Tóm tắt Luận án Assessment toxicity and efficacity of ich goi khang in knee osteoarthritis treatment

tis diagnosis According to American College of Rheumatology 1991 (ACR 1991) 1.1.3. Treatment The mainstay of treatment was relieving pain and motion range promote. There are multi-treatments. In order to relieve the pain: ACR recommends stage by stage, starting with acetaminophen (paracetamol, efferalgan), then NSAIDs. In combination with SYSADOA group, and others local treatment such as: PRP and stem cells. Surgical treatments: lavage, replacement Physical therapies: lose weight, cast 1.2. Traditional medicine therapy 1.2.1. Diagnosis and mechanism In traditional medicine, the knee OA is called Hac Tat Phong. It is imagined as the obstruction causing the disease. The main mechanisms are: trauma, exotic and intrinsic factors. 1.2.2. Discussion of the clinical types There are three types:”Phong han thap ty”, “Phong han thap ty and can than hu”, “phong thap nhiet ty and can than hu”. The treatment for the first type: “khu phong, tan han, tru thap, chi thong”, 4 for the second type: “khu phong, tan han, tru thap, chi thong, bo can than”, for the third type “thanh nhiet, khu phong, tru thap, chi thong, bo can than”. 1.3. The model to evaluate the toxicity and treatment efficatity of Knee osteoarthritis Origin of the Prunus persica and Ramulus Cinnamomi with additional ingredients, the ingredients could be modified, therefore Ich goi khang need to be qualified the toxicity and another further investigation. The evaluation was the acute and sub-acute toxicity; inflammatory effect, pain reliever in the studied model. 1.4. The previous knowledge of Knee osteoarthritis There was various researches of knee osteoarthritis (OA) in the world. Liu (2014) was conducted a research of pain characteristics in knee OA patients which showed that intermittent pain in 46% cases, continuous pain in 97%. Dinh Thi Dieu Hang (2013) in the research of OA in Hai Duong: mechanical pain 92,8%; nocturne pain 71,9%; radiograph characteristics 78% narrow joint, osteophyte 65%. In Vietnam, there is lacking of the OA research. In term of modern and traditional medicine combination, Dinh Thi Lam (2011) conducted a research in 60 OA patients treated by glucosamine and Agelica laxiflora and Herba taxilli, the proportion of improved patients after 60 days according to the Lequesne score was 26.67%. Nguyen Thu Thuy (2014) in the thesis “Efficacity assessment of Herba taxilli in combination with the pulse wave” showed the pain reliever efficacity in both patients groups. 1.5. The general description of the studied medication 1.5.1. Origin and effect It based on the two ingredients Ramulus cinnamomi and Prunus persica of Truong Ngoc Ban – China. This medication includes: Semen Pruni, Flos Carthami tinctorii, Gummi resina Olibanum, Herba Asari, Radix Angelicae sinensis, Radix Angelicae pubescentis, 5 Radix Saposhnikoviae divaricatae, Rhizoma Atractylodis, Resina myrrhae, Radix Notopterygii, Radix Clematidis, Frutus Chanomelis. This medication aims to treat the patients caused by the exotique factors, especially in knee osteoarthritis patients. Ich goi khang received the result of tradition medicine and added on some others ingredients. 1.5.2 The ingredients and the function of each ingredient Ingredients: Semen Pruni 40g Cortex Schefflerae heptaphyllae 50g Flos Carthami tinctorii 20g Radix Angelicae pubescentis 40g Herba Loranthi Gracifilolii 50g Rhizoma Atractylodis 40g Ramulus Cinnamomi 20g Herba Siegesbeckiae 50g Radix et Rhizoma Glycyrrhizae 20g Rhizoma Homalomenae occultae 40g Gummi resina Olibanum 20g Radix Achyranthis bidentatae 50g Caulis Tinosporae tomentosae 50g Cortex Eucommiae 40g Radix Angelicae sinensis 40g Semen Coicis 50g Radix Paeoniae lactiflorae 40g Radix Saposhnikoviae divaricatae 50g Analysis In this medication, the ingredients: Semen Pruni, Flos Carthami