Naotaifang Formula Regulates Drp1-Induced Remodeling of Mitochondrial Dynamics Following Cerebral Ischemia-Reperfusion Injury

Abstract

Cerebral ischemia-reperfusion injury (CIRI) has emerged as a hindrance for rehabilitation of ischemic stroke patients. Naotaifang (NTF) exhibits beneficial efficacy in alleviating inflammation and ferroptosis in vitro during CIRI. While the potential role of NTF in regulating mitochondrial dynamics in CIRI are not elucidated. This study aimed to explore the mechanism of NTF against CIRI by regulating the dynamin‐related protein 1 (Drp1)-dependent mitochondrial fission/fusion. Modeling middle cerebral artery occlusion/reperfusion (MCAO/R) in vivo to evaluate the effects of NTF on the MCAO/R-damaged neurons and the structure, dynamics and function of mitochondria. An oxygen-glucose deprivation/reperfusion (OGD/R) cell model was established to evaluate the role of NTF in OGD/R-damaged cells. Function of Drp1 in CIRI and the neuroprotection of NTF through the mitochondrial fission/fusion pathway were investigated in vivo and in vitro. The results revealed that in vivo, NTF alleviated neuron injury in a dose-dependent manner, down-regulated Drp1 and fission protein 1 (Fis1) levels, upregulated optic atrophy 1 (Opa1), mitofusin 1/2 (Mfn1 and Mfn2), facilitated mitochondrial fusion and inhibited mitochondrial fission to rescue cells from CIRI. In vitro, Drp1 overexpression inhibited mitochondrial fusion and activated mitochondrial fission, while silencing of Drp1 exhibited the opposite result. NTF rebalanced mitochondrial dynamic in the OGD/R cell model. NTF could alleviate neuron injury following CIRI by regulating the balance of mitochondrial fission and fusion. Targeting Drp1-dependent mitochondrial dynamics may represent a viable treatment strategy for addressing the issues of CIRI post ischemic stroke.

Note:
Funding declaration: This work was supported by the National Natural Science Foundation of China (82174167), the Key Project of Hunan Province Education Department (20A366), the Project of Natural Science Foundation of Hunan Province (2021JJ30499, 2023JJ30464), the Young Qihuang Scholar Support Project of National Administration of Traditional Chinese Medicine (2022) and Shenzhen Polytechnic University Research Fund (6024310022K).

Conflict of Interests: The authors declare no competing financial interests.

Ethical Approval: Animals were bred at the Experimental Animal Center of the Science and Technology Innovation Center of Hunan University of Chinese Medicine, operating under animal laboratory certificate number SYXK (Hunan) 2019-0009 and ethics number 202105280001.

Keywords: Cerebral ischemia-reperfusion injury, Ischemic stroke, Naotaifang, Mitochondrial fission/fusion, Drp1