Impact of therapeutic hypothermia onset and duration on survival, neurologic function, and neurodegeneration after cardiac arrest

Abstract

Objective: Post-cardiac-arrest therapeutic hypothermia improves outcomes in comatose cardiac arrest survivors. This study tests the hypothesis that the efficacy of post-cardiac-arrest therapeutic hypothermia is dependent on the onset and duration of therapy.

Design: Prospective randomized laboratory investigation.

Setting: University research laboratory.

Subjects: A total of 268 male Long Evans rats.

Interventions: Post-cardiac-arrest therapeutic hypothermia.

Measurements and main results: Adult male Long Evans rats that achieved return of spontaneous circulation after a 10-min asphyxial cardiac arrest were block randomized to normothermia (37°C ± 1°C) or therapeutic hypothermia (33°C ± 1°C) initiated 0, 1, 4, or 8 hrs after return of spontaneous circulation and maintained for 24 or 48 hrs. Therapeutic hypothermia initiated 0, 1, 4, and 8 hrs after return of spontaneous circulation resulted in 7-day survival rates of 45%*, 36%*, 36%*, and 14%, respectively, compared to 17% for normothermic controls and survival with good neurologic function rates of 24%*, 24%*, 19%*, and 0%, respectively, compared to 2% for normothermic controls (*p < .05 vs. normothermia). These outcomes were not different when therapeutic hypothermia was maintained for 24 vs. 48 hrs. In contrast, hippocampal CA1 pyramidal neuron counts were 53% ± 27%*, 53% ± 19%*, 51% ± 24%*, and 65% ± 16%* of normal, respectively, when therapeutic hypothermia was initiated 0, 1, 4, or 8 hrs after return of spontaneous circulation compared to 9% in normothermic controls (*p < .01 vs. normothermia). Furthermore, surviving neuron counts were greater when therapeutic hypothermia was maintained for 48 hrs compared to 24 hrs (68% ± 15%* vs. 42% ± 22%, *p < .0001).

Conclusions: In this study, post-cardiac-arrest therapeutic hypothermia resulted in comparable improvement of survival and survival with good neurologic function when initiated within 4 hrs after return of spontaneous circulation. However, histologic assessment of neuronal survival revealed a potentially broader therapeutic window and greater neuroprotection when therapeutic hypothermia was maintained for 48 vs. 24 hrs.

Figure 1. Temperature Regulation

Temperature curves (mean ± SD) for sham injured rats (A), normothermic post-cardiac arrest rats (B) and post-cardiac arrest rats treated with TH beginning 0 (C), 1, (D), 4 (E), or 8 (F) hours after ROSC. The time scale in minutes shows asphyxia onset (-10 min), CPR onset (0 min) and the first 60 minutes after ROSC. The remainder of the time scale in hours shows the time course of therapeutic hypothermia. All cardiac arrest rats developed a spontaneous hypothermia from 37.0° (at -15 min) to 35.5° (at 15 min), and were rewarmed to normothermia except for those in the immediate cooling groups (C). The induction of TH in all the hypothermia groups was achieved in 30 minutes (see insert graphs). After 24 hours (solid line) or 48 hours (dash line) of hypothermia, rats were rewarmed to normothermia over 8 hours at a rate of 0.5°C/hour.

Figure 2. Impact of Post-Cardiac Arrest Therapeutic Hypothermia on Survival

A Kaplan-Meier curve demonstrating improved survival with post-cardiac arrest TH compared to normothermia. B. Kaplan-Meier curve demonstrating equivalent improvement in survival with 24-hour and 48-hour duration TH compared to normothermia. C. Kaplan-Meier curve demonstrating improved survival with TH initiated 0, 1 or 4 hours after ROSC but not 8 hours after ROSC.

Figure 3. Impact of Post-Cardiac Arrest Therapeutic Hypothermia on Neurologic Function

A. Post-cardiac arrest TH significantly improves survival with good neurologic function (GNF) (16.67% NFS≥450) compared to normothermia (2.3% NFS≥450; Fisher's exact test, *p<0.05). B. 48-hour duration TH (16.67% NFS≥450) is equivalent to 24-hour duration TH (16.67% NFS≥450) in improving survival with GNF (Pearson's chi-square test, p=1). C. TH initiated at 0, 1, and 4 hours (23.81%, 23.81% and 19.05% NFS≥450) but not 8 hours (0% NFS≥450) after ROSC significantly improved survival with GNF compared to normothermia (2% NFS≥450; Fisher's Exact Test, *p<0.05).

Figure 4. Neuroprotection with Post-Cardiac Arrest Therapeutic Hypothermia

Stereologic quantification of hippocampal sector CA1 pyramidal neurons in sham injured rats and post-cardiac arrest rats at 7 days after ROSC. The results are displayed on the original scale while the comparisons were performed after arcsine transforming the proportions, which was used to normalize the data and stabilize the variances. A. Comparison of sham (100%), normothermia (8.81%) and combined hypothermia (53.4%) groups demonstrates improved neuron survival with post-cardiac arrest TH compared to rats maintained at normothermia (One way ANOVA, p<0.0001). B. Comparison of sham (100%), normothermia (8.8%), combined 24-hour duration TH group (41.6%) and combined 48-hour duration TH (67.6%) group demonstrates 48-hour duration TH results in superior neuroprotection compared to 24-hour TH (One way ANOVA, p<0.0001). C. Comparison of sham (100%), normothermia (8.8%) and 0-hour (52.6%), 1-hour (52.6%), 4-hour (50.5%), or 8-hour (65%) post-ROSC TH onset indicates that TH initiated between 0-8 hours after ROSC results in equivalent neuroprotection among 7-day survivors (Two way ANOVA, p=0.5058). Two-way ANOVA was adopted because that the hypothermia duration was significant.

Figure 5. Hippocampal Histology after Post-Cardiac Arrest Therapeutic Hypothermia

Representative hippocampal sections from sham injured rats or rats 7 days after 10-minute asphyxial cardiac arrest followed by normothermia or TH initiated 1 hour after ROSC and maintained for 24 or 48 hours. A. Sections immunolabeled for NeuN (red) and counterstained with Hoechst (blue). Degeneration of CA1 pyramidal neurons is characterized by loss of NeuN immunolabeling (red) and condensed shrunken nuclei (blue). B. Sections stained by the H&E method. Classic ischemic neuronal change in the CA1 pyramidal layer is characterized by shrunken dark-staining pyknotic nuclei (blue) and eosinophilic cytoplasm (pink) see in the high power images. Scale bar 500μm for low power images and 50μm for high power images.