关节损伤对软骨蛋白酶基因表达影响的研究

Research on the influence of joint injuries on gene expressions of articular cartilage proteases

目的本研究旨在探索膝关节损伤后特定的透明软骨退化酶,如基质金属蛋白酶(MMP)、蛋白聚糖酶(Agg)的基因表达是否增加。方法试验纳入138例因关节组织如半月板、前交叉韧带(anterior cruciate ligament,ACL)、透明软骨损伤而行膝关节镜检查的患者,男57例,女81例,平均年龄38.8岁。检查前采集全血样本,检查时采集滑膜样本。利用RT-PCR和分光光度法处理样本。结果患者中,有56例ACL损伤,65例内侧半月板损伤,5例外侧半月板损伤,软骨损伤者根据Outerbridge标准进行分级。实验室检测显示,ACL撕裂和基因表达(MMP1、MMP2、MMP8、MMP9、MMP13、MMP14、Agg1、Agg2、IL1、TNFα和TIMP2,除外血浆TIPM1)显著相关:ACL患者滑膜中基因表达水平较非ACL患者滑膜中的对应基因表达水平显著上调:MMP1 0.920±0.068 vs 0.794±0.061,MMP2 1.075±0.053 vs 0.668±0.035,MMP8 0.951±0.047 vs0.766±0.045,MMP9 1.354±0.032 vs 0.947±0.042,MMP13 1.148±0.058 vs 0.991±0.058,MMP14 1.379±0.049vs 0.777±0.036,Agg1 1.309±0.071 vs 0.647±0.034,Agg2 1.043±0.072 vs 0.684±0.069,IL1 1.320±0.054 vs0.857±0.049,TNFα1.101±0.050 vs 0.802±0.039,TIMP1 1.197±0.060 vs 1.035±0.071,TIMP2 1.110±0.048vs 0.861±0.048(P值均<0.05)。除TIMP1外(1.092±0.053 vs 1.081±0.033,P=0.32),ACL患者血液中基因表达水平较非ACL患者血液中的对应基因表达水平显著上调:MMP1 1.163±0.030 vs 0.948±0.049,MMP20.918±0.056 vs 0.827±0.049,MMP8 1.172±0.029 vs 0.783±0.034,MMP9 1.304±0.059 vs 0.94±0.04,MMP131.155±0.060 vs 0.824±0.043,MMP14 1.095±0.058 vs 0.759±0.033,Agg1 1.270±0.077 vs 0.676±0.038,Agg21.244±0.037 vs 0.871±0.037,IL1 1.128±0.056 vs 0.912±0.045,TNFα1.018±0.076 vs 0.724±0.047,TIMP11.092±0.053 vs 1.081±0.033,TIMP2 0.906±0.056 vs 0.875±0.049(P值均<0.05)。所有基因在血浆和滑膜中的表达具有相关性(Agg1:r=0.89,MMP1:r=0.88,MMP2:r=0.87,P<0.001)。结论 ACL损伤引起蛋白酶基因表达增加,而其它损伤造成的改变尚不确切。滑膜与血浆中的基因表达存在相关性。

Objective To explore whether injuries of the knee joint tissues will increase gene expressions of selected hyaline cartilage degenerating enzymes such as matrix metaloproteinases（ MMP） and aggreacaneses（ Agg）. Methods A total of 138 patients with joint tissue lesions such as menisci, anterior cruciate ligament（ ACL） and hyaline cartilage were admitted for knee arthroscopy. There were 81 females and 57 males with a mean age of 38.8 years. Full blood samples were collected preoperatively and synovium samples intraoperatively. Real time polymerase chain reaction（ PCR） and spectrophotometric analysis were performed. Results ACL lesions were found in 56 patients, medial menisci（ MM） lesions in 65 patients and lateral menisci（ LM） lesions in 5 patients. Chondral lesions were estimated according to Outerbridge＇s grading system. In laboratory tests, significant correlation was seen between ACL tears and gene expressions（ MMP1, MMP2, MMP8, MMP9, MMP13, MMP14, Agg1, Agg2, IL1, TNFα and TIMP2, except TIMP1）（ P 0.05）. The gene expression level in synovium of the patients with ACL lesions was significantly elevated compared with that of the patients without ACL lesions： MMP1 0.920 ± 0.068 vs 0.794 ± 0.061; MMP2 1.075 ± 0.053 vs 0.668 ± 0.035; MMP8 0.951 ± 0.047 vs 0.766 ± 0.045; MMP9 1.354 ± 0.032 vs 0.947 ± 0.042; MMP13 1.148 ± 0.058 vs 0.991 ± 0.058; MMP14 1.379 ± 0.049 vs 0.777 ± 0.036; Agg1 1.309 ± 0.071 vs 0.647 ± 0.034; Agg2 1.043 ± 0.072 vs 0.684 ± 0.069; IL1 1.320 ± 0.054 vs 0.857 ± 0.049; TNFα 1.101 ± 0.050 vs 0.802 ±0.039; TIMP1 1.197 ± 0.060 vs 1.035 ± 0.071; TIMP2 1.110 ± 0.048 vs 0.861 ± 0.048（ P 0.05）. Except TIMP1（ 1.092 ± 0.053 vs 1.081 ± 0.033, P = 0.32）, the gene expression level in serum of the patients with ACL lesions was significantly elevated compared with that of the patients without ACL lesions： MMP1 1.163 ± 0.030 vs 0.948 ± 0.049; MMP2 0.918 ± 0.056 vs 0.827 ± 0.049; MMP8 1.172 ± 0.029 vs 0.783 ± 0.034; MMP9 1.304 ± 0.059 vs 0.94 ± 0.04; MMP13 1.155 ± 0.060 vs 0.824 ± 0.043; MMP14 1.095 ± 0.058 vs 0.759 ± 0.033; Agg1 1.270 ± 0.077 vs 0.676 ± 0.038; Agg2 1.244 ± 0.037 vs 0.871 ± 0.037; IL1 1.128 ± 0.056 vs 0.912 ± 0.045; TNFα 1.018 ± 0.076 vs 0.724 ± 0.047; TIMP1 1.092 ± 0.053 vs 1.081 ± 0.033; TIMP2 0.906 ± 0.056 vs 0.875 ± 0.049（ P 0.05）. All of the gene expressions were correlated in the serum and synovium（ Agg1： r = 0.89, MMP1： r = 0.88, MMP2： r = 0.87, P 0.001）. Conclusions An ACL lesion provokes elevation in the gene expression of proteases, while the influence of other lesions remains elusive. Gene expression in the synovium is correlated with that in the peripheral blood.