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Objective: Spinal cord injury (SCI) often leads to severe physiological and pathological changes in patients. Erxian Decoction (EXD) is effective in the postoperative treatment of spinal cord injury, but its specific mechanism of action is poorly defined. Methods: Network pharmacology and molecular docking were used to predict the potential mechanisms of EXD in SCI. In vivo studies were used to validate the above predictions. For in vivo study, the rats were pretreated with or without EXD (5.76 g/kg, by intragastric gavage). Multiple molecular biological test methods to identify molecular mechanisms. One-way analysis of variance (ANOVA) was used with Bonferroni's post-hoc test to identify the differences between groups. Results: In vivo studies have shown that EXD improved motor function at 7dpi in SCI rats (P < 0.0001), significantly reduced spinal cord edema (P = 0.0139), upregulated 5-HT, GFAP, and TMEM119 expression. Through network pharmacology analysis, we found that Akt1 in EXD plays a role in treating SCI. The underlying mechanism may be the inhibition of apoptosis after activation of Akt1 phosphorylation. Molecular docking revealed that the key compounds could spontaneously bind to the Akt1 protein. Pharmacological inhibition of Akt1 activation by MK-2206, attenuated the anti-apoptotic effect of EXD on SCI in rats (P < 0.0001). Conclusions: EXD inhibits apoptosis by activating Akt1, reduce spinal cord edema and restore behavioral function after SCI in rats.

Objective: Spinal cord injury (SCI) is a devastating disease resulting in lifelong disability, but the molecular mechanism remains unclear. Our study was designed to observe the role of excision repair cross-complementing group 6 (ERCC6) following SCI and to determine the underlying mechanism.Methods: SCI mouse models and LPS-induced microglia cell models were established. ERCC6 expression was blocked by ERCC6-siRNA-carrying lentivirus. Nissl staining was utilized for detecting neuronal damage, and apoptosis was analyzed with TUNEL and Western blotting (apoptotic markers). Immunofluorescence was used for measuring macrophage markers (CD68 and F4/80) and astrocyte and microglia markers (GFAP and Iba-1). Pro-inflammatory cytokines (TNF-α, IL-1β, and IL-6) were measured via ELISA. Senescent cells were estimated via SA-β-Gal staining as well as Western blot (senescent markers p21 and p27). Oxidative stress was investigated by detecting the expression of 4-HNE, Nrf2, and Keap1, and intracellular ROS levels.Results: ERCC6 expression was remarkably upregulated both in the spinal cord of SCI mice and LPS-induced microglia cells. ERCC6 deficiency alleviated neuronal damage and apoptosis. Macrophage infiltration and inflammatory response were suppressed by si-ERCC6 treatment. Moreover, ERCC6 blockage ameliorated astrocyte and microglia activation and cell senescence in the damaged spinal cord. Excessive oxidative stress was significantly decreased by ERCC6 knockdown in SCI.Conclusion: Collectively, ERCC6 exerts crucial functions in mediating physiological processes (apoptosis, inflammation, senescence, and oxidative stress), implying that ERCC6 might act as a prospective therapeutic target against SCI.

Objective: Spinal cord injury (SCI) is a debilitating disease that cannot be cured at present. Kaempferol (KPL) has proven neuroprotective, antioxidant, and antibacterial properties. However, the neuroprotective effect of KPL on SCI and its potential mechanism are still unclear. Methods: Network pharmacology and molecular docking were used to determine the potential mechanisms of KPL on SCI. In vitro studies were used to validate the results. Hydrogen peroxide (H2O2)-exposed PC12 cells were pretreated with or without KPL. The safe concentrations of KPL, H2O2, and MK2206 in PC12 cells were determined via cell counting kit-8 assays. Then, terminal deoxynucleotidyl transferase dUTP nick end labeling staining and Annexin V-FITC/PI double staining were used to detect apoptotic cells. In addition, Western blotting was utilized to measure apoptosis-related gene and protein expression levels. Results: Network pharmacology and molecular docking data demonstrated an interaction between KPL and Akt1 and identified apoptosis as a potential target of the KPL/Akt1 complex. The data verified that KPL could inhibit mitochondrial apoptosis in vivo by inducing Akt1 phosphorylation. Subsequently, MK2206, a pharmacological Akt1 inhibitor, was used to assess the role of Akt1 in this process, and we found that MK2206 attenuated the antiapoptotic effect of KPL on PC12 cells. Conclusion: This study demonstrates that KPL could inhibit mitochondrial apoptosis by inducing the phosphorylation of Akt1, suggesting that KPL is a very promising candidate for SCI treatment. [ABSTRACT FROM AUTHOR]