Clinical Study On Guang Jiekang Treating Knee OA And Experimental Study On Its Influence On The Dedifferentiated Cartilaginous Cells

ⅠLiterature studyWe realize that the etiological root factor of the knee osteoarthritis of TCM is the deficiency of liver and kidney and the external symptom of it is the stagnation of blood and vessels after studying the TCM literature about the knee osteoarthritis. The principle of treating the disease is replenishing the kidney and liver, nourishing qi and promoting blood circulation.The clinical reports about the effect of this type of treating are very positive.However,.as to its mechanism,we rarely know.According to the modern medical research,the etiological factors of knee osteoarthritis are the dedifferentiation of cartilaginous cells in cartilage and the disorder in the cartilage-secretive anabolism.In order to prove the effect of the treating by replenishing the kidney and liver,nourishing qi and promoting blood circulation and to probe into its mechanism,we have designed the experiment.ⅡClinical study1 Objectives:Through clinical studying the effect of the replenishing kidney and promoting blood circulation coupled with articulation lens on treating knee osteoarthritis,we expect to know the preventative effect and the practical characteristics of this method in treating knee osteoarthritis at different phase.2 Methods:46 knee OA sufferers resulted from the deficiency of liver and kidney and the stagnation of blood and vessels,who live up to Chinese and Western clinical diagnosis standards,are randomly divided into two groups:Guanjiekang and Guanjiekang+articulation lens.The treatment lasts 36 weeks.Graded by the different phase of the disease,the patients will be assessed in terms of their pain symptoms,daily function obstacles and their respectively general scores.All the data are analyzed by paired-samples T test and one-way ANOVA and processed by the SPSS11.0 software package.3 Results:After treatment,the average scores of the symptom,daily life dysfunction and general scores of the two groups are notable higher than before with the mitigating knee joint pain and improving articulation function.According to the statistics,the scores of daily life activities of knee joint(namely pain symptom average score,daily life dysfunction average score and the general scores)are significantly higher than before(P＜0.05).No difference existed between the two groups(P＞0.05)who have received 4 and 12 weeks' treatment. But after 32 weeks' treatment,distinct differentiation appears between the two groups with the pain symptom average score,daily life dysfunction average score(P＜0.05)and general scores(P＜0.01).4 Conelusions:Clinical study shows that articulation lens treatment effectively works on the OA patients in short term,which greatly improves functions of patients' joints after 4 weeks' treatment but decreases the magic as time goes on. Thus this treatment can improve the functions of patients' joints,mitigate pains and delay the replacement of artificial joints. On the other hand,Guanjiekang treatment(i.e.using Chinese medicine to replenish kidney and promote blood circulation)doesn't work so effectively on OA patients in short term.But it achieves more satisfactory results in long term. In all,the combination of Chinese medical treatment and articulation lens method is in common use to treat the knee OA,a die-hard disease on which many single treatment methods do not work well.ⅢExperimental study 11 Objectives:Observe the external cultivated and dedifferentiated cartilaginous cells and the change in its biological characters and functions after the cells being cultivated by Chinese medicine serum.Probe into the methods of re-differentiation of the dedifferentiated cartilaginous cells and find a proper external cultivation way for the engineering seedling cells of cartilage.2 Methods:Group enzymatic-digested cartilaginous cells in bacteria-flee rabbit cartilage organization into two:one is comparison group for the single generational cultivation;the other is experimental group for three -generation conventional cultivation which thereafter continues to be cultivated in two weeks by Chinese medicine serum.Observe and compare the cell shapes of the two groups.When the cells come onto the patch,dye the cells with Alcian Blue and test them withⅡcollagen immune tissue so as to find out the surface change in the cells.3.Results:rabbit cartilaginous cells,which are cultivated externally after three generations,dedifferentiate rapidly but proliferate slowly.After added with Chinese medicine serum,the dedifferentiation slows down obviously.And two weeks later,the shape of cells well recover and Alcian blue andⅡcollagen immune tissue both show they are positive.4.Conclusions:The compound cultivation of Chinese medicine serum,which has functions of replenishing kidney and promoting blood circulation,and cartilaginous cells can prolong the dedifferentiation of the cells,advance its re-dedifferentiation,produceⅡcollagen and sugar protein and retain the shape and function of the cells.It is expected to be used in such fields as prevention of dedifferentiation of the cartilaginous cells in external cultivation and advancement in re-dedifferentiation of the dedifferentiated cartilaginous cells.ⅣExperimental study 21.Objectives:Apply the technology of stem cell committed differentiation. Directed differentiate the adipose tissue-derived stem cells(ADSCs)into adipose cells,osteoblast cells and cartilaginous cells and check them up.All these are done to prepare cartilaginous tissue project for acquiring a large amount of seedling cells.2.Methods:cultivate the lipid tissue which is donated voluntarily by the adult patients after liposuction in a generational way.Assess the cell activity and draw the proliferation curve line with tetrazolium salt XTT method.Check up the stem cell mark with flow cell surface antigen analysis.Induce the ADSCs into mature adipose cells,osteoblast cells and cartilaginous cells and then use oil red O, Alizarin red and Alcian blue mark them.Assess the Induced ADSCs withⅡcollagen immune tissue.3.Results:The ADSCs can proliferate externally in a very fast way.Thus its generational cultivation can acquire a large amount of differentiated cells which can be doubled every 50 hours on the average and maintain this stable productive ratio up to 13-15 generations.Among them is there a very small portion of declining and dying cells.The flow cell surface antigen analyzer showing CD29 positive,CD44positive,says that the cells have the characteristics of stem cells which is further testified by the inducement of ADSCs into mature adipose cells, osteoblast cells and cartilaginous cells and check-up by oil red O,Alizarin red and Alcian blue.The positive test byⅡcollagen immune tissue on the differentiated cartilaginous cells shows that the differentiated cells have normal cartilaginous cells' functions.ⅤConclusions:The result-of this study has again confirmed the dedifferentiation of the cartilaginous cells.Besides,the external experiment of using Chinese medicine serum has also confirmed the effective re-dedifferentiation of the dedifferentiated cartilaginous cells,which serves a sound evidence to apply Chinese medicine on repairing OA.However,the specific mechanism is not yet known.We have succeeded in directed differentiation of ADSCs into mature adipose cells,osteoblast cells and cartilaginous cells and check-up by oil red O, Alizarin red and Alcian blue.Under the designated circumstances,after being induced into differentiated cartilaginous cells,the ADSCs serve as reliable seedling cells for the cartilaginous tissue project which may come out a much better result than the pure cartilaginous cells.