Poly(ADP-Ribose)synthetase activation and NAD depletion after traumatic brain injury in rats

Margaret A. Satchell, Robert S B Clark, Minzhi Chen, Melick John, Csaba Szabo, Patrick M. Kochanek

Research output: Contribution to journalArticle

Abstract

Introduction: Poly(ADP-Ribose) synthetase (PARS; also known as PARP) deficient mice are dramatically protected from functional deficits after traumatic brain injury (TBI)1. PARS activation generates poly(ADP-ribose)polymers (PADPR) on damaged DNA and nuclear proteins consuming NAD in the process; accordingly, we hypothesized that PARS activation contributes to energy failure after TBI. Methods: Anesthetized adult rats were subjected to controlled cortical impact with imposed secondary insult using our standard model. Rats were killed at 6, 8, 24, and 72 h after TBI; naïve rats were used as controls (n = 3-5/group). To determine whether the potent trigger of PARS activation, peroxynitrite, is generated after TBI, Western Wots were analyzed for nitrotyrosine residues on proteins. In addition, brain tissue was examined for evidence of oxidative DNA damage using DNA polymerase mediated dATP nick translation (PANT). Activation of PARS was measured indirectly using Western blot and immunohistochemistry to detect PADPR. NAD levels in cortex and hippocampus were measured using an enzymatic cycling method. Results: Nitrotyrosine residues were detected on ∼30 and ∼80kD protein bands from ipsilateral cortex at 8, 24 and 72 h after TBI but not in controls. PANT showed single-strand DNA nicks in ipsilateral cortex and hippocampus at 6 and 24 h after TBI. An increase in PADPR on multiple protein bands was seen at 8 and 24 h but not in controls. By 72 h, PADPR on proteins were reduced (vs. 24 h). PADPR were also detected immunohistochemically in injured brain. NAD levels in ipsilateral cortex and hippocampus were depleted by 86% at 24 h after TBI vs. control. Conclusions: These data support the hypothesis that NAD depletion after TBI is a consequence of PARS activation triggered by peroxynitrite-mediated DNA damage. Thus, PARS activation after TBI may exacerbate energy failure in injured brain, which might in turn explain the protective effects observed in PARS deficient mice. Further study using clinically relevant PARS inhibitors after TBI is warranted. 1Whalen et al, J Cereb Blood Flow Metab 1999;19:825-42.

Original languageEnglish (US)
JournalCritical Care Medicine
Volume27
Issue number12 SUPPL.
StatePublished - 1999
Externally publishedYes

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Poly Adenosine Diphosphate Ribose
Polymers
NAD
Hippocampus
Peroxynitrous Acid
DNA Damage
Brain
Proteins
Traumatic Brain Injury
NAD+ synthase
Single-Stranded DNA Breaks
DNA-Directed DNA Polymerase
Ligases
Nuclear Proteins
Western Blotting
Immunohistochemistry

ASJC Scopus subject areas

  • Critical Care and Intensive Care Medicine

Cite this

Satchell, M. A., Clark, R. S. B., Chen, M., John, M., Szabo, C., & Kochanek, P. M. (1999). Poly(ADP-Ribose)synthetase activation and NAD depletion after traumatic brain injury in rats. Critical Care Medicine, 27(12 SUPPL.).

Poly(ADP-Ribose)synthetase activation and NAD depletion after traumatic brain injury in rats. / Satchell, Margaret A.; Clark, Robert S B; Chen, Minzhi; John, Melick; Szabo, Csaba; Kochanek, Patrick M.

In: Critical Care Medicine, Vol. 27, No. 12 SUPPL., 1999.

Research output: Contribution to journalArticle

Satchell, MA, Clark, RSB, Chen, M, John, M, Szabo, C & Kochanek, PM 1999, 'Poly(ADP-Ribose)synthetase activation and NAD depletion after traumatic brain injury in rats', Critical Care Medicine, vol. 27, no. 12 SUPPL..
Satchell, Margaret A. ; Clark, Robert S B ; Chen, Minzhi ; John, Melick ; Szabo, Csaba ; Kochanek, Patrick M. / Poly(ADP-Ribose)synthetase activation and NAD depletion after traumatic brain injury in rats. In: Critical Care Medicine. 1999 ; Vol. 27, No. 12 SUPPL.
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abstract = "Introduction: Poly(ADP-Ribose) synthetase (PARS; also known as PARP) deficient mice are dramatically protected from functional deficits after traumatic brain injury (TBI)1. PARS activation generates poly(ADP-ribose)polymers (PADPR) on damaged DNA and nuclear proteins consuming NAD in the process; accordingly, we hypothesized that PARS activation contributes to energy failure after TBI. Methods: Anesthetized adult rats were subjected to controlled cortical impact with imposed secondary insult using our standard model. Rats were killed at 6, 8, 24, and 72 h after TBI; na{\"i}ve rats were used as controls (n = 3-5/group). To determine whether the potent trigger of PARS activation, peroxynitrite, is generated after TBI, Western Wots were analyzed for nitrotyrosine residues on proteins. In addition, brain tissue was examined for evidence of oxidative DNA damage using DNA polymerase mediated dATP nick translation (PANT). Activation of PARS was measured indirectly using Western blot and immunohistochemistry to detect PADPR. NAD levels in cortex and hippocampus were measured using an enzymatic cycling method. Results: Nitrotyrosine residues were detected on ∼30 and ∼80kD protein bands from ipsilateral cortex at 8, 24 and 72 h after TBI but not in controls. PANT showed single-strand DNA nicks in ipsilateral cortex and hippocampus at 6 and 24 h after TBI. An increase in PADPR on multiple protein bands was seen at 8 and 24 h but not in controls. By 72 h, PADPR on proteins were reduced (vs. 24 h). PADPR were also detected immunohistochemically in injured brain. NAD levels in ipsilateral cortex and hippocampus were depleted by 86{\%} at 24 h after TBI vs. control. Conclusions: These data support the hypothesis that NAD depletion after TBI is a consequence of PARS activation triggered by peroxynitrite-mediated DNA damage. Thus, PARS activation after TBI may exacerbate energy failure in injured brain, which might in turn explain the protective effects observed in PARS deficient mice. Further study using clinically relevant PARS inhibitors after TBI is warranted. 1Whalen et al, J Cereb Blood Flow Metab 1999;19:825-42.",
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T1 - Poly(ADP-Ribose)synthetase activation and NAD depletion after traumatic brain injury in rats

