Regeneration of tissue interfaces: Biomaterial controlled morphogen gradients for tissue patterning in regeneration

关节中骨软骨缺损的愈合仍然是整形外科医生面临的挑战，目前的技术都没有实现骨和软骨阶段的完全重建和功能。治疗的有限临床成功的一个原因是骨和软骨组织在骨软骨连接处的紧密相互作用，这是一种典型的独特组织模式。通过所提出的项目，我们希望使用骨软骨缺损作为示例系统来验证再生医学中不同组织模式的新概念。我们的概念基于发育生物学的基本原理，其中包括局部激活剂和抑制剂紧密地作用在一起以形成不同的组织模式。为此目的，合作开发了一种多层构建体（基于微球的PLG支架），其允许刺激剂及其拮抗剂从不同层的受控释放。我们计划采用我们的生物力学控制良好的骨软骨缺损模型在绵羊体内验证的概念，并比较内源性因子释放。具体地，所述层状支架由含有TGF β和抗BMP 4的软骨区组成，含有抗BMP和抗TGF β抗体的中间缓冲区和由BMP 4和抗TGF β组成的成骨诱导层。我们的假设是，来自PLG支架的刺激因子和相反因子的空间控制释放可以引导组织图案化（软骨样组织-潮标-软骨下骨）在绵羊骨软骨缺损模型中的作用。我们分析的基础是通过早期血肿形成评价骨软骨缺损的内源性反应。为此，我们将研究在植入不含生长因子或仅含TGF β 3或BMP 4的支架后的第一个小时内启动的炎症反应级联反应。我们将主要研究新形成的骨软骨缺损中血肿中的细胞因子模式和炎症细胞反应。在下一步中，将通过植入负载有TGF β或BMP-4的支架来分析TGF β和BMP-4对该相的可吸收性。总之，拟定的分析将允许判断基于生物材料的策略在不同组织连接处（如骨软骨修复）局部控制组织形成的可行性。此外，如果这种方法有助于克服目前通过生长因子TGF β和BMP 4进行骨软骨修复的临床缺点，将允许获得理解。一旦得到验证，这种不同组织形成的概念可以应用于多个其他甚至更复杂的器官结构。

The healing of ostechondral defects in the joint still remains a challenge for orthopaedic surgeons and none of the current techniques achieve full reconstruction and functionality of the bone and cartilage phases. A reason for the limited clinical success of treatments is the tight interplay of bone and cartilage tissues at the osteochondral junction, a classical distinct tissue pattern. With the proposed project we want to validate a new concept for distinct tissue patterning in regenerative medicine using osteochondral defects as an example system. We base our concept on a fundamental principle of developmental biology, which includes locally activators and inhibitors acting tightly together to form distinct tissue patterns. For that purpose a multi-layered construct (microsphere-based PLG scaffold) that allows for the controlled release of both stimulatory agents and their antagonists from distinct layers ws developed in cooperation. We plan to employ our biomechanically well controlled osteochondral defect model in sheep to validate the concept in vivo and compare to endogenous factor release. Specifically, the layered scaffold is composed of a cartilage zone containing TGFbeta and anti-BMP4, a middle buffer zone containing anti-BMP and anti-TGFbeta antibodies and an osteogenic inductive layer composed of BMP4 and anti-TGFbeta It is our hypothesis that spatially controlled release of both stimulatory and opposing factors from the PLG scaffold can steer tissue patterning (cartilage like tissue-tidemark-subchondral bone) in an osteochondral defect model in sheep. Basis to our analyses will be the evaluation of the endogenous response to an osteochondral defect by means of the early hematoma formation. Towards that aim, we will investigate the inflammatory response cascade initiated within the first hours after implantation of a scaffold containing either no growth factors or TGFbeta3 or BMP4 alone. We will primarily investigate the cytokine pattern and inflammatory cell response in the hematoma that develops in the freshly created osteochondral defect. In the next step, the suggestibility of this phase by TGFbeta and BMP-4 will be analyzed by implanting the scaffold loaded either with TGFbeta or BMP-4. In summary, the proposed analyses will allow judging on the feasibility of a biomaterial-based strategy to control locally tissue formation at distinct tissue junctions such as in osteochondral repair. Further, it will allow gaining understanding if such approach is helpful to overcome the current clinical shortcomings in osteochondral repair by the growth factors TGFbeta and BMP4. Once validated, this concept of distinct tissue formation may be applied to multiple other even more complex organ structures.