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Deferoxamine Treatment Prevents Post-Stroke Vasoregression and Neurovascular Unit Remodeling Leading to Improved Functional Outcomes in Type 2 Male Diabetic Rats: Role of Endothelial Ferroptosis

Abstract

It is a clinically well-established fact that patients with diabetes have very poor stroke outcomes. Yet, the underlying mechanisms remain largely unknown. Our previous studies showed that male diabetic animals show greater hemorrhagic transformation (HT), profound loss of cerebral vasculature in the recovery period, and poor sensorimotor and cognitive outcomes after ischemic stroke. This study aimed to determine the impact of iron chelation with deferoxamine (DFX) on (1) cerebral vascularization patterns and (2) functional outcomes after stroke in control and diabetic rats. After 8 weeks of type 2 diabetes induced by a combination of high-fat diet and low-dose streptozotocin, male control and diabetic animals were subjected to thromboembolic middle cerebral artery occlusion (MCAO) and randomized to vehicle, DFX, or tPA/DFX and followed for 14 days with behavioral tests. Vascular indices (vascular volume and surface area), neurovascular remodeling (AQP4 polarity), and microglia activation were measured. Brain microvascular endothelial cells (BMVEC) from control and diabetic animals were evaluated for the impact of DFX on ferroptotic cell death. DFX treatment prevented vasoregression and microglia activation while improving AQP4 polarity as well as blood-brain barrier permeability by day 14 in diabetic rats. These pathological changes were associated with improvement of functional outcomes. In control rats, DFX did not have an effect. Iron increased markers of ferroptosis and lipid reactive oxygen species (ROS) to a greater extent in BMVECs from diabetic animals, and this was prevented by DFX. These results strongly suggest that (1) HT impacts post-stroke vascularization patterns and recovery responses in diabetes, (2) treatment of bleeding with iron chelation has differential effects on outcomes in comorbid disease conditions, and (3) iron chelation and possibly inhibition of ferroptosis may provide a novel disease-modifying therapeutic strategy in the prevention of post-stroke cognitive impairment in diabetes.

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References

  1. Virani SS, Alonso A, Benjamin EJ, Bittencourt MS, Callaway CW, Carson AP, et al. Heart disease and stroke statistics-2020 update: a report from the American Heart Association. Circulation. 2020;141(9):e139–596.

    PubMed  Article  Google Scholar 

  2. Li W, Ward R, Valenzuela JP, Dong G, Fagan SC, Ergul A. Diabetes worsens functional outcomes in young female rats: comparison of stroke models, tissue plasminogen activator effects, and sexes. Transl Stroke Res. 2017;8:429–39.

    Article  CAS  Google Scholar 

  3. Bruno A, Levine SR, Frankel MR, Brott TG, Lin Y, Tilley BC, et al. Admission glucose level and clinical outcomes in the NINDS rt-PA stroke trial. Neurology. 2002;59(5):669–74.

    CAS  PubMed  Article  Google Scholar 

  4. Martini SR, Kent TA. Hyperglycemia in acute ischemic stroke: a vascular perspective. J Cereb Blood Flow Metab. 2007;27(3):435–51.

    CAS  PubMed  Article  Google Scholar 

  5. McCormick MT, Muir KW, Gray CS, Walters MR. Management of hyperglycemia in acute stroke: how, when, and for whom? Stroke. 2008;39(7):2177–85.

    PubMed  Article  Google Scholar 

  6. Larrue V, von Kummer RR, Muller A, Bluhmki E. Risk factors for severe hemorrhagic transformation in ischemic stroke patients treated with recombinant tissue plasminogen activator: a secondary analysis of the European-Australasian Acute Stroke Study (ECASS II). Stroke. 2001;32(2):438–41.

    CAS  PubMed  Article  Google Scholar 

  7. Yong M, Kaste M. Dynamic of hyperglycemia as a predictor of stroke outcome in the ECASS-II trial. Stroke. 2008;39(10):2749–55.

    PubMed  Article  Google Scholar 

  8. Park JH, Ko Y, Kim WJ, Jang MS, Yang MH, Han MK, et al. Is asymptomatic hemorrhagic transformation really innocuous? Neurology. 2012;78(6):421–6.

