Bridging the Gap between Nano and Macroscale Hierarchies in Collagen Assembly

胶原蛋白作为一种生物材料和支架在再生组织替代策略中起着核心作用。肌腱和韧带等负重组织的手术每年有数十万例，与它们要替代的组织相比，现有的胶原蛋白合成类似物的生物力学性能非常差。这个缺点部分是由于在纤维层以上的层次顺序中缺乏定向。该项目将通过以下方式提高合成胶原结构的强度和粘弹性，以达到与天然胶原结构相匹配的水平：a)采用非常规的电化学工艺，实现前所未有的分子排列水平和贯穿所有结构层次的分子填充密度；2)使用仿生类decorin连接分子来控制纤维间的附着。拟议研究的第一阶段将通过阐明胶原蛋白溶液在直接应用于溶液的弱电作用下实现长期有序的机制来优化结构的机械强度和刚度。研究电流振幅和胶原蛋白浓度对胶原蛋白层次组织的影响，优化合成工艺。通过确定戊二醛、genipin、去二氢愈创木酸（NDGA）或核糖之间交联的适当类型和浓度，可以提高所得到的定向胶原凝胶的强度。第二阶段将调节定向和交联凝胶的粘弹性性能，通过由皮肤硫酸酯组成的decorin模拟物附着在选择性结合I型胶原蛋白分子的肽基序上。合成结构的机械性能将分别通过宏观力学测试和原子力显微镜在束和纤维水平上进行评估，并与大鼠肌腱（一种参考自然组织）进行比较。第三阶段将评估肌腱成纤维细胞在体外定向胶原结构三维网络中的表型和基因型反应，并通过评估体外构建物的非酶和酶降解率。该项目将包括使少数民族中学生熟悉新兴的生物医学工程领域的外联部分。这一目标将通过暑期活动来实现，在此期间，学生将通过与普渡大学少数民族工程项目的协调，在生物医学工程领域进行实践项目。通过整合拟议的研究成果，在本科生生物力学/生物材料实验室中创建一个实验室模块，并通过普渡大学夏季本科生奖学金计划（SURF）在项目期间容纳9名本科生进行夏季研究，将进一步加强更广泛的影响。总的来说，这项研究将为设计一种新的生物材料开发一种新的制造工艺，这种材料可能在创造替代组织（如肌腱、韧带、皮肤、角膜和血管壁）的策略中发挥关键作用。

CBET-0754442AkkusCollagen plays a central role as a biomaterial and as a scaffold in the regenerative tissue replacement strategies. Surgeries of load bearing tissues such as tendons and ligaments are occurring by hundreds of thousands annually and existing synthetic analogs of collagen have extremely poor biomechanical properties in comparison to the tissues they are targeted to replace. This shortcoming is due, in part, to the lack of orientation in hierarchical orders above the level of fibers. This project will improve the strength and viscoelasticity of synthetic collagenous constructs to match those of natural counterparts by: a) an unconventional electrochemical process to attain an unprecedented level of molecular alignment and molecular packing density persistent across all levels of structural hierarchies, and, 2) the control of interfibrillar attachment by use of a biomimetic decorin-like linkage molecule. Phase 1 of proposed studies will optimize the mechanical strength and stiffness of the construct by elucidating the mechanisms by which collagen solutions achieve long-range order under the effect of weak currents applied directly to the solutions. The effects of electric current amplitude and collagen concentration on the hierarchical organization of collagen will be investigated to optimize the synthesis process. The strength of resulting oriented collagen gels will be improved by identifying the appropriate type and concentration of crosslinking amongst glutaraldehyde, genipin, nordihydroguaiaretic acid (NDGA) or ribose. Phase 2 will modulate the viscoelastic properties of oriented and crosslinked gels by decorin mimics consisting of dermatan sulfate attached to peptide motifs which selectively bind to type I collagen molecules. Mechanical properties of resulting synthetic constructs will be assessed at the bundle and the fiber levels by macroscale mechanical tests and atomic force microscopy, respectively, and compared to those of rat tendon, a reference natural tissue. The third phase is going to assess the phenotypic and genotypic response of tendon fibroblasts seeded in three-dimensional networks of the oriented collagenous construct in vitro, and, by assessing the non-enzymatic and enzymatic degradation rates of constructs in vitro. The project will include the outreach component of familiarizing the minority middle-school student population with the emerging field of biomedical engineering. This aim will be attained by a summer activity during which students will conduct hands-on projects in the area of biomedical engineering through coordination with the Minority Engineering Program at Purdue University. Broader impacts will be further strengthened by creation of a laboratory module in an undergraduate biomechanics/biomaterials laboratory by incorporating outcomes of the proposed research and by way of accommodating 9 undergraduates for summer research during the course of the project through Summer Undergraduate Fellowship program (SURF) at Purdue. In the overall, the proposed study will develop a novel fabrication process towards the design of a new biomaterial which may play a key role in creating strategies towards replacement of tissues such as tendons, ligaments, skin, cornea and vascular walls.