| Chronic pain is a common symptom that affects human health physically and psychological. Now it has been accepted as a kind of disease as well as a symptom, most of which are chronic inflammatory pain. Chronic inflammatory pain is one of important problems that confront clinical subjects.At present, COXs inhibitors (NSAIDs) are used for the therapy of chronic inflammatory pain mostly, the side effects of which prevent them from further application. Acupuncture is an effective method of Traditional Chinese Medicine, especially prominent for chronic inflammatory pain, and with few side effects only if properly operated. Great progress has been made about mechanism of electroacupuncture (EA) analgesia. Because of the complexity of transmission and modulation of pain, there are still a lot of phenomenons that have not clarified.It has long been recognized the prostaglandin E2 (PGE2) is involved in the peripheral and central sensitization of chronic inflammatory pain. COX-1 and COX-2 are the key enzyme in the biosynthesis of PGE2. Number of studies showed the expression of COX-2 increased greatly in inflammatory pain, while still had opposite results. COX-1 might also participate in chronic inflammation according to a few reports. It needs further exploration for the potential roles of COX-1 and COX-2 in the occurrence and development of chronic inflammatory pain.A large amount of previous studies in our lab suggested that EA had good immediate and accumulative analgesic effect. There was report that EA could directly regulate the expression of COX-2 of peritoneal macrophage in mice, as well as inhibited PGE2 in spinal cord obviously in inflammatory pain. Does EA operate through inhibition of spinal cord COX-2? Does COX-1 is involved in the anti-inflammatory pain effect of EA? Compared with NSAIDs, what differences do they have on spinal cord COXs?Astrocytes are activated under the chronic inflammatory paincondition. The activation of astrocytes is necessary for the generation and maintenance of pathological pain. They could express and release a great quantity of PGE2 by COX-2. Whether spinal cord astrocytes is involved in the EA analgesic effect?On monoarthritic rat model that injection of CFA into the tibio-tarsal joint of rat produced monoarthritic model, by using behavioural, HE staining, immuneohistochemistry and RT-PCR techniques, we first observed the changes of COX-1 and COX-2 protein expression in spinal dorsal horn to make sure of role of COXs played in chronic inflammatory pain. Then the analgesic effects of EA and Diclofenac on chronic inflammation pain and COXs expression in spinal cord are investigated. We also observed the changes of astrocytes in spinal cord in EA and Diclofenac treatment.1. Changes of COX-1 and COX-2 in spinal cord of the rat model of CFA-induced arthritis1. 1 Changes of COX-1 protein in spinal dorsal horn of the rat model of CFA-induced arthritisThe results showed that COX-1 immunoreactive cells exist in spinal cord of normal rats and relatively in constant expression. The amount of COX-1 immunoreactive cells in spinal dorsal horn of rat model remarkably increased on day 4, then quickly regressed, and on day 6, there was no significant difference compared to normal group until on day 14, suggesting COX-1 protein might be involved in early stage of inflammatory pain.1. 2 Changes of COX-2 protein in spinal dorsal horn of the rat model of CFA-induced arthritisThe results showed that COX-2 immunoreactive cells exist in spinal cord of normal rats and relatively in constant expression. The amount of COX-2 immunoreactive cells in spinal dorsal horn of rat model remarkably increased on day 4, then slowly regressed, but on day 14 it was still above the level of normal group, suggesting that continuous increase of COX-2 expression in spinal cord might be involved in hyperalgesia.1. 3 Changes of the expression of COX-1 mRNA in spinal cord of the rat model of CFA-induced arthritisThe results showed that COX-1 mRNA exists in spinal cord of normal rats and relatively in constant expression. The amount of COX-1 mRNA in spinal cord of model rat remarkably increased on day 4, then quickly regressed, and on day 6, there was no significant difference compared to normal group until on day 14, suggesting COX-1 mRNA might be involved in early stage of inflammatory pain.1. 4 Changes of the expression of COX-2 mRNA in spinal cord of the rat model of CFA-induced arthritisThe results showed that COX-2 mRNA exist in spinal cord of normal rats and relatively in constant expression. The amount of COX-2 mRNA in spinal cord of rat model remarkably increased on day 4, then slowly regressed, but on day 14 it was still above the level of normal group (P<0.05), suggesting that continuous increase of COX-2 mRNA in spinal cord might be involved in hyperalgesia.1.5 Changes of hyperalgesic score of the rat model of CFA-induced arthritisThe results showed that there is no difference between left and right PWL. On day 4 after CFA-induced arthritis, the ipsilateral PWL decreased significantly compared to normal paw, until day 14, suggesting that rats were still on hyperalgesic state.Summary:On day 4 following CFA-induced arthritis, the expression of spinal cord COX-1 and COX-2 protein and mRNA greatly increased, while the rats were hyperalgesia to thermal. On day 14, the expression of COX-2 protein and mRNA still above normal group as well as hyperalgesic score. While the expression of COX-1 Protein and mRNA withdraw to normal level, and kept afterwards. The results suggested that COX-2 might play an important role in occurrence, development and maintenance of chronic inflammatory pain. Continuous increase of COX-2 expression in spinal cord might beinvolved in continue hyperalgesic state. COX-1 might be involved in early stage of inflammatory pain.2. Analgesic effect of electroacupuncture (EA) and diclofenac on the rat model of CFA-induced arthritis and COXs expression in spinal cord2. 