The Role of Oxidative Stress in Microvascular Disturbances after Experimental Subarachnoid Hemorrhage


Oxidative stress was shown to play a crucial role in the diverse pathogenesis of early brain injury (EBI) after subarachnoid hemorrhage (SAH). Microcirculatory dysfunction is thought to be an important and fundamental pathological change in EBI. However, other than blood-brain barrier (BBB) disruption, the influence of oxidative stress on microvessels remains to be elucidated. The aim of this study was to investigate the role of oxidative stress on microcirculatory integrity in EBI. SAH was induced in male Sprague-Dawley rats using an endovascular perforation technique. A free radical scavenger, edaravone, was administered prophylactically by intraperitoneal injection. SAH grade, neurological score, brain water content, and BBB permeability were measured at 24 h after SAH induction. In addition, cortical samples taken at 24 h after SAH were analyzed to explore oxidative stress, microvascular mural cell apoptosis, microspasm, and microthrombosis. Edaravone treatment significantly ameliorated neurological deficits, brain edema, and BBB disruption. In addition, oxidative stress-induced modifications and subsequent apoptosis of microvascular endothelial cells and pericytes increased after SAH induction, while the administration of edaravone suppressed this. Consistent with apoptotic cell inhibition, microthromboses were also inhibited by edaravone administration. Oxidative stress plays a pivotal role in the induction of multiple pathological changes in microvessels in EBI. Antioxidants are potential candidates for the treatment of microvascular disturbances after SAH.

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  1. 1.

    Macdonald RL, Schweizer TA. Spontaneous subarachnoid haemorrhage. Lancet. 2017;389(10069):655–66.

    PubMed  Google Scholar 

  2. 2.

    Macdonald RL. Origins of the concept of vasospasm. Stroke. 2016;47(1):e11–5.

    PubMed  Google Scholar 

  3. 3.

    Macdonald RL, Kassell NF, Mayer S, Ruefenacht D, Schmiedek P, Weidauer S, et al. Clazosentan to overcome neurological ischemia and infarction occurring after subarachnoid hemorrhage (conscious-1): randomized, double-blind, placebo-controlled phase 2 dose-finding trial. Stroke. 2008;39(11):3015–21.

    CAS  PubMed  Google Scholar 

  4. 4.

    Macdonald RL, Higashida RT, Keller E, Mayer SA, Molyneux A, Raabe A, et al. Clazosentan, an endothelin receptor antagonist, in patients with aneurysmal subarachnoid haemorrhage undergoing surgical clipping: a randomised, double-blind, placebo-controlled phase 3 trial (conscious-2). Lancet Neurol. 2011;10(7):618–25.

    CAS  PubMed  Google Scholar 

  5. 5.

    Sehba FA, Pluta RM, Zhang JH. Metamorphosis of subarachnoid hemorrhage research: from delayed vasospasm to early brain injury. Mol Neurobiol. 2011;43(1):27–40.

    CAS  PubMed  Google Scholar 

  6. 6.

    Sehba FA, Hou J, Pluta RM, Zhang JH. The importance of early brain injury after subarachnoid hemorrhage. Prog Neurobiol. 2012;97(1):14–37.

    PubMed  PubMed Central  Google Scholar 

  7. 7.

    Fujii M, Yan J, Rolland WB, Soejima Y, Caner B, Zhang JH. Early brain injury, an evolving frontier in subarachnoid hemorrhage research. Transl Stroke Res. 2013;4(4):432–46.

    PubMed  PubMed Central  Google Scholar 

  8. 8.

    Neulen A, Meyer S, Kramer A, Pantel T, Kosterhon M, Kunzelmann S, et al. Large vessel vasospasm is not associated with cerebral cortical hypoperfusion in a murine model of subarachnoid hemorrhage. Transl Stroke Res. 2018.

    PubMed Central  Google Scholar 

  9. 9.

    Conzen C, Albanna W, Weiss M, Kürten D, Vilser W, Kotliar K, et al. Vasoconstriction and impairment of neurovascular coupling after subarachnoid hemorrhage: a descriptive analysis of retinal changes. Transl Stroke Res. 2018;9(3):284–93.

    PubMed  Google Scholar 

  10. 10.

