【作者】 唐华；

【导师】 徐志飞；

【作者基本信息】 第二军医大学 ， 外科学， 2010， 博士

【摘要】 研究背景胸壁肿瘤、放射性溃疡、感染、外伤及先天性畸形等疾病的广泛胸壁切除及胸部创伤均可造成胸壁缺损。大块胸壁缺损的重建对于外科医生仍然是一个挑战,人工材料应用于胸壁缺损修复,大大拓展了可选材料来源,推动了胸壁重建外科的发展。许多材料曾被尝试应用于胸壁重建,然而到目前为止,还没有一种完全理想的胸壁重建材料。目前临床上可用的材料仍然存在各种问题：与组织结合能力差,难以与组织有机融合,后期常可松动、移位、活动甚至脱出；长期植入后易导致血清肿、瘘管形成、疼痛、血管破坏及引发感染等并发症,部分病人后期需要再次手术取出。而对于重建方式来说,目前应用最多的仍是MARLEX网+骨水泥+MARLEX网的三明治结构,但是其不能进行解剖和功能的双重重建,往往会影响胸廓的美观或者影响术后对于内脏器官的观察。目前,组织工程技术的临床应用已经取得了举世瞩目的成绩,包括骨、软骨、皮肤等多种组织的再生。组织工程技术主要有两个关键因素,即种子细胞和支架。骨髓基质干细胞作为种子细胞已经得到了公认,其可向多种细胞分化,而对于支架材料的应用,目前种类和方式较多,但是到目前为止没有一种完全理想的材料和方法,对于骨缺损来说,目前只能修复短缺损,而对于长段缺损仍然束手无策。研究目的一、在前期的研究基础上,优化PDO补片,探索新编织方法,使其带有管道结构,为后期的仿生型人工胸壁提供软组织修复支架；同时筛选出符合要求的组织工程人工肋骨支架,为人工胸壁硬组织修复提供支架；二、利用多段支架结合组织工程技术修复长段肋骨缺损,证明分段融合定向成骨修复长段骨缺损的可行性；三、利用PDO补片和分段融合定向成骨方法修复大面积胸壁缺损,证明其可行性。研究内容与方法一、仿生型人工胸壁补片的选材、设计与编织我们在前期的研究工作基础上,继续采用PDO单丝作为补片的原材料,探索其新编织方法,使其能满足以下条件：1)补片大小为8cm×5.5cm；2)补片带有两个横向的管道结构；3)补片具有较好的孔径大小。二、组织工程人工肋骨三维支架的筛选按照医疗器械生物学评价IS010993-1998和GB／T16886-2001标准和要求,对三种待试材料进行了体内外生物相容性和安全性试验研究,包括细胞毒性试验、急性全身毒性试验、溶血试验、热源试验,体内降解试验等,同时对其结构和组份进行测试,筛选出更符合要求的肋骨支架。三、分段融合定向成骨修复长段骨缺损的实验研究建立犬单根肋骨长段缺损动物模型,并利用双段支架结合骨髓基质干细胞和PDO管道来修复此缺损,分别于术后各时间点行影像学检查,并于24周时处死所有实验犬,利用组织学检查观察肋骨修复情况,证明分段融合定向成骨修复长段骨缺损的可行性。四、仿生型复合式人工胸壁动物模型的建立建立犬胸壁缺损动物模型,应用自行设计制备的仿生型人工胸壁重建胸壁缺损,并设置相应对照组,动态观察胸壁组织的再生过程,包括软组织和硬组织,同时进行大体标本、影像学、组织学和力学检查进行验证,探讨利用补片结合分段融合定向成骨修复大面积胸壁缺损的的可行性。研究结果一、利用穿综方法编织出了带有双管道结构的PDO补片,其大小为8cm×5.5cm,管径为1.25cm,两管道间间隔1cm,孔径大小为250um×250um,补片能够进行任意裁剪。二、脱钙骨、多孔磷酸钙(CPC)、和PLGA/HA三种待试材料均具有较好的生物相容性,安全无毒；但是从体内降解实验来看,PLGA/HA材料8周时已经完全失去强度,降解速度太快,与骨组织再生速度不匹配,而其它两种材料均具有较好的力学强度；结构测试表明脱钙骨和CPC同时具有较好的孔径、孔隙率和HA含量,但CPC材料脆性较脱钙骨大,因此脱钙骨是较好的组织工程肋骨支架。三、实验结果表明,我们所制备的4cm肋骨缺损超出了自身的再生能力,可以认为是长段骨缺损,影像学检查和组织学检查证明PDO管道/DBM/BMSCs组的肋骨无论是在原肋骨与支架之间还是支架与支架之间均存在着骨连接,分段融合定向成骨能够修复长段骨缺损。四、除空白组外,所有实验犬均存活。对于软组织重建而言,PDO补片于术后6月时已经完全降解,原PDO补片为纤维结缔组织所替代,其厚度与正常肋间隙的厚度几乎一致,统计学无差异(P>0.05)；而对于肋骨重建而言,影像学检查和组织学检查均证明有新生骨存在,支架间均为骨性连接,而在其他两个组均为纤维连接,没有力学强度,且其弧度与原来肋骨具有相同的弧度；力学测试也证明重建的肋骨强度达到了正常肋骨强度的70%左右,统计学没有差异(P>0.05)。实验结论异体脱钙骨具有良好的生物相容性和力学强度,是组织工程骨理想的支架材料；利用多段支架结合组织工程技术能够修复长段骨缺损,这为长段骨缺损的修复提供了新思路和新方法；利用我们所制备的仿生型复合式人工胸壁能够进行胸壁缺损解剖和功能的双重重建,其能进行胸壁软组织和硬组织的同时重建,是一种具有较好前景的胸壁缺损修复方法。更多还原

