Reduction of caspase-8 and -9 cleavage is associated with increased c-FLIP and increased binding of Apaf-1 and Hsp70 after neonatal hypoxic/ischemic injury in mice overexpressing Hsp70

Yasuhiko Matsumori, Frances Northington, Shwuhuey M. Hong, Takamasa Kayama, R. Ann Sheldon, Zinaida S. Vexler, Donna M. Ferriero, Philip R. Weinstein, Jialing Liu

Research output: Contribution to journalArticle

Abstract

Background and Purpose - Caspase-8 and caspase-9 are essential proteases of the extrinsic and intrinsic apoptotic pathways, respectively. We investigated whether neuroprotection associated with overexpression of heat-shock protein 70 (Hsp70), a natural cellular antiapoptotic protein, is mediated by caspase-8 and caspase-9 signaling in the neonatal mouse brain after hypoxia/ischemia (H/I) injury. Methods - Postnatal day 7 transgenic mice overexpressing rat Hsp70 (Hsp70 Tg) and their wild-type (Wt) littermates underwent unilateral common carotid artery ligation followed by 30 minutes of exposure to 8% O2. The expression of apoptotic proteins was quantified by Western blot analysis, and the specific interaction between Hsp70 and apoptotic protease activating factor 1 (Apaf-1) was determined by coimmunoprecipitation. Results - Hsp70 overexpression reduced cytosolic translocation of cytochrome c without affecting the levels of Apaf-1 and pro-caspase-9 24 hours after H/I. The expression of these apoptotic proteins in the naïve neonatal brains was also not affected by Hsp70 overexpression. Reduced caspase-9 cleavage occurred in Hsp70 Tg mice compared with Wt littermates 24 hours after H/I and correlated with increased binding of Hsp70 and Apaf-1. Increased cellular Fas-associated death domain-like interleukin-1β-converting enzyme inhibitory protein (FLIP) expression and decreased caspase-8 cleavage were also observed in Hsp70 Tg compared with Wt mice 24 hours after H/I. Conclusions - Our results suggest that the extrinsic and intrinsic apoptotic pathways mediate the neuroprotective effects of Hsp70 overexpression in neonatal H/I, specifically by upregulating FLIP and sequestering Apaf-1, leading to reduced cleavage of caspase-8 and caspase-9.

Original languageEnglish (US)
Pages (from-to)507-512
Number of pages6
JournalStroke
Volume37
Issue number2
DOIs
StatePublished - Feb 2006

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Apoptotic Protease-Activating Factor 1
HSP70 Heat-Shock Proteins
Caspase 9
Caspase 8
Wounds and Injuries
Proteins
Ischemia
Brain Hypoxia-Ischemia
Caspase 1
Common Carotid Artery
Neuroprotective Agents
Cytochromes c
Transgenic Mice
Ligation

Keywords

  • Apoptosis
  • Mitochondria
  • Stress proteins

ASJC Scopus subject areas

  • Cardiology and Cardiovascular Medicine
  • Neuroscience(all)

Cite this

Reduction of caspase-8 and -9 cleavage is associated with increased c-FLIP and increased binding of Apaf-1 and Hsp70 after neonatal hypoxic/ischemic injury in mice overexpressing Hsp70. / Matsumori, Yasuhiko; Northington, Frances; Hong, Shwuhuey M.; Kayama, Takamasa; Sheldon, R. Ann; Vexler, Zinaida S.; Ferriero, Donna M.; Weinstein, Philip R.; Liu, Jialing.

In: Stroke, Vol. 37, No. 2, 02.2006, p. 507-512.

Research output: Contribution to journalArticle

Matsumori, Yasuhiko ; Northington, Frances ; Hong, Shwuhuey M. ; Kayama, Takamasa ; Sheldon, R. Ann ; Vexler, Zinaida S. ; Ferriero, Donna M. ; Weinstein, Philip R. ; Liu, Jialing. / Reduction of caspase-8 and -9 cleavage is associated with increased c-FLIP and increased binding of Apaf-1 and Hsp70 after neonatal hypoxic/ischemic injury in mice overexpressing Hsp70. In: Stroke. 2006 ; Vol. 37, No. 2. pp. 507-512.
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abstract = "Background and Purpose - Caspase-8 and caspase-9 are essential proteases of the extrinsic and intrinsic apoptotic pathways, respectively. We investigated whether neuroprotection associated with overexpression of heat-shock protein 70 (Hsp70), a natural cellular antiapoptotic protein, is mediated by caspase-8 and caspase-9 signaling in the neonatal mouse brain after hypoxia/ischemia (H/I) injury. Methods - Postnatal day 7 transgenic mice overexpressing rat Hsp70 (Hsp70 Tg) and their wild-type (Wt) littermates underwent unilateral common carotid artery ligation followed by 30 minutes of exposure to 8{\%} O2. The expression of apoptotic proteins was quantified by Western blot analysis, and the specific interaction between Hsp70 and apoptotic protease activating factor 1 (Apaf-1) was determined by coimmunoprecipitation. Results - Hsp70 overexpression reduced cytosolic translocation of cytochrome c without affecting the levels of Apaf-1 and pro-caspase-9 24 hours after H/I. The expression of these apoptotic proteins in the na{\"i}ve neonatal brains was also not affected by Hsp70 overexpression. Reduced caspase-9 cleavage occurred in Hsp70 Tg mice compared with Wt littermates 24 hours after H/I and correlated with increased binding of Hsp70 and Apaf-1. Increased cellular Fas-associated death domain-like interleukin-1β-converting enzyme inhibitory protein (FLIP) expression and decreased caspase-8 cleavage were also observed in Hsp70 Tg compared with Wt mice 24 hours after H/I. Conclusions - Our results suggest that the extrinsic and intrinsic apoptotic pathways mediate the neuroprotective effects of Hsp70 overexpression in neonatal H/I, specifically by upregulating FLIP and sequestering Apaf-1, leading to reduced cleavage of caspase-8 and caspase-9.",
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T1 - Reduction of caspase-8 and -9 cleavage is associated with increased c-FLIP and increased binding of Apaf-1 and Hsp70 after neonatal hypoxic/ischemic injury in mice overexpressing Hsp70