tinctorii, Gummi resina Olibanum, Radix Achyranthis bidentatae help to pain reliever, anti-inflammation. And the others ingredients which help enhance organs functions: Cortex Eucommiae, Radix Achyranthis bidentatae, Herba Loranthi Gracifilolii. And others ingredients promoting circulation: Radix Angelicae sinensis, Radix Paeoniae lactiflorae. Some ingredients are believed as good for rheumatologic diseases: Radix Saposhnikoviae divaricatae, Herba Siegesbeckiae, Rhizoma Homalomenae occultae, Rhizoma Atractylodis, Semen Coicis, Cortex Schefflerae heptaphyllae, Caulis Tinosporae tomentosae, Radix Angelicae pubescentis, Herba Loranthi Gracifilolii help to reduce the synovial fluid, and pain 6 reliever. Ramulus Cinnamomi helps to promote circulation. Radix et Rhizoma Glycyrrhizae helps to combine all ingredients. Chapter 2 MATERIALS, SUBJECTS, AND METHODOLOGY 2.1. Materials - Materials of Ich goi khang: Mentioned earlier, all ingredients was treated to the liquid material, in the bottle 250 ml, produced in the Institute of Traditional Medicine with the standardization. - The control material: Agelica laxiflora and Herba taxilli: Radix Gentianae 12g, Radix Rehmanniae 12g, Herba Loranthi Gracifilolii 16 g, Ramulus Cinnamomi 4g, Radix et Rhizoma Glycyrrhizae 6g, Herba Asari 6g, Rhizoma Ligustici wallichii 8g, Poria 12g, Radix Angelicae sinensis 12g, Radix Paeoniae lactiflorae 12g, Radix Codonopsis 12g, Radix Angelicae pubescentis 12g, Radix Achyranthis bidentatae 12g, Cortex Eucommiae 12g, Radix Saposhnikoviae divaricatae 12g. It was produced at the Institute of Traditional Medicine 2.2. The subjects 2.2.1. The experiment subjects: White mouse Wistar, and White mouse Swiss, healthy, both male and female selected. Location: Pharmacology Department of Hanoi Medical University. 2.2.2. The clinical subjects: Sample size was measured by the formula of WHO, in 120 patients at Traditional Medical Institute, matched with the criteria * Selection criteria: Not depending on the age, gender, occupational status. Patient consent was necessary. Diagnosis criteria according to ACR 1991 knee OA, stage classification Kellgren Lawrence (1987). In traditional medicine, the diagnosis was confirmed as “phong han thap ty, can than hu”. * Elimination criteria: NSAIDs treatment around three recent days, local corticoid injection, local infection or systemic infection, 7 organ damages, others chronic diseases such as acute hepatitis, cirrhosis 2.3. Methodology 2.3.1. Experimental study 2.3.1.1. Acute and sub-acute toxicity * Acute toxicity: Definition of LD50 of Ich goi khang oral admission in the white mouse by the Litchfield Wilcoxon method. The mouse was given gradually the medication to define the minimum fatal dose of 100% mouse and the highest non-fatal dose. General observation, the variation after toxic signs and the mortality after 72 hours of admission was performed. Autopsy the death mouse, figure out the LD50 map, continually observing until 7 th day. * Sub-acute toxicity: According to WHO recommendation, mouse was separated as three groups: the control (water admission); the treatment group 1 (Ich goi khang 17.04 g/kg/day); the treatment group 2 (Ich goi khang 51.12 g/kg/day). Admission in 8 continuous weeks, in the morning, then observe and evaluate in term of liver and kidney function, compare between case and control. 