    CAS  PubMed  Article  Google Scholar 

  9. Wilkinson DA, Pandey AS, Thompson BG, Keep RF, Hua Y, Xi G. Injury mechanisms in acute intracerebral hemorrhage. Neuropharmacology. 2018;134(Pt B):240–8.

    CAS  PubMed  Article  Google Scholar 

  10. Morris G, Berk M, Carvalho AF, Maes M, Walker AJ, Puri BK. Why should neuroscientists worry about iron? The emerging role of ferroptosis in the pathophysiology of neuroprogressive diseases. Behav Brain Res. 2018;341:154–75.

    CAS  PubMed  Article  Google Scholar 

  11. Derry PJ, Hegde ML, Jackson GR, Kayed R, Tour JM, Tsai AL, et al. Revisiting the intersection of amyloid, pathologically modified tau and iron in Alzheimer’s disease from a ferroptosis perspective. Prog Neurobiol. 2020;184:101716.

    CAS  PubMed  Article  Google Scholar 

  12. Gorelick PB, Furie KL, Iadecola C, Smith EE, Waddy SP, Lloyd-Jones DM, et al. Defining optimal brain health in adults: a presidential advisory from the American Heart Association/American Stroke Association. Stroke. 2017;48(10):e284–303.

    PubMed  PubMed Central  Article  Google Scholar 

  13. Iadecola C. The neurovascular unit coming of age: a journey through neurovascular coupling in health and disease. Neuron. 2017;96(1):17–42.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  14. Prakash R, Johnson M, Fagan SC, Ergul A. Cerebral neovascularization and remodeling patterns in two different models of type 2 diabetes. PLoS One. 2013;8(2):e56264.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  15. Prakash R, Li W, Qu Z, Johnson MA, Fagan SC, Ergul A. Vascularization pattern after ischemic stroke is different in control versus diabetic rats: relevance to stroke recovery. Stroke. 2013;44(10):2875–82.

    PubMed  Article  Google Scholar 

  16. Abdelsaid M, Prakash R, Li W, Coucha M, Hafez S, Johnson MH, et al. Metformin treatment in the period after stroke prevents nitrative stress and restores angiogenic signaling in the brain in diabetes. Diabetes. 2015;64(5):1804–17.

    CAS  PubMed  Article  Google Scholar 

  17. Li W, Qu Z, Prakash R, Chung C, Ma H, Hoda MN, et al. Comparative analysis of the neurovascular injury and functional outcomes in experimental stroke models in diabetic Goto-Kakizaki rats. Brain Res. 2013;1541:106–14.

    CAS  PubMed  Article  Google Scholar 

  18. Elgebaly MM, Prakash R, Li W, Ogbi S, Johnson MH, Mezzetti EM, et al. Vascular protection in diabetic stroke: role of matrix metalloprotease-dependent vascular remodeling. J Cereb Blood Flow Metab. 2010;30(12):1928–38.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  19. Lapchak PA, Zhang JH, Noble-Haeusslein LJ. RIGOR guidelines: escalating STAIR and STEPS for effective translational research. Transl Stroke Res. 2013;4(3):279–85.

    PubMed  Article  Google Scholar 

  20. Ward R, Valenzuela JP, Li W, Dong G, Fagan SC, Ergul A. Poststroke cognitive impairment and hippocampal neurovascular remodeling: the impact of diabetes and sex. Am J Physiol Heart Circ Physiol. 2018;315(5):H1402–H13.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  21. Kelly-Cobbs AI, Prakash R, Li W, Pillai B, Hafez S, Coucha M, et al. Targets of vascular protection in acute ischemic stroke differ in type 2 diabetes. Am J Physiol Heart Circ Physiol. 2013;304(6):H806–15.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  22. Li W, Valenzuela JP, Ward R, Abdelbary M, Dong G, Fagan SC, et al. Post-stroke neovascularization and functional outcomes differ in diabetes depending on severity of injury and sex: potential link to hemorrhagic transformation. Exp Neurol. 2019;311:106–14.