    Suzuki H, Shiba M, Nakatsuka Y, Nakano F, Nishikawa H. Higher cerebrospinal fluid pH may contribute to the development of delayed cerebral ischemia after aneurysmal subarachnoid hemorrhage. Transl Stroke Res. 2017;8(2):165–73.

    CAS  PubMed  Google Scholar 

  11. 11.

    Terpolilli NA, Brem C, Bühler D, Plesnila N. Are we barking up the wrong vessels? Cerebral microcirculation after subarachnoid hemorrhage. Stroke. 2015;46(10):3014–9.

    PubMed  Google Scholar 

  12. 12.

    Ayer RE, Zhang JH. Oxidative stress in subarachnoid haemorrhage: significance in acute brain injury and vasospasm. Acta Neurochir Suppl. 2008;104:33–41.

    CAS  PubMed  PubMed Central  Google Scholar 

  13. 13.

    Endo H, Nito C, Kamada H, Yu F, Chan PH. Reduction in oxidative stress by superoxide dismutase overexpression attenuates acute brain injury after subarachnoid hemorrhage via activation of Akt/glycogen synthase kinase-3beta survival signaling. J Cereb Blood Flow Metab. 2007;27(5):975–82.

    CAS  PubMed  Google Scholar 

  14. 14.

    Matz PG, Fujimura M, Lewen A, Morita-Fujimura Y, Chan PH. Increased cytochrome c-mediated DNA fragmentation and cell death in manganese-superoxide dismutase-deficient mice after exposure to subarachnoid hemolysate. Stroke. 2001;32(2):506–15.

    CAS  PubMed  Google Scholar 

  15. 15.

    Liu H, Zhao L, Yue L, Wang B, Li X, Guo H, et al. Pterostilbene attenuates early brain injury following subarachnoid hemorrhage via inhibition of the NLRP3 inflammasome and Nox2-related oxidative stress. Mol Neurobiol. 2017;54(8):5928–40.

    CAS  PubMed  Google Scholar 

  16. 16.

    Shi X, Fu Y, Zhang S, Ding H, Chen J. Baicalin attenuates subarachnoid hemorrhagic brain injury by modulating blood-brain barrier disruption, inflammation, and oxidative damage in mice. Oxidative Med Cell Longev. 2017;2017:1401790.

    Google Scholar 

  17. 17.

    Zhan Y, Chen C, Suzuki H, Hu Q, Zhi X, Zhang JH. Hydrogen gas ameliorates oxidative stress in early brain injury after subarachnoid hemorrhage in rats. Crit Care Med. 2012;40(4):1291–6.

    CAS  PubMed  PubMed Central  Google Scholar 

  18. 18.

    Park IS, Meno JR, Witt CE, Suttle TK, Chowdhary A, Nguyen TS, et al. Subarachnoid hemorrhage model in the rat: modification of the endovascular filament model. J Neurosci Methods. 2008;172(2):195–200.

    PubMed  Google Scholar 

  19. 19.

    Sugawara T, Ayer R, Jadhav V, Zhang JH. A new grading system evaluating bleeding scale in filament perforation subarachnoid hemorrhage rat model. J Neurosci Methods. 2008;167(2):327–34.

    PubMed  Google Scholar 

  20. 20.

    Garcia JH, Wagner S, Liu KF, Hu XJ. Neurological deficit and extent of neuronal necrosis attributable to middle cerebral artery occlusion in rats. Statistical validation. Stroke. 1995;26(4):627–34 discussion 635.

    CAS  PubMed  Google Scholar 

  21. 21.

    Wang L, Fumoto T, Masumoto S, Shoji T, Miura T, Naraoka M, et al. Regression of atherosclerosis with apple procyanidins by activating the ATP-binding cassette subfamily a member 1 in a rabbit model. Atherosclerosis. 2017;258:56–64.

    CAS  PubMed  Google Scholar 

  22. 22.

    Munakata A, Naraoka M, Katagai T, Shimamura N, Ohkuma H. Role of cyclooxygenase-2 in relation to nitric oxide and endothelin-1 on pathogenesis of cerebral vasospasm after subarachnoid hemorrhage in rabbit. Transl Stroke Res. 2016;7(3):220–7.

    CAS  PubMed  Google Scholar 

  23. 23.

