| Objective:To investigate the effect of moxibustion and thermosensitive moxibustion on the connective tissue of Knee osteoarthritis(KOA)model rats and its mechanism.Methods:1.Rat KOA model was prepared by papain injection.SPF class rats were randomly divided into groups.After skin preparation anesthesia,4% papain solution was injected into the bilateral knee cavity of the rats to establish the rat KOA model.The morphologic effects of thermosensitive moxibustion on articular cartilage of rats were observed by HE staining and saffron-solid green staining.2.The contents of IL-1β and TGF-β1 in serum of rats were determined by enzyme-linked immunosorbent assay(ELISA),and the content of NO in joint fluid was determined by nitrate reductase assay.3.HE staining was used to observe the effect of thermosensitive moxibustion on rat synovial tissue.4.HE and Masson staining were used to observe the effect of thermal sensitive moxibustion on the collagen fibers of the superficial fascia of rats.5.Immunohistochemistry(SP)and western blot(WB)were used to observe the expression of ERK1/2 and P-ERK1/2 protein in fascia and synovium of rats under acupoints.Results:1.Morphological changes of articular cartilage in rats:In the normal group,the cartilage surface was smooth,the cartilage layer was thick,the chondrocytes were many and orderly,the tide line was clear and coherent,and the structure was clear.In the model group,the cartilage surface was rough and broken,the cells were reduced,and the tidal line was interrupted disorderly.In other words,papain injection into knee joint can effectively establish the model of knee osteoarthritis in rats.2.Changes of serum IL-1β,TGF-β1 and NO content in the joint fluid of rats:Compared with the normal group,the contents of IL-1β and NO were increased in the model group,moxibustion group and thermosensitive moxibustion group,while the content of TGF-β1 was decreased.Compared with model group,the contents of IL-1β and NO in thermosensitive moxibustion group and moxibustion group were significantly decreased(P<0.01),TGF-β1 content was significantly increased(P<0.01).Compared with moxibustion group,the contents of IL-1β and NO in serum of rats in thermosensitive moxibustion group decreased significantly(P<0.01),TGF-β1 content was significantly increased(P<0.01).3.Effect of heat-sensitive moxibustion on synovium of rats by HE staining:Compared with the normal group,synovial cell proliferation,inflammatory cell infiltration,fibrous tissue hyperplasia and vascular hyperplasia were significantly increased in the model group(P<0.01).Compared with model group,synovial cell proliferation,inflammatory cell infiltration,fibrous tissue hyperplasia and vascular hyperplasia were significantly decreased in moxibustion group and thermosensitive moxibustion group(P<0.01).Compared with the moxibustion group,synovial hyperplasia,fibrous tissue hyperplasia and inflammatory cell infiltration were all improved in the thermosensitive moxibustion group(P<0.01).4.Effect of thermosensitive moxibustion on superficial Fascia of rats by HE and Masson staining:HE staining: in the normal group and the model group,the fibroblast synovial cells were "star" or "fusiform",with fuzzy boundaries,dark and small nuclei,and less distribution of plasmacytic synovial cells.The cell body resembled the round shape,with large and round nuclei,darker staining,and disordered arrangement of collagen fibers.After the treatment of moxibustion,the collagen fibers were distributed in a wavy way,and the cross was a net.The long fiber cells were flat or "fusiform",and the plasma cells were more distributed.Masson staining: the collagen fibers in the normal group showed a longitudinal distribution,and after moxibustion treatment,they showed a wavy distribution,and the content of collagen fibers increased significantly.5.Immunohistochemistry and Western-blot:Immunohistochemistry showed that the binding area was brownish yellow,and the positive binding area was mostly concentrated in fibroblasts,and the cytoplasm and nucleus were stained.ERK1/2 was expressed in fascial tissue and synovial tissue under acupoints in the acupoint area of rats in each group to different degrees,and the contents of ERK1/2 and p-ERK1/2 were increased,and the p-ERK1/2 content was increased more obviously,and the effect of thermosensitive moxibustion was significantly better than that of moxibustion group,the difference was statistically significant(P<0.01).Conclusion:1.The model of knee osteoarthritis can be effectively prepared by injecting papain into the knee cavity of rats.2.Moxibustion method can reduce the level of NO in the blood IL-1β and joint fluid,and increase the level of TGF-β1 in the blood,so as to relieve the symptoms of knee osteoarthritis.3.Moxibustion method may achieve its therapeutic effect by affecting the morphological changes of connective tissue at the local acupoints and lesions(synovium),and its further mechanism may be realized by activating the ERK1/2-MAPK signaling pathway. |
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