| Objective The Prevention and treatment of chronic joint diseases throu gh Long term oral medicine decoction is a specific embodiment o f the traditional Chinese medicine theory of "treating the dise ase with no cure".Considering the knee osteoarthritis due to " deficiency in origin and excess in superficiality " triggers,We choose the prescription of Duhuojisheng decoction as the stu dy object from many in the treatment of osteoarthritis.We put the “Consolidating Basis and Qu Xie” as the theoretical basis of treatment.After the establishment of animal model of OA,we use Duhuojisheng Decoction on experimental animal intervention.In the 8 week after modeling,the cartilage specimens were col lected to observe the histomorphology.To detect the content of Prostaglandin 2(PGE2),interleukin-1?(IL-1?),tumor necrosi s factor-?(TNF-?),interleukin-6(IL-6)in the synovial flui d;To detect the expression of matrix metalloproteinase-3(MMP-3)m RNA and tissue inhibitor of metalloproteinase-1(TIMP-1)m RNA in OA chondrocytes.To detect the expression of type II col lagen and extracellular matrix protein in Polysaccharides;And t o explore the main effect of Duhuojisheng Decoction in the prevention and treatment of OA in the process of target point.Prov ide experimental evidence for Duhuojisheng Decoction in the pre vention and treatment of OA.Methods 40 New Zealand rabbits were divided into five groups at ran dom:the normal group(A),model control group(B),Duhuojisheng Dec ocion of low dose groups(C),Duhuojisheng Decoction middle dose group(D)and the Duhuojisheng Decoction of high dose group(E),8 for each group.These animals were established models with plas ter cast unilateral hindlimb straighten place breaking method e xcept for the rabbits in group A,and intervention was used afte r molding on the second day.Duhuojisheng Decoction group was gi ven medicine by concentrated liquid,once a day,the normal grou p and model group were gastriced with isodose physiological sal ine,once a day,8 weeks in all.After the end of the treatment,al l the experimental rabbits were fasting 12 hours,and then all o f them were killed by Air embolism under anesthesia.During this period,we gave the rabbit knee joint X-ray films.Synovial flu id was taken before opening the rabbit knee joint.1 Concentrations of IL-1?,IL-6,PGE2 and TNF-a in joint syno vial fluid were detected by Enzyme-linked immunosorbent assay(E LISA),and compared the differences between each two groups.2 After opening the rabbit knee joint in experimental side,o bserved and scorded the articular cartilage of the rabbit‘s kn ee joint according to previous literature and compared the diff erences between each two groups.The histological examinations o f rabbits knee joint cartilages were performed by HE staining u nder Light microscopy,and compared the differences between each two groups.3 The MMP-3 and TIMP-1 m RNA expression levels of rabbits kne e joint cartilages were performed by RT-PCR technology,and com pared the differences between each two groups.4 The GAG and Collagen II expression levels of rabbits knee joint cartilages were measured by Immunohistochemical method co mbined with Western blot method,and compared the differences b etween each two groups.Results 1 After the establishment of rabbit knee joint model,Visibl e normal animal knee joint appearance without obvious swelling and deformity,and there was no significant thickening of the p osterior articular capsule,Synovial morphology was normal witho ut obvious hyperplasia,The Articular cartilage was complete,fl exible and without defect area;The appearance of joint swellin g significantly in model group.We could see the knee joint caps ule thickening after opening it,the color of articular cartilag e is dim,the wear of articular cartilage is relatively serious,and surface ulcer was formed,part of the surface of the cartil age defect.The low dose group showed slight swelling of joint a ppearance,we could see the joint capsule was mild hyperplasia-after opening it,the articular cartilage surface was slightly rough and have a few cracks,there was no obvious defect and ulcer surface of articular cartilage.There is no cartilage def ects and cracks in the middle and high dose group.the joint cap sule and synovial without hyperplasia,the articular cartilage surface was smooth,The blank group Mankin score was the lowest,On the contrary the highest score belong to the model group,The blank group and Duhuojisheng Decoction in low,medium and hi gh dose group was compared with the model group,showing statis tical difference between them was significant(P<0.01)2 Double antibody sandwich enzyme-linked immunosorbent assa y(ELISA)shows: Duhuojisheng Decoction group IL-1?,tumor nec rosis factor,IL-6,PGE2 and other inflammatory factors were si gnificantly lower than model group,the difference was statisti cally significant(p<0.01).The IL-1?,tumor necrosis factor,I L-6,PGE2 in middle and high dose group is lower than low dose group,and the difference was also statistically significant(p<0.01).The IL-1?,tumor necrosis factor,IL-6,PGE2 in middle group is higher than high dose group,There were no significant differences between the groups in middle and high dose(p>0.05).3 Realtime-PCR showed that after different doses of Duhuoji sheng decoction treatment,it could reduce the MMP-3 m RNA in ca rtilage cells of OA animal model,however TIMP-1 m RNA high expr ession.The expression of MMP-3 m RNA in articular cartilage of model group was higher,but the TIMP-1 m RNA was significantly l ower than that in model group.There was a significant differen ce of Duhuojisheng Decoction groups and model group(p<0.01).4 After the intervention,it shows that there was significan t difference between the expression of Duhuojisheng Decoction o f type II collagen in the middle and high dose group and model group(p<0.01).There was no significant difference between low dose group and model group(p>0.05).Between the middle and hi gh dose group was not statistically significant(p>0.05).The c ontent of GAG in middle and high dose group was significantly higher than that in model group,the difference was statisticall y significant(p<0.01).The content of GAG in low dose group wa s higher than that in model group,the difference was statistic ally significant(p<0.05).The content of GAG in middle and hig h dose group was significantly higher than that in low dose gro up(p<0.01).The content of GAG in the middle dose group was lo wer than that in the high dose group,and there was no signific ant difference between the two groups(p>0.05).Conclusions 1 The model of rabbit's knee OA can be successful establis hed with the modified fixed extension type for 8 weeks.2 The Duhuojisheng Decocion can decrease the level of infla mmatory factors in joint synovial fluid contents of IL-1?,IL-6,PGE2 and TNF-a.It also can increase the expression of TIMP-1 m R NA and reduce the MMP-3 m RNA expression.The effective concentr ation of Duhuojisheng Decocion can promote the synthesis of typ e II collagen and delay the degradation process,through the wa y it can increase expression of GAG content in OA cartilage tis sue to delay the progression of the disease.3 Duhuojisheng decoction is to remove cold dampness,nouris hing liver and kidney,supplementing qi and blood and supplemen ted by drugs,in the treatment of KOA,Medicine Duhuojisheng De coction prescription in the use of each other better than singl e drug effect.Animal experiments have proved that the treatmen t of Duhuojisheng decoction could protect the articular cartila ge of rabbit knee OA. |
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