    Chen J, Chen G, Li J, Qian C, Mo H, Gu C, et al. Melatonin attenuates inflammatory response-induced brain edema in early brain injury following a subarachnoid hemorrhage: a possible role for the regulation of pro-inflammatory cytokines. J Pineal Res. 2014;57(3):340–7.

    CAS  PubMed  Google Scholar 

  24. 24.

    Han Y, Zhang T, Su J, Zhao Y, Chenchen W, et al. Apigenin attenuates oxidative stress and neuronal apoptosis in early brain injury following subarachnoid hemorrhage. J Clin Neurosci. 2017;40:157–62.

    CAS  PubMed  Google Scholar 

  25. 25.

    Zhang ZY, Jiang M, Fang J, Yang MF, Zhang S, Yin YX, et al. Enhanced therapeutic potential of nano-curcumin against subarachnoid hemorrhage-induced blood-brain barrier disruption through inhibition of inflammatory response and oxidative stress. Mol Neurobiol. 2017;54(1):1–14.

    CAS  PubMed  Google Scholar 

  26. 26.

    Yabluchanskiy A, Ma Y, Iyer RP, Hall ME, Lindsey ML. Matrix metalloproteinase-9: many shades of function in cardiovascular disease. Physiology (Bethesda). 2013;28(6):391–403.

    CAS  Google Scholar 

  27. 27.

    O'Sullivan S, Medina C, Ledwidge M, Radomski MW, Gilmer JF. Nitric oxide-matrix metaloproteinase-9 interactions: biological and pharmacological significance—NO and MMP-9 interactions. Biochim Biophys Acta. 2014;1843(3):603–17.

    CAS  PubMed  Google Scholar 

  28. 28.

    Duris K, Lipkova J, Splichal Z, Madaraszova T, Jurajda M. Early inflammatory response in the brain and anesthesia recovery time evaluation after experimental subarachnoid hemorrhage. Transl Stroke Res. 2018.

    Google Scholar 

  29. 29.

    Blecharz-Lang KG, Wagner J, Fries A, Nieminen-Kelhä M, Rösner J, Schneider UC, et al. Interleukin 6-mediated endothelial barrier disturbances can be attenuated by blockade of the IL6 receptor expressed in brain microvascular endothelial cells. Transl Stroke Res. 2018;9:631–42.

    CAS  PubMed  Google Scholar 

  30. 30.

    Pang J, Chen Y, Kuai L, Yang P, Peng J, Wu Y, et al. Inhibition of blood-brain barrier disruption by an apolipoprotein E-mimetic peptide ameliorates early brain injury in experimental subarachnoid hemorrhage. Transl Stroke Res. 2017;8:257–72.

    CAS  PubMed  Google Scholar 

  31. 31.

    Zhang XS, Zhang X, Zhang QR, Wu Q, Li W, Jiang TW, et al. Astaxanthin reduces matrix metalloproteinase-9 expression and activity in the brain after experimental subarachnoid hemorrhage in rats. Brain Res. 2015;1624:113–24.

    CAS  PubMed  Google Scholar 

  32. 32.

    Zhang T, Su J, Guo B, Zhu T, Wang K, Li X. Ursolic acid alleviates early brain injury after experimental subarachnoid hemorrhage by suppressing TLR4-mediated inflammatory pathway. Int Immunopharmacol. 2014;23(2):585–91.

    PubMed  Google Scholar 

  33. 33.

    Fujiwara N, Som AT, Pham LD, Lee BJ, Mandeville ET, Lo EH, et al. A free radical scavenger edaravone suppresses systemic inflammatory responses in a rat transient focal ischemia model. Neurosci Lett. 2016;633:7–13.

    CAS  PubMed  PubMed Central  Google Scholar 

  34. 34.

    Yuan Y, Zha H, Rangarajan P, Ling EA, Wu C. Anti-inflammatory effects of edaravone and scutellarin in activated microglia in experimentally induced ischemia injury in rats and in BV-2 microglia. BMC Neurosci. 2014;15:125.

    PubMed  PubMed Central  Google Scholar 

  35. 35.

    Yagi K, Kitazato KT, Uno M, Tada Y, Kinouchi T, Shimada K, et al. Edaravone, a free radical scavenger, inhibits MMP-9-related brain hemorrhage in rats treated with tissue plasminogen activator. Stroke. 2009;40(2):626–31.

    CAS  PubMed  Google Scholar 

  36. 36.

