The Research Of Blocking SDF-1/CXCR4 Signaling Pathway To Reduce The Degeneration Of Articular Cartilage With AMD3100

Part 1The influence on human articular cartilage histology after blocking SDF-1/CXCR4 signaling pathway with AMD3100Objective:To observed the influence on human articular cartilage histology after blocking SDF-1/CXCR4 signaling pathway with AMD3100.Methods:1 The cartilage tissue were acquired in size of 3×3×1mm from the knee cartilage surface (Mankin score of 0 or 1).12 cases which diagnosed as OA were taken from total knee replacement surgery. Another 12 normal cases were taken from traumatic amputation. The cartilage tissue were cultured in the nutrient solution containing of SDF-1 100ng/ml, added AMD3100, MAB310, MAB002 1000nM respectively,37℃CO2 incubator for 2 or 4 days. The cartilage tissue were classfied 4 groups:(1) the experimental group:AMD3100+SDF-1, (2)the experimental control group1:MAB310+SDF-1,(3) the experimental control group 2:MAB002+SDF-1, (4)the blank control group:nothing+SDF-1.2 The cartilage tissue were cultured for 2 or 4 days. Then the cartilage tissue were taken for histology examination(HE and Safranin-O staining).Results:Examinated by histology(HE and Safranin-O staining) showed:The experimental group was lower in Mankin score and cartilage degeneration lighter than the experimental control group and the blank control group. Conclusions:1. SDF-1 could induce the articular cartilage degeneration thruogh the SDF-1/CXCR4 signaling pathway in vitro culture.2 AMD3100 could blocked SDF-1/CXCR4 signaling pathway and reduced the degeneration of articular cartilage.3 AMD3100 couldn't reverse the degeneration of cartilage into normal cargilage.Part 2The influence on MMP-3,9,13 level of human articular cartilage cells after blocking SDF-1/CXCR4 signaling pathway with AMD3100Objective:To observed the influence on MMP-3,9,13 level of human articular cartilage cells after blocking SDF-1/CXCR4 signaling pathway with AMD3100 and defined the function mechanism.Methods:1 The cartilage tissue were acquired in size of 3×3×1mm from the knee cartilage surface (Mankin score of 0 or 1).12 cases which diagnosed as OA were taken from total knee replacement surgery. Another 12 normal cases were taken from traumatic amputation. The cartilage tissue were cultured in the nutrient solution containing of SDF-1 100ng/ml, added AMD3100, MAB310, MAB002 1000nM respectively,37℃CO2 incubator for 2 or 4 days. The cartilage tissue were classfied 4groups:(1) the experimental group:AMD3100+SDF-1,(2)the experimental control group 1:MAB310+SDF-1,(3) the experimental control group 2:MAB002+SDF-1,(4)the blank control group:nothing+SDF-1.2 The cartilage tissue were cultured for 2 or 4 days. ELISA was used to measure the level of MMP-3,-9,-13 in the culture medium; RT-PCR was taken to test the expression of MMP-3,-9,-13mRNA in the cartilage tissue;. Excel and SPSS 17.0 were used for data analysis.Results:1 ELISA:In the culture medium of experimental group, the level of MMP-3,-9,-13 was lower than in the experimental control group and blank control group, the differences was statistically significant. 2 RT-PCR:In the experimental group the expression of MMP-3,-9,-13 mRNA was lower than that in the experimental control group and the blank control group,the differences was statistically significant.Conclusions:1. SDF-1 could induced the articular cartilage overexpressed and released MMP-3,-9,-13 thruogh the SDF-1/CXCR4 signaling pathway.2 AMD3100 could block the SDF-1/CXCR4 signaling pathway and reduce cartilage cell to express and secrete MMP-3,-9,-13mRNA.3 AMD3100 couldn't keep OA cartilage to secrete MMP-3,-9,-13 to return to normal levels.Part 3The influence on human articular cartilage cells secretion of typeⅡcollagen and aggrecan after blocking SDF-1/CXCR4 signaling pathway with AMD3100Objective:To observe the influence on human articular cartilage secretion of type II collagen and aggrecan after blocking SDF-1/CXCR4 signaling pathway with AMD3100 and further confirmed that it could reduced the degeneration of articular cartilage.Methods:1 The cartilage tissue were acquired in size of 3×3×1mm from the knee cartilage surface (Mankin score of 0 or 1).12 cases which diagnosed as OA were taken from total knee replacement surgery. Another 12 normal cases were taken from traumatic amputation. The cartilage tissue were cultured in the nutrient solution containing of SDF-1 100ng/ml, added AMD3100, MAB310, MAB002 1000nM respectively,37℃CO2 incubator for 2 or 4 days. The cartilage tissue were classfied 4 groups:(1) the experimental group:AMD3100+SDF-1, (2)the experimental control group1:MAB310+SDF-1,(3) the experimental control group 2:MAB002+SDF-1, (4)the blank control group:nothing+SDF-1.2 The cartilage tissue were cultured for 2 or 4 days. Then the cartilage tissue were taken for RT-PCR was taken to test the secretion of typeⅡcollagen and aggrecan mRNA in the cartilage tissue; Western blotting was taken to test the typeⅡcollagen of the cartilage tissue.Excel and SPSS 17.0 were used for data analysis.Results:1 RT-PCR:In the experimental control group and the blank control group cartilage, the expression of typeⅡcollagen and aggrecan mRNA was higher than that in the experimental group, the difference was statistically significant;2 Western blotting:The level of typeⅡcollagen in the experimental group was higher than that of experimental control group and blank control group, the difference was statistically significant.Conclusions:1. SDF-1 could induce the articular excessive degradation cartilage typeⅡcollagen and aggrecan thruogh the SDF-1/CXCR4 signaling pathway.2 AMD3100 could block SDF-1/CXCR4 signaling pathway and reduce typeⅡcollagen and aggrecan degradation, slow down the degeneration of articular cartilage.3 AMD3100 couldn't recover OA cartilage typeⅡcollagen and aggrecan return to the normal levels...