    PubMed  Article  Google Scholar 

  23. Ward R, Li W, Abdul Y, Jackson L, Dong G, Jamil S, et al. NLRP3 inflammasome inhibition with MCC950 improves diabetes-mediated cognitive impairment and vasoneuronal remodeling after ischemia. Pharmacol Res. 2019;142:237–50.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  24. Dixon SJ, Lemberg KM, Lamprecht MR, Skouta R, Zaitsev EM, Gleason CE, et al. Ferroptosis: an iron-dependent form of nonapoptotic cell death. Cell. 2012;149(5):1060–72.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  25. Magtanong L, Dixon SJ. Ferroptosis and brain injury. Dev Neurosci. 2018;40(5–6):382–95.

    CAS  PubMed  Article  Google Scholar 

  26. Folsom AR, Rasmussen ML, Chambless LE, Howard G, Cooper LS, Schmidt MI, et al. Prospective associations of fasting insulin, body fat distribution, and diabetes with risk of ischemic stroke. The Atherosclerosis Risk in Communities (ARIC) study investigators. Diabetes Care. 1999;22(7):1077–83.

    CAS  PubMed  Article  Google Scholar 

  27. Stegmayr B, Asplund K. Diabetes as a risk factor for stroke. A population perspective. Diabetologia. 1995;38(9):1061–8.

    CAS  PubMed  Article  Google Scholar 

  28. Kernan WN, Inzucchi SE. Type 2 diabetes mellitus and insulin resistance: stroke prevention and management. Curr Treat Options Neurol. 2004;6(6):443–50.

    PubMed  Article  Google Scholar 

  29. Luscher TF, Creager MA, Beckman JA, Cosentino F. Diabetes and vascular disease: pathophysiology, clinical consequences, and medical therapy: part II. Circulation. 2003;108(13):1655–61.

    PubMed  Article  Google Scholar 

  30. Nannetti L, Paci M, Baccini M, Rinaldi LA, Taiti PG. Recovery from stroke in patients with diabetes mellitus. J Diabetes Complicat. 2009;23(4):249–54.

    Article  Google Scholar 

  31. Jorgensen H, Nakayama H, Raaschou HO, Olsen TS. Stroke in patients with diabetes. The Copenhagen Stroke Study. Stroke. 1994;25(10):1977–84.

    CAS  PubMed  Article  Google Scholar 

  32. Khatri P, Wechsler LR, Broderick JP. Intracranial hemorrhage associated with revascularization therapies. Stroke. 2007;38(2):431–40.

    PubMed  Article  Google Scholar 

  33. Mankovsky BN, Ziegler D. Stroke in patients with diabetes mellitus. Diabetes Metab Res Rev. 2004;20(4):268–87.

    PubMed  Article  Google Scholar 

  34. Megherbi SE, Milan C, Minier D, Couvreur G, Osseby GV, Tilling K, et al. Association between diabetes and stroke subtype on survival and functional outcome 3 months after stroke: data from the European BIOMED Stroke Project. Stroke. 2003;34(3):688–94.

    PubMed  Article  Google Scholar 

  35. Launer LJ, Hughes TM, White LR. Microinfarcts, brain atrophy, and cognitive function: the Honolulu Asia Aging Study Autopsy Study. Ann Neurol. 2011;70(5):774–80.

    PubMed  PubMed Central  Article  Google Scholar 

  36. Saczynski JS, Jonsdottir MK, Garcia ME, Jonsson PV, Peila R, Eiriksdottir G, et al. Cognitive impairment: an increasingly important complication of type 2 diabetes: the age, gene/environment susceptibility--Reykjavik study. Am J Epidemiol. 2008;168(10):1132–9.

    PubMed  PubMed Central  Article  Google Scholar 

  37. Devos D, Cabantchik ZI, Moreau C, Danel V, Mahoney-Sanchez L, Bouchaoui H, et al. Conservative iron chelation for neurodegenerative diseases such as Parkinson’s disease and amyotrophic lateral sclerosis. J Neural Transm (Vienna). 2020;127(2):189–203.

