030 - Therapeutic Hypothermia with Adenosine and/or Creatinine Phosphate (CrP) for Reducing Brain Injury and Inflammatory Prostanoids in a Neonatal Hypoxic-Ischemic Encephalopathy (HIE) Rat Model

Publication Number: 30.6906

Onur Taparli, State University of New York Downstate Medical Center College of Medicine, BROOKLYN, NY, United States; Charles L. Cai, SUNY Downstate, Brooklyn, NY, United States; Vadim Bronshtein, Suny Downstate Health Sciences University, Brooklyn, NY, United States; Mayan Kolb, Westchester Medical, River Edge, NJ, United States; Ivan L. Hand, Kings County Hospital/SUNY Downstate School of Medicine, Great neck, NY, United States; Jacob V V. Aranda, State University of New York Downstate Medical Center College of Medicine, BROOKLYN, NY, United States; Kay Beharry, SUNY Downstate Health Sciences University, Brooklyn, NY, United States

Background: Therapeutic hypothermia (TH) is an established intervention for hypoxic-ischemic encephalopathy (HIE). However, its effectiveness is partial, as many infants still suffer severe neurodevelopmental outcomes. Creatinine phosphate (CrP) and adenosine (Ad) have distinct neuroprotective roles—maintaining ATP levels and inhibiting glutamate release, respectively—which have demonstrated brain-protective effects across various pathologies.

Objective: To test the hypothesis that combining TH with Ad and/or CrP provides short- and long-term synergistic neuroprotection. Additionally, we aimed to evaluate the role of inflammatory prostanoids in HIE using a neonatal rat model.

Design/Methods: Postnatal day 7 (P7) rats underwent unilateral carotid artery ligation followed by hypoxia (8% O₂, 92% N₂) for 100 minutes at 37°C, using the Rice-Vannucci model. Immediately post-hypoxia, a single intraperitoneal dose of saline (Sal) or treatment was administered. Treatment groups included: 1) Sham-operated (no treatment); 2) HIE+Sal; 3) HIE+Sal+TH; 4) HIE+Ad (0.4 mg/kg); 5) HIE+Ad+TH; 6) HIE+CrP (100 mg/kg); 7) HIE+CrP+TH; 8) HIE+Ad+CrP; and 9) HIE+Ad+CrP+TH. After 1 hour of recovery, TH (34°C) was applied for 24 hours. Rats were euthanized either at 24 hours post-HIE (P8) or at 5 weeks (5W). Assessments included histopathology, morphometry, infarct size, apoptosis, myelination, oxidative stress, COX-2 immunoreactivity, and inflammatory prostanoids in the cerebral cortex. Data are presented as mean±SEM.

Results: At P8, brain width increased in groups 5 and 9 compared to sham (p < 0.01). Cortical layer III and V thickness was increased (p < 0.01) across all groups versus sham. By 5W, persistent increases in brain width were observed in groups 3–6 and 8, with sustained layer III (group 4) and V (group 3) thickening. TTC staining revealed reduced viability in HIE+Sal and HIE+Ad+CrP+TH groups. Significant apoptosis was noted in all groups at P8, except in group 4 and 5, with no reduction at 5W. This was associated with elevated 8-isoPGF2α, inflammatory prostanoids, and COX-2 immunoreactivity.

Conclusion(s): Our results did not show any statistically significant neuroprotective effect of combination Ad+CrP with or without TH compared to sham group. Oxidative stress and COX-2-related inflammation play role in HIE pathophysiology. Cerebral injury and swelling concurrent with induction of inflammatory prostanoids suggest edema and osmotic cell death. Findings highlight a potentially deleterious interaction between oxidative stress and the COX-2 inflammatory pathway in HIE, which may be central to developing future therapeutic approaches.