Effect of electroacupuncture of "Weizhong" (BL40) on iron metabolism disorder in rats with lumbar multifidus muscle injury

Abstract

To observe the effect of electroacupuncture (EA) of "Weizhong" (BL40) on the disorder of iron metabolism and the level of oxidative stress after lumbar multifidus muscle injury (LMMI), so as to explore its mechanisms underlying promoting the repair of LMMI. Male SD rats were randomly divided into normal, model and EA groups (6 rats in each group). The LMMI model was established by injecting 0.5% bupivacaine (BPVC) solution (400 μL) into the lumbar multifidus muscle with the syringe-needle close to the spinous process (L4-L5). Twenty-four hours after successful establishment of the model, EA (2 Hz/15 Hz, 2 mA) was applied to bilateral BL40 for 30 min, once a day for 2 days. Histopathological changes of the multifid muscle were observed under microscope after H.E. staining, and the iron granules in the multifid muscle tissue observed after Prussian blue staining. The expression of glutathione synthase (GSS) was detected by Western blot, and the expressions of iron regulatory protein 1 (IRP1), ferroportin (Fpn), ferritin heavy chain 1 (FTH1, iron metabolism-related proteins) and gluta-thione peroxidase 4 (GPX4, functions in protecting cells against detrimental lipid peroxidation and governing a novel form of regulated necrotic cell death, called ferroptosis) mRNAs were detected by quantitative real-time PCR. The contents of glutathione (GSH) and malondialdehyde (MDA) were measured by biochemical methods. H.E. staining showed large areas of necrosis and breakage of muscle fibers, disordered arrangement of muscle fibers, widened muscle cell space, accompanying with a large number of inflammatory cell infiltration in the multifidus muscle tissue of the model group, which was relatively milder in the EA group. Outcomes of Prussian blue staining showed that compared with the normal group, there were more iron particles in the multifidus muscle tissue and enlarged muscle fiber gaps, which was also milder in the EA group. Compared with the normal group, the expression level of IRP1 mRNA and content of MDA were significantly increased (P<0.001), the expression levels of Fpn, FTH1 and GPX4 mRNAs and GSS protein, and the content of GSH were considerably decreased (P<0.001) in the model group. In comparison with the model group, the increase of IRP1 mRNA expression and MDA content, as well as the decrease of Fpn, FTH1 and GPX4 mRNAs expressions and GSH content were reversed in the EA group (P<0.001,P<0.05,P<0.01). EA of BL40 has a protect effect in BPVC-induced injury of lumbar multifidus muscle in rats, which may be related to its functions in improving iron metabolism to reduce oxidative damage by regulating expression of IRP1, Fpn and FTH1.