Elevated microRNA-214-3p level ameliorates neuroinflammation after spinal cord ischemia-reperfusion injury by inhibiting Nmb/Cav3.2 pathway

Guo-Qiang Xia; Miao Xu; Cong Sun; Zai-Li Zhang; Xiao-Qian Li

doi:10.1016/j.intimp.2024.112031

Elevated microRNA-214-3p level ameliorates neuroinflammation after spinal cord ischemia-reperfusion injury by inhibiting Nmb/Cav3.2 pathway

Int Immunopharmacol. 2024 Apr 16:133:112031. doi: 10.1016/j.intimp.2024.112031. Online ahead of print.

Authors

Guo-Qiang Xia¹, Miao Xu², Cong Sun², Zai-Li Zhang³, Xiao-Qian Li⁴

Affiliations

¹ Department of Anesthesiology, The First Hospital of China Medical University, Shenyang 110001, Liaoning, China.
² Department of Pain Medicine, The First Hospital of China Medical University, Shenyang 110001, Liaoning, China.
³ Department of Anesthesiology, The First Hospital of China Medical University, Shenyang 110001, Liaoning, China. Electronic address: zhangzaili333@sina.com.
⁴ Department of Anesthesiology, The First Hospital of China Medical University, Shenyang 110001, Liaoning, China. Electronic address: shirley037305@hotmail.com.

PMID: 38631219
DOI: 10.1016/j.intimp.2024.112031

Abstract

Background: Neuromedin B (Nmb) plays a pivotal role in the transmission of neuroinflammation, particularly during spinal cord ischemia-reperfusion injury (SCII). However, the detailed molecular mechanisms underlying this process remain elusive.

Methods: The SCII model was established by clamping the abdominal aorta of male Sprague-Dawley (SD) rats for 60 min. The protein expression levels of Nmb, Cav3.2, and IL-1β were detected by Western blotting, while miR-214-3p expression was quantified by qRT-PCR. The targeted regulation between miR-214-3p and Nmb was investigated using a dual-luciferase reporter gene assay. The cellular localization of Nmb and Cav3.2 with cell-specific markers was visualized by immunofluorescence staining. The specific roles of miR-214-3p on the Nmb/Cav3.2 interactions in SCII-injured rats were explored by intrathecal injection of Cav3.2-siRNA, PD168368 (a specific NmbR inhibitor) and synthetic miR-214-3p agomir and antagomir in separate experiments. Additionally, hind-limb motor function was evaluated using the modified Tarlov scores.

Results: Compared to the Sham group, the protein expression levels of Nmb, Cav3.2, and the proinflammatory factor Interleukin(IL)-1β were significantly elevated at 24 h post-SCII. Intrathecal injection of PD168368 and Cav3.2-siRNA significantly suppressed the expression of Cav3.2 and IL-1β compared to the SCII group. The miRDB database and dual-luciferase reporter gene assay identified Nmb as a direct target of miR-214-3p. As expected, in vivo overexpression of miR-214-3p by agomir-214-3p pretreatment significantly inhibited the increases in Nmb, Cav3.2 and IL-1β expression and improved lower limb motor function in SCII-injured rats, while antagomiR-214-3p pretreatment reversed these effects.

Conclusions: Nmb protein levels positively correlated with Cav3.2 expression in SCII rats. Upregulating miR-214-3p ameliorated hind-limb motor function and protected against neuroinflammation via inhibiting the aberrant Nmb/Cav3.2 interactions and downstream IL-1β release. These findings provide novel therapeutic targets for clinical prevention and treatment of SCII.

Keywords: Neuroinflammation; Neuromedin B; Spinal cord ischemia–reperfusion injury; T-type Ca(2+) channel; miR-214-3p.