Study Of Correlation Between Apoptosis And Osteoarthritis

【Objective】To investigate the expression and significance of apoptosis control genes(FAS,BAX,BAK,BCL-XL),XIAP,caspases,uPA/uPAR,YKL-40,α2M in articular cartilage and synovium of osteoarthritic animal model and study the relation of the expressions with apoptosis of articular chondrocytes and synovial cell.【Methods】Specimens of the articular cartilage and synovium obtained from osteoarthritic animal model and osteoarthritic patients respectively.Immunohistochemistry was used to detect the expression of apoptosis control genes(FAS,BAX,BAK,BCL-XL),XIAP,caspases,uPA/uPAR,YKL-40,α2M in articular cartilage and synovium of animal model.TUNEL assay was used to detect the apoptosis of the articular chondrocytes and synovial cell.The relation of the expression of apoptosis control genes(FAS,BAX,BAK,BCL-XL),XIAP,caspases,uPA/uPAR,YKL-40,α2M with apoptosis of articular chondrocytes and synovial cell was studied.1.animal modelThe left knees of 30 rabbits were immobilized in full extension using plaster cast. Specimens of the articular cartilage and synovium obtained from 4 rabbits that had been immobilized for 1,2,4,6 weeks were studied by using histopathologic techniques so that the effect of the immobilization would be observed.After six weeks immobilization,the samples of the articular cartilage and synovium obtained from the left knees of 26 rabbits and that from the right knees which was used to be control group.The fresh samples were fixed in 10%formalin for 48 hours and decalcificated in 10%EDTA for 6-7 weeks.TUNEL assay was used to detect the apoptosis of the articular chondrocytes and synovial cell.2.expression of apoptosis control genesThe fresh samples were fixed in 10%formalin for 48 hours and decalcificated in 10%EDTA for 6-7 weeks.Immunohistochemistry was used to detect the expression of FAS,BAX,BAK,BCL-XL proteins in articular cartilage and synovium of animal model.TUNEL assay was used to detect the apoptosis of the articular chondrocytes and synovial cell.The relation of the expression of FAS,BAX,BAK,BCL-XL proteins with apoptosis of articular chondrocytes and synovial cell was studied.3.expression of uPA/uPARThe fresh samples were fixed in 10%formalin for 48 hours and decalcificated in 10%EDTA for 6-7 weeks.Immunohistochemistry was used to detect the expression of uPA/uPAR proteins in articular cartilage and synovium of animal model.TUNEL assay was used to detect the apoptosis of the articular chondrocytes and synovial cell. The relation of the expression of uPA/uPAR proteins with apoptosis of articular chondrocytes and synovial cell was studied.4.expression ofα2MThe fresh samples were fixed in 10%formalin for 48 hours and decalcificated in 10%EDTA for 6-7 weeks.Immunohistochemistry was used to detect the expression ofα2M proteins in articular cartilage and synovium of animal model.TUNEL assay was used to detect the apoptosis of the articular chondrocytes and synovial cell.The relation of the expression ofα2M proteins with apoptosis of articular chondrocytes and synovial cell was studied.5.expression of caspasesThe fresh samples were fixed in 10%formalin for 48 hours and decalcificated in 10%EDTA for 6-7 weeks.Immunohistochemistry and TUNEL assay was used to detect the expression of caspases in articular cartilage and the apoptosis of the articular chondrocytes cell in 22 cases.6.expression of XIAPThe fresh samples were fixed in 10%formalin for 48 hours and decalcificated in 10%EDTA for 6-7 weeks.Immunohistochemistry and TUNEL assay was used to detect the expression of XIAP in articular cartilage and the apoptosis of the articular chondrocytes cell in 22 cases.7.expression of YKL-40The fresh samples were fixed in 10%formalin for 48 hours and decalcificated in 10%EDTA for 6-7 weeks.Immunohistochemistry and TUNEL assay was used to detect the expression of YKL-40 in articular cartilage and the apoptosis of the articular chondrocytes cell in 22 cases.【Results】1.animal modelThe results showed that apoptosis specific morphology were detected on superficial and middle layer of the articular cartilage from one to two weeks after immobilization,and these changes progressed until 4 weeks.At six weeks,the typical histopathologic character was seen in the articular cartilage.The apoptosis index(AI)of osteoarthritic and normal cartilage was 6.708±4.278 and 1.583±1.613 respectively.There was statistically significant difference of AI between the two groups(P＜0.01).The AI of osteoarthritic and normal synovium was 2.667±2.036 and 2.917±2.185 respectively.There was no statistically significant difference of AI between the two groups(P＞0.05).2.expression of apoptosis control genesPositive expression of Fas protein was detectable in chondrocytes and synovium of both osteoarthritic and normal groups and the positive expression rats were 66.67%, 58.33%,37.50%and 37.50%respectively.Fas of chondrocytes and synovium of osteoarthritic group was overexpressed compared with that of the normal controls. There was statistically significant difference between chondrocytes of osteoarthritic and normal groups(P＜0.05)but was no statistically significant difference between synovium of the two groups(P＞0.05).Positive expression of Bcl-xl protein was detectable in chondrocytes and synovium of both