基于网络药理学和实验验证探讨脉络舒通丸对大鼠股骨骨折所致后肢肌肉肿胀的作用机制

Mechanism of Mailuo Shutong Pills on posterior limb muscle swelling caused by femoral fracture in rats based on network pharmacology and experimental verification

旨在通过网络药理学及动物实验探讨脉络舒通丸(MLST)对股骨骨折后肢体肿胀(SCFF)的作用机制。利用液质联用技术分析MLST入血成分,网络药理学预测其入血成分对SCFF的靶点和信号通路,分子对接进行验证。通过动物实验建立SCFF模型大鼠,给予不同剂量的MLST治疗,检测其肢体肿胀程度,苏木精-伊红(HE)染色检测肌肉组织病理变化,酶联免疫吸附测定法(ELISA)测定外周血白细胞介素-6(IL-6)、白细胞介素-1β(IL-1β)及肿瘤坏死因子-α(TNF-α)含量,采用Western blot测定相关信号通路表达。网络药理学结果显示,MLST与SCFF疾病作用靶点共计153个,关键靶点有IL-6、TNF等,涉及丝裂原活化蛋白激酶(MAPK)信号通路、磷脂酰肌醇3-激酶(PI3K)/蛋白激酶B(AKT)信号通路等;对主要入血成分和关键靶点进行分子对接,结合能均小于-7.0 kcal·mol^(-1),说明网络分析结果可靠。动物实验结果表明,与模型组相比,MLST可以减轻SCFF大鼠后肢肌肉肿胀度及其病理损伤,降低SCFF大鼠血清中IL-6、IL-1β及TNF-α水平,降低SCFF大鼠中p-AKT、p-PI3K、p-NF-κB、p-p38 MAPK、p-ERK蛋白表达。综上,MLST可能通过调节PI3K/AKT与MAPK相关信号通路蛋白的表达,降低血清中炎症因子含量,改善大鼠SCFF。

This study aims to investigate the mechanism of Mailuo Shutong Pills(MLST)on posterior limb muscle swelling caused by femoral fracture(SCFF)through network pharmacology and animal experiments.The plasma components of MLST were analyzed by LC-MS,and the target and signal pathway of SCFF were predicted by network pharmacology and verified by molecular docking.SCFF model rats were established through animal experiments,and different doses of MLST were administered to detect the degree of limb swelling.Hematoxylin-eosin(HE)staining was used to observe pathological changes in muscle tissue,and interleukin-6(IL-6),interleukin-1β(interleukin-1β),and tumor necrosis factor-α(TNF-α)in peripheral blood were determined by enzyme-linked immunosorbent assay(ELISA).The expression of relevant signaling pathways was measured by Western blot.Network pharmacological results showed that MLST and SCFF had a total of 153 disease targets,and the key targets were IL-6,TNF,etc.,involving mitogenactivated protein kinase(MAPK)signaling pathway,phosphatidylinositol 3-kinase(PI3K)/protein kinase B(AKT)signaling pathway,etc.The binding energies of the main components and key targets were lower than-7.0 kcal·mol^(-1),indicating that the network analysis results were reliable.The results of animal experiments showed that MLST could reduce the swelling degree and pathological damage of the posterior limb muscles of SCFF rats compared with the model group.ELISA results showed that MLST could reduce the levels of IL-6,IL-1β,and TNF-αin the serum of SCFF rats.Western blot results showed that MLST can reduce the expression of p-AKT,p-PI3K,p-NF-κB,p-p38 MAPK,and p-ERK in SCFF rats.MLST may reduce the content of inflammatory factors in serum by regulating the expression of PI3K/AKT and MAPK-related signaling pathway protein and improving posterior limb muscle SCFF in rats.