| Osteoarthritis is a common degenerative joint disease, more common and frequently in the elderly, in three of the most common diseases by elderly patients (the most common cardiovascular disease, osteoporosis, osteoarthritis), the incidence rate of osteoarthritis in the first place in the world.Osteoarthritis occurs in the weight-bearing larger spine, knee, hip and other parts, and the main lesion is degeneration of articular cartilage and secondary bone hyperplasia, clinical manifestations of slowly progressive development of joint pain, swelling, stiffness, with limited mobility.Severely affected patients in daily life and work, and even can cause disability. With the population aging process accelerated, this problem becomes increasingly prominent. Therefore, the prevention and treatment of osteoarthritis has received widespread attention in the community, and the osteoarthritis research has important practical significance.Currently the etiology and pathogenesis of this disease is not clear.Due to weight bearing of knee joint and activity of it is larger, so the knee osteoarthritis (KOA) is one of the most common bone arthritis, generally conservative treatment is more difficult and easy recurrence.For more than30years, acupotomy as a new, minimally invasive technique of traditional Chinese medicine,had better curative effect in the treatment of KOA, but basic research remains to be in-depth.This study through experiments to explore possible mechanisms of acupotomy for treating knee osteoarthritis, lay a foundation for further research, and provide theoretical basis for clinical treatment.ObjectiveThe experiment used acupotomy intervention and compared with electric acupuncture group, to observe acupotomy intervention on animal models of osteoarthritis articular cartilage tissue type â…¡collagen (Col-â…¡), aggrecan (Aggrecan), matrix metalloproteinases-3(MMP-3) gene and protein expression, to explore the possible mechanism of knee osteoarthritis acupotomy intervention from the perspective of molecular biology, to lay the foundation for the later research, and also provide a theoretical basis for clinical treatment of knee osteoarthritis.MethodsSix-month-old healthy adult New Zealand rabbits(equal between genders) were divided into four groups randomly:normal control group, model group, acupotomy group,electroacupunctur(EA) group. The left hind limb immobilization in extension position braking method (Vidman method) for preparing animal models of knee osteoarthritis. Acupotomy group and electro-acupuncture group began to intervene after one week at the end of molding, acupotomy group intervention for one time per week, a total of three weeks of intervention, electric acupuncture group was treated one time on alternate days, treatment three times in one weeks, a total of three weeks of treatment. Before and after treatment knee joint behavior index were observed in rabbit; gross observed pathological manifestations of the articular cartilage and synovial membrane; pathology of rabbit cartilage slices were observed under optical microscope; Real-time PCR method to detect Col-â…¡mRNA, AggrecanmRNA, MMP-3mRNA expression level, and Western blot method to detect Col-â…¡,Aggrecan, MMP-3protein expression level.Results1General observation:Normal control group rabbits knee joints were not swelling, congestion, joint fluids transparent, articular cartilages pale blue, translucent, smooth. Synovial membranes was without edema, hypertrophy.Model group rabbits knee joints were significantly swelling or congestion. Joint fluid was pale yellow and slightly turbid.Articular cartilages were gray, dull, not smooth, and not opaque.There were defect.Ulcers and fissures could be seen clearly. Some subchondral bone was exposed, medial tibial condyle more obvious, synovial hyperemia, edema, hypertrophy.Acupotomy group rabbits the left knee joints mild swelling and had no significant congestion, cartilage pale color, bright color, mild defect, and the obvious ulcers was not seen.Joint fluid was transparent and slightly yellow.Compared with the model group,edema and hyperemia was significantly ease. EA group articular cartilages transparency reduced and were less shin. Some visible cracks could be seen.Synovial hyperplasia reduced,and there was not significant congestion.Indicating acupotomy, electro-acupuncture treatment can relieve cartilage damage,and joint edema.2Optical microscope observation:Knee joint articular cartilages of normal control group were structural integrity.Articular cartilage surface was smooth.Thickness was normal. Cartilage cells arranged in neat rows, and layered clear, orderly. Cartilage matrix was homogencously,and tide line was clear and complete, no pannus formation. In model group, the cartilage surface was not smooth.The surface of cartilage cells were scarce.Cells arranged irregularly. Levels were disturbance.Sometimes visible peeling defect was as deep as the tide