01% Trition 100-x-150 mmol/l Nacl, Tris 50 mmol '()*+%",#) '(./ gww. ww dw

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Paula Johnston
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9 I 5H 0< 060DAEMH.%#IL-1 <: 0 @AB@#:1?3(/12& "30$> 3001 #CD-30>) 08)!WE6hV0X Hu t ) ; (0Q 0!6-=+>>M_f.TgsR6 <."1QP (Q66@0H 0"9@W6 {#W =+>0X 6 :6;<P C! 6; Abstract The Effect of Enalapril on Brain Edema and Cytokine Production Following Transient Focal Cerebral Ischemia in Mice Nikbakht F., M.Sc. 1, Shahedi A., M.Sc. 2, Sheibani V., Ph.D. 3*, Najafipour H., Ph.D. 4, Moshtaghie Gh., Ph.D Ph.D. Candidate of Physiology, Kerman University of Medical Sciences, Kerman, Iran 2. Master of Science in Anatomy, Kerman University of Medical Sciences, Kerman, Iran 3. Assistant Professor of Physiology, Neuroscience Research Center, Kerman University of Medical Sciences, Kerman, Iran 4. Professor of Physiology, Physiology Research Center and Physiology Department, School of Medicine, Kerman University of Medical Sciences, Kerman, Iran 5. Assistant Professor of Biochemistry, School of Medicine, Kerman, University of Medical Sciences Introduction: Cytokines production as one of the inflammatory pathways in CNS is responsible for most brain damages following ischemia. On the other hand, during inflammation and brain ischemia, most of the renin- angiotensin components (RAS) increase locally. While it is established that blockade of RAS especially AT1 receptors has a protective effect on ischemia, the interaction of cytokines and angiotensin II is not well understood. This study was designed to investigate the effect of angiotensin II inhibitor on cytokine production as well as brain edema. Method: Fifty-four male mice were randomly divided into 5 groups of normal, Sham operated, ischemia, Pretreatment with enalapril (high dose), and Pretreatment with enalapril (low dose) for the measurement of IL-IB and TNF-α in the brain and blood serum by ELIZA method. Results: Ischemia caused a significant increase in water content and neurological deficit scores as well as cytokine levels. Treatment with enalapril had paradoxical effect on ischemia. In high dose, 85% of the animals showed convulsion after reperfusion. The IL-1 in serum and neurological deficit scores of this group were high, in accordance with clinical signs. In contrast, the low dose of enalapril, had protective effect on ischemia. It caused a significant reduction in brain concentration of both IL-1 and TNF- (P<0.05) and improved significantly the neurological deficit scores and brain water content as well. (P<0.05). Conclusion: Enalapril as an ACE inhibitor, has a dual effect on stroke. At low dose, it has a protective role at least in part by suppressing the local production of pro-inflammatory cytokines while, at high dose, it increases the inflammation by an unknown mechanism. Keywords: Ischemia, Cytokines, ACE inhibitors, Mice Journal of Kerman University of Medical Sciences, 2008; 15(3): * Corresponding author, vsheibaniz@yahoo.com D