2.3.1.2. The research of pain reliever and anti-inflammation of Ich goi khang in the osteoarthritis model The white mouse was separated randomly into 10 slots. Slot 1 (biologic control): Saline injection, water drinking 1ml/100g mice. Slot 2 (model): local injection MIA 3mg/joint, water drinking 1ml/100 g mouse. Slot 3 (case): MIA 3mg/joint injection, diclofenac 3mg/kg. Slot 4 (trial): 3mg MIA / joint injection, Ich goi khang 17.04 g/kg/day. Slot 5 (trial): MIA injection 3mg / joint, Ich goi khang 51.12 g/kg/day. MIA injection in right knee was performed to cause the OA. The biologic control was injected with saline. The slots 1, 2 and 3 was treated with diclofenac 3mg/kg; slots 4 and 5 was treated 8 with Ich goi khang 17.04 g/kg/day and 51.12 g/kg/day. All slots drank water 1 time/day in 6 consecutive weeks. Valued index: the knee diameter, pain reliever efficacity (Von Frey needle measurement), and Randall Sellito method, Interleukin index, pathology of knee. Randomly assessing in 30% mouse at the MIA slots after 6 weeks, the right knee biopsy was performed and the specimen was conserved in Formaldehyd 10% in term of OA evaluation 2.3.2. Clinical research Study design: Clinical trial, case-control in term of before and after treatment 2.3.2.1. The protocol Patients information was performed as the common chart. They was divided into two groups: + The research group: 60 patients with Ich goi khang in 1 month. + The control group: 60 patients with Angelica laxiflora and Herba taxilli, one time per day in one month. If the patients had serious pain (VAS 7), they would be prescribed Meloxicam 7.5 mg with 2 tablets per day. 2.3.2.2. Evaluation criteria - Subjects characteristics: Age, gender, occupation, BMI, symptoms - Ich goi khang: comparison between before and after treatment. Pain reliever (D0 to D30): VAS, WOMAC score. Knee range of motion: range of motion, heel-buttock index. - The secondary effects before and after treatment. 2.3.2.3. Data analysis: All data was analysed by statistical medicine SPSS 22.0. The statistical significance was defined as p ≤ 0.05. 9 2.3.2.4 Morality of research: The moral council of Hanoi Medical University and National Institute of Tradition Medicine Chapter 3. THE RESULTS 3.1. The experimental study results 3.1.1. Acute, sub-acute toxicity of Ich goi khang * Acute toxicity: The dose from 255.6g/kg to 426 g/kg caused no abnormality. Continuous observation noted no other abnormal evidence, no mice died after 72 hours and in continuous 7 days. LD50 toxicity was not defined. * Sub-acute toxicity: the three model of mouse was normal during the research. Ich goi khang 17.04 kg/kg and 51.12 g/kg in 4-8 weeks could not harm to mice circulation and hepatic, renal function. 3.1.2. The study of treatment efficacity in knee osteoarthritis mouse * The knee diameter of white mouse: Ich goi khang dose 17.04 g/kg helped to decrease knee diameter at the third days, p<0.01. The increase of swelling knee after 3 weeks was observed even though no statistical significance p > 0.05. With the dose of 51.12 g/kg, the increase of knee diameter was significantly decreased in all slots (p<0.05) Role of Ich goi khang (IGK) in term of cytokines concentration 10 Graph 3.1. Cytokines concentration in the slots It showed that the slots treated with two types of dose of Ich goi khang had the IL-1B lower than others (p<0.01). The dose of 51.12 g/kg showed the higher decrease in comparison with diclofenac 3mg/kg (p<0.05); however comparing with the dose 17.04 g/kg showed no difference (p>0.05). Both doses slots showed the decrease of TNF-alpha concentration (p<0.001), this effect was stronger than diclofenac 3 mg/kg (p<0.01)  Pain relieve potential - The needle Vonfrey was utilized to measure the pain relieve effect. Table 3.1 The effect of Ich goi khang in the pain measured by the Vonfrey needle Slot n Force causing pain (g) Before After 1 w After 2 w After 3 w After 4 w After 5 w After 6 w Slot 1 (Biologic slot) 10 22.57 ± 3.60 20.06 ± 4.61 23.41 ± 7.58 22.61 ± 7.14 22.67 ± 6.39 22.39 ± 3.98 23.37 ± 3.67 Slot 2 (Model) 10 22.85 ± 6.87 15.07 ± 3.64 20.51 ± 4.60 18.37 ± 6.72 24.83 ± 4.66 23.16 ± 4.95 27.68 ± 4.06* Slot 3 (Diclofena c 3mg/kg) 10 22.41 ± 2.99 23.37 ± 3.47 22.59 ± 6.57 18.49 ± 5.20 22.77 ± 5.24 22.94 ± 5.78 23.60 ± 4.38 Slot 4 (IGK 17.04g/kg) 10 21.77 ± 5.35 22.89 ± 5.75 27.36 ± 7.00 32.79 ± 10.66 29.13 ± 7.62 27.25 ± 7.54 26.53 ± 8.89 Slot 5 (IGK 51.12g/kg) 10 20.19 ± 5.71 