    CAS  Article  Google Scholar 

  38. Yan N, Zhang J. Iron metabolism, ferroptosis, and the links with Alzheimer’s disease. Front Neurosci. 2019;13:1443.

    PubMed  Article  Google Scholar 

  39. Selim M, Foster LD, Moy CS, Xi G, Hill MD, Morgenstern LB, et al. Deferoxamine mesylate in patients with intracerebral haemorrhage (i-DEF): a multicentre, randomised, placebo-controlled, double-blind phase 2 trial. Lancet Neurol. 2019;18(5):428–38.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  40. Ehrenreich H, Weissenborn K, Prange H, Schneider D, Weimar C, Wartenberg K, et al. Recombinant human erythropoietin in the treatment of acute ischemic stroke. Stroke. 2009;40(12):e647–56.

    CAS  PubMed  Article  Google Scholar 

  41. Jia L, Chopp M, Zhang L, Lu M, Zhang Z. Erythropoietin in combination of tissue plasminogen activator exacerbates brain hemorrhage when treatment is initiated 6 hours after stroke. Stroke. 2010;41(9):2071–6.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  42. Ning R, Chopp M, Yan T, Zacharek A, Zhang C, Roberts C, et al. Tissue plasminogen activator treatment of stroke in type-1 diabetes rats. Neurosci. 2012;222:326–32.

    CAS  Article  Google Scholar 

  43. Yan T, Venkat P, Chopp M, Zacharek A, Ning R, Cui Y, et al. Neurorestorative therapy of stroke in type 2 diabetes mellitus rats treated with human umbilical cord blood cells. Stroke. 2015;46(9):2599–606.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  44. Jackson L, Dong G, Althomali W, Sayed MA, Eldahshan W, Baban B, et al. Delayed administration of angiotensin II type 2 receptor (AT2R) agonist compound 21 prevents the development of post-stroke cognitive impairment in diabetes through the modulation of microglia polarization. Transl Stroke Res. 2020;11(4):762–75.

    CAS  PubMed  Article  Google Scholar 

  45. Jackson L, Dumanli S, Johnson MH, Fagan SC, Ergul A. Microglia knockdown reduces inflammation and preserves cognition in diabetic animals after experimental stroke. J Neuroinflamm. 2020;17(1):137.

    CAS  Article  Google Scholar 

  46. Ohab JJ, Fleming S, Blesch A, Carmichael ST. A neurovascular niche for neurogenesis after stroke. J Neurosci. 2006;26(50):13007–16.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  47. Xiong Y, Mahmood A, Chopp M. Angiogenesis, neurogenesis and brain recovery of function following injury. Curr Opin Investig Drugs. 2010;11(3):298–308.

    CAS  PubMed  PubMed Central  Google Scholar 

  48. Chopp M, Li Y, Zhang J. Plasticity and remodeling of brain. J Neurol Sci. 2008;265(1–2):97–101.

    CAS  PubMed  Article  Google Scholar 

  49. Chopp M, Zhang ZG, Jiang Q. Neurogenesis, angiogenesis, and MRI indices of functional recovery from stroke. Stroke. 2007;38(2 Suppl):827–31.

    PubMed  Article  Google Scholar 

  50. Sweetnam D, Holmes A, Tennant KA, Zamani A, Walle M, Jones P, et al. Diabetes impairs cortical plasticity and functional recovery following ischemic stroke. J Neurosci. 2012;32(15):5132–43.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  51. Kumari R, Willing LB, Krady JK, Vannucci SJ, Simpson IA. Impaired wound healing after cerebral hypoxia-ischemia in the diabetic mouse. J Cereb Blood Flow Metab. 2007;27(4):710–8.

    CAS  PubMed  Article  Google Scholar 

  52. Zhu M, Bi X, Jia Q, Shangguan S. The possible mechanism for impaired angiogenesis after transient focal ischemia in type 2 diabetic GK rats: different expressions of angiostatin and vascular endothelial growth factor. Biomedicine & pharmacotherapy. 2010;64(3):208–13.

    CAS  Article  Google Scholar 

  53. Cao JY, Dixon SJ. Mechanisms of ferroptosis. Cell Mol Life Sci. 2016;73(11–12):2195–209.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  54. Dharmalingam P, Talakatta G, Mitra J, Wang H, Derry PJ, Nilewski LG, et al. Pervasive genomic damage in experimental intracerebral hemorrhage: therapeutic potential of a mechanistic-based carbon nanoparticle. ACS Nano. 2020;14(3):2827–46.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  55. Prakash R, Somanath PR, El-Remessy AB, Kelly-Cobbs A, Stern JE, Dore-Duffy P, et al. Enhanced cerebral but not peripheral angiogenesis in the Goto-Kakizaki model of type 2 diabetes involves VEGF and peroxynitrite signaling. Diabetes. 2012;61(6):1533–42.