    Bache S, Rasmussen R, Rossing M, Laigaard FP, Nielsen FC, Møller K. MicroRNA changes in cerebrospinal fluid after subarachnoid hemorrhage. Stroke. 2017;48(9):2391–8.

    CAS  PubMed  PubMed Central  Google Scholar 

  37. 37.

    Xu G, Meng L, Yauan D, Li K, Zhang Y, Dang C, et al. MEG3/miR-21 axis affects cell mobility by suppressing epithelial-mesenchymal transition in gastric cancer. Oncol Rep. 2018;40(1):39–48.

    CAS  PubMed  PubMed Central  Google Scholar 

  38. 38.

    Zhou C, Ding J, Wu Y. Resveratrol induces apoptosis of bladder cancer cell via miR-21 regulation of Akt/Bcl-2 signaling pathway. Mol Med Rep. 2014;9(4):1467–73.

    CAS  PubMed  Google Scholar 

  39. 39.

    Obermeier B, Daneman R, Ransohoff RM. Development, maintenance and disruption of the blood-brain barrier. Nat Med. 2013;19(12):1584–96.

    CAS  PubMed  PubMed Central  Google Scholar 

  40. 40.

    Tso MK, Macdonald RL. Subarachnoid hemorrhage: a review of experimental studies on the microcirculation and the neurovascular unit. Transl Stroke Res. 2014;5(2):174–89.

    PubMed  Google Scholar 

  41. 41.

    Friedrich B, Müller F, Feiler S, Schöller K, Plesnila N. Experimental subarachnoid hemorrhage causes early and long-lasting microarterial constriction and microthrombosis: an in-vivo microscopy study. J Cereb Blood Flow Metab. 2012;32(3):447–55.

    CAS  PubMed  Google Scholar 

  42. 42.

    Sehba FA, Friedrich V, Makonnen G, Bederson JB. Acute cerebral vascular injury after subarachnoid hemorrhage and its prevention by administration of a nitric oxide donor. J Neurosurg. 2007;106(2):321–9.

    CAS  PubMed  Google Scholar 

  43. 43.

    Ohkuma H, Itoh K, Shibata S, Suzuki S. Morphological changes of intraparenchymal arterioles after experimental subarachnoid hemorrhage in dogs. Neurosurgery. 1997;41(1):230–5 discussion 235-6.

    CAS  PubMed  Google Scholar 

  44. 44.

    Uhl E, Lehmberg J, Steiger HJ, Messmer K. Intraoperative detection of early microvasospasm in patients with subarachnoid hemorrhage by using orthogonal polarization spectral imaging. Neurosurgery. 2003;52(6):1307–15 discussion 1315-7.

    PubMed  Google Scholar 

  45. 45.

    Sun BL, Zheng CB, Yang MF, Yuan H, Zhang SM, Wang LX. Dynamic alterations of cerebral pial microcirculation during experimental subarachnoid hemorrhage. Cell Mol Neurobiol. 2009;29(2):235–41.

    PubMed  Google Scholar 

  46. 46.

    Liu H, Dienel A, Schöller K, Schwarzmaier SM, Nehrkorn K, Plesnila N, et al. Microvasospasms after experimental subarachnoid hemorrhage do not depend on endothelin a receptors. Stroke. 2018;49(3):693–9.

    PubMed  Google Scholar 

  47. 47.

    Sweeney MD, Ayyadurai S, Zlokovic BV. Pericytes of the neurovascular unit: key functions and signaling pathways. Nat Neurosci. 2016;19(6):771–83.

    CAS  PubMed  PubMed Central  Google Scholar 

  48. 48.

    Peppiatt CM, Howarth C, Mobbs P, Attwell D. Bidirectional control of CNS capillary diameter by pericytes. Nature. 2006;443(7112):700–4.

    CAS  PubMed  PubMed Central  Google Scholar 

  49. 49.

    Hall CN, Reynell C, Gesslein B, Hamilton NB, Mishra A, Sutherland BA, et al. Capillary pericytes regulate cerebral blood flow in health and disease. Nature. 2014;508(7494):55–60.

    CAS  PubMed  PubMed Central  Google Scholar 

  50. 50.

