To observe the effect of electroacupuncture (EA) of bilateral "Jiaji" (EX-B2) on autophagy, apoptosis and phosphoinositide 3-kinase (PI3K)/protein kinase B (AKT)/mammalian target of rapamycin (mTOR) signaling pathway of nucleus pulposus cells in rabbits with intervertebral disk degeneration (IVDD), so as to explore its possible mechanisms underlying improvement of IVDD.

A total of 18 male New Zealand rabbits were randomly and equally divided into sham operation, model and EA groups. The IVDD model was established by axial disc compression, and the modeling effect was confirmed by magnetic resonance imaging (MRI). EA (2 Hz/15 Hz, 1 mA) was applied to bilateral "Jiaji" (EX-B2) of L4—L5 for 20 min, once daily, 6 times a week for 4 weeks. The general condition and body weight change in each group were recorded. The morphology of nucleus pulposus was observed after H.E. staining. The ultrastructure of nucleus pulposus cells was observed by using transmission electron microscope. The apoptosis of nucleus pulposus cells was detected by terminal deoxynucleotidyl transferase (TdT) dUTP nick-end labeling (TUNEL) assay. The expressions of Bax, Bcl-2, Beclin1, light chain 3 (LC3), autophagy-related protein 5 (Atg5), p62 and PI3K/AKT/mTOR signaling pathway proteins were detected by Western blot.

Compared with the sham operation group, the general condition of IVDD rabbits was poorer, with decreased MRI signal of intervertebral disc, destruction and necrosis of nucleus pulposus cells, damage of nucleus pulposus cell matrix, and blurred organelles, and the apoptosis rate, expression levels of Bax, Beclin1 and Atg5 proteins and the ratio of LC3-Ⅱ/LC3-Ⅰ were significantly increased (P<0.01, P<0.05), while the body weight, expressions of Bcl-2 and p62, p-PI3K/PI3K, p-AKT/AKT and p-mTOR/mTOR considerably decreased (P<0.01) in the model group. In contrast to the model group, the general condition of the rabbits was improved after EA, with enhanced MRI signal of intervertebral disc, fewer mitochondrial autophagosomes, slightly normal mitochondrial construction, etc., and modeling-induced increase of apoptosis rate, the expression levels of Bax, Beclin1 and Atg5 proteins and the ratio of LC3-Ⅱ/LC3-Ⅰ, and decrease of body weight, the expression levels of Bcl-2, p62, and p-PI3K/PI3K, p-AKT/AKT and p-mTOR/mTOR were all reversed in the EA group (P<0.01, P<0.05).

EA stimulation of EX-B2 can delay IVDD by regulating apoptosis and autophagy of nucleus pulposus cells in the intervertebral disc in IVDD rabbits, which may be related to its function in activating PI3K/AKT/mTOR signaling pathway.

To observe the protective function and mechanism of electroacupuncture (EA) “Jiaji” (EX-B2) on dorsal root ganglion (DRG) in rabbits with pressure-induced lumbar intervertebral disc degeneration (IVDD).

Twenty-five male New Zealand rabbits with mature bones were divided into control, sham operation, model, EA, and acupuncture groups, with 5 rabbits in each group. The IVDD model was established by inserting kirschner wires to the vertebral bone surface between the lumbar (L) 4 and L5 vertebrae, followed by applying continuous axial pressure for 28 days. EA (2 Hz/15 Hz, 1 mA) or acupuncture (only insertion of acupuncture needles into bilateral EX-B2, but without electrical stimulation) was applied to bilateral EX-B2 for 20 min, once daily for 28 days.The general conditions and weight changes of rabbits in each group were observed, and the free walking pain score was calculated after interventions. The morphology and structure of DRG cells were observed by HE staining. The activities of superoxide dismutase (SOD), glycine peroxidase (GSH-Px) catalase (CAT) and content of malondialdehyde (MDA) in DRG were detected by ELISA. The concentration of calcium ions in DRG was measured by flow cytometry. The protein expression levels of brain-derived neurotrophic factor (BDNF), nuclear factor E2 related factor 2 (NRF2), heme oxygenase-1 (HO-1) and NAD (P) quinone oxidoreductase 1 (NQO1) in DRG were detected by Western blot.

Compared with the sham operation group, the free walking pain score of rabbifs in the model group was significantly increased (P<0.01), and the concentration of calcium ion and content of MDA in DRG were significantly increased (P<0.01), the contents of SOD, GSH-PX and CAT and the protein expressions of BDNF, NRF2, NQO1 and HO-1 in DRG were significantly decreased (P<0.01). Compared with the model group, the free walking pain score of rabbits in the EA group was significantly decreased (P<0.01), the concentration of calcium ions and the content of MDA in DRG were decreased (P<0.01), the contents of SOD, GSH-PX and CAT and the protein expressions of BDNF, NRF2, NQO1 and HO-1 were significantly increased (P<0.01). The effect of EA was better than acupuncture (P<0.01, P<0.05). HE staining showed atrophied cell bodies, disordered cellular arrangement, dissolved nuclei, and indistinct borders of DRG cells in the model group, which was milder in both acupuncture and EA groups.

EA at EX-B2 may inhibit oxidative stress in DRG cells by regulating BDNF/NRF2 signaling pathway, protect DRG cell damage, reduce nerve pain, thus delay the progression of IVDD.