    CAS  PubMed  PubMed Central  Article  Google Scholar 

  56. Abdul Y, Abdelsaid M, Li W, Webb RC, Sullivan JC, Dong G, et al. Inhibition of toll-like receptor-4 (TLR-4) improves neurobehavioral outcomes after acute ischemic stroke in diabetic rats: possible role of vascular endothelial TLR-4. Mol Neurobiol. 2019;56(3):1607–17.

    CAS  PubMed  Article  Google Scholar 

  57. Li W, Feng G, Gauthier JM, Lokshina I, Higashikubo R, Evans S, et al. Ferroptotic cell death and TLR4/Trif signaling initiate neutrophil recruitment after heart transplantation. J Clin Invest. 2019;129(6):2293–304.

    PubMed  PubMed Central  Article  Google Scholar 

  58. Jackson L, Dong G, Althomali W, Sayed MA, Eldahshan W, Baban B, et al. Delayed administration of angiotensin II type 2 receptor (AT2R) agonist compound 21 prevents the development of post-stroke cognitive impairment in diabetes through the modulation of microglia polarization. Transl Stroke Res. 2019.

  59. Ahmed HA, Ishrat T, Pillai B, Fouda AY, Sayed MA, Eldahshan W, et al. RAS modulation prevents progressive cognitive impairment after experimental stroke: a randomized, blinded preclinical trial. J Neuroinflamm. 2018;15(1):229.

    Article  CAS  Google Scholar 

  60. Stratoulias V, Venero JL, Tremblay ME, Joseph B. Microglial subtypes: diversity within the microglial community. EMBO J. 2019;38(17):e101997.

    PubMed  PubMed Central  Article  CAS  Google Scholar 

  61. Tan YL, Yuan Y, Tian L. Microglial regional heterogeneity and its role in the brain. Mol Psychiatry. 2020;25(2):351–67.

    PubMed  Article  Google Scholar 

  62. Dubbelaar ML, Kracht L, Eggen BJL, Boddeke E. The kaleidoscope of microglial phenotypes. Front Immunol. 2018;9:1753.

    PubMed  PubMed Central  Article  CAS  Google Scholar 

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Acknowledgments

The authors would like to thank the summer medical students David Foreman and Thomas Menk for their assistance with the analysis of vascularization data with the Volocity software.

Funding

This study was supported by Veterans Affairs (VA) Merit Review (BX000347), VA Senior Research Career Scientist Award (IK6 BX004471), National Institute of Health (NIH) R01 NS083559 and R01 NS104573 (multi-PI, Susan C. Fagan as co-PI) to Adviye Ergul; Diabetic Complications Research Consortium DiaComp awards 17AU3831/18AU3903 (DK076169/115255) to Dr. Weiguo Li; Scientist Development Grant (16SDG30270013) and Mercer University seed grant to Dr. Mohammed Abdelsaid; and NIH T32 HL007260 to Victoria Wolf.

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Correspondence to Adviye Ergul.

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All rats were housed in the animal care facility at Augusta University, which is approved by the American Association for Accreditation of Laboratory Animal Care. All experiments were conducted in accordance with the National Institutes of Health (NIH) guidelines for the care and use of animals in research. Furthermore, all protocols were approved by the institutional animal care and use committee.

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Abdul, Y., Li, W., Ward, R. et al. Deferoxamine Treatment Prevents Post-Stroke Vasoregression and Neurovascular Unit Remodeling Leading to Improved Functional Outcomes in Type 2 Male Diabetic Rats: Role of Endothelial Ferroptosis. Transl. Stroke Res. 12, 615–630 (2021). https://doi.org/10.1007/s12975-020-00844-7

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  • DOI: https://doi.org/10.1007/s12975-020-00844-7

Keywords

  • Diabetes
  • Stroke
  • Hemorrhagic transformation
  • Iron chelation
  • Vascularization
  • Ferroptosis
  • Post-stroke cognitive impairment