【摘要】 目的观察在转化生长因子-β1(transforming growth factor beta1,TGF-β1)作用下,损伤的前交叉韧带(anteri-or cruciate ligament,ACL)和内侧副韧带(medial collateral ligament,MCL)中骨形态发生蛋白-1(bone morphogenetic protein-1,BMP-1)基因的表达,找出TGF-β1、BMP-1之间的关系,揭示机械损伤后ACL和MCL细胞中BMP-1的表达差异。方法采用反转录PCR(RT-PCR)和实时荧光定量PCR方法检测1、5、50 ng/ml TGF-β1处理后2 h受损的ACL和MCL细胞中BMP-1的表达以及5 ng/ml TGF-β1作用2、6、12、24 h受损的ACL和MCL细胞中BMP-1的表达;Western blot检测5 ng/mlTGF-β1处理48 h后受损的ACL和MCL细胞中BMP-1的表达。结果受损的ACL和MCL细胞中BMP-1的基因表达比正常状态下偏高,并随着TGF-β1浓度的增大而增高,在MCL中的增高程度比在ACL中高出近1倍(P<0.05);与正常组相比,在5 ng/ml TGF-β1处理24 h后,ACL细胞中BMP-1的表达在24 h达到最高比例(约为6.1倍),而在MCL中12 h达到最高比例(约为9.84倍,P<0.05)。5 ng/ml TGF-β1处理48 h后BMP-1蛋白也明显上调,与无TGF-β1处理的对照组相比,ACL细胞中BMP-1上调2.32倍,MCL细胞中BMP-1上调3.84倍(P<0.05)。结论 TGF-β1刺激BMP-1的变化可能直接影响到细胞外基质中活性赖氨酰氧化酶的表达,对损伤ACL和MCL的修复有极其重要的参考价值和临床意义。更多还原

【Abstract】 Objective To investigate the differential expression of bone morphogenetic protein-1(BMP-1) in injured anterior cruciate ligament(ACL) and medial collateral ligament(MCL) fibroblasts induced by TGF-β1,and to find out the differences between the poorly self-healing ACL and well functionally self-healing MCL fibroblasts.Methods Fibroblasts were primarily cultured from clinical ACL and MCL samples,and then given 12% mechanical stretch injury and treated by 1,5 and 50 ng/ml TGF-β1 for 2 h,or by 5 ng/ml TGF-β1 for 2,6,12 and 24 h respectively at the same time.The expression of BMP-1 in the above treated and untreated fibroblasts were detected by reversed-transcript PCR and real-time quantitative PCR.Western blotting was used to detect the expression of BMP-1 in the injured fibroblasts induced by 5 ng/ml TGF-β1 for 48 h.Results TGF-β1 treatment resulted in an increased mRNA expression of BMP-1 in the injured fibroblasts in a dose-depended manner,especially in the cells from MCL than in those from ACL(about 1 times higher,P<0.05).Compared with the fibroblasts without TGF-β1 treatment,5 ng/ml TGF-β1 treatment for 24 h made the expression of BMP-1 reach the summit in ACL(6.1 folds higher,P<0.05) and in MCL(9.84 folds higher,P<0.05).Compared with the control,the protein expression of BMP-1 were elevated to 2.32 and 3.84 folds higher in ACL and MCL respectively after 5 ng/ml TGF-β1 treatment for 48 h(P<0.01).Conclusion TGF-β1 affects the expression of BMP-1 in the injured fibroblasts,and then directly affects activities of lysyl oxiadse in the extracellular matrix,implying its potential significant value and clinical usage in repair of injured ACL.更多还原