类弹性蛋白多肽作为温敏型液体栓塞剂治疗食管胃底静脉曲张的实验研究

Acute variceal bleeding is a fatal complication in patients with liver cirrhosis. Endoscopic variceal obturation with an injection of histoacryl is the most appropriate treatment. However, histoacryl injection may cause some serious complications, including embolization into systemic vessels, excreted from the body and bleeding, mucosal necrosis, obstruction of the endoscopic channel and needle sticking in the varix. Moreover, the material costs a great deal. Therefore, it is mandatory to search a safe, effective, and inexpensive agent for endoscopic variceal obturation. Elastin-like polypeptides (ELPs) are a unique class of genetically engineered, thermosensitive peptide polymers that have several attractive properties, such as exquisite control of the phase transition property, biocompatible and easy to obtain at low cost. We have confirmed that ELPs are injectable by using this material as submucosal injection solution in endoscopic submucosal dissection successfully. ELPs displayed a satisfactory embolic effect by embolizing the simulation embolic model and rabbit’s femoral artery. Because of its properties, ELPs have great potential as an embolic agent. In this study, we will determine the optimal amino acid composition and molecular weight of ELPs by evaluate the phase transition property. The gastroesophageal varices of dogs will be endoscopic embolized with ELPs radiolabeled with 131I. Endoscopic ultrasonography, radiography and histology examination will be performed to evalulate the embolic outcome. In a word, the aim of this study is to evaluate the feasibility of ELPs used as a thermosensitive liquid embolic agent for treatment of gastroesophageal varices.

食管胃底静脉曲张破裂出血是肝硬化最为凶险的并发症。内镜下注射组织黏合剂为重要的治疗方法，但存在异位栓塞、排胶出血、黏膜坏死等并发症，并常因黏合剂泄漏致内镜孔道堵塞、注射针与血管粘连，且费用高昂。因此，寻找更安全、有效、经济的栓塞剂迫在眉睫。类弹性蛋白多肽(ELPs)是一类温敏自组装生物多肽，其相变特性精确可控、组织相容性好且成本低廉。我们已将其作为分离剂用于内镜下黏膜剥离治疗，证实其经内镜可注射；通过体外栓塞模拟装置及兔股动脉栓塞初步证实其可栓塞性。本研究将从基因水平合成含有不同氨基酸组成及长度的ELPs，评价其相变特性，筛选出具有最佳栓塞性能的ELPs；以131I标记ELPs，经内镜栓塞治疗造模犬食管胃底曲张静脉，通过超声胃镜、影像学和组织病理学方法评价栓塞后血管的组织病理学动态演变过程、ELPs的病理转归及血管重建情况。旨在探讨ELPs作为温敏型液体栓塞剂治疗食管胃底静脉曲张的可行性。

食管胃底静脉曲张破裂出血是肝硬化最为凶险的并发症。内镜下注射组织黏合剂为重要的治疗方法，但存在异位栓塞、排胶出血、黏膜坏死等并发症，并常因黏合剂泄漏致内镜孔道堵塞、注射针与血管粘连，且费用高昂。因此，寻找更安全、有效、经济的栓塞剂迫在眉睫。类弹性蛋白多肽是一类由五肽重复序列单元串联而成的多肽聚合物，源自哺乳动物弹性蛋白原的疏水区域，其相变特性精确可控、组织相容性好且成本低廉，在生物医学材料方面展现出广阔的应用前景。本课题组基于其基因的特殊性，根据大肠杆菌密码子的偏好性和简并性，结合依次插入单体基因法、递归定向连接法、重叠延伸滚动循环扩增法，获得了具有不同客座氨基酸组成和分子量的多肽基因库，通过重组菌发酵、诱导过量表达以及可逆相变循环法分离纯化，获得高均一性重组类弹性蛋白多肽。系统测定了内部因素 (客座氨基酸组成、分子量) 和外部因素 (多肽浓度、离子浓度) 对多肽相变温度的影响，初步探索其变化规律：客座残基的疏水性越强，相变温度越低；亲水性越强，相变温度越高；分子量大小与相变温度成负相关；随着多肽浓度和离子浓度的升高，相变温度逐渐降低。选择前期构建合成的相变温度介于25~30℃的类弹性蛋白多肽，通过栓塞性能评价，优化出具有栓塞性能的类弹性蛋白多肽（以缬氨酸和半胱氨酸组成的多肽V80C20）。通过门腔静脉分流联合血管缩窄环方法成功构建门脉高压食管胃底静脉曲张动物模型。以重组类弹性蛋白多肽作为新型原位凝胶化液体栓塞剂成功用于食管胃底曲张静脉的栓塞治疗，证实了类弹性蛋白多肽作为温敏型液体栓塞材料经内镜栓塞治疗食管胃底曲张静脉的可行性、安全性和有效性，拓展了类弹性蛋白多肽在消化内镜治疗领域的应用，为内镜下栓塞治疗提供了一种更为安全、有效、经济的栓塞材料。