[Visual acupotomy intervention mitigates pain reaction by improving intervertebral disc degeneration and inhibiting apoptosis of nucleus pulposus cells in rabbits with cervical spondylosis].

OBJECTIVE: To investigate the effect of visual acupotomy intervention on intervertebral disc degeneration, nucleus pulposus cell apoptosis and expression of apoptosis related proteins in rabbits with cervical spondylosis (CS), so as to explore its mechanism underlying improvement of CS. METHODS: A total of 48 male New Zealand rabbits were randomly divided into blank control, model, acupotomy and medication (meloxicam) groups, with 12 rabbits in each group. The neck type CS model was established by forcing the rabbit to make a neck flexion for 5 hours in a restrained chamber, once daily for 12 weeks. Rabbits of the medication group received an intramuscular injection of meloxicam (0.35 mg/kg), once daily for 4 consecutive weeks, and those of the acupotomy group received ultrasound-guided acupotomy intervention, once a week for 4 weeks. The pain threshold (PT) was measured by using a VonFrey electronic pain detector. The levels of prostaglandin E2 (PGE2), 5-hydroxytryptamine (5-HT) and substance P (SP) in serum were detected by ELISA. The severity of intervertebral disc degeneration was observed by using magnetic resonance imaging (MRI) and given scores in accordance with Suzuki's and colleague's "new classification system of cervical disk degeneration". The apoptosis of nucleus pulposus cells was analyzed by TUNEL staining. The protein expression levels of apoptosis-related protein Fas, cysteinyl aspartate-specific protease-3 (Caspase-3), B-cell lymphoma-2 asso-ciated X protein (Bax) and B-cell lymphoma-2 protein (Bcl-2) were measured by Western blot. RESULTS: Compared with the blank control group, the PT and Bcl-2 expression and MRI score were significantly down-regulated (P<0.01, P<0.001), whereas the contents of serum PGE2, 5-HT and SP, ratios of TUNEL-positive cells, and expression of Fas, Caspase-3 and Bax were considerably up-regulated (P<0.001, P<0.05, P<0.01) in the model group. In contrast to the model group, both the medication and acupotomy groups had an obvious increase in the levels of PT and Bcl-2 expression and MRI score (P<0.05, P<0.01), and a significant decrease in the contents of serum PGE2, 5-HT, SP, ratios of TUNEL-positive cells, and expression of Fas, Caspase-3 and Bax proteins (P<0.05). No significant differences were found between the medication and acupotomy groups in all the indexes mentioned above (P>0.05). CONCLUSION: Visual acupotomy intervention can mitigate the pain state of CS rabbits, which may be related to its functions in improving the intervertebral disc degeneration, reducing inflammatory reactions and apoptosis of nucleus pulposus cells.

[Effect of acupotomy intervention on cervicomuscular apoptosis in cervical spondylosis rabbits].

OBJECTIVE: To observe the effect of acupotomy therapy on cervicomuscular apoptosis and apoptosis regulator Bax protein expression in cervical spondylosis (CS) rabbits so as to investigate its mechanisms underlying improvement of CS. METHODS: Twenty-four New Zealand rabbits were randomly divided into normal control, model, acupotomy and electroacupuncture (EA) groups, with 6 rabbits in each group. The CS model was made by forced head-bowing for 5 hours in a restrained chamber, once daily for 12 weeks. Acupotomy was performed at the starting point of trapezius, the mastoid process attaching point of sternocleidomastoid, the cerverical vertebrae joint process or the local induration or cord-like mass (2 or 3 points of them were used as the needle-knife entering points), once a week for 3 weeks. For animals of the EA group, EA (2 Hz/100 Hz, 2 mA) was applied to bilateral "Tianzhu" (BL 10), "Jingbailao" (EX-HN 15), "Dazhu" (BL 11) for 20 min, once daily and 3 times a week for 3 weeks. The number of apoptotic cells in the cervical muscle was observed by light microscope after TUNEL staining and muscular Bcl-2 and Bax protein expression was detected by Western blot. RESULTS: In comparison with the control group, the number of cervicomuscular apoptotic cells, and the expression level of cervicomuscular Bax protein were significantly increased, and the Bcl-2/Bax was obviously decreased in the model group (P < 0.01, P < 0.05). Compared to the model group, the number of apoptotic cells and the expression level of muscular Bax protein were notably decreased in the acupotomy group (P < 0.01, P < 0.05), while the ratio of Bcl-2 and Bax was apparently increased in the acupotomy group (P < 0.05). The effects of acupotomy were significantly superior to those of EA in lowering apoptotic cell number and in up-regulating Bcl-2/Bax (P < 0.01, P < 0.05). No significant differences were found between the EA and model groups in the apoptotic cell number and among the four groups in Bcl-2 protein expression levels (P > 0.05). CONCLUSION: Acupotomy therapy can reduce cervicomuscular cellular apoptosis and Bax protein expression in CS rabbits, which may be one of its mechanism underlying improving CS.

