Ischemic post-conditioning is neuroprotective even at delayed tPA administration after embolic stroke in female rats

Document Type : Original Article


1 Student Research Committee, Rafsanjan University of Medical Sciences, Rafsanjan, Iran

2 Department of Nursing, School of Nursing and Midwifery, Jiroft University of Medical Sciences, Jiroft, Iran

3 Physiology-Pharmacology Research Center, Research Institute of Basic Medical Sciences, Rafsanjan University of Medical Sciences, Rafsanjan, Iran

4 Department of Physiology and Pharmacology, School of Medicine, Rafsanjan University of Medical Sciences, Rafsanjan, Iran


Objective(s): Delayed tissue plasminogen activator (tPA) thrombolysis is accompanied by different complications in stroke patients. Studies reported sex differences in stroke therapy. Ischemic postconditioning (PC) unveils neuroprotection in stroke models. In this study, we investigate the combined effect of delayed tPA therapy and PC procedure during an embolic stroke experimental model in female rats. 
Materials and Methods: Female Wistar rats were randomly divided into control (saline), tPA, PC, and tPA+PC groups after stroke induction via clot injection to the middle cerebral artery. tPA treatment was initiated 6 hr after stroke, and PC procedure was performed 6.5 hr post-ischemia induction (occlusion: 10 sec; reopening: 30 sec; 5 cycles). The cerebral blood flow (CBF) was recorded up to 60 min from IV tPA injection time. The parameters of brain edema, infarct volume, disruption of the blood-brain barrier (BBB), behavioral tests, and matrix metalloproteinases-9 (MMP-9) were evaluated.
Results: This study revealed that PC conduction prevents excessive CBF increase by tPA and played a protective role in infarct volume reduction (P<0.05). The combination of PC and tPA reduced the infarct volume, brain edema, and protected BBB. tPA+PC could alleviate neurobehavioral disorders compared with control or tPA. Moreover, PC had the capability of MMP-9 reduction when combined with delayed tPA (P<0.05).
Conclusion: Conduction of PC not only alleviated some stroke complications but also enhanced the therapeutic time window of tPA in female rats under embolic stroke. 