20.15 ± 2.71 30.20 ± 8.53 21.92 ± 6.60 26.76 ± 6.60 27.31 ± 9.62 25.75 ± 7.43 p p1-2,3,4,5; p2-3,4,5; p3-4,5; p4-5; > 0.05 p1-2; p4-5 < 0.05 p1-3,4,5; p3-4,5 > 0.05 p2-3; < 0.001 p1-2,3,4,5; p2-3; p3-4,5 > 0.05 p2-4; p4-5 < 0.05 p1-2,3,5; p2-3,5; p3-5 > 0.05 p1-4; p3- 4; p4-5 p1-2,3,4,5; p2-3,4,5; p3-5; p4- 5 > 0.05 p3-4 < 0.05 p1-2,3,4,5 p2-3,4,5; p3-4,5; p4-5 > 0.05 p1-2,3,4,5; p2-3,4,5; p3-4,5; p4-5; > 0.05; p2-3 < 0.05 11 Slot n Force causing pain (g) Before After 1 w After 2 w After 3 w After 4 w After 5 w After 6 w p2-4,5 < 0.01 p2-5 < 0.01 < 0.05 p2-4 < 0.01 In the slots with IGK, after 6 weeks of MIA injection, the force causing pain was in descending trend in comparison with model (p>0.05) - The pain relieve effect measured by Randall Selitto method Graph 3.2. Force causing pain in the knee according to time The result showed the IGK 17.04 g/kg slots need more force to cause the pain than the model slot, at the point after 3 weeks (p<0.001). This effect was equal to diclofenac 3mg/kg. In the IGK 51.12 g/kg slot need more force to cause pain, especially at the 3 rd and 6 th week (p<0.005; p<0.01).  IGK in the pathology of knee  Studied slot Pathologic characteristics Image 12 Studied slot Pathologic characteristics Image Slot 1: Biologic control Slight degeneration cartilage: No sub-condral damage, no lose of proteoglycan dyed layer, normal synovial cells. Cellular invasion in the synovial area (+). (6 th mice) (HE x 100) Slot 2: Model (MIA 3mg/khớp) Severe chondral degeneration: Severe damage of sub-chondral bone, average loss of proteoglycan dyed layer, degenerative synovial cells. Inflammatory cells invasion of synovial area. Cartilage damage level from average to severe. Osteophytes and loss of joint space (23 rd mice) (HE x 100) Slot 3: Diclofenac 3mg/kg Average chondral degeneration: average sub-chondral damage, proteoglycan dyed class lost from light to severe, synovial cells degeneration. Inflammatory cells invasion level average. Light to average chondral damage. Joint space loss. Osteophyte. Fibrosis and chondrocyte hyperplasia. ( 25 th mice) (HE x 100) Slot 4: IGK 17.04g/kg Slight degeneration cartilage: Slight subchondral damge, light loss of proteoglycan. Slight degeneration of synovial cells, hyperplasia, slight invasion of inflammatory cells in synovial tissue. Slight loss of articular space. Degeneration and necrosis of cartilage surface sligh to severe. Osteophyte. (82 th mice) (HE x 100) 13 Studied slot Pathologic characteristics Image Slot 5: IGK 51.12g/kg Slight degenerative cartilage: Slight sub-chondral damge, slight loss of proteoglycan layer. Slight degenerative synovial cells, hyperplasia synovial cells, slight inflammatory cells invasion. Slight loss of articular space. Slight necrosis of cartilage surface. Osteophyte. (75 th mice) (HE x 100) 3.2 The clinical study results 3.2.1 Clinical characteristics Age, gender, occupational, BMI, and affected joint, clinical characteristics: There was no difference between two groups about VAS, WOMAC score, heel-buttock index, radigraphic damage, US damage (p>0.05). 3.2.2 The effect of treatment 3.2.2.1 Pain relieve evaluation, range of motion improve after treatment * Pain relieve effect according VAS score Table 3.2. VAS score in two groups Time VAS (points) ( ± SD) p Control (1) (n= 60) Case (2) (n=60) D0 5.53 ± 1.87 5.52 ± 1.8 0.96 D15 4.33 ± 1.47 4.03 ± 1.5 0.27 D30 3.33 ± 1.39 2.63 ± 1.09 0.003 Reduction of productivity D15 - D0 -1.2 ± 1.36 -1.48 ± 1.47 0.275 D30 -D15 -1.00 ± 1.12 -1.4 ± 1.09 0.048 D30 - D0 -2.2 ± 1.62 -2.88 ± 1.63 0.023 After treatment, the average pain point of patients at the D30 of control groups 3.33 ± 1.39 higher than the case 2.63 ± 1.09, statistical significance p<0.05. The productivity at the two time pont D30-D0 of control group was -2.2 ± 1.62, of case group was -2.88 ± 1.63, statistically significant p<0.05 * Level of relieving pain in