Electroacupuncture inhibits annulus fibrosis cell apoptosis in vivo via TNF-α-TNFR1-caspase-8 and integrin ß1/Akt signaling pathways.

OBJECTIVE: To examine whether electroacupuncture (EA) treatment inhibited cell apoptosis of intervertebral annulus fibrosis (AF) via tumor necrosis factor-α (TNF-α)-tumor necrosis factor receptor 1 (TNFR1)-caspase-8 and integrin ß1/Akt signaling pathways in a rat model of cervical intervertebral disc degeneration caused by unbalanced dynamic and static forces. METHODS: Thirty-two Sprague-Dawley rats were included in this study, of which 24 rats underwent surgery to induce cervical intervertebral disc degeneration, while eight rats received EA treatment at Dazhui (GV 14). Immunohistochemical staining was used to detect TNF-α, TNFR1, and caspase-8. Apoptosis of AF cells was examined with terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL) staining. The mRNA and protein expression levels of integrin ß1 and Akt were evaluated with real-time polymerase chain reaction and western blot analysis, respectively. RESULTS: Treatment with EA decreased TUNEL-positive AF cells and lowered TNF-α, TNFR1 and caspase-8 positive cells compared with control groups. EA treatment also increased integrin ß1 and Akt mRNA and protein levels compared with controls. CONCLUSION: Treatment with EA inhibits AF cell apoptosis through suppression of the TNF-α-TNFR1-caspase-8 signal pathway and increases the expression of integrin ß1 and Akt. EA may be a good alternative therapy for treating cervical spondylosis.

[Effects of electroacupuncture on Wnt-ß-catenin signal pathway in annulus fibrosus cells in intervertebral disc in rats with cervical spondylosis].

OBJECTIVE: To observe the effects of electroacupuncture (EA) at "Dazhui" (GV 14) on Wnt-ß-catenin signal pathway in annulus fibrosus cells in intervertebral disc in rats with cervical spondylosis. METHODS: Forty SD rats were randomized into a control group, a model group, an EA group and a medication group, 10 rats in each one. Rats in the control group were treated with sham operation, only incision on local skin; rats in the remaining groups were made into cervical spondylosis models. After model establishment, rats in the control group and model group received fixed treatment under identical condition; rats in the EA group were treated with EA at "Dazhui" (GV 14), 30 min per treatment; rats in the medication group were treated with intragastric administration of meloxicam tablets. Treatments were both given once a day, and 14 days were taken as one session; there was an interval of 2 days between two sessions, and totally two sessions were given. After the treatments, immunohistochemistry was applied to measure the expression of Wnt, glycogen synthase kinase-3ß (GSK-3ß) and Axin in annulus fibrosus cells; western blot was used to test the expression of P-ß-catenin. RESULTS: In the control group, there were more positive cells of Wnt, GSK-3ß and Axin, which were intensively distributed, deeply colored, and strongly positive; In the model group, there were less positive cells of Wnt, GSK-3ß and Axin, which were sparsely distributed and weakly positive. The expression of Wnt, GSK-3ß, Axin and P-ß-catenin in the model group was less than that in the control group (all P < 0.05); expression of Wnt, GSK-3ß, Axin and P-ß-catenin in the EA group and medication group was higher than that in the model group (all P < 0.05); expression of Wnt, GSK-3ß, Axin and P-ß-catenin was not significantly different between EA group and medication group (all P > 0.05). CONCLUSION: EA could delay the degeneration of intervertebral disc, which may be related to EA inhibiting signal pathway of Wnt-ß-catenin.