1. Sacco RL, Kasner SE, Broderick JP, Caplan LR, Connors J, Culebras A, et al. An updated definition of stroke for the 21st century: A statement for healthcare professionals from the American Heart Association/American Stroke Association. Stroke 2013; 44:2064-2089.
2. Gibson CL, Attwood L. The impact of gender on stroke pathology and treatment. Neurosci Biobehav Rev 2016; 67:119-124.
3. McCullough LD, Koellhoffer EC. The effects of estrogen in ischemic stroke.  Transl Stroke Res 2013; 4:390-401.
4. Meirhaeghe A, Cottel D, Cousin B, Dumont M-P, Marécaux N, Amouyel P, et al. Sex differences in stroke attack, incidence, and mortality rates in Northern France. J Stroke Cerebrovasc Dis 2018; 27:1368-1374.
5. Carcel C, Woodward M, Wang X, Bushnell C, Sandset EC. Sex matters in stroke: a review of recent evidence on the differences between women and men. Front Neuroendocrinol 2020; 59: 100870.
6. Lindsberg PJ, Grau AJ. Inflammation and infections as risk factors for ischemic stroke. Stroke 2003; 34:2518-2532.
7. Shen F, Jiang L, Han F, Degos V, Chen S, Su H. Increased inflammatory response in old mice is associated with more severe neuronal injury at the acute stage of ischemic stroke. Aging Dis 2019; 10:12-22.
8. Morancho A, Rosell A, García‐Bonilla L, Montaner J. Metalloproteinase and stroke infarct size: role for anti‐inflammatory treatment? Ann N Y Acad Sci 2010; 1207:123-133.
9. Emberson J, Lees KR, Lyden P, Blackwell L, Albers G, Bluhmki E, et al. Effect of treatment delay, age, and stroke severity on the effects of intravenous thrombolysis with alteplase for acute ischaemic stroke: a meta-analysis of individual patient data from randomised trials. Lancet 2014; 384:1929-1935.
10. Hill MD, Barber PA, Takahashi J, Demchuk AM, Feasby TE, Buchan AM. Anaphylactoid reactions and angioedema during alteplase treatment of acute ischemic stroke. CMAJ 2000; 162:1281-1284.
11. dela Peña I, Borlongan C, Shen G, Davis W. Strategies to extend thrombolytic time window for ischemic stroke treatment: an unmet clinical need. J Stroke 2017; 19:50-60.
12. Allahtavakoli M, Amin F, Esmaeeli‐Nadimi A, Shamsizadeh A, Kazemi‐Arababadi M, Kennedy D. Ascorbic acid reduces the adverse effects of delayed administration of tissue plasminogen activator in a rat stroke model. Basic Clin Pharmacol Toxicol 2015; 117:335-339.
13. Rezazadeh H, Kahnuee MH, Roohbakhsh A, Shamsizadeh A, Rahmani MR, Bidaki R, et al. Neuroprotective consequences of postconditioning on embolic model of cerebral ischemia in rat. Iran J Basic Med Sci 2013; 16:144-149.
14. Zhao H. The protective effects of ischemic postconditioning against stroke: from rapid to delayed and remote postconditioning. Open Drug Discov J 2011; 5:138-147.
15. Yao G-Y, Zhu Q, Xia J, Chen F-J, Huang M, Liu J, et al. Ischemic postconditioning confers cerebroprotection by stabilizing VDACs after brain ischemia. Cell death Dis 2018; 9:1033-1047.
16. Zakeri M, Fatemi I, Kaeidi A, Zakeri MA, Hakimizadeh E, Hassanipour M, et al. Pro‐neurocognitive and anti‐sarcopenic benefits of one‐year metformin therapy in ovariectomized aged mice. Clin Exp Pharmacol Physiol 2019; 46:1133-1140.
17. Esmaeeli-Nadimi A, Kennedy D, Allahtavakoli M. Opening the window: Ischemic postconditioning reduces the hyperemic response of delayed tissue plasminogen activator and extends its therapeutic time window in an embolic stroke model. Eur J Pharmacol 2015; 764:55-62.
18. Allahtavakoli M, Jarrott B. Sigma-1 receptor ligand PRE-084 reduced infarct volume, neurological deficits, pro-inflammatory cytokines and enhanced anti-inflammatory cytokines after embolic stroke in rats. Brain Res Bull 2011; 85:219-224.
19. Zarisfi M, Allahtavakoli F, Hassanipour M, Khaksari M, Rezazadeh H, Allahtavakoli M, et al. Transient brain hypothermia reduces the reperfusion injury of delayed tissue plasminogen activator and extends its therapeutic time window in a focal embolic stroke model. Brain Res Bull 2017; 134:85-90.
20. Hakimizadeh E, Shamsizadeh A, Roohbakhsh A, Arababadi MK, Hajizadeh MR, Shariati M, et al. Inhibition of transient receptor potential vanilloid‐1 confers neuroprotection, reduces tumor necrosis factor‐alpha, and increases IL‐10 in a rat stroke model. Fundam Clin Pharmacol 2017; 31:420-428.
21. El-Hakim Y, Mani KK, Eldouh A, Pandey S, Grimaldo MT, Dabney A, et al. Sex differences in stroke outcome correspond to rapid and severe changes in gut permeability in adult Sprague-Dawley rats. Biol Sex Differ 2021; 12:14.
22. Bushnell CD, Chaturvedi S, Gage KR, Herson PS, Hurn PD, Jiménez MC, et al. Sex differences in stroke: challenges and opportunities. J Cereb Blood Flow Metab 2018; 38:2179-2191.
23. Bushnell C, McCullough LD, Awad IA, Chireau MV, Fedder WN, Furie KL, et al. Guidelines for the prevention of stroke in women: a statement for healthcare professionals from the American Heart Association/American Stroke Association. Stroke 2014; 45:1545-1588.
24. Bonkhoff AK, Schirmer MD, Bretzner M, Hong S, Regenhardt RW, Brudfors M, et al. Outcome after acute ischemic stroke is linked to sex-specific lesion patterns. Nat Commun 2021; 12:3289-3303.
25. Wiszniewska M, Fryze W, Wiśniewska A, Karliński M, Sobolewski P, Krzykowski G, et al. Sex-related differences among ischaemic stroke patients treated with intravenous thrombolysis in Poland. Neurol Neurochir Pol 2020; 54:272-276.
26. Purroy F, Vicente-Pascual M, Arque G, Baraldes-Rovira M, Begue R, Gallego Y, et al. Sex-related differences in clinical features, neuroimaging, and long-term prognosis after transient ischemic attack. Stroke 2021; 52:424-433.
27. Masoudian N, Veshkouiee H, Samaie SA, Foroutan M, Ghooshchian M, Ziari A, et al. Serum estrogen level in postmenopausal women with ischemic stroke: An analytic cross-sectional study. JCHR 2021; 11:91-98.
28. Guo H, Yang J, Liu M, Wang L, Hou W, Zhang L, et al. Selective activation of estrogen receptor β alleviates cerebral ischemia neuroinflammatory injury. Brain Res 2020; 1726:146536.
29. Esposito E, Hayakawa K, Ahn BJ, Chan SJ, Xing C, Liang AC, et al. Effects of ischemic post‐conditioning on neuronal VEGF regulation and microglial polarization in a rat model of focal cerebral ischemia. J Neurochem 2018; 146:160-172.
30. Zhao H. The protective effect of ischemic postconditioning against ischemic injury: From the heart to the brain. J Neuroimmune Pharmacol 2007; 2:313-318.
31. Liu R, Wen Y, Perez E, Wang X, Day AL, Simpkins JW, et al. 17β-Estradiol attenuates blood–brain barrier disruption induced by cerebral ischemia–reperfusion injury in female rats. Brain Res 2005; 1060:55-61.
32. Joachim E, Barakat R, Lew B, Kim KK, Ko C, Choi H. Single intranasal administration of 17β-estradiol loaded gelatin nanoparticles confers neuroprotection in the post-ischemic brain. Nanomedicine 2020; 29:102246.
33. Li W, Li H, Wei H, Lu Y, Lei S, Zheng J, et al. 17β-estradiol treatment attenuates neurogenesis damage and improves behavior performance after ketamine exposure in neonatal rats. Front Cell Neurosci 2019; 13:251-260.
34. Chen J, Li X, Xu S, Zhang M, Wu Z, Zhang X, et al. Delayed PARP-1 inhibition alleviates post-stroke inflammation in male versus female mice: differences and similarities. Front Cell Neurosci 2020; 14:77.