负载塞来昔布的静电纺PELA纤维膜防止肌腱粘连及相关机制

发布时间：2018-03-19 06:39

本文选题：塞来昔布　切入点：ERK　出处：《上海交通大学》2015年博士论文　论文类型：学位论文

【摘要】：目的肌腱损伤术后粘连的防治一直是困扰临床医生的一大难题,肌腱粘连的主要原因之一是外源性成纤维细胞的过度增殖以及胶原纤维的过度沉积。以往研究证实细胞外信号调节激酶1/2(ERK1/2)和SMAD2/3是外源性成纤维细胞的过度增殖以及胶原纤维的过度合成的关键靶点,而塞来昔布能够通过抑制ERK1/2和SMAD2/3的磷酸化进而抑制体外成纤维细胞的增殖和胶原蛋白的分泌。本课题拟采用静电纺丝技术制成负载塞来昔布的聚乳酸聚乙二醇共聚物(PELA)纤维膜及基于负载塞来昔布PELA纤维膜的多层电纺纤维膜,研究其对肌腱粘连的抑制作用并探索其机制,为防止肌腱粘连促进患肢功能康复提供新的治疗方法及理论依据。方法通过共混电纺法制备载不同含量塞来昔布的PELA电纺纤维膜,观察纤维膜的表征和体外药物缓释情况,对比不同载药量的PELA电纺纤维膜对成纤维细胞和肌腱细胞的生理学行为影响;其次,构建新西兰兔肌腱粘连模型,利用载药膜预防屈趾肌腱粘连,通过大体观察、生物力学测试、组织切片观察,和对照组比较,分析防治粘连效果及对肌腱愈合的影响。同时,分别检测粘连组织中ERK1/2和SMAD2/3磷酸化水平,分析塞来昔布能否抑制粘连组织中ERK1/2和SMAD2/3的磷酸化进而抑制成纤维细胞的增殖和胶原蛋白的分泌,研究其预防肌腱粘连形成的可能作用机理。在此基础上制备多层仿生纤维膜,观察体内防治粘连效果及对肌腱愈合的影响。结果体外表征显示负载塞来昔布的PELA电纺纤维均匀、光滑,具有良好的药物缓释能力。体外细胞实验发现随塞来昔布浓度增加,负载塞来昔布的PELA纤维膜明显抑制了成纤维细胞和肌腱细胞的粘附和增殖。在兔肌腱修复模型中,对比PELA纤维膜和对照组,负载塞来昔布的PELA纤维膜明显减轻了腱周炎症,下调了粘连组织中ERK1/2和SMAD2/3的磷酸化(p0.05),抑制了I型和III型胶原的合成(p0.05),抑制了肌腱粘连,但部分损伤了肌腱愈合;而多层仿生纤维膜的体内试验显示其明显抑制了肌腱粘连且没有损伤肌腱愈合。结论负载塞来昔布的PELA纤维膜通过调节MAPK和SMAD信号通路,下调粘连组织中ERK1/2和SMAD2/3的磷酸化,抑制胶原蛋白合成和成纤维细胞的增殖,从而抑制了肌腱粘连。而多层仿生纤维膜模仿腱鞘结构,优化了负载塞来昔布的PELA纤维膜,预防了粘连的形成,同时避免了内源性愈合的损害。负载塞来昔布的PELA纤维膜不但揭示了肌腱粘连的机制,同时也为解决肌腱粘连这一临床难题提供了新的治疗策略,具有广阔的应用前景。
[Abstract]:Objective the prevention and treatment of postoperative adhesions after tendon injury has been a difficult problem for clinicians. One of the main causes of tendon adhesion is excessive proliferation of exogenous fibroblasts and excessive deposition of collagen fibers. Previous studies have confirmed that extracellular signal-regulated kinase 1 / 2 ERK1 / 2) and SMAD2/3 are excessive proliferation of exogenous fibroblasts. And the key targets for the over-synthesis of collagen fibers, Celecoxib can inhibit the proliferation of fibroblasts and the secretion of collagen by inhibiting the phosphorylation of ERK1/2 and SMAD2/3. Poly (PELA) fiber membrane and multilayer electrospun fiber membrane based on celecoxib PELA fiber membrane, To study its inhibitory effect on tendon adhesion and its mechanism, to provide a new therapeutic method and theoretical basis for preventing tendon adhesion and promoting the functional rehabilitation of affected limbs. Methods PELA electrospun fiber membranes with different contents of celecoxib were prepared by blending electrospinning. To observe the characterization of fiber membrane and drug release in vitro, to compare the physiological behavior of fibroblasts and tendon cells by PELA electrospun fiber membrane with different drug loading. Secondly, to establish a new Zealand rabbit tendon adhesion model. In order to prevent adhesion of flexor digitorum tendon with drug loaded film, the effect of preventing adhesion and its influence on tendon healing were analyzed by gross observation, biomechanical test, tissue section observation and control group. The phosphorylation levels of ERK1/2 and SMAD2/3 in adhesions were detected, and whether celecoxib could inhibit the phosphorylation of ERK1/2 and SMAD2/3 in adhesions and then inhibit the proliferation of fibroblasts and the secretion of collagen were analyzed. To study the possible mechanism of preventing tendon adhesion, and to prepare multilayer biomimetic fiber membrane, observe the effect of preventing adhesion and tendon healing in vivo. Results the PELA electrospun fiber loaded with celecoxib was uniform in vitro. In vitro, with the increase of celecoxib concentration, the PELA fiber membrane loaded with celecoxib significantly inhibited the adhesion and proliferation of fibroblasts and tendon cells. Compared with the control group, the PELA fiber membrane loaded with celecoxib significantly alleviated peritendinous inflammation, down-regulated the phosphorylation of ERK1/2 and SMAD2/3 in adhesion tissue, inhibited the synthesis of type I and III collagen, and inhibited tendon adhesion. In vivo experiments of multilayer bionic fiber membrane showed that it inhibited tendon adhesion and did not damage tendon healing. Conclusion Celecoxib loaded PELA fiber membrane regulates MAPK and SMAD signaling pathway. Down-regulation of phosphorylation of ERK1/2 and SMAD2/3 in adhesion tissue, inhibition of collagen synthesis and proliferation of fibroblasts, thereby inhibiting tendon adhesion. Multilayer biomimetic fibrous membrane mimics tendon sheath structure and optimizes PELA fiber membrane loaded with celecoxib. The PELA fiber membrane loaded with celecoxib not only reveals the mechanism of tendon adhesion, but also provides a new treatment strategy for solving the clinical problem of tendon adhesion. It has broad application prospect.
【学位授予单位】：上海交通大学
【学位级别】：博士
【学位授予年份】：2015
【分类号】：R687.2

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