缓慢释放特性的纳米球载工程化microRNA基因治疗减少肌腱粘连促进滑动的离体和动物实验研究

The formation of adhesion during healing process of injured tendon is still a difficult problem in clinical practice. In this study, we used hydrogel system containing the engineered microRNA-based plasmid-loaded nanoparticles to reduce tendon adhesion and improve tendon gliding. We will construct the engineered miRNA expression plasmid to silence expression of the COX-1 and COX-2 gene, which are considered to play a key role in the inflammatory reactions, in order to inhibit the inflammatory reactions of the tendon tissue and surrounding tissues and to reduce tendon adhesions. The present study is a continuation of our effort in prevention of tendon adhesion using nanotechnology therapy in the past 6 years. Based on the RNA interference technique to inhibit the expression of the cyclooxygenases, we adopt nanoparticle with biocompatibility and safety as the treatment vector, and combined with hydrogel with sustained release function to explore and attempt in the study of tendon injury. The termination time and safety of this miRNA nanoparticle hydrogel system will be explored to provide the basis of biomechanics and molecular biology for clinical use. The purpose of this study is to explore a new approach to solve the clinical problem of formation of adhesion during tendon healing using the advanced nanotechnology.

肌腱损伤愈合过程中粘连形成仍是目前临床上难以解决的问题。本研究立足于前沿生物技术，采用负载工程化微小RNA(miRNA)质粒纳米球的缓释水凝胶系统来减少鞘内肌腱粘连和滑动阻力。构建工程化miRNA表达载体，抑制对炎症反应起关键作用的环氧合酶（Cox-1和Cox-2）的基因表达，以期抑制肌腱组织及周围组织的炎症反应来减少肌腱粘连形成。本研究是过去6年系统探索纳米基因治疗减少肌腱粘连的延续，以RNA干扰技术抑制环氧合酶表达的预实验为依据，采用生物相容性和安全性更高的纳米球为治疗载体，并结合到缓释水凝胶中在肌腱损伤愈实验中进行探索和尝试。通过活体动物模型探索工程化微小miRNA纳米球水凝胶缓释系统作用的终止时间及安全性，为临床使用提供生物力学和分子生物学依据。本研究是有望最终应用于临床肌腱修复时防止粘连的基础研究部分，研究的宗旨是为解决临床上手指屈肌腱愈合时粘连形成影响手功能的难题提供有效方法。

肌腱损伤愈合过程中粘连的形成一直是临床上的难题。抑炎基因的局部传递能够提高局部的基因浓度，降低损伤肌腱微环境的炎症反应，减少全身副作用，提高体内的疗效。在这项研究中，我们设计了一种新型的、局部的、持续的基因传递系统，通过将环氧合酶（COX-1和COX-2）工程化的miRNA质粒/纳米球包裹进入透明质酸（HA）水凝胶中来减少屈肌腱粘连的形成。在体外实验中，我们证实了肌腱损伤后肌腱与皮下组织环氧合酶的高表达，设计并筛选了最具沉默效果的COX-1和COX-2工程化的miRNA质粒，并成功将miRNA质粒/纳米球转染进入肌腱细胞中；在体内实验中，我们发现透明质酸水凝胶载miRNA质粒/纳米球能够转染至组织中，而且这种局部基因持续传递系统显著下调了肌腱组织和周围皮下组织中COX-1和COX-2的表达，显著减少了粘连组织的形成，同时提高了肌腱的愈合强度。这种方法提供了一种有效的治疗策略，通过直接针对损伤肌腱微环境中COX-1和COX-2表达的下调来减少肌腱粘连。