecid Protects Against Transient Focal Cerebral Ischemic Injury by Inhibiting HMGB1 Release and Attenuating AQP4 Expression in Mice. Neurochem Res 39, 216–224 (2014). https://doi.org/10.1007/s11064-013-1212-z

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11064-013-1212-z

Keywords

Search

Navigation