| OBJECTIVE:To explore the effects and mechanisms of electro-acupuncture (EA) on different acupoint modules in the rat model of inflammatory pain induced by autologus nucleus pulposus, we established a rat model and evaluated the effects of EA on pain-related behaviors, pathological changes of spinal cord and expression of microglia (MG) and cyclooxygenase 2(COX2) in the rat model of inflammatory pain.METHODS:In this study, a rat model of low back pain was establishedby autologous nucleus pulposus obtained from the coccygeal intervertebral discs of the amputated tail. Rats were randomly divided into normal, model, sham, distal-proximal acupoints, proximal acupoints, distal acupoints. In distal-proximal acupoints group, rats were needled at bilateral L5 Jiajixue (Ex-B2), Dachangshu (BL 25), Weizhong (BL 40) and Kunlun (BL 60). In proximal acupoints group, rats were needled at bilateral L5 Ex-B2 and BL 25. In distal acupoints group, rats were needled at bilateral BL 40 and BL 60. Rats were stimulated at a frequency of 2/100 Hz daily with EA for 20 min during 1 week consecutively. Behavioral tests including assessment of motor function, thermal withdrawal latency (TWL), and mechanical withdrawal threshold (MWT) were performed at 1 day preoperatively and at 3,5 and 7 days postoperatively. After 7 days of intervention, hematoxylin and eosin (HE) method was applied to observe the pathological changes of spinal cord. The spinal COX2 mRNA and EP2 mRNA were detected by RT-PCR technique. Immunohistochemistry and double immunofluorescence staining technique were used to detect microglia and COX2 in the spinal cord.RESULTS:1 Behaviors resultsAfter transplantation of autologus nucleus pulposus, TWL and MWT of rats in model group decreased at 3 days postoperatively, compared with that in the normal group (P<0.05, P< 0.01). It last for 7 days postoperatively. TWL and MWT in EA groups were much higher than that in the model group at 5 and 7 days postoperatively (P<0.01). After administration of EA, TWL among EA groups had no significant differences. MWT in distal-proximal and proximal acupoint group had no obvious differences. MWT in distal-proximal and proximal acupoint group were higher than that in the distal acupoint group at 5 days postoperatively (P<0.01). MWT in distal-proximal acupoint group was higher than that in distal acupoint group at 7 days postoperatively (P<0.05).2 Hematoxylin and eosin staining resultsPathological changes of the spinal cord were evaluated by hematoxylin and eosin (HE) stained at 7 days postoperatively. The inflammatory edema of the spinal cord can be observed in model group. The nerve myelin sheath disintegrates and necrosis. The nerve ganglion cells had numerous cytoplasms. Some nucleus disappeared. Autologus nucleus pulposus triggered the extent of damage of the spinal cord. Although the damages in spinal cord could be observed in three EA groups, they have been prevented to some extent by EA treatment.3 immunohistochemistry ResultsCompared with normal group, the microglia in model group had been highly stained. Cell body was bigger and the positive cell number increased. The optical density was obviously upregulated (P<0.05). The optical density of microglia had no statistical difference between normal and sham group (P>0.05). There were no significant differences between EA and model group (P>0.05). But that in EA groups had a tendency to decline. There is no obvious difference among EA groups. Compared with normal group, COX2 protein in model group was highly expressed and COX2 optical density increased significantly (P<0.01). There is no statistical significance between sham group and normal group (P>0.05). That in EA groups had obvious difference compared with model group (P<0.01). There was no difference among that in EA groups (P> 0.05). But the downward trend of that in distal-proximal and proximal acupoint group was superior to distal acupoint group.It is shown that EA on distal-proximal and proximal acupoints had a better effect on microglia and COX2 than EA on distal acupoints.4 immunofluorescence ResultsIt is showed that COX2 was expressed at a very low level in normal and sham group. The expression of COX2 was enhanced in modle group. The expression of COX2 in three EA groups decreased compared with model group. Microglia (marked by OX42), as resident cells, can be observe in normal spinal cord. OX42 was highly expressed in model group 7 days after operation, whereas it was lowly expressed in EA groups. The expression of COX2 and OX42 has common areas.5 RT-PCR ResultsIt is showed that COX2 mRNA was expressed at a very low level in normal and sham group. COX2 mRNA expression were increased in model group (P<0.05). EA markedly decreased its expression in distal-proximal and proximal acupoint group compared with model group (P <0.05). EA on distal acupoints can not change it significantly (P>0.05). There was no statistical difference between distal-proximal and proximal acupoint group (P>0.05). EA on distal-proximal and proximal acupoints had better effect than distal acupoints. In normal group the expression of EP2 mRNA showed basal constitutive expression in spinal. EP2 mRNA expression were increased in spinal on day 7 after established the model (P<0.05). There were no significant differences between sham group and normal group (P>0.05). Compared with model group, the expression of EP2 mRNA had not seen obvious decrease, but has a downward trend after EA treatment.CONCLUSION:EA can play an analgesic and anti-inflammatory role in the treatment of low back pain. EA on distal-proximal, proximal and distal acupoints can all reverse the up-regulation of microglia and COX2 level in spinal cord and alleviate hyperalgesia to some extent. The effect of EA on distal-proximal acupoints and proximal acupoints was superior to distal acupoints. |
PDF Full Text Request