Combined Treatment of Xenon and Hypothermia in Newborn Rats - Additive or Synergistic Effect?

Hemmen Sabir,John Dingley,Lars Walløe,Elisa Smit,Marianne Thoresen,Xun Liu,Lucio Annunziato

doi:10.1371/journal.pone.0109845

Hemmen Sabir, John Dingley + Show 5 more

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https://doi.org/10.1371/journal.pone.0109845

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Abstract

BackgroundBreathing the inert gas Xenon (Xe) enhances hypothermic (HT) neuroprotection after hypoxia-ischemia (HI) in small and large newborn animal models. The underlying mechanism of the enhancement is not yet fully understood, but the combined effect of Xe and HT could either be synergistic (larger than the two effects added) or simply additive. A previously published study, using unilateral carotid ligation followed by hypoxia in seven day old (P7) rats, showed that the combination of mild HT (35°C) and low Xe concentration (20%), both not being neuroprotective alone, had a synergistic effect and was neuroprotective when both were started with a 4 h delay after a moderate HI insult. To examine whether another laboratory could confirm this finding, we repeated key aspects of the study.Design/MethodsAfter the HI-insult 120 pups were exposed to different post-insult treatments: three temperatures (normothermia (NT) NT37°C, HT35°C, HT32°C) or Xe concentrations (0%, 20% or 50%) starting either immediately or with a 4 h delay. To assess the synergistic potency of Xe-HT, a second set (n = 101) of P7 pups were exposed to either HT35°C+Xe0%, NT+Xe20% or a combination of HT35°C+Xe20% starting with a 4 h delay after the insult. Brain damage was analyzed using relative hemispheric (ligated side/unligated side) brain tissue area loss after seven day survival.ResultsImmediate HT32°C (p = 0.042), but not HT35°C significantly reduced brain injury compared to NT37°C. As previously shown, adding immediate Xe50% to HT32°C increased protection. Neither 4 h-delayed Xe20%, nor Xe50% at 37°C significantly reduced brain injury (p>0.050). In addition, neither 4 h-delayed HT35°C alone, nor HT35°C+Xe20% reduced brain injury. We found no synergistic effect of the combined treatments in this experimental model.ConclusionsCombining two treatments that individually were ineffective (delayed HT35°C and delayed Xe20%) did not exert neuroprotection when combined, and therefore did not show a synergistic treatment effect.

Highlights

Therapeutic hypothermia (HT) has been shown to be safe and reduces brain injury after hypoxia ischemia (HI) in human newborns [1,2,3,4,5,6] and animal models [7,8]
We found no synergistic effect of the combined treatments in this experimental model
When repeating our previously published experiments and using brain area loss instead of a neuropathology score as an outcome parameter, we found that mean (6SD) area loss in the NT37uC group was 54.2% (65.56)

Summary

Introduction

Therapeutic hypothermia (HT) has been shown to be safe and reduces brain injury after hypoxia ischemia (HI) in human newborns [1,2,3,4,5,6] and animal models [7,8]. A previously published study, using unilateral carotid ligation followed by hypoxia in seven day old (P7) rats, showed that the combination of mild HT (35uC) and low Xe concentration (20%), both not being neuroprotective alone, had a synergistic effect and was neuroprotective when both were started with a 4 h delay after a moderate HI insult. To examine whether another laboratory could confirm this finding, we repeated key aspects of the study

Methods

Results

Conclusion