Xiyuan Hospital, China Academy of Chinese Medical Sciences, Beijing 100091, China; Postdoctoral Research Station, China Academy of Chinese Medical Sciences, Beijing 100700, China.
Xiyuan Hospital, China Academy of Chinese Medical Sciences, Beijing 100091, China; NMPA Key Laboratory for Clinical Research and Evaluation of Traditional Chinese Medicine, Beijing 100091, China; National Clinical Research Center for Chinese Medicine Cardiology, Beijing 100091, China.
Phytomedicine. 2021 Nov;92:153749. doi: 10.1016/j.phymed.2021.153749. Epub 2021 Sep 16.
MicroRNA-155(miR-155) is closely associated with diabetic peripheral neuropathy (DPN). Astragaloside IV (AST) is a significant extract of Astragalus membranaceus, which has been found to be effective in the treatment of DPN. However, whether astragaloside IV alleviate DPN via regulating miR-155-mediated autophagy remains unclear.
This study was designed to evaluate the effects of AST on DPN myelin Schwann cells injury and explore the mechanism of AST in treating DPN for the first time.
GK rats fed with high-fat diet and RSC96 cells cultured in high glucose were used to establish DPN Schwann cells injury in vivo and in vitro model. The effects of AST on DPN were explored through blood glucose detection, nerve function detection, pathological detection and the expression of Neuritin detected by immunohistochemical. To study the effect of AST on the DPN Schwann cells autophagy and the upstream PI3K/Akt/mTOR pathway, the expressions of beclin-1 and LC3 were detected by western blot (WB) in sciatic nerves and by immunofluorescence (IFC) in RSC96 cells. The real-time polymerase chain reaction (RT-PCR) was applied to detect the expressions of miR-155, ATG5, ATG12 both in vivo and in vitro. The binding effect of miR-155 and target gene PI3KCA was verified by luciferase reporter gene assay. The expressions of PI3K, p-Akt/Akt, p-mTOR/mTOR were detected by WB and the expressions of PI3KCA were detected by RT-PCR in vitro. The apoptosis was detected by flow cytometry. Meanwhile, the influence of miR-155 overexpression and knocked down on the above indicators was also detected in RSC96 cells. At last, further mechanism experiments were conducted to verify the mechanism of AST regulating the autophagy and apoptosis of RSC96 cells.
AST reduced blood glucose levels, alleviated peripheral nerve myelin sheath injury, and improved neurological function in DPN rats. In addition, AST enhanced the autophagy activity and alleviated the apoptosis in RSC96 cell. Mechanism study shown that AST promote autophagy via regulating miR-155-mediated PI3K/Akt/mTOR signaling pathways. AST reduced RSC96 cells apoptosis by promoting autophagy.
AST alleviate the myelin sheath injury of DPN caused by the apoptosis of Schwann cells via enhancing autophagy, which was attributed to inhibiting the activation of the PI3K/Akt/mTOR signaling pathway by upregulating miR-155 expression.
微小 RNA-155（miR-155）与糖尿病周围神经病变（DPN）密切相关。黄芪甲苷（AST）是黄芪的重要提取物，已被证明对 DPN 的治疗有效。然而，AST 是否通过调节 miR-155 介导的自噬来减轻 DPN 尚不清楚。
本研究旨在评估 AST 对 DPN 雪旺细胞髓鞘损伤的影响，并首次探讨 AST 治疗 DPN 的机制。
使用高脂饮食喂养的 GK 大鼠和高糖培养的 RSC96 细胞建立体内和体外 DPN 雪旺细胞损伤模型。通过血糖检测、神经功能检测、病理检测和免疫组化检测 Neuritin 的表达来研究 AST 对 DPN 的影响。通过 Western blot（WB）检测坐骨神经中 beclin-1 和 LC3 的表达以及 RSC96 细胞中的免疫荧光（IFC），研究 AST 对 DPN 雪旺细胞自噬及上游 PI3K/Akt/mTOR 通路的影响。通过实时聚合酶链反应（RT-PCR）检测体内和体外 miR-155、ATG5、ATG12 的表达。通过荧光素酶报告基因检测验证 miR-155 与靶基因 PI3KCA 的结合效应。通过 WB 检测 PI3K、p-Akt/Akt、p-mTOR/mTOR 的表达，通过 RT-PCR 检测 PI3KCA 的表达，通过流式细胞术检测细胞凋亡。同时，在 RSC96 细胞中检测 miR-155 过表达和敲低对上述指标的影响。最后，进行进一步的机制实验以验证 AST 调节 RSC96 细胞自噬和凋亡的机制。
AST 降低了 DPN 大鼠的血糖水平，减轻了周围神经髓鞘损伤，改善了神经功能。此外，AST 增强了 RSC96 细胞的自噬活性并减轻了细胞凋亡。机制研究表明，AST 通过调节 miR-155 介导的 PI3K/Akt/mTOR 信号通路促进自噬。AST 通过促进自噬减少 RSC96 细胞凋亡。
AST 通过增强自噬减轻 DPN 引起的雪旺细胞凋亡导致的髓鞘损伤，这归因于通过上调 miR-155 表达抑制 PI3K/Akt/mTOR 信号通路的激活。
