Erythropoietin alleviates astrocyte pyroptosis by targeting the miR-325-3p/Gsdmd axis in rat spinal cord injury.

Abstract

Neuroinflammation plays an important role in spinal cord injury (SCI), and an increasing number of studies have focused on the role of astrocytes in neuroinflammation. Pyroptosis is an inflammation-related form of programmed cell death, and neuroinflammation induced by astrocytes in the form of pyroptosis has been widely reported in many central nervous system diseases. Recent studies have found that erythropoietin has significant anti-inflammatory and neuroprotective effects in SCI; however, it has not been reported whether erythropoietin can reduce neuroinflammation by inhibiting neural cell pyroptosis in SCI. A GEO dataset (GSE153720) was used to analyse the expression of pyroptosis-related genes in sham astrocytes and astrocytes 7days, 1month and 3months after SCI. TargetScan and miRDB databases were used to predict the miRNA that could bind to the 3'UTR of rat Gsdmd. Primary rat spinal astrocytes were used for in vitro experiments, and the modified version of Allen's method was used to establish the rat SCI model. Western blotting, quantitative real-time polymerase chain reaction, flow cytometry, immunofluorescence, lactate dehydrogenase release assay and propidium iodide staining were used to detect the pyroptosis phenotype. A dual luciferase reporter gene assay was used to verify that miR-325-3p can bind to the 3'UTR of Gsdmd. We found that pyroptosis-related genes mediated by the canonical NLRP3 inflammasome were highly expressed in astrocytes in an SCI animal model by bioinformatic analysis. We also observed that erythropoietin could reduce astrocyte pyroptosis in vivo and in vitro. In addition, we predicted miRNAs that regulate Gsdmd, the pyroptosis executor, and verified that erythropoietin inhibits astrocyte pyroptosis in SCI through the miR-325-3p/Gsdmd axis. We demonstrated that erythropoietin can inhibit astrocyte pyroptosis through the miR-325-3p/Gsdmd axis. This study is expected to provide a new mechanism for erythropoietin in the treatment of SCI and a more reliable theoretical basis for clinical research.