481 - A Novel BDNF-Peptidomimetic Potentiator, dSyn3, Promotes Sustained Neuroprotection after Neonatal Hypoxic-Ischemic Brain Injury

Publication Number: 4472.481

Hang Xing, Women & Infants Hospital of Rhode Island, Providence, RI, United States; Benjamin Lee, Women & Infants Hospital of Rhode Island, White Plains, NY, United States; Bingqing Wang, Women & Infants Hospital of Rhode Island, PROVIDENCE, RI, United States; Zexi Zang, The Warren Alpert Medical School of Brown University, Providence, RI, United States; Barbara S.. Stonestreet, Brown Univ, Barrington, RI, United States; John Marshall, Brown University, Providence, RI, United States; Xiaodi F.. Chen, Women & Infants Hospital of Rhode Island, Providence, RI, United States

Background: Hypoxic-ischemic (HI) brain injury is a major cause of neurological disabilities in newborns. Effective pharmacological agents do not exist to treat HI encephalopathy. We have shown that dSyn3, a novel BDNF-peptidomimetic potentiator, confers short-term neuroprotection after HI in neonatal rats when HI was induced on postnatal (P) day 7. dSyn3 treatment reduced brain tissue volume atrophy and neuroinflammation when examined at P10, a developmental stage similar to the human near term brain. However, its long-term neuroprotective efficacy after HI has not been examined.

Objective: To determine whether treatment with dSyn3 exerts durable neuroprotective effects at P21 after exposure of rats to HI on P7.

Design/Methods: P7 rats were randomly assigned to Sham, placebo (HI-PL), or HI-dSyn3 groups. HI was induced by right carotid artery ligation and 8% O2 for 90 min. HI animals received phosphate-buffered saline (PL) or dSyn3 (1 mg/kg, intraperitoneally) at 0, 24, and 48 h after HI; Sham received PL without HI. Behavioral outcomes examined included the righting reflex (P8-P10, coordination), small open field (P13, activity level), negative geotaxis (P14, posture), wire hang (P16, strength), and rotarod (P20-P21, coordination) tests, and were recorded and analyzed with EthoVision XT 11. On P21, brains were perfused, fixed, and cryosectioned (20 µm coronal). Cresyl violet-stained sections were analyzed with ImageJ (NIH) to determine hemispheric volume atrophy, expressed as the percent ratio of damaged to total hemispheric area, corrected for edema and integrated across serial slices.

Results: dSyn3 treatment improved righting reflex performance on P8 compared with HI+PL rats (P < 0.05; Fig. A). On P20 (Day 1, training), HI+PL rats showed an increased number of falls and shorter latency to first fall on the rotarod (P < 0.05; Fig. B-I, top and lower panels), indicative of motor impairment. dSyn3 treatment reduced the number of falls and prolonged latency, similar to Sham levels. On P21 (Day 2, testing), dSyn3-treated rats exhibited greater latency to fall on the accelerating rotarod task (Fig.B-II, top panel) and higher fall velocity (Fig. B-II, lower panel) compared to HI+PL (P < 0.05). dSyn3 decreased hemispheric tissue volume atrophy at P21 (P < 0.05; Fig.C), demonstrating its sustained neuroprotective efficacy after HI in neonates.

Conclusion(s): dSyn3 provides durable neuroprotection and functional recovery after HI in neonates. Our findings support its potential use as an effective pharmacological adjunctive or alternative treatment for therapeutic hypothermia.

dSyn3 treatment improves neurobehavioral performance and reduces hemispheric atrophy after neonatal hypoxic-ischemic injury
Figure A. Righting reflex performance. dSyn3 treatment (red box) significantly improved righting reflex performance on P8 compared with HI+PL rats (blue box; * P < 0.05). Each pup was placed supine on a flat surface, and the latency to right itself to a prone position with all four paws touching the surface was recorded (maximum 30 s per trial). Three consecutive trials were averaged per animal. Data were analyzed using lognormal Brown-Forsythe and Welch ANOVA followed by Dunnett's T3 multiple comparisons test.
Figure B. Rotarod performance over two days. On Day 1 (P20), rats were habituated on a stationary rod and trained for 3 min at 16 rpm. The number of falls (B-I, top) and latency to first fall (B-I, bottom) were recorded. On Day 2 (P21), rats were tested on an accelerating rod (2-20 rpm over 5 min). Latency to fall (B-II, top) and rotarod speed at first fall (B-II, bottom) were measured. Each rat completed three trials per day. Statistical analysis used Kruskal-Wallis with Dunn's post hoc tests (* P < 0.05).
Figure C. Hemispheric tissue atrophy. Percent hemispheric tissue atrophy was quantified for Sham, HI-PL, and HI-dSyn3 groups. Statistics were performed using Kruskal-Wallis with Dunn's tests (* P < 0.05).
Note: Sample sizes (males and females combined) are shown. Data are presented as box plots (empty box = Sham; red = HI-PL; blue = HI-dSyn3) with medians, interquartile ranges (25th/75th), 10th/90th percentiles, and individual data points; n values are indicated in the panels. Statistical analyses and plotting were performed using GraphPad Prism 10 and R.