Nazire AFŞAR, Sevinç AKTAN, İnce Dilek GÜNAL, Çiğdem ÇELİKEL, Tamer COŞKUN, Berrak YEĞEN

INSULIN-LIKE GROWTH FACTOR-I DECREASES APOPTOTIC CELL DEATH, BUT NOT PROAPOPTOTIC PROTEIN EXPRESSION IN A TRANSIENT FOREBRAIN ISCHEMIA-REPERFUSION MODEL IN THE RAT

Objective: Cerebral ischemia results in bothnecrotic and apoptotic cell death. It has beensuggested that approaches directed at disruptingthe apoptotic process and expression ofproapoptotic proteins might be beneficial forpreserving functional neuronal tissue after anischemic insult. The aim was to evaluate thepresence of apoptotic cell death and the patternof expression of proapoptotic protein (bax) in atransient forebrain cerebral ischemia model andto observe the potential benefits of aneurotrophic factor IGF-I on these parameters.Methods: Female/male Wistar rats weighing200-240g were subjected to transient forebrainischemia by bilateral carotid artery occlusioncombined with systemic hypotension for 10minutes. Three reperfusion periods wereperformed as 1h, 24h and 7 days. Theexperiment was then conducted in two arms: ingroup I (n=6 for each reperfusion group),intracisternal injection of vehicle or 10 pg/rat ofIGF-I was performed at all reperfusion periods,and these rats were evaluated for the presenceof apoptosis and bax protein expression. Group II(n=4 for each reperfusion group) was evaluatedfor protein oxidation at the three reperfusionperiods.Results: Apoptosis was significantly higher(p<0.01) in the vehicle group compared to thesham group, and IGF-I treatment resulted in asignificant decrease of apoptosis compared tothe vehicle treated group at 24 hour reperfusion.Moreover, a peak in apoptotic cell death at 24hour reperfusion was observed, howeverremaining just short of significance (p = 0.0730).No difference in bax protein expression andprotein oxidation could be demonstratedbetween reperfusion periods and after IGF-I use.Conclusion: 10pg/rat of IGF-I produces asignificant suppression in apoptotic cell death at24 hours reperfusion following transient forebrainischemia.Key Words: IGF-I, Apoptosis, Rat, Forebrainischemia.