knee based on VAS score X 14 Level of knee pain by VAS score Control (1) (n = 60) Case (2) (n = 60) Total (n = 120) n TL(%) n TL(%) N TL(%) Before treatment D0 Slight (1 - 3 points) 17 28.3 16 26.7 33 27.5 Moderate (4 - 6 points) 34 56.7 38 63.3 72 60 Severe (7 - 10 points) 9 15 6 10 15 12.5 p1-2 0.653 > 0.05 After treatment D30 Slight (1 - 3 points) 47 78.3 56 93.3 103 85.8 Moderate (4 - 6 points) 13 21.7 4 6.7 17 14.2 p1-2 0.018 < 0.05 PD30-D0 < 0.001 < 0.001 After 30 days, the pain score in two groups was significantly improved, there was no severe pain, 93.3% slight pain; 6.7% average pain. In case group 78.3% patients with slight pain, and 21.7% average pain (p<0.05) * Assessment the pain reliever effect according to WOMAC score Table 3.3. The assessment of the treatment outcome according to WOMAC score Time of study WOMAC score ( ± SD) p1-2 Control (1) (n = 60) Case (2) (n = 60) D0 45.10 ± 12.62 45.15 ± 13.22 0.983 > 0.05 D15 38.05 ± 12.12 35.45 ± 12.00 0.240 > 0.05 D30 33.65 ± 9.76 24.95 ± 10.28 < 0.001 D30 - D0 -11.45 ± 6.56 -20.20 ± 8.45 < 0.001 X 15 PD30-D0 <0.001 <0.001 At the D30, the WOMAC in the case group was lower than control group, p<0.001. The substraction of WOMAC score between D30 and D0 had no difference between the case and control study p<0.001. * The effect of Range of Motion (ROM) improvement Table 3.4. The improvement of knee at the observation Time Knee flexion ( 0 ) ( ± SD) p1-2 Control (1) (n = 60) Case (2) (n = 60) D0 115.42 ± 16.96 115.67 ± 18.56 0.939 D15 124.28 ± 13.59 122.75 ± 14.45 0.551 D30 128.42 ± 9.46 133.08 ± 7.34 0.003 Increase of productivity D15 - D0 8.87 ± 10.46 7.08 ± 21.06 0.558 D30 - D15 4.13 ± 7.92 10.33 ± 12.03 0.001 D30 - D0 13 ± 13.28 17.42 ± 19.42 0.149 The knee flexion of case group was higher than control group at the D30. statistically significant (p<0.05). The productivity of knee flexion at D30-D15 in 2 groups was statistically different (p<0.05), in the case group the productivity increased 10.33 ± 12.03 than control group 4.13 ± 7.92. Table 3.5. The change of heel-buttock index at the observation point Time Heel-buttock (cm) ( ± SD) p1-2 Control (1) (n = 60) Case (2) (n = 60) D0 19.58 ± 9.27 18.88 ± 7.38 0.648 D15 16.77 ± 7.04 15.38 ± 5.63 0.237 D30 14.95 ± 5.87 12.43 ± 4.00 0.007 Productivity D15 - D0 -2.82 ± 4.37 -3.50 ± 4.70 0.411 D30 - D15 -1.82 ± 2.73 -2.95 ± 3.18 0.038 X X 16 D30 - D0 -4.63 ± 5.24 -6.45 ± 5.65 0.07 The heel-buttock index at the D0, D15 in two groups had no difference at the D30 (p=0.007, <0.05). The productivity of heel- buttock point change at D30-D15 in case group -2.95 ± 3.18 was lower than the control group -1.82 ± 2.73, statistically significant (p<0.05). 3.2.2.2. The clinical study results * The change of Erythrocyte Sedimentation Rate (ESR) before and after treatment Table 3.6. The comparison of ESR between two groups Time of study Erythrocyte Sedimentation Rate (mm/h) ( ± SD) p1-2 Control (1) (n = 60) Case (2) (n = 60) D0 30.6 ± 21.85 28.02 ± 20.02 0.501 D30 23.22 ± 16.50 15.49 ± 12.5 0.005 Substraction D30-0 -7.38 ± 16.05 -12.5 ± 21.06 0.135 PD30-D0 0.001 < 0.05 0.001 < 0.05 After treatment, the substraction of ESR between day 30 th and day 0 was lowering trend, however no statistically significant p>0.05. 3.2.3. The side effects * Clinical scope: After 30 days, all studied patients had no side effect * Sub-clinical scope: There was no statistically significant change, p>0.05 X 17 Chapter 4. DISCUSSION 4.1. Experiment results 4.1.1. Acute and sub-acute toxicity of “Ich goi khang” * Acute: At the highest clinical proposed dose 426 g/kg, there was no mice died after 72 hours, therefore, it could not define the toxic dose LD50 in the white mouse. In WHO recommendation, IGK is safe for clinical use. * Sub-acute toxicity The common status of every slot was normal after 4-8 weeks. The weight of mouse in three slots increased after 8 weeks, there was no difference between each slot. It supposed that all mouse was in the adult period. At the slot with the dose higher three times, the mouse had the slight diarrhea. It was supposed to be caused by Radix Achyranthis bidentatae, betaine, achyrathine, aminoacid ... and viscous. Viscous is a carbohydhydrate originating from plants, non- absorptable. There was no other secondary effect. “Ich goi khang” (IGK) did not affect the general status and weight of the mouse. Both two doses of IGK caused no harm presented in subclinical results. 