SMAD7调控内皮间质转化可预防跟腱损伤后的异位骨化

SMAD7 prevents heterotopic ossification by regulating endothelial-mesenchymal transition after Achilles tendon imjury

背景:现有的研究结果证明,内皮细胞向间质细胞转化(End MT)在异位骨化病程中起主要作用。目的:基于SMAD7基因在阻止肌成纤维细胞转化等内皮-间质转化进展中的潜能,验证其是否是异位骨化潜在的治疗靶点。方法:构建了过表达SMAD7的慢病毒颗粒,并在大鼠主动脉内皮细胞中测定了其最佳滴度和转染效率。随后,将该病毒颗粒注入构建的大鼠跟腱损伤模型中,对照组注射生理盐水。使用qPCR和蛋白印迹实验分析了损伤处内皮和间质标志物的表达,采用X射线和组织染色观察异位骨化病程转化。结果与结论:(1)局部注射转染SMAD7基因的病毒载体可引起内皮细胞标志物(CD31,VE-cadherin)发生上调,同时间质细胞标志物(N-cadherin,vimentin)发生下调,提示SMAD7的局部高表达阻断了内皮-间质转化改变。这种表达差异在造模后10周时比在造模后6周时更明显;(2)X射线片和组织染色显示,相比对照组,注入表达SMAD7的慢病毒后,肌腱组织中骨化结构缺失;(3)因此认为,促进局部SMAD7的高表达可用来预防术后异位骨化形成,而不影响正常的伤口愈合过程。

BACKGROUND： The endothelial-mesenchymal transition is known to play a central role in the pathological process of heterotopic ossification. OBJECTIVE： To verify the inhibitory effect of SMAD7 on endothelial-mesenchymal transition such as myofibroblast transformation and to explore whether it is a potential target for heterotopic ossification.METHODS： A lentivirus overexpressing SMAD7 was contructed and the optimal titre and transfection efficiency in rat aortic endothelial cells were determined. The lentivirus was then injected into a rat model of Achilles tendon injury, while the controls were given the injection of normal saline. Expressions of endothelial and mesenchymal markers at the injured site were analyzed by quantitative PCR and western blot assay. The heterotopic ossification was observed radiologically and histologically.RESULTS AND CONCLUSION： Local injection of SMAD7-delivering lentivirus resulted in an upregulation of CD31 and VE-cadherin, and a downregulation of N-cadherin and vimentin, suggesting that the endothelial-mesenchymal transition is blocked due to local SMAD7 overexpression. The inhibitory effect became more evident at 10 weeks than at 6 weeks after modeling. Radiology and histological staining further confirmed that the ossified structures in the tendon tissue disappeared after injection of SMAD7-delivering lentivirus, as opposed to the control group. These data suggest that local overexpression of SMAD7 can prevent postoperative heterotopic ossification with no effect on wound healing.