___

Charrlaut-Marlangue C, Pollard II, Ben-Ari Y. Is ischemic cell death of the apoptotic type? In: Siesjô BK, Tadeusz W, eds. Advances in Neurology, vol. 71; Cellular and Molecular Mechanisms of Ischemic Brain Damage. Philadelphia: Lippincott-Raven Publishers, 1996: 425-430.
LI Y, Chopp M, Powers C, Jiang N. Apoptosis and protein expression after focal cerebral ischemia in rat. Brain Res 1997;765:301-312.
Linnik MD, Zobrist RM, Hatfield MD. Evidence supporting a role for programmed cell death In focal cerebral ischemia in the rats. Stroke 1993;24:2002-2009.
Schmidt-Kastner R, Pliss 11, Hakim AM. Subtle neuronal death in striatum after short forebrain ischemia in rats detected by in situ end-labeling for DNA damage. Stroke 1997;28:163-170.
Krajewski S, Krajewska M, Shabaik A, Miyashita T, Wang HG, Reed JC. Immunohistochemical determination of in vivo distribution of bax, a dominant inhibitor ofbcl-2. Am J Pathol 1994; 145:1323-1336.
Krajewski S, Mai JK, Krajewska M, Sikorska M, Mossakowski MJ, Reed JC. Upregulation of bax protein levels In neurons following cerebral ischemia. J Neurosci 1995; 15:6364- 6376.
Nicotera P, Ankarcrona M, Bonfoco E,
Orrenius S, Lipton 5/1. Neuronal necrosis and apoptosis: two distinct events induced by exposure to glutamate or oxidative stress. In: Seil EJ, ed. Advances in Neurology, vol. 72; Neuronal Regeneration, Reorganization, and Repair. Philadelphia: Lippincott-Raven
Publishers, 1997: 95-101.
Oppenheim RW. Related mechanisms of action of growth factors and antioxidants in apoptosis: an overview. In: Seil EJ, ed. Advances in Neurology, vol. 72; Neuronal Regeneration, Reorganization, and Repair. Philadelphia: Lippincott-Raven Publishers, 1997: 69-78.
Nikolics K, Hefti E, Thomas GR, Gluckman PD. Trophic factors and their role in the
postischemic brain. In: Siesjô BK, Tadeusz W, eds. Advances in Neurology, vol. 71; Cellular and Molecular Mechanisms of Ischemic Brain Damage. Philadelphia: Lippincott-Raven
Publishers, 1996: 389-402.
Zhu CZ, Auer RN. Intraventricular administration of insulin and IGE-I In transient forebrain ischemia. J Cereb Blood Flow Metab 1994:14:237-242.
Gluckman P, Klempt N, Guan J, et at A role for IGE-I in the rescue of CNS neurons following hypoxic-ischemic injury. Biochem Biophys Res Commun 1992; 182:593-599.
Guan J, Williams C, Gunning M, Mallard C, Gluckman P. The effects of IGE-I treatment after hypoxic-ischemic brain injury in adult rats. J Cereb Blood Flow Metab 1993; 13:609- 616.
Ginsberg MD, Busto R. Rodent models of cerebral Ischemia. Stroke 1989:20:1627- 1642.
Bergstedt K, Wieloch T. Changes in insulin-like growth factor receptor density after transient cerebral ischemia in the rat. Lack of protection against ischemic brain damage following injection of insulin-like growth factor I. J Cereb Blood Flow Metab 1993; 13:895- 898.
Li Y, Sharov VG, Jiang N, Zaloga C, Sabbah HN, Chopp M. Ultrastructural and light microscopical evidence of apoptosis after middle cerebral artery occlusion in the rat. Am J Pathol 1995; 146:1045-1051.
Hara A, Iwai T, Niwa M, et aL Immunohistochemical detection of Bax and Bcl-2 proteins in gerbil hippocampus following transient forebrain ischemia. Brain Res 1996;711:249-253.
Levine RL, Garland D, Oliver CN.
Determination of carbonyl content in oxidatively modified proteins. Meth Enzym 1990;186:464-475.
Upret GS, Rosemary AR, Riches RC. Protein estimation in tissues containing high levels of lipid. Modification to Lowry's method of protein determination. Analytical Biochem 1988;168:421-427.
Fisher M, Garcia JH. Evolving stroke and the ischemic penumbra. Neurology 1996;47:884- 888.
Hakim AM. Ischemic penumbra: the
therapeutic window. Neurology
:51 (Suppl 3):S44-S46.
Héron A, Pollard H, Dessl F, et al. Regional variability in DNA fragmentation after global
IGF-1 decreases apoptotic cell death, but not proapoptotic protein expression
ischemia evidenced by combined histological and gel electrophoresis observations in the rat brain. J bieurochem I993;6:1973-1976.
tiitatori T, Sato hi, Waguri S, et a!. Delayed neuronal death in the CA I pyramidal cell layer of the gerbil hippocampus following transient ischemia is apoptosis. J bieurosci 1995;15:1001-101 I.
Guglielmo MA, Chan FT, Cortez S, et al. The temporal profile and morphologic features of neuronal death in human stroke resemble those observed in experimental forebrain ischemia: the potential role of apoptosis. Fieurol Res 1998;20:283-296.
Shigeno T, Yamasaki Y, Rato O, et al. Reduction of delayed neuronal death by inhibition of protein synthesis. bieurosci Lett 1990,120:1 17-1 19.
Goto R, Ishige A, Sekiguchi R, et al. Effects of cycloheximide on delayed neuronal death in rat hippocampus. Brain Res 1990:534:299- 302.
Papas S, Crépel V, Hasboun D, Jorquera P, Chinestra P, Ben-Ari Y. Cycloheximide reduces the effects of anoxic insult in vivo and in vitro. Eur J bieurosci 1992;4:758-765.
MacManus JP, Linnik MD. Gene expression induced by cerebral ischemia: an apoptotic perspective. J Cereb Blood Plow Metab 1997;17:815-832.
Isenmann S, Stoll G, Schroeter M, Rrajewski S, Reed JC, Bahr M. Differential regulation of bax, bcl-2, and bcl-x proteins in focal cortical ischemia in the rat. Brain Pathol ¡998:8:49- 62.
Li Y, Chopp M, Powers C, Jiang bi. Apoptosis and protein expression after focal cerebral ischemia in rat. Brain Res 1997:765:301-312.
Deshpande J, Bergstedt R, Linden T, Ralimo
H, Wieloch T. Ultrastructural changes in the hippocampal CAI region following transient cerebral ischemia: evidence against
programmed cell death. Exp Brain Res 1992;88:91-105.
Iwai T, ftara A, biiwa M, et al. Temporal profile of nuclear DblA fragmentation in situ in gerbil hippocampus following transient forebrain ischemia. Brain Res 1995;671:305-308.
Colbourne E, Sutherland GR, Auer Rbl. Electron microscopic evidence against apoptosis as the mechanism of neuronal death in global ischemia. J bieurosci 1999;19:4200-4210.
Einklestein SP, Remmou A, Caday CD, Berlove DJ. Basic fibroblast growth factor protects cerebrocortical neurons against excitatory amino acid toxicity in vitro. Stroke ¡993:24 (suppl l):l-141-1-143.
Mattson MP, Cheng B. Growth factors protect neurons against excitotoxic/ischemic damage by stabilizing calcium homeostasis. Stroke 1993;24 (suppl I):I-136-1-140.
Mattson MP, Mark RJ. Excitotoxicity and
excitoprotection in vivo. In: Siesjd BR,
Tadeusz W, eds. Advances in bieurology, vol. 71; Cellular and Molecular Mechanisms of Ischemic Brain Damage. Philadelphia: Lippincott-Raven Publishers, 1996:1 -20.
Yuan J. The genes that regulate programmed cell death: from worm to mammal. In: Moskowitz MA, Caplan LR, eds. Cerebrovascular Diseases, 19th Princeton Stroke Conference, blew York: Butterworth- fleinemann, 1995:199-216.