line. Chondrocytes nested.Tide line blurred or double tide line was seen, part of the cartilage pannus formation and vascular. Acupotomy group and electro-acupuncture group cartilage cells arranged in neat rows. Cartilage fracture was rare.But local cells arranging in disorder still can be seen. Repetitive tide line was occasionally seen.Cartilage surface was smooth,without pannus formation. The sections were scored according to Mankin method.Four groups overall score were significantly different (P<0.01). Acupotomy group comparing with model group had significant difference (P<0.01); and compareing with the control group had significant difference (P<0.05).There was not significant difference as compared with electroacupuncture group (P>0.05). EA group comparing with the control group had significant difference (P<0.01), and no significant difference as compared with the model group (P>0.05). Acupotomy method had cartilage protective effect.3Behavior changes:the model group and acupotomy group, acupuncture group Lequesne MG score before treatment had no significant statistical difference (P>0.05).The score difference before and after treatment, and score after treatment had significant difference (P<0.05).Acupotomy therapy is effective in improving the rabbit KOA joint pain, swelling and gait, activity.4Real-time PCR experimental results showed Col-â…¡mRNA content expression of acupotomy group increased more significantly than that of model group, and there were significant differences between the two (P<0.05). Compared with model group, Col-â…¡mRNA content expression of electro-acupuncture group significantly increased, and there were significant differences between them(P<0.05). Electroacupuncture group had significant difference with normal control group (P<0.05). Cartilage AggrecanmRNA content expression of model group was significantly decreased,compared with control group,and there were significant differences between the two compared (P<0.01).The expression of MMP-3mRNA in model group was significantly increased compared with normal control group, and there were significant differences between the two (P<0.05).Each group decreased as compared with the model group, but no significant difference compared with the normal control group (P>0.05).Acupotomy group decreased as compared with the model group, but no significant difference between the two (P>0.05). The results of Real-time PCR tests showed that acupotomy could effective up-regulated expression of Col-â…¡mRNA and AggrecanmRNA, down-regulated MMP-3mRNA expression, and the effect of acupotomy group was better than electroacupuncture group.5Western blot experimental results showed Coll-â…¡protein expression in cartilage of model group significantly decreased than that of normal control group group, and there were significant differences between the two (P<0.01). Acupotomy group increased significantly compared with model group, the two had very significant difference (P<0.01). There was significant difference between acupotomy group and electroacupuncture group (P<0.05).The expression of aggrecan protein in acupotomy group was higher than that of model group, and there was significant differences between the two (P<0.05. Electroacupuncture group and model group had no significant difference (P>0.05). The expression of MMP-3protein in model group was significantly increased compared with normal control group, and there was significant differences between the two(P<0.01), Acupotomy group was significantly lower than the model group, and the two had significant differences(P<0.01).There was significant difference between acupotomy group and the electroacupuncture group (P<0.05). The results of Real-time PCR tests showed that acupotomy could effectively increase expression of Col-â…¡protein and Aggrecan protein,reduced expression of MMP-3protein, and the effect of acupotomy group was better than control group.Conclusions1Gene and protein expression of MMP-3in articular cartilage model group rabbits significantly increased, while the gene and protein content expression in the cartilage matrix of Coll-â…¡and Aggrecan decreased obviously, sand howed that matrix metalloproteinases could well reflect the changes of knee cartilage metabolism, It had a role in the degradation of the extracellular matrix of articular cartilage, and could accelerate the degeneration of cartilage.2Acupotomy can make the gene and protein content expression of MMP-3in articular cartilage decreased, and gene and protein content expression in the matrix of Col-â…¡and Aggrecan elevated, showing that acupotomy was inhibition of extracellular matrix collagen degradation, protection to promote and stimulate cartilage repair function.Provided the basis for the experimental study of acupotomy treatment of osteoarthritis of the knee, and reveaed the scientific connotation of acupotomy in treating knee joint osteoarthritis. |
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