국소뇌허혈 모델에서 열충격단백질70의 카스파아제-3 활성화 및 디옥시리보핵산 분절화 억제에 의한 신경보호효과
Heat-Shock Protein 70 Attenuates Caspase-3 Activation and DNA Fragmentation Following Focal Cerebral Ischemia
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Background: Recently, HSP70 has been shown to act as an inhibitor of apoptotic pathways in the cell culture following heat shock; however, little information is available on the mechanism of neuroprotection after cerebral ischemia. In this study, our purpose is to investigate whether the HSP70 protein can protect apoptotic cell death after focal cerebral ischemia. Methods: hsp70.1 knockout (KO) and wild-type (WT) mice were subjected to transient middle cerebral artery occlusion for 2 hours. At 22 hours, we measured infarction volumes, and detected DNA fragmentation with TUNEL staining. HSP70 and hsp70.1 mRNA expression were analyzed by Western blots and Northern blots, respectively. Caspase-3 activation was examined with Western blots and caspase-3 activity assay. Results: hsp70.1 mRNA was not detected in hsp70.1 KO mice after ischemia, and HSP70 expression was markedly suppressed in KO mice versus WT mice. The infarction volume was significantly larger in the KO (82.1±9.5 mm3) than in the WT (58.4±10.3 mm3; p<0.05) 24hours later. Caspase-3 activation was also significantly enhanced in KO mice versus WT mice, as evidenced by higher levels of activated caspase-3 and cleaved gelsolin, as determined by Western blotting and caspase-3 activity assay. TUNEL-positive apoptotic cells were much higher in the KO (1.94±0.61×103/mm2) than in the WT (1.05±0.35 ×103/mm2; p<0.05) in the cortex, but not in the striatum. Conclusions: We conclude that that HSP70 acts as a strong inhibitor of apoptosis via blocking caspase-3 activation following focal cerebral ischemia.
Abstract
Background: Recently, HSP70 has been shown to act as an inhibitor of apoptotic pathways in the cell culture following heat shock; however, little information is available on the mechanism of neuroprotection after cerebral ischemia. In this study, our purpose is to investigate whether the HSP70 protein can protect apoptotic cell death after focal cerebral ischemia. Methods: hsp70.1 knockout (KO) and wild-type (WT) mice were subjected to transient middle cerebral artery occlusion for 2 hours. At 22 hours, we measured infarction volumes, and detected DNA fragmentation with TUNEL staining. HSP70 and hsp70.1 mRNA expression were analyzed by Western blots and Northern blots, respectively. Caspase-3 activation was examined with Western blots and caspase-3 activity assay. Results: hsp70.1 mRNA was not detected in hsp70.1 KO mice after ischemia, and HSP70 expression was markedly suppressed in KO mice versus WT mice. The infarction volume was significantly larger in the KO (82.1±9.5 mm3) than in the WT (58.4±10.3 mm3; p<0.05) 24hours later. Caspase-3 activation was also significantly enhanced in KO mice versus WT mice, as evidenced by higher levels of activated caspase-3 and cleaved gelsolin, as determined by Western blotting and caspase-3 activity assay. TUNEL-positive apoptotic cells were much higher in the KO (1.94±0.61×103/mm2) than in the WT (1.05±0.35 ×103/mm2; p<0.05) in the cortex, but not in the striatum. Conclusions: We conclude that that HSP70 acts as a strong inhibitor of apoptosis via blocking caspase-3 activation following focal cerebral ischemia.
참고문헌 (32)
- "Role of the major heat shock proteins as molecular chaperones", 9, pp.601~634 (1993)
- "Transgenic mice expressing the human heat shock protein 70 have improved post-ischemic myocardial recovery", 95, pp.1854~1860 (1995)
- "Mice overexpressing rat heat shock protein 70 are protected against cerebral infarction", 47, pp.782~791 (2000)
- "Targeted hsp70.1 disruption increases infarction volume after focal cerebral ischemia in mice", 32, pp.2905~2912 (2001)
- "Heat shock proteins and neuroprotection", 513, pp.281~299 (2002)
- "Differential neuroprotection from human heat shock protein 70 overexpression in in vitro and in vivo models of ischemia and ischemia-like conditions", 170, pp.129~139 (2001)
- "Lack of neuroprotection by heat shock protein 70 overexpression in a mouse model of global cerebral ischemia", 154, pp.442~449 (2004)
- "Many mechanisms for hsp70 protection from cerebral ischemia", 16, pp.53~61 (2004)
- "Programmed cell death:alive and well in the new millennium", 11, pp.526~534 (2001)
- "Intact,injured,necrotic and apoptotic cells after focal cerebral ischemia in the rat", 156, pp.119~132 (1998)
- "Ischemic intensity influences the distribution of delayed infarction and apoptotic cell death following transient focal cerebral ischemia in rats", 956, pp.14~23 (2002)
- "Targeted disruption of hsp70.1 sensitizes to osmotic stress", 3, pp.857~861 (2002)
- "Reversible middle cerebral artery occlusion without craniectomy in rats", 20, pp.84~91 (1989)
- "A cautionary note on the use of the TUNEL stain to determine apoptosis", 7, pp.61~64 (1995)
- "Correlation between cerebral blood flow and histologic changes in a new rat model of middle cerebral artery occlusion", 20, pp.1037~1043 (1989)
- "Multislice diffusion mapping for 3-D evolution of cerebral ischemia in a rat stroke model", 45, pp.172~177 (1995)
- "Ischemic penumbra in a model of reversible middle cerebral artery occlusion in the rat", 89, pp.67~78 (1992)
- "Electron microscopic evidence against apoptosis as the mechanism of neuronal death in global ischemia", 19, pp.4200~4210 (1999)
- "The stress protein response in cultured neurons:characterization and evidence for a protective role in excitotoxicity", 7, pp.1053~1060 (1991)
- "Astrocyte survival and HSP70 heat shock protein induction following heat shock and acidosis", 17, pp.147~159 (1996)
- "Over-expression of HSP-70 protects astrocytes from combined oxygen-glucose deprivation", 7, pp.429~432 (1996)
- "HSP70 protects murine astrocytes from glucose deprivation injury", 224, pp.9~12 (1997)
- "Transfection with hsp70i protects rat dorsal root ganglia neurones and glia from heat stress", 1(1), pp.65~ (1994)
- "Defective herpes simplex virus vectors expressing the rat brain stress-inducible heat shock protein 72 protect cultured neurons from severe heat shock", 68, pp.961~969 (1997)
- "Overexpression of the rat inducible 70-kD heat stress protein in a transgenic mouse increases the resistance of the heart to ischemic injury", 95, pp.1446~1456 (1995)
- "Role of the human heat shock protein hsp70 in protection against stress-induced apoptosis", 17, pp.5317~5327 (1997)
- "Hsp70 exerts its anti-apoptotic function downstream of caspase-3-like proteases", 17, pp.6124~6134 (1998)
- "The chaperone function of hsp70 is required for protection against stress-induced apoptosis", 20, pp.7146~7159 (2000)
- "Negative regulation of the Apaf-1 apoptosome by Hsp70", 2, pp.476~483 (2000)
- "Heat-shock protein 70 inhibits apoptosis by preventing recruitment of procaspase-9 to the Apaf-1 apoptosome", 2, pp.469~475 (2000)
- "Overexpression of rat heat shock protein 70 is associated with reduction of early mitochondrial cytochrome C release and subsequent DNA fragmentation after permanent focal ischemia", 23, pp.718~727 (2003)
- "Multiple molecular penumbras after focal cerebral ischemia", 20, pp.1011~1032 (2000)