纳米绒球状细胞外基质源性微载体高效扩增软骨细胞并维持其表型的作用机制

Tissue engineering technology is an effective method for the treatment of large articular cartilage defects. How to get enough high-quality chondrocytes in a short time has become a key point in cartilage tissue engineering. We successfully prepared pompon-like microcarriers derived from articular cartilage extracellular matrix(CEDMs) . The preliminary research results showed that these microcarriers have large surface and good biocompatibility for its nano-structure surface and are rich in extracelluler matrix of cartilage. Under Microgravity culture conditions, the chondrocytes cultured by our micrrocarriers express more cartilage-related gene compared with that of Cytodex3, an traditional microcarriers.. Based on these results, we detect not only the molecular biological mechanism of maintaining chondrocytes phenotye and proliferation of human articular chondrocytes on pompon-like microcarriers under microgravity culture condition,but also the role of redifferentiation of the dedifferentiated chondrocytes.As a kind of scaffolds, CEDMs with their cultured chondrocytes can be reconstructed into injectable tissue engineering cartilage to repair the articular cartilage defects without trypsin digestion, which is another porpose of our study.

组织工程技术是治疗大面积软骨损伤有效方法，但如何在短时间内获得足够的高质量的软骨细胞成为组织工程研究的重点工程之一。本项目前期研究成功制备出绒球状关节软骨细胞外基质源性微载体，该微载体表面呈微绒毛纳米结构，富含II 型胶原和GAG，表面积大；微重力环境下，该微载体除了可以在短时间内大量扩增细胞外，所培养的软骨细胞更多地表达成软骨基因（Aggrecan、Ⅱ型胶原、Ⅹ型胶原和SOX-9基因表达量分别是传统微载体培养软骨细胞表达量的4倍、1.9倍、2.4倍和4.9倍）。.本项目以前期研究结果为基础，①研究在微重力培养条件下，这种绒球状微载体扩增软骨细胞、维持软骨细胞表型的分子生物学机制；②观察该微载体对去分化软骨细胞再分化方面的作用；③在不损失细胞外基质的情况下，以微载体为组织工程支架，构建可注射型组织工程软骨，并对这种微载体-细胞复合体修复大动物关节软缺损的效果进行评价。

组织工程技术是治疗大面积软骨损伤有效方法，但如何在短时间内获得足够的高质量的软骨细胞成为组织工程研究的重点工程之一。本项目通过优化预实验过程中关节软骨源性微载体（CEDMs）制备方法，最终得到具有70-150微米绒毛覆盖表面、大小均一、形状较规则、无DNA残留的关节软骨源性微载体；对关节软骨源性微载体进行组织学及免疫组化染色观察，发现该种微载体保留了软骨组织特有细胞外基质成分（GAG、II型胶原）；在对软骨源性微载体的细胞相容性观察中发现，该微载体能够促进人软骨细胞的增殖；在增殖过程中与商业化微载体Cytodex-3相比，关节软骨源性微载体表达更多的软骨细胞相关基因，抑制软骨细胞的再分化（dedifferentiation）；通过将软骨源性微载体与人软骨细胞复合体注射到裸鼠皮下，发现软骨源性微载体具有异位成软骨能力且所成软骨组织的弹性模量更接近天然软骨组织，为将来应用该软骨源性微载体作为天然支架体内修复软骨缺损提供了有力的理论基础。所得成果共发表SCI论文3篇，国内核心期刊2篇；共培养博士研究生生1名，硕士研究生2名。