基于柠檬酸的新型生物支架修复大段骨缺损及其促进骨矿化的机制研究

Complicated fractures combined with large bone defects have always been one of the most challengeable diseases in orthopedics. However, with the increasingly development of tissue engineering technology, biodegradable polymer materials have achieved good effects in the treatment of these diseases, highly biomimetic, complete osseointegration, and side-effect-free biodegradation in vivo are the main problem faced with present biomaterials. Recently, we developed a biodegradable, mechanically strong, citrate-based polymer blend/HA (CBPBHA) composite with a compressive strength of 116.23 ± 5.37 MPa similar to that of human cortical bone without altering their osteoinductivity and osteoconductivity. In our preliminary experiment, the composites perform well integrated with the host bone with almost no fibrous tissue encapsulation after 6 weeks of implantation into the lateral femoral condyles of rabbit knees. What's more, we also found the composites can greatly up-regulate the expression of ALP and Osterix mRNA, which are the key factors in osteogenesis, during the osteogenic differentiation of C2C12. Besides, what makes we more interesting is that, the citric acid can obviously enhance the bone mineralization in the osteogenic differentiation of bone marrow stromal cells. The discovery of CBPBHA composites bridges the gap in previous bone biomaterial designs in that the role of citrate molecules was inadvertently overlooked. The CBPBHA composites represent a new generation of bone biomaterials which offers new opportunities to tackle the challenges in bone tissue engineering and in the treatment of orthopedics persistent diseases. In this research, we intend to further study the short and long-term effects of CBPBHA scaffolds combined with bone marrow stromal cells transplantation in the management of goats' large bone defects, and to further explore the mechanism of its bone mineralization enhancement.

伴大段骨缺损的复杂性骨折是骨科顽症之一，随着组织工程技术的日新月异，可降解高分子材料在修复骨缺损中已取得较好疗效。但高度仿生性、完全骨整合能力及在体降解等问题困扰当前骨生物材料的发展。本课题组最近研制出一种基于柠檬酸的新型可降解高分子材料，首次将天然骨中5%柠檬酯成分纳入材料设计，且能复合65%羟基磷灰石，最大程度模拟天然骨的构成。我们预实验发现：该材料模量和强度可与人体皮质骨相媲美且降解可调，植入兔体内6周后不产生慢性炎症反应，具有很强的骨诱导和骨整合能力；且能显著提高成骨分化早期碱性磷酸酶（ALP）和Osterix mRNA表达水平，并极大促进骨髓基质细胞(BMSCs)成骨分化过程中钙结节形成。本项目拟使用该材料联合BMSCs细胞移植，在大动物（山羊）水平，进一步研究其修复大段骨缺损的近期和远期疗效及其作用机制，并在体外探讨柠檬酸促进成骨分化的分子机制。为该材料今后进入临床提供实验依据.

在全世界范围内，每年大约有220万的骨移植手术，已经成为及输血之后的第二大移植手术，大部分的骨移植手术都能完全愈合，但是也有一小部分骨移植手术不愈合或者是延迟愈合。其中最重要的原因是缺乏合适的骨移植材料，所以移植材料的选择对于骨移植手术是极其重要的。目前移植物主要有三种：自体骨移植、同种异体骨移植以及组织工程骨生物材料移植等。在几十年来的发展历程中，已有大量人工组织工程生物材料先后问世，在临床修复治疗中取得了令人欣慰的疗效。因此，组织工程骨生物材料在将来有望取代自体骨移植和同种异体骨移植，骨移植手术的首选方法之一。 .本课题组研发的基于柠檬酸的新型可降解高分子无机复合材料（ CBPBHA） 由氨基掺杂聚酯弹性体及聚二醇柠檬酸酯构成高分子共混物与羟基磷灰石复合。利用柠檬酸的多羟基结构特点，能和羟基磷灰石的钙紧密螯合，极大提高了该材料复合 HA 的能力（最多能复合 65%的 HA）及其生物力学强度。该材料的模量与强度可与人体皮质骨相媲美且强度与降解可调，动物实验表明该材料具有很强的骨诱导能力，与大多数合成材料不同，该材料在体外能显著促进 BMSCs 成骨分化过程中钙结节形成，在体内 6 周后不产生慢性炎症反应，且能和周围天然骨充分融合。动物实验中，将支架材料移植于兔子的桡骨缺损处，15周后缺损处基本愈合；将支架材料模拟椎间融合器移植于兔子椎体间能够使上下椎体融合。细胞实验结果表明该材料能显著促进小鼠 C2C12 细胞成骨分化早期碱性磷酸酶（ ALP）和 Osterix mRNA 的表达水平。结果表明，此种生物材料具有很强的骨诱导性、骨传导性和良好的生物相容性，而且能够提供足够的支撑强度。