4.1.2. The effect of “Ich goi khang” in the mouse model with knee osteoarthritis The model of knee osteoarthritis (OA) caused by MIA injection was first time introduced in Vietnam. The mechanism of MIA in cartilage was inhibition of glyceraldehyde-3-phosphatedehydrogenase, leading the interpretion of energy from glucose, apoptosis, invasion of inflammatory cells. In this thesis, the white mouse was used, the judgement based on: 18 - The inflammatory symptoms, Range of Motion (ROM) - Cytokine concentrations - Pathology: gold standard, and stage of disease 4.1.2.1. Anti-inflammatory effect It showed that both doses of IGK presented the protective effect in knees. And the dose of 51.12 g/kg was better, and improving the range of motion, reduce knee damage, decrease interleukin production, promote the cartilage structure. It supposed that the main effect of IGK was inhibition of inflammatory mediators, anti- inflammation and anti-degeneration. CML-1 extracted from Cortex Eucommiae, Ramulus Cinnamomi, Radix et Rhizoma Glycyrrhizae, Radix Paeoniae lactiflorae, could help to relieve the pain in mouse. Weg showed the effect of polysaccharid in Radix Achyranthis bidentatae in mouse by the mechanism of cartilage regeneration and collagen type II expression. The effect of Radix Angelicae pubescentis and Herba Asari was showed by Xu Y in 2014, it was believed to inhibite the destruction of cartilage, inflammatory reaction, decrease IL-1B and TNF-alpha. The inflammatory index was announced through the diameter of mouse knee. Both doses showed the effect of decrease diameter of knee with p<0.05. This conclusion fits in our propose of anti-inflammatory and pain relieve effect of IGK. 4.1.2.2. The pain relieve effect * The effect of Ich goi khang in model with pain mesurement The needle of Von Frey was used to pain stimulation in right plantar of mouse. Form this point, evaluation of pain relieve effect and range of motion was estimated. From this results, the mouse slots treated with both doses of IGK had the force causing pain and the 19 reaction time lower than model slots, non-statistically different (p>0,05). This decrease explains the effect of medication. It supposed by the role of B-eudesmol in Rhizoma Atractylodis and Radix Achyranthis bidentatae which was proved in the model of acute pain caused by acid acetic. Some ingredients in Radix Achyranthis bidentatae was proved with the reduction of serotonin in mouse brain, anti-inflammation. * The pain relieve assessed by the Randall Selitto This model causes gradually the pain in the feet of mice until it caused a reflexion of withdral. The results of this study showed that both doses of IGK could increase the possibiliity of pain suffering and increase response time, significantly different (p>0,05). 4.1.2.3. The pathologic change In the IGK slot, the slight degeneration, slight decoloration of proteoglycan, hyperplasia, invasion of inflammatory cells was seen. Narrow joint space, osteophyte was also observed. This showed the benifit of anti-inflammation of IGK. Therefore, it could raise a conclusion that IGK not only has the cartilage protection, but also inhibition of pro-inflammatory cytokines such as IL-1B and TNF- alpha, or in the mechanism which relates to the exudation presented by diclofenac, however diclofenac itself does not show this effect. It could be benefited from s