Effects of electroacupuncture at "Jiaji"(EX-B2) on autophagy and endoplasmic reticulum stress in spinal cord injury mice

Abstract

To observe the effect of electroacupuncture (EA) at "Jiaji" (EX-B2) on the levels of autophagy and endoplasmic reticulum stress in mice with spinal cord injury (SCI), so as to explore its mechanism underlying improvement of SCI. A total of 60 female C57BL/6 mice were randomly divided into sham operation, model and EA groups， which were further divided into 7 d and 14 d subgroups (10 mice in each subgroup). The SCI model was established by pressing the exposed spinal cord (L1) with a vascular clamp for 15 s. EA was applied to bilateral EX-B2 3 h after modeling, once a day for 7 and 14 d, respectively. Basso Mouse Scale(BMS) for locomotion was used to evaluate hindlimb motor function on day 7 and 14 after SCI. H.E. staining was used to observe histopathologic changes of the injured spinal cord tissue, and Western blot employed to detect the expression of glucose regulatory protein-78 (GRP78), Caspase-12, microtubule-associated protein light chain 3 II (LC-II) and P62(also known as sqstm1/Sequestome1) proteins. Immunofluorescence staining was used to detect the immunoacti-vities of spinal CCAAT/enhancer-binding protein (C/EBP) homologous protein (CHOP, an endoplasmic reticulum stress-inducible protein) and P62. On the 7th and 14th day after SCI, the BMS scores and expression levels of LC3II protein were significantly down-regulated (P<0.05), and the expression levels of P62, GRP78 and Caspase-12 proteins, the immunoactivities of CHOP and P62 were all significantly up-regulated on both day 7 and 14 in the model group than in the sham operation group (P<0.05).Compared with the model group, the BMS scores and the expression levels of LC3II protein were significantly increased on both day 7 and 14 (P<0.05), while the expression levels of P62, GRP78 and Caspase-12 proteins, and the immunoactivities of CHOP and P62 were obviously decreased on day 7 and 14 in the EA group (P<0.05). Outcomes of H.E. stain showed that the cells with nuclei pyknosis and swelling and the necrotic cells appeared in the model group, which was relatively fewer in the EA group. EA of EX-B2 can improve the locomotor function in SCI mice, which may be related to its effects in up-regulating the expression of LC3II (to promote cell autophagy), and down-regulating the expression of P62, GRP78, Caspase-12 and CHOP proteins (to inhibit endoplasmic reticulum stress) in the spinal cord tissue.