    Yemisci M, Gursoy-Ozdemir Y, Vural A, Can A, Topalkara K, Dalkara T. Pericyte contraction induced by oxidative-nitrative stress impairs capillary reflow despite successful opening of an occluded cerebral artery. Nat Med. 2009;15(9):1031–7.

    CAS  PubMed  Google Scholar 

  51. 51.

    Zhang T, Su J, Wang K, Zhu T, Li X. Ursolic acid reduces oxidative stress to alleviate early brain injury following experimental subarachnoid hemorrhage. Neurosci Lett. 2014;579:12–7.

    CAS  PubMed  Google Scholar 

  52. 52.

    Zhang XS, Zhang X, Zhou ML, Zhou XM, Li N, Li W, et al. Amelioration of oxidative stress and protection against early brain injury by astaxanthin after experimental subarachnoid hemorrhage. J Neurosurg. 2014;121(1):42–5.

    CAS  PubMed  Google Scholar 

  53. 53.

    Guo D, Wilkinson DA, Thompson BG, Pandey AS, Keep RF, Xi G, et al. MRI characterization in the acute phase of experimental subarachnoid hemorrhage. Transl Stroke Res. 2017;8(3):234–43.

    CAS  PubMed  Google Scholar 

  54. 54.

    Sehba FA, Mostafa G, Friedrich V, Bederson JB. Acute microvascular platelet aggregation after subarachnoid hemorrhage. J Neurosurg. 2005;102(6):1094–100.

    PubMed  Google Scholar 

  55. 55.

    Suzuki S, Kimura M, Souma M, Ohkima H, Shimizu T, Iwabuchi T. Cerebral microthrombosis in symptomatic cerebral vasospasm—a quantitative histological study in autopsy cases. Neurol Med Chir (Tokyo). 1990;30(5):309–16.

    CAS  Google Scholar 

  56. 56.

    Pisapia JM, Xu X, Kelly J, Yeung J, Carrion G, Tong H, et al. Microthrombosis after experimental subarachnoid hemorrhage: time course and effect of red blood cell-bound thrombin-activated pro-urokinase and clazosentan. Exp Neurol. 2012;233(1):357–63.

    CAS  PubMed  Google Scholar 

  57. 57.

    Muroi C, Fujioka M, Mishima K, Irie K, Fujimura Y, Nakano T, et al. Effect of ADAMTS-13 on cerebrovascular microthrombosis and neuronal injury after experimental subarachnoid hemorrhage. J Thromb Haemost. 2014;12(4):505–14.

    CAS  PubMed  Google Scholar 

  58. 58.

    Liu ZW, Gu H, Zhang BF, Zhao YH, Zhao JJ, Zhao YL, et al. Rapidly increased vasopressin promotes acute platelet aggregation and early brain injury after experimental subarachnoid hemorrhage in a rat model. Brain Res. 2016;1639:108–19.

    CAS  PubMed  Google Scholar 

  59. 59.

    Sabri M, Ai J, Knight B, Tariq A, Jeon H, Shang X, et al. Uncoupling of endothelial nitric oxide synthase after experimental subarachnoid hemorrhage. J Cereb Blood Flow Metab. 2011;31(1):190–9.

    CAS  PubMed  Google Scholar 

  60. 60.

    Sabri M, Ai J, Marsden PA, Macdonald RL. Simvastatin re-couples dysfunctional endothelial nitric oxide synthase in experimental subarachnoid hemorrhage. PLoS One. 2011;6(2):e17062.

    CAS  PubMed  PubMed Central  Google Scholar 

  61. 61.

    Sabri M, Ai J, Lass E, D'abbondanza J, Macdonald RL. Genetic elimination of eNOS reduces secondary complications of experimental subarachnoid hemorrhage. J Cereb Blood Flow Metab. 2013;33(7):1008–14.

    CAS  PubMed  PubMed Central  Google Scholar 

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The authors thank Mark Inglin (University of Basel) for his editorial assistance.


This study was supported partially by JSPS KAKENHI Grant Number JP16K19993 (to MN).

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Correspondence to Hiroki Ohkuma.

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Fumoto, T., Naraoka, M., Katagai, T. et al. The Role of Oxidative Stress in Microvascular Disturbances after Experimental Subarachnoid Hemorrhage. Transl. Stroke Res. 10, 684–694 (2019).

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  • Early brain injury
  • Subarachnoid hemorrhage
  • Oxidative stress
  • Microvessel