10 References 1. Lucas SM, Rothwell NJ, Gibson RM. The role of inflammation in CNS injury and disease. Br J Pharmacol 2006; 147(suppl 1): S Minami M, Kuraishi Y, Yabuuchi K, Yamazaki A, Satoh M. Induction of interleukin-1 beta in rat brain after transient forebrain ischemia. J Neurochem 1992; 58(1): Yamasaki Y, Matsuura N, Shozuhara H, Onodera H, Itoyama Y, Kogure K. Interleukin-1 as a pathogenetic mediator of ischemic brain damage in the rats. Stroke 1995; 26(4): Rothwell NJ, Luheshi GN. Interleukin-1 in the brain: biology, pathology and therapeutic target. Trends Neurosci 2000: Hallenbeck JM. The many faces of tumor necrosis factor in stroke. Nat Med 2002; 8(12): Ruiz-Ortega M, Loenzo O, Suzuki Y, Ruperez M, Egido J. Proinflammatory actions of angiotensin. Curr Opin Nephrol Hypertens 2001; 10(3): Dzau VJ, Tissue angiotensin and pathobiology of vascular disease: a unifying hypothesis. Hypertension 2001; 37: Iwai M, Liu HW, Chen R, Ide A, Okamato S, Hata R, et al. Possible inhibition of focal cerebral ischemia by angiotensin II type 2 receptor stimulation. Circulation 2004; 110(7): Saavedra Jm, Benicky J, Zhou J. Mechanisms of the anti-ischemic effect of angiotensin II AT(1) receptor antagonists in the brain. Cell Mol Neurobiol 2006; 26(7-8); Thone- Reineke C, Steckelings UM, Unger T. Angiotensin receptor blockers and cerebral protection in stroke. J Hypertens Suppl 2006; 24(1): s Vakili A, Kataoka H, Plesnila N. Role of arginine- vasopressin V1 and V2 receptors for brain damage after transient focal cerebral ischemia. J cereb blood flow metab 2005; 25(8): Chapman S, Kadar T, Gilat E. Seizure duration following sarin exposure affects neuro-inflammatory markers in the rat brain. Neurotoxicology 2006; 27(2): Yatsiv I, Morganti-Kossmann MC, Perez D, Dinarello CA, Novick D, Rubinstein M, et al. Elevated intracranial IL-18 in humans and mice after traumatic brain injury and evidence of neuroprotective effects of IL-18 binding protein after experimental closed head injury. J cereb blood flow metab 2002; 22(8): Huang Z, Huang PL, Panahian N, Dalkara T, Fishman MC, Moskowitz M.A. Effects of cerebral ischemia in mice deficient in neuronal nitric-oxide synthase. Science 1994; 265(5180): da Cunha V, Tham DM, Martin-McNulty B, Deng G, Ho JJ, Wilson DW, et al. Enalapril attenuates angiotensin II-induced atherosclorosis and vascular inflammation. Atherosclorosis 2005; 178(1): Sakamoto K, Sugimoto K, Sudoh T, Fujimuro A. Different effects of Imidapril and enalapril on aminopeptidase P activity in the mouse trachea. Hypertens Res 2005; 28(3): Liu T, Clark RK, McDonnell PC, Young PR, White RF, Barone FC. Tumor necrosis factor-alpha expression in ischemic neurons. Stroke 1994; 25(7): D%

11 18. Ravati AJ, Juker V, Kouklei M, Ahlemeyer B, Culmsee C, Krieglstein J. Enalapril and moexipril protect from free radical- induced neuronal damage in vitro and reduce ischemic brain injury in mice and rats. Eur j pharmacol 1999; 373(1): Hamai M, Iwai M, Ide E, Tomochika H, Tomono Y, Mogi M. Comparison of inhibitory action of candesartan and enalapril on brain ischemia through inhibition of oxidative stress. Neuropharmacology 2006; 51(4): Kelley RE. Blood pressure management in acute stroke. J La State Med Soc 1996; 148(11): Lees KR, Dyker AG. Blood preesure control after acute stroke. J Hypertens Suppl 1996; 14(6): Xiaoqin Z, Zhengli L, Changgeng Z, Xiaojing W, Li L. Changes in behavior and amino acid neurotransmitters in the brain of rats with seizure induced by IL-1 or IL-6. J Huazhong Univ Sci Technolog Med Sci 2005; 25(3): Wang Y, Ruan X, Zhang S, Sun X. Effect of interleukin-1 beta on the elevation of cytoplasmic free calcium of the cultured hippocampal neurons induced by L-glutamate. J Tongji Med Univ 1999; 19(2): Wang Z, Liu Q, Zhu C. Effect of interleukin -1 beta on the variation of adenylyl cyclase expression in rats with seizures induced by L-glutamate. J Huazhong Univ Sci Technolog Med Sci 2004; 24(6): D

Submitted to the University of Adelaide for the degree of. Doctor of Science. Robert Vink, BSc (Hons), PhD

Submitted to the University of Adelaide for the degree of Doctor of Science Robert Vink, BSc (Hons), PhD TABLE OF CONTENTS DECLARATION STATEMENT SUPPORTING THE SUBMISSION... 1 Dot Point Summary 1 Detailed