AU - Satchell, Margaret A.

AU - Clark, Robert S B

AU - Chen, Minzhi

AU - John, Melick

AU - Szabo, Csaba

AU - Kochanek, Patrick M.

PY - 1999

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N2 - Introduction: Poly(ADP-Ribose) synthetase (PARS; also known as PARP) deficient mice are dramatically protected from functional deficits after traumatic brain injury (TBI)1. PARS activation generates poly(ADP-ribose)polymers (PADPR) on damaged DNA and nuclear proteins consuming NAD in the process; accordingly, we hypothesized that PARS activation contributes to energy failure after TBI. Methods: Anesthetized adult rats were subjected to controlled cortical impact with imposed secondary insult using our standard model. Rats were killed at 6, 8, 24, and 72 h after TBI; naïve rats were used as controls (n = 3-5/group). To determine whether the potent trigger of PARS activation, peroxynitrite, is generated after TBI, Western Wots were analyzed for nitrotyrosine residues on proteins. In addition, brain tissue was examined for evidence of oxidative DNA damage using DNA polymerase mediated dATP nick translation (PANT). Activation of PARS was measured indirectly using Western blot and immunohistochemistry to detect PADPR. NAD levels in cortex and hippocampus were measured using an enzymatic cycling method. Results: Nitrotyrosine residues were detected on ∼30 and ∼80kD protein bands from ipsilateral cortex at 8, 24 and 72 h after TBI but not in controls. PANT showed single-strand DNA nicks in ipsilateral cortex and hippocampus at 6 and 24 h after TBI. An increase in PADPR on multiple protein bands was seen at 8 and 24 h but not in controls. By 72 h, PADPR on proteins were reduced (vs. 24 h). PADPR were also detected immunohistochemically in injured brain. NAD levels in ipsilateral cortex and hippocampus were depleted by 86% at 24 h after TBI vs. control. Conclusions: These data support the hypothesis that NAD depletion after TBI is a consequence of PARS activation triggered by peroxynitrite-mediated DNA damage. Thus, PARS activation after TBI may exacerbate energy failure in injured brain, which might in turn explain the protective effects observed in PARS deficient mice. Further study using clinically relevant PARS inhibitors after TBI is warranted. 1Whalen et al, J Cereb Blood Flow Metab 1999;19:825-42.

AB - Introduction: Poly(ADP-Ribose) synthetase (PARS; also known as PARP) deficient mice are dramatically protected from functional deficits after traumatic brain injury (TBI)1. PARS activation generates poly(ADP-ribose)polymers (PADPR) on damaged DNA and nuclear proteins consuming NAD in the process; accordingly, we hypothesized that PARS activation contributes to energy failure after TBI. Methods: Anesthetized adult rats were subjected to controlled cortical impact with imposed secondary insult using our standard model. Rats were killed at 6, 8, 24, and 72 h after TBI; naïve rats were used as controls (n = 3-5/group). To determine whether the potent trigger of PARS activation, peroxynitrite, is generated after TBI, Western Wots were analyzed for nitrotyrosine residues on proteins. In addition, brain tissue was examined for evidence of oxidative DNA damage using DNA polymerase mediated dATP nick translation (PANT). Activation of PARS was measured indirectly using Western blot and immunohistochemistry to detect PADPR. NAD levels in cortex and hippocampus were measured using an enzymatic cycling method. Results: Nitrotyrosine residues were detected on ∼30 and ∼80kD protein bands from ipsilateral cortex at 8, 24 and 72 h after TBI but not in controls. PANT showed single-strand DNA nicks in ipsilateral cortex and hippocampus at 6 and 24 h after TBI. An increase in PADPR on multiple protein bands was seen at 8 and 24 h but not in controls. By 72 h, PADPR on proteins were reduced (vs. 24 h). PADPR were also detected immunohistochemically in injured brain. NAD levels in ipsilateral cortex and hippocampus were depleted by 86% at 24 h after TBI vs. control. Conclusions: These data support the hypothesis that NAD depletion after TBI is a consequence of PARS activation triggered by peroxynitrite-mediated DNA damage. Thus, PARS activation after TBI may exacerbate energy failure in injured brain, which might in turn explain the protective effects observed in PARS deficient mice. Further study using clinically relevant PARS inhibitors after TBI is warranted. 1Whalen et al, J Cereb Blood Flow Metab 1999;19:825-42.

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