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Heliox Preconditioning Exerts Neuroprotective Effects on Neonatal Ischemia/Hypoxia Injury by Inhibiting Necroptosis Induced by Ca2+ Elevation

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Abstract

Our previous studies have indicated that heliox preconditioning (HePC) may exert neuroprotective effects on neonatal hypoxic-ischemic encephalopathy (HIE). The present study was to investigate whether HePC alleviates neonatal HIE by inhibiting necroptosis and explore the potential mechanism. Seven-day-old rat pups were randomly divided into Sham group, HIE group, HIE + HePC group, HIE + Dantrolene (DAN) group, and HIE + Necrostatin-1 (Nec-1) group. HIE was induced by common carotid artery ligation and subsequent hypoxia exposure. The neurological function, brain injury, and molecular mechanism were evaluated by histological staining, neurobehavioral test, Western blotting, Ca2+, immunofluorescence staining, co-immunoprecipitation (Co-IP), and transmission electron microscopy (TEM). Results supported that the expression of necroptosis markers and p-RyR2 in the brain increased significantly after HIE. HePC, DAN, or Nec-1 was found to improve the neurological deficits after H/I and inhibit neuronal necroptosis. Interestingly, both HePC and DAN inhibited the increases in cytoplasmic Ca2+ and CaMK-II phosphorylation in the brain secondary to HIE, but Nec-1 failed to affect Ca2+. In conclusion, our results suggest HePC may alleviate cytoplasmic Ca2+ overload by regulating p-RyR2, which inhibits the necroptosis in the brain, exerting neuroprotective effects on HIE.

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The original contributions presented in the study are included in the article/supplementary materials; further inquiries can be directed to the corresponding authors.

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Funding

This work was supported by the National Natural Science Foundation of China (No. 81873789) and the Medical-Engineering Cross Fund of Shanghai Jiao Tong University (No. YG2019GD03).

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Author notes

  1. Weijie Zhong and Juan Cheng have contributed equally to this work and share first authorship.

Authors and Affiliations

  1. Department of Neurosurgery, Ninth People Hospital Affiliated to Shanghai Jiao Tong University School of Medicine, Shanghai, 200011, People’s Republic of China

    Weijie Zhong, Xiaosheng Yang & Yi Li

  2. Department of Ultrasound, Xinhua Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, People’s Republic of China

    Juan Cheng

  3. Naval Characteristic Medical Center Diving and Hyperbaric Medicine Research Laboratory, Shanghai, 200433, People’s Republic of China

    Wenwu Liu

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  1. Weijie Zhong
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Contributions

WZ, JC, and XY made the idea and experimental design of this study. WZ, JC, WL, and YL performed most of the experiments. WZ and WL drafted the manuscript. WZ and YL approved the final version of the manuscript on behalf of all the authors. All authors did the critical revision and language modification of the manuscript.

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Correspondence to Wenwu Liu or Yi Li.

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All the experimental protocols were approved by the Ethics Committee of Shanghai Jiaotong University School of Medicine. This article does not contain any studies with human participants performed by any of the authors.

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Supplementary A

Experimental schematics. B Proposed Ca2+ -mediated signaling cascade following H/I. (PPTX 28462 kb)

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Zhong, W., Cheng, J., Yang, X. et al. Heliox Preconditioning Exerts Neuroprotective Effects on Neonatal Ischemia/Hypoxia Injury by Inhibiting Necroptosis Induced by Ca2+ Elevation. Transl. Stroke Res. 14, 409–424 (2023). https://doi.org/10.1007/s12975-022-01021-8

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