双相纳米支架复合干细胞构建组织工程椎间盘

Tissue-engineered intervertebral disc implants provide a potentional solution to intervertebral disc (IVD) degeneration. Bone marrow-derived mesenchymal stem cells (MSCs) are a potential autologous stem cell source for IVD regeneration. However, one of the key issue of constructing a functional tissue-engineered IVD is to create unique niches that can "direct" a single MSC population to differentiate into the nucleus pulposus or anulus fibrosus zones of the IVD. The extracellular matrix (ECM) plays an important role in stem cell niche, a good scaffolding material should mimic the advantageous features of the natural ECM. Since the natural ECM is mainly composed of 50-500 nm nanofibers, a scaffold with synthetic nanofibers is needed..It has been reported that electrospun and oriented nanofibers could direct the MSCs differentiating into the fibrous phenotype. Therefore, the elespinning nanofibrous scaffold is suitable for annulus fibrosus engineering. However, the low cell penetration rate of electrospun nanofibrous scaffold limits their further application in nucleus pulposus regeneration. Previously, using phase separation, we developed a 3D scaffold with highly inter-connected macropores, nanofibrous matrix with a fiber diameter on the scale of 100 nm and a high surface to volume ratio. Combined with the low oxygen tension, the extracellular matrix-mimicking nanofibrous architecture could further mimic the physiological environment existing in the IVD, providing suitable stem cell niche for nucleus pulposus differentiation. Therefore, based on our previous work and result from literature, we focus on constucting a biphasic scaffold with nanofibers to regenerate the IVD. The outer phase of the scaffold was a ring-shaped electrospinning nanofibers, which mimicks the type I collagen orientation and ligamentous properties of anulus fibrosus. The inner phase of the scaffold was a porous nanofibrous scaffolds from phase separation, to recapitulate the nucleus pulposus, which is rich in type II collagen and proteoglycan. Using histology, gene expression assay, biochemical analysis, biomechanical test, We test the performance of the tissue-engineered IVD on three different model: in vitro, in vivo and rabbit lumbar repair model. Our final goal is to creating a single multi-layered sturcture with zone-specific biomaterials for the simultaneous differentiation of MSCs into structurally organized IVD tissue.

干细胞复合支架材料构建的组织工程椎间盘为椎间盘退变的治疗提供了新方法。然而，目前该领域的一大挑战在于创建适当的干细胞微环境"指导"同一群落的干细胞分化为椎间盘不同区域（髓核/纤维环）细胞表型。细胞外基质是干细胞微环境的重要组成部分，作为人工细胞外基质，支架材料应有效模拟天然细胞外基质（主要由直径50-500纳米的纤维组成）。因此，构建与椎间盘结构相适应的纳米支架材料，是目前亟需解决的问题。.电纺可编织同向排列的纳米纤维诱导干细胞分化为成纤维细胞并形成纤维组织，是构建纤维环的理想支架。而我们前期研究表明，相分离编织出的纳米支架联合低氧可成功模拟髓核微环境，构建组织工程髓核。在前期工作的基础上，本项目拟通过相分离联合电纺，编织纳米尺度的双相支架，营造分别与髓核/纤维环分化对应的干细胞微环境，"指导"干细胞在支架不同区域自发分化为相应细胞表型，从而形成具有生理功能的组织工程椎间盘。

干细胞复合支架材料构建的组织工程椎间盘为椎间盘退变的治疗提供了新方法。 然而,目前该领域的一大挑战在于创建适当的干细胞微环境“指导”同一群落的干细胞分化 为椎间盘不同区域(髓核/纤维环)细胞表型。细胞外基质是干细胞微环境的重要组成部分, 作为人工细胞外基质,支架材料应有效模拟天然细胞外基质(主要由直径 50-500 纳米的纤 维组成)。因此,构建与椎间盘结构相适应的纳米支架材料,是目前亟需解决的问题。.电纺可编织同向排列的纳米纤维诱导干细胞分化为成纤维细胞并形成纤维组织,是构建 纤维环的理想支架。而我们前期研究表明,相分离编织出的纳米支架联合低氧可成功模拟髓 核微环境,构建组织工程髓核。在前期工作的基础上,本项目拟通过相分离联合电纺,编织 纳米尺度的双相支架,营造分别与髓核/纤维环分化对应的干细胞微环境,“指导”干细胞在 支架不同区域自发分化为相应细胞表型,从而形成具有生理功能的组织工程椎间盘。