【Abstract】 BackgroundReconstruction of chest wall defects remains a great challenge for the reconstructive surgeons. Extensive chest wall defects, which result primarily from tumor, radiation necrosis, infection, or trauma, often require prosthetic repair to prevent flail chest and paradoxical breathing. The artificial prostheses have gained increasing popularity as it provides better stability with shorter and easier surgical procedure. Over the years, many different types of materials have been introduced and used to repair the chest wall. However, no product possesses the properties of the ideal material and late wound complications such as prosthesis dislocation, infection, fistulas or dense scar tissue formation, occur frequently in the chest wall reconstruction, due to the properties of the employed materials. As to the method of reconstruction, the sandwich structure "marlex mesh+bone cement+marlex mesh" is the most popular method in clinic use, but it can’t reconstruct the chest wall defect both in anatomy and function which result in affecting the shape of chest wall and the observation of the organ in the chest cage.Now, tissue engineering technique has achieving tremendous success in clinic use, including bone, cartilage,skin reconstruction. Tissue engineering technique contains two critical factors, seed cell and scaffold. Bone marrow stem cell(BMSC) which can been induced to various cell is an acknowleded seed cell. As to the use of scaffolds, there are many different types and forms, but to now there is no ideal material and method. And long bone defect reconstruction is still a challenge problem.Objective1.On the base of early research, explore new method of weavement to improve the PDO mesh which would have two cages and to be the scaffold of soft tissue reconstruction; Choose the most suitable tissue engineered rib scaffold to reconstruct the rib defect;2. To prove the possibility of the method that using multiple scaffolds to reconstruct the long defect of rib,3. To prove the possibility of chest wall reconstruction using PDO mesh and tissue engineering ribMethods 1. The design, preparation of bionic chest wall prosthesisOn the base of early research, PDO suture will still be used as raw material for mesh. New method of weavement will be explored and the improved mesh will satisfy the following requirement:1) The size of the mesh is 8cm×5.5cm; 2) the mesh has two cages; 3)the mesh has many pores which allow the adhesion of tissue2. The selection of tissue engineering rib scaffoldthe biocompatibility and biological safety of the three materials for chest wall prosthesis were evaluated comprehensively in accordance with ISO10993-1:1998 and GB/T16886.1-2001, including in vitro cytotoxicity test, acute systemic toxicity test, hemolysis test, and pyrogen test, and in vivo biodegradation test. In the while, the structure and components of the scaffolds will be analyzed to select the most suitable scaffold.3. the research of long defect of rib reconstruction using multiple scaffoldsThe compound of the PDO cage containing two demineralized bone matrix(DBM) which seeded with osteogenically induced BMSCs was used to reconstruction the single long defect of rib defect. The radiographic examination and histological examination were used to observe the new bone regeneration.4. Establishing the animal model of the chest wall reconstruction using bionic,multiplex prothesisEstablish the animal model of chest wall defect, then reconstruct it with the bionic artificial chest wall designed by ourselves. Observe the regeneration of chest wall tissue including soft tissue and hard tissue by gross view of the sample, radiographic examination, histological examination and mechanical test. Explore the possibility of reconstruction of chest wall defect with biodegradable mesh and tissue engineering rib.ResultsPart 1 We successfully weaved a new PDO mesh which had two cages. The mesh’s size is 8cm×5.5cm. The cages diameter is 1.25cm. The intervals between the two cages is 1cm in width. The mesh had a lot of pores whose size is 250um×250um. Besides, the mesh also can be optionally cut out.Part 2 DBM, CPC and PLGA/HA have good biocompatible and they are all nontoxic. The in vivo biodegradation test showed that the degradable speed of PLGA/HA was so fast that it lost its strength 8 weeks after it was inplant into the animal body, in the mean while, the other two materials had good mechanical strength. The structure test showed that DBM and CPC had good pore and HA content. But CPC is more fragile than DBM. So DBM is the most suitable scaffold for tissue engineering rib.Part 3 The result showed that the 4-cm long defect of rib is acknowleded as the long defect of bone as it exceeded the natural regeneration capability. Radiographic and histological examination showed that there were bone union not only in the connection of two scaffolds but also in the connection of scaffold and primary rib. Reconstruction of the long defect of bone using multiple pieces of scaffolds was a feasible method.Part 4 All the dogs survived after surgery while dogs in blank group died. As to the soft tissue reconstruction, PDO mesh completely degraded 6 months after surgery and the primary mesh was replaced by fibrous tissue whose thickness was nearly equal to the normal one.(No significant difference were found between the two groups (P>0.05). As to the rib reconstruction, Radiographic and histological examination showed that there were bone union not only in the connection of two scaffolds but also in the connection of scaffold and primary rib in the experimental group while they were fibrous union in other two groups which had no mechanical strength. The reconstruction rib’ s radian was analogus to the primary rib. The result of mechanical test also proved that the strength of reconstructed rib was familiar to the primary one(No significant difference were found between the two groups (P>0.05).ConclusionsXenograft-DBM is a ideal scaffold for tissue engineering tib as it has good compatible and mechanical strength; Reconstucting the long defect of rib with multiple pieces of scaffolds seed with BMSCs is a promising method for the long defect of bone reconstruction; The bionic prothesis can reconstruct the chest wall not only in anatomy but also in function. It can reconstruct the soft tissue and hard tissue in the same time which make it a new and promising method for chest wall reconstruction.更多还原