可注射原位成型葡甘聚糖材料用于牙周组织修复的研究

The rapid progress of biomaterials science provides new hopes for the unmet clinical needs of periodontal tissue regeneration. Periodontal defects are usually irregular in shape and their regeneration involves multiple cell types of varying tissue origins. A biomaterial aimed at regenerating periodontal tissue should ideally be biocompatible, injectable and in-situ shape-forming; it is also expected to facilitate cell adhesion and growth, as well as regulate the regenerative functions of cells. In his previous studies, the applicant has found a naturally-derived glucomannan with a proper molecular size, a composition mimicking that of certain extracellular matrix components and favourable bio-activities. Particularly, the affinity between the sugar units of this glucomannan and the c-type lectins receptors on the surface of monocytes/macrophages enables the glucomannan to induce these cells to release tissue-trophic cytokines, in addition to the glucomannan’s capability in stimulating endothelial cell proliferation. Chemical modification of the glucomannan may lead to successful development of an injectable scaffold to facilitate regenerative cell adhesion and growth and thereby promote tissue restoration. As such, this study aims to acetylate the glucomannan, optimise the parameters to achieve desirable injectability and in-situ solidifying property in the acetyl glucomannan, investigate its capability to regulate the functions of various cells involved in periodontal regeneration and the underlying biological mechanism, and finally test its repairing performance in a rat periodontal defect model. Taken together, these efforts would provide important insights into the evaluation of the potential of the acetyl glucomannan as a novel injectable biomaterial for periodontal tissue regeneration.

生物材料学的迅速发展为牙周组织修复这一迫切的临床需求带了新的希望。牙周组织缺损的特点，如形状不规则，涉及细胞类型多，组织差异性大等，对生物材料的理化性质和生物学活性都提出了较高的要求：应生物相容性好、可注射并于原位成型、可促进细胞黏附生长且调控细胞修复活性。申请人的前期研究发现，天然来源的葡甘聚糖具有合适的分子量与类似细胞外基质的化学组成，其糖单元结构与单核/巨噬细胞表面的C型凝集素受体具有亲和性，可活化此类细胞释放帮助伤口愈合的细胞因子，及具有促进血管内皮细胞增殖的活性。因此，对此类葡甘聚糖进行化学修饰，有望开发可促进牙周前体细胞黏附生长、引导组织再生的可注射性材料。本项目拟就此进行立项，对葡甘聚糖进行乙酰化修饰，确定其在生理环境下的可注射性与原位成型性，研究其调节各类牙周组织细胞生长和活性的生物学机理，并建立大鼠动物牙周缺损模型进行实验，探讨其作为新型可注射牙周修复材料的可能性。

本项目针对牙周组织缺损这一临床巨大需求，根据生物化学、细胞生物学与免疫学原理，结合生物材料工程技术，开发一款可注射生物活性材料。该材料为富含葡萄糖与甘露糖单元的多糖聚合物（葡甘聚糖）材料，其特性是可通过调节损伤局部的巨噬细胞活性，促进伤口愈合和组织修复。研究主要内容为：葡甘聚糖的提取纯化与化学修饰、葡甘聚糖材料的理化特性与可注射性表征，以及葡甘聚糖材料的体外和体内生物学评价。本研究发现，针对一定分子量的天然葡甘聚糖进行适当乙酰化修饰后，可于特定溶剂（二甲基亚砜）溶解且注射至生理环境（如生理缓冲液）后迅速形成具有多孔结构的三维支架。由于单核/巨噬细胞表面富含可识别和结合上述两种糖单元的受体，这类细胞可有效富集、黏附于所得葡甘聚糖材料表面，表达促进组织修复再生的相关基因（如血管内皮生长因子VEGF，血小板衍生生长因子PDGF，促进成骨的oncostatin M等等）。数据表明，这类黏附活化与巨噬细胞表面的toll-like receptor及integrin家族受体介导的信号通路密切相关。在体外共培养体系与体内异位成骨模型中，所得乙酰葡甘聚糖支架可通过原位激活巨噬细胞活性促进成骨前体细胞（包括成骨细胞osteoblast与间充质干细胞MSC）的生长、分化与成熟，以碱性磷酸酶（ALP）和骨钙素（OCN）为代表的成骨基因标志物表达上调。同时，植入材料无毒，无刺激，动物无异样。综上所述，本研究通过一系列实验探索，开发出新型的、具有“通过原位调节巨噬细胞功能从而促进骨组织修复和再生”活性的可注射生物材料支架。本研究作为初期探索性研究顺利完成，研究结果支持进一步在特定疾病缺损模型中展开深入探索。