Total flavonoids of hawthorn leaves protect spinal motor neurons via promotion of autophagy after spinal cord injury

Qiong Zhang; Mingfu Liu; Haibin Nong; Yanan Zhang; Yiguang Bai; Pan Liu; Shaohui Zong; Gaofeng Zeng

doi:10.3389/fphar.2022.925568

Total flavonoids of hawthorn leaves protect spinal motor neurons via promotion of autophagy after spinal cord injury

Front Pharmacol. 2022 Aug 22:13:925568. doi: 10.3389/fphar.2022.925568. eCollection 2022.

Authors

Qiong Zhang¹, Mingfu Liu², Haibin Nong², Yanan Zhang³, Yiguang Bai², Pan Liu², Shaohui Zong^{2

4}, Gaofeng Zeng¹

Affiliations

¹ School of Public Health of Guangxi Medical University, Nanning, China.
² Department of Spine Osteopathia, The First Affiliated Hospital of Guangxi Medical University, Nanning, China.
³ Collaborative Innovation Center of Guangxi Biological Medicine, Guangxi Medical University, Nanning, China.
⁴ Research Centre for Regenerative Medicine and Guangxi Key Laboratory of Regenerative Medicine, Guangxi Medical University, Nanning, China.

Abstract

The death of spinal motor neurons (SMNs) after spinal cord injury (SCI) is a crucial cause, contributing to a permanent neurological deficit. Total flavonoids of hawthorn leaves (TFHL) have been confirmed to have potentially therapeutic for SCI. Nonetheless, the roles and mechanisms of TFHL in recovering neuromotor function and regenerating axons of SMNs have not been fully elucidated. In this study, TFHL was applied to treat rats with SCI and injured SMNs for 7 days. In vivo experiment, rats with SCI were evaluated by a BBB (Basso-Beattie-Bresnahan) score to assess their motor functional recovery. The morphology, microstructure, apoptosis, Nissl bodies, and autophagy of SMNs in spinal cord tissue were detected by Hematoxylin-eosin (HE) staining, transmission electron microscopy, TUNEL staining, Nissl staining, and immunohistochemistry respectively. In vitro experiment, the co-culture model of SMNs and astrocytes was constructed to simulate the internal environment around SMNs in the spinal cord tissue. The cell morphology, microstructure, axonal regeneration, and autophagy were observed via optical microscope, transmission electron microscopy, and immunofluorescence. The content of neurotrophic factors in the cell culture medium of the co-culture model was detected by ELISA. Moreover, the expression of axon-related and autophagy-related proteins in the spinal cord tissue and SMNs was measured by Western Blot. We demonstrated that TFHL improved the neuromotor function recovery in rats after SCI. We then found that TFHL significantly promoted injured spinal cord tissue repair, reduced apoptosis, and improved the functional status of neurons in spinal cord tissue in vivo. Meanwhile, the cell morphology, microstructure, and axonal regeneration of damaged SMNs also obviously were improved, and the secretion of neurotrophic factors was facilitated after treatment with TFHL in vitro. Further, we revealed that TFHL promoted autophagy and related protein expression in vivo and vitro. Taken together, our study suggested that TFHL might facilitate autophagy and have neuroprotective properties in SMNs to enhance the recovery of neuromotor function of rats with SCI.

Keywords: autophagy; axon regeneration; neuroprotection; spinal cord injury; spinal cord motor neurons; total flavonoids of hawthorn leaves.