1 Effects of EA and diclofenac on hyperalgesic score of the rat model of CFA-induced arthritisThe results showed that on day 4 following CFA-induced arthritis, the latency of injected hind paw was significantly shorter than that of the contralateral hind paw, which remained the same as before CFA injection. EA and diclofenac can significantly decrease the hyperalgesic score, suggesting EA and diclofenac had remarkable accumulative analgesic effects.2.2 Effect of EA on inflammatory tissue repairThe results showed that on day 4 following CFA-induced arthritis, there were severe destruction of the synovium, infiltration of mononuclear cells to sub-synovial tissue, edema, and inflammatory leakage in joint. The tissue pathological changes were significantly reduced by EA, suggesting the tissue repair function of EA.2. 3 Effects of EA and diclofenac on COX-1 and COX-2 in spinal cord of the rat model of CFA-induced arthritis2. 3. 1 Effects of EA and diclofenac on COX-1 protein in spinal dorsal horn of the rat model of CFA-induced arthritisThe results showed that the amount of COX-1 immunoreactive cells in spinal dorsal horn of rat model remarkably increased on day 4, and then quickly regressed, and on day 6, there was no significant difference compared to normal group. There were no difference between EA group and normal group about the number of COX-1 like immunoreactive cells in course of treatment. The number of COX-1 like immunoreactive cells of diclofenac group was lower than that of normal group on day 10 (P<0. 05), suggesting that EA had no obvious influence on expression of COX-1 protein inarthritis; while diclofenac could inhibit the expression of COX-1 proteinsignificantly.2. 3. 2 Effects of EA and diclofenac on COX-2 protein in spinal dorsal hornof the rat model of CFA-induced arthritisThe results showed that the number of COX-2 like immunoreactive cells increased significantly following CFA-induced arthritis, both EA and diclofenac could lowered the COX-2 like immunoreactive cells gradually with accumulative treatments. However after 10 days treatment, the number of COX-2 like immunoreactive cells in diclofenac group was lower than that of normal group (P<0.01), while there were no significant different between EA group and normal group, suggesting that EA could regulate COX-2 protein expression of spinal cord, while diclofenac could inhibit the expression of COX-1 protein significantly.2. 3. 3 Effects of EA and diclofenac on COX-1 mRNA expression in spinal cord of the rat model of CFA-induced arthritisThe results showed that the amount of COX-1 mRNA expression in spinal dorsal horn of rat model remarkably increased on day 4, then quickly regressed, and on day 8, there was no significant difference compared to normal group. There were no difference between EA group and normal group for COX-1 mRNA in course of treatment. The level of COX-1 mRNA in diclofenac group was lower than that of normal group on day 10 (P<0. 05), suggesting that EA had no obvious influence on expression of COX-1 mRNA in arthritis, While diclofenac could inhibit the expression of COX-1 mRNA significantly.2. 3. 4 Effects of EA and diclofenac on COX-2 mRNA expression in spinal cord of the rat model of CFA-induced arthritisThe results showed that the level of COX-2 mRNA increased significantly following CFA-induced arthritis, both EA and diclofenac could lowered the COX-2 mRNA expression gradually with accumulative treatments. However after 10 days treatment, the level of COX-2 mRNA in diclofenac group were lower than that of normal group (P<0.01), while there were no significant different between EA group and normal group(P>0. 05), suggesting that EA could regulate COX-2 protein expression of spinal cord, while diclofenac could inhibit the expression of COX-1 protein significantly.Summary: EA had remarkable analgesic effects on CFA-induced arthritis. In addition to prominent accumulative effect of repeated EA, EA could also promote the repair of inflammatory tissue preferably. EA could adjust the increased expression of COX-2 protein and mRNA to normal, and did no effect on unchanged expression of COX-1 protein and mRNA. Diclofenac could greatly inhibit expression of spinal cord COX-2 and COX-1 protein and mRNA. These were the important differences between EA analgesia and diclofenac.3. The changes of astrocytes in the rat model of CFA-induced arthritis and the analgesic effects of EA and diclofenac on itGFAP like immunoreactive cells existed in spinal cord of normal rats and relatively in constant expression. The number of GFAP like immunoreactive cells began to increase 10 days after CFA-induced arthritis, and kept this tendency on day 14. EA could regulat it to normal group level, while diclofenac inhibited it strongly.Summary: The number of GFAP like immunoreactive cells increased significantly in chronic inflammatory pain. EA could regulate it, while diclofenac inhibited it strongly.
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