Ajit A. Sarnaik

Influence of PARP-1 Polymorphisms in Patients after Traumatic Brain Injury

Poly(ADP-ribose) polymerase-1 (PARP-1) plays an important role in the cellular response to stress and DNA damage. However, excessive activity of PARP-1 exacerbates brain injury via NAD+ depletion and energy failure. The purpose of this study was to determine if tagging single nucleotide polymorphisms (tSNPs) covering multiple regions of the PARP-1 gene are related to outcome after traumatic brain injury (TBI) in humans. DNA from 191 adult patients with severe TBI was assayed for four tSNPs corresponding to haplotype blocks spanning the PARP-1 gene. Categorization as favorable or poor outcome was based on Glasgow Outcome Scale (GOS) score assigned at 6 months. PARP-1 enzyme activity was indirectly evaluated by quantifying poly-ADP-ribose (PAR)-modified proteins in cerebrospinal fluid (CSF) using an enzyme-linked immunosorbent assay. In multiple logistic regression analysis controlling for age, initial Glasgow Coma Scale score, and gender, the AA genotype of SNP rs3219119 was an independent predictor of favorable neurologic outcome. This SNP tags a haplotype block spanning the automodification and catalytic domains of the PARP-1 gene. SNP rs2271347 correlated with CSF PAR-modified protein level. This SNP, which did not correlate with outcome, tags a haplotype block spanning the promoter region of the PARP-1 gene. We conclude that after severe TBI in humans, a PARP-1 polymorphism within the automodification-catalytic domain is associated with neurological outcome, while a polymorphism within the promoter region was associated with CSF PAR-modified protein level. These findings must be replicated in a prospective study before the relevance of PARP-1 polymorphisms after TBI can be established.

Quantification of Poly(ADP-Ribose)-Modified Proteins in Cerebrospinal Fluid from Infants and Children after Traumatic Brain Injury

Poly-ADP-ribosylation (PAR) of proteins by poly(ADP-ribose) polymerases (PARP) occurs after experimental traumatic brain injury (TBI) and modulates neurologic outcome. Several promising pharmacological PARP inhibitors have been developed for use in humans, but there is currently no clinically relevant means of monitoring treatment effects. We therefore used an enzyme-linked immunosorbent assay to measure PAR-modified proteins in cerebrospinal fluid (CSF). Cerebrospinal fluid samples from 17 pediatric TBI patients and 15 controls were plated overnight and then incubated with polyclonal antibody against PAR. Histone-1, a PARP substrate, was incubated with active PARP, NAD, and nicked DNA, and served as the standard. Both peak and mean CSF PAR-modified proteins were increased in TBI patients versus controls. Peak CSF PAR-modified protein levels occurred on day 1 and levels remained increased on day 2 after TBI. Increases in peak CSF PAR-modified protein concentrations were independently associated with age and male sex, but not initial Glasgow Coma Scale score, Glasgow outcome score, or mechanism of injury. The increase in PAR-modified proteins in CSF after TBI may be because of increased PARP activation, decreased PAR degradation, or both. As PAR-modified protein concentration correlated with age and male sex, developmental and sex-dependent roles for PARP after TBI are implicated.

Views of pediatric intensive care physicians on the ethics of organ donation after cardiac death

Objective:Donation after cardiac death has been endorsed by professional organizations, including the American Academy of Pediatrics as a means of increasing the supply of transplantable organs. However, ethical concerns have been raised about donation after cardiac death, especially in children. This study explores the views of pediatric intensive care physicians on the ethics of pediatric donation after cardiac death. Design:Internet survey. Subjects:Physician members of the American Academy of Pediatrics Section of Critical Care. Interventions:Physicians were emailed an anonymous survey consisting of four demographic items and 16 items designed to assess their views on the ethics of pediatric donation after cardiac death. Responses to ethics items were rated on a 5-point scale ranging from strongly disagree to strongly agree. Physicians were also given the opportunity to provide free-text comments regarding their views. Measurements and Main Results:Of the 598 eligible physicians, 264 (44.1%) responded to the survey. Of these, 193 (73.4%) were practicing in a transplant center and 160 (60.6%) participated in at least one donation after cardiac death procedure at the time of survey completion. Two hundred twenty (83.4%) agreed or strongly agreed that regarding donation after cardiac death, parents should be able to make decisions based on the best interests of their child. Two hundred twenty-two (84.1%) agreed or strongly agreed that it is not acceptable to harvest organs from a child before the declaration of death, consistent with the Dead Donor Rule. However, only 155 (59.1%) agreed or strongly agreed that the time of death in donation after cardiac death can be conclusively determined. Twenty-nine (11.0%) agreed or strongly agreed that the pediatric donation after cardiac death donor may feel pain or suffering during the harvest procedure. Conclusions:Most pediatric intensive care physicians agree that the Dead Donor Rule should be applied for donation after cardiac death and that donation after cardiac death can be consistent with the best interest standard. However, concerns about the ability to determine time of death for the purpose of organ donation and the possibility of increasing donor pain and suffering exist.