AU - Matsumori, Yasuhiko

AU - Northington, Frances

AU - Hong, Shwuhuey M.

AU - Kayama, Takamasa

AU - Sheldon, R. Ann

AU - Vexler, Zinaida S.

AU - Ferriero, Donna M.

AU - Weinstein, Philip R.

AU - Liu, Jialing

PY - 2006/2

Y1 - 2006/2

N2 - Background and Purpose - Caspase-8 and caspase-9 are essential proteases of the extrinsic and intrinsic apoptotic pathways, respectively. We investigated whether neuroprotection associated with overexpression of heat-shock protein 70 (Hsp70), a natural cellular antiapoptotic protein, is mediated by caspase-8 and caspase-9 signaling in the neonatal mouse brain after hypoxia/ischemia (H/I) injury. Methods - Postnatal day 7 transgenic mice overexpressing rat Hsp70 (Hsp70 Tg) and their wild-type (Wt) littermates underwent unilateral common carotid artery ligation followed by 30 minutes of exposure to 8% O2. The expression of apoptotic proteins was quantified by Western blot analysis, and the specific interaction between Hsp70 and apoptotic protease activating factor 1 (Apaf-1) was determined by coimmunoprecipitation. Results - Hsp70 overexpression reduced cytosolic translocation of cytochrome c without affecting the levels of Apaf-1 and pro-caspase-9 24 hours after H/I. The expression of these apoptotic proteins in the naïve neonatal brains was also not affected by Hsp70 overexpression. Reduced caspase-9 cleavage occurred in Hsp70 Tg mice compared with Wt littermates 24 hours after H/I and correlated with increased binding of Hsp70 and Apaf-1. Increased cellular Fas-associated death domain-like interleukin-1β-converting enzyme inhibitory protein (FLIP) expression and decreased caspase-8 cleavage were also observed in Hsp70 Tg compared with Wt mice 24 hours after H/I. Conclusions - Our results suggest that the extrinsic and intrinsic apoptotic pathways mediate the neuroprotective effects of Hsp70 overexpression in neonatal H/I, specifically by upregulating FLIP and sequestering Apaf-1, leading to reduced cleavage of caspase-8 and caspase-9.

AB - Background and Purpose - Caspase-8 and caspase-9 are essential proteases of the extrinsic and intrinsic apoptotic pathways, respectively. We investigated whether neuroprotection associated with overexpression of heat-shock protein 70 (Hsp70), a natural cellular antiapoptotic protein, is mediated by caspase-8 and caspase-9 signaling in the neonatal mouse brain after hypoxia/ischemia (H/I) injury. Methods - Postnatal day 7 transgenic mice overexpressing rat Hsp70 (Hsp70 Tg) and their wild-type (Wt) littermates underwent unilateral common carotid artery ligation followed by 30 minutes of exposure to 8% O2. The expression of apoptotic proteins was quantified by Western blot analysis, and the specific interaction between Hsp70 and apoptotic protease activating factor 1 (Apaf-1) was determined by coimmunoprecipitation. Results - Hsp70 overexpression reduced cytosolic translocation of cytochrome c without affecting the levels of Apaf-1 and pro-caspase-9 24 hours after H/I. The expression of these apoptotic proteins in the naïve neonatal brains was also not affected by Hsp70 overexpression. Reduced caspase-9 cleavage occurred in Hsp70 Tg mice compared with Wt littermates 24 hours after H/I and correlated with increased binding of Hsp70 and Apaf-1. Increased cellular Fas-associated death domain-like interleukin-1β-converting enzyme inhibitory protein (FLIP) expression and decreased caspase-8 cleavage were also observed in Hsp70 Tg compared with Wt mice 24 hours after H/I. Conclusions - Our results suggest that the extrinsic and intrinsic apoptotic pathways mediate the neuroprotective effects of Hsp70 overexpression in neonatal H/I, specifically by upregulating FLIP and sequestering Apaf-1, leading to reduced cleavage of caspase-8 and caspase-9.

KW - Apoptosis

KW - Mitochondria

KW - Stress proteins

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