Ligustilide attenuates ischemic stroke injury by promoting Drp1-mediated mitochondrial fission via activation of AMPK

Abstract

Background: Ischemic stroke is a major global cause of death and permanent disability. Studies have suggested that mitochondria play a critical role in maintaining cellular energy homeostasis and inevitably involved in neuronal damage during cerebral ischemic. Ligustilide is the main active ingredient of Angelica sinensis and Ligusticum chuanxiongs with neuroprotective activity.

Purpose: These study sought to exlopre the role of LIG in improving mitochondrial function and the relationship between LIG induced mitochondrial fission and mitophagy in ischemic stroke.

Methods: Cerebral I/R injury was established by the model of Oxygen-glucose deprivation/reperfusion (OGD/R) in HT22 cells and middle cerebral artery occlusion (MCAO) in rats. Mitochondrial functions of were detected by flow cytometry and immunofluorescence, and mitochondrial fission were detected by western blots. Furthermore, we studied the role of AMPK pathway in the neuroprotective effect of LIG.

Results: LIG treatment significantly increased the MMP and ATP production, decreased the reactive oxygen species (ROS) generation and Ca²⁺ overload, and further induced mitochondrial fission and mitophagy. Moreover, we found that blocking mitochondrial fission by mdivi-1 resulted in accumulation of damaged mitochondria mainly through selectively blocking mitophagy, thereby inhibiting viability of HT-22 cells after OGD/R. Also, Drp-1 inhibitor mdivi-1 increased the infarct volume and aggravated the neurological deficits after MCAO operation in vivo. Additionally, LIG triggered AMP-activated protein kinase (AMPK) pathway. AMPKα2 knockdown attenuated LIG-induced mitochondrial fission through inhibiting the expression of Drp1 and Fis1, and led to nerve cell apoptosis.

Conclusion: Our study indicate that LIG attenuated the injury of ischemic stroke by improving mitochondrial function and highlight the critical role of LIG in the regulation of LIG-induced mitochondrial fission and mitophagy via an AMPK-dependent manner. These findings indicate that LIG protects nerve damage against ischemic stroke by inducing Drp1-mediated mitochondrial fission via activation of AMPK signaling pathway in vivo and in vitro.

Keywords: AMPK; ligustilide; mitochondrial fission; mitophagy.