osteoarthritic and normal groups and the positive expression rats were 20.83%,54.17%,33.33%and 12.50%respectively.The positive expression rat of Bcl-xl protein in osteoarthritic chondrocytes was lower than that in normal chondrocytes,there was no statistically significant difference between the two groups(P＞0.05).The positive expression rat of Bcl-xl protein in osteoarthritic synovium was higher than that in normal synovium,and there was statistically significant difference between the two groups(P＜0.01). Positive expression of Bax protein was detectable in chondrocytes and synovium of both osteoarthritic and normal groups and the positive expression rats were 75.00%, 16.67%,29.17%and 25.00%respectively.The positive expression rat of Bax protein in osteoarthritic chondrocytes was higher than that in normal chondrocytes,and there was statistically significant difference between the two groups(P＜0.01).The positive expression rat of Bax protein in osteoarthritic synovium was lower than that in normal synovium,but there was no statistically significant difference between two groups(P＞0.05).Positive expression of Bak protein was detectable in chondrocytes and synovium of both osteoarthritic and normal groups and the positive expression rats were 54.17%, 45.83%,20.83%and 37.50%respectively.The positive expression rat of Bak protein in osteoarthritic chondrocytes was higher than that in normal chondrocytes,and there was statistically significant difference between the two groups(P＜0.05).The positive expression rat of Bak protein in osteoarthritic synovium was higher than that in normal synovium,but there was no statistically significant difference between two groups(P＞0.05).There was a positive correlation between degrees of the expression of FAS,BAX, BAK and chondrocytes apoptosis in osteoarthritic articular cartilage(P＜0.01),but a negative correlateion between degree of the expression of Bcl-xl and chondrocytes apoptosis in osteoarthritic articular cartilage(P＜0.05)3.expression of uPA/uPARThe positive expression rat of uPA in osteoarthritic chondrocytes was higher than that in normal chondrocytes,and there was statistically significant difference between the two groups(P＜0.05).The positive expression rat of uPAR in osteoarthritic chondrocytes was higher than that in normal chondrocytes,and there was statistically significant difference between the two groups(P＜0.05).There was a positive correlation between degrees of the expression of uPA/uPAR and chondrocytes apoptosis in osteoarthritic articular cartilage(P＜0.01).4.expression ofα2MThe positive expression rat ofα2M in osteoarthritic chondrocytes was higher than that in normal chondrocytes,and there was statistically significant difference between the two groups(P＜0.05).There was a negative correlation between degrees of the expression ofα2M and chondrocytes apoptosis in osteoarthritic articular cartilage(P＜0.05).5.expression of caspasesThe positive expression rat of caspase3 in osteoarthritic chondrocytes were higher than that in normal chondrocytes,and there were statistically significant difference between the two groups(P＜0.01).The positive expression rat of caspase6 in osteoarthritic chondrocytes were higher than that in normal chondrocytes,and there were statistically significant difference between the two groups(P＜0.05).The positive expression rat of caspase8 in osteoarthritic chondrocytes were higher than that in normal chondrocytes,and there were statistically significant difference between the two groups(P＜0.01).There was a positive correlation between degrees of the expression of caspases and chondrocytes apoptosis in osteoarthritic articular cartilage(P＜0.05).6.expression of XIAPThe positive expression rat of XIAP in osteoarthritic chondrocytes was higher than that in normal chondrocytes,but there wasn't statistically significant difference between the two groups(P＞0.05).There was a negative correlateion between degree of the expression of XIAP and chondrocytes apoptosis in osteoarthritic articular cartilage(P＜0.05).7.expression of YKL-40The positive expression rat of YKL-40 in osteoarthritic chondrocytes was higher than that in normal chondrocytes,and there was statistically significant difference between the two groups(P＜0.05).There was a positive correlation between degrees of the expression of YKL-40 and chondrocytes apoptosis in osteoarthritic articular cartilage(P＜0.05).【Conclusion】1.It is an ideal model that immobilize the knees of rabbit in full extension using plaster cast to get osteoarthritic animal model. 2.The apoptosis of chondrocytes in OA was higher than that in normal.It might be one of the important causations of OA.3.There was a consanguineous correlation between degrees of the expression of FAS,BAX,BAK,BCL-XL and chondrocytes apoptosis in osteoarthritic articular cartilage.It demonstrated that the abnormal expression of FAS,BAX,BAK, BCL-XL closely related to the biological behaveior of OA such as occurrence, progress,et al.4.There was a consanguineous correlation between degrees of the expression of caspases,XIAP,uPA/uPAR,YKL-40,α2M and chondrocytes apoptosis in osteoarthritic articular cartilage.It demonstrated that the abnormal expression of caspases,XIAP,uPA/uPAR,YKL-40,α2M closely related to the biological behaveior of OA such as occurrence,progress,et al.