构建聚乳酸-羟基乙酸电纺丝-壳聚糖电喷微球牙周仿生膜

Construction of periodontal biomimetic membrane with electrospun poly(lactic-co-glycolic acid) nanofibers and electrosprayed chitosan microspheres

背景:当牙齿脱离牙槽窝后,牙周膜断裂,残留在脱位牙根表面的牙周膜由三维变成二维,丧失了支架膜的作用,导致脱位牙再植后根骨粘连。如何研发一种能黏附牙根表面具有一定厚度及强度的三维缓释支架材料,是脱位牙牙周膜再生成功的关键之一。目的:构建可黏附脱位牙根表面的缓释生长因子的三维仿生膜。方法:采用静电纺丝技术制备聚乳酸-羟基乙酸电纺膜,研究电纺溶剂二氯甲烷与二甲基甲酰胺混合溶液、六氟异丙醇、三氯甲烷对电纺膜的影响,筛选最佳的电纺溶剂。采用电喷技术与离子交联法制备壳聚糖微球,研究壳聚糖相对分子质量(5万、10万)与质量浓度(10,20g/L)、接受液三聚磷酸钠浓度(2%,5%,10%)、电压(14,28 kV)对壳聚糖微球的影响,筛选最佳的参数。构建含基质细胞衍生因子1壳聚糖微球(最优参数设计),检测其体外释放基质细胞衍生因子1α的速率。首先将聚乳酸-羟基乙酸电纺膜裹在牙齿根表面,然后在其表面滴加壳聚糖微球,在其外层裹一层薄薄的聚乳酸-羟基乙酸电纺膜,从而形成聚乳酸-羟基乙酸-壳聚糖微球-聚乳酸-羟基乙酸膜。结果与结论:(1)利用电纺溶剂六氟异丙醇制备的聚乳酸-羟基乙酸电纺膜平均直径最小、空隙率最大;(2)当壳聚糖相对分子质量为5万、质量浓度为20 g/L时,微球的大小基本一致,平均直径366.6μm,单分散性好、饱满、稳定;28 kV电压下形成的壳聚糖微球更符合脱位牙仿生膜的要求;利用5%三聚磷酸钠制备的壳聚糖微球表面微观结构孔径居中,最有利于临床牙周膜再生;壳聚糖微球可持续释放基质细胞衍生因子1α1个月左右;(3)实验创建了一种黏附牙齿表面的具有缓释效能的聚乳酸-羟基乙酸-壳聚糖微球-聚乳酸-羟基乙酸三维仿生膜并筛选出构建此仿生膜的最佳参数,可在此模型基础上进一步研究组织工程手段对脱位牙再植的效果及机制。

BACKGROUND: When the teeth are separated from the alveolar fossa, the periodontal membrane breaks, and the residual periodontal membrane on the avulsed tooth root surface changes from three-dimensional to two-dimensional, thus losing the role of scaffold, and leading to root bone adhesion after replantation of avulsed tooth. How to develop a three-dimensional sustained-release scaffold material that can adhere to the root surface with a certain thickness and strength is one of the key factors for successful regeneration of avulsed tooth periodontal membrane. OBJECTIVE: To construct a three-dimensional periodontal biomimetic membrane that can adhere to the avulsed tooth root surface and allow sustained-release of growth factors. METHODS: Poly(lactic-co-glycolic acid)(PLGA) membrane was prepared using electrospinning technique. The effects of dichloromethane and dimethylformamide mixture, hexafluoroisopropanol, and trichloromethane on electrospun membrane were investigated t o obtain the optimal electrospinning solvent. Chitosan microspheres were prepared by electrospray and ion cross-linking techniques. The effects of molecular weight(50,000,100,000) and mass concentration(10, 20 g/L) of chitosan, sodium tripolyphosphate concentration(2%, 5%, 10%) and voltage(14, 28 kV) on chitosan microspheres were studied to screen the optimum parameters. Chitosan microspheres containing stromal cell-derived factor-1(optimal parameter design) were constructed. The release rate of stromal cell-derived factor-1 alpha in vitro was determined. First, the root surface of teeth was wrapped with electrospun PLGA membrane, then chitosan microspheres were dripped on the surface, and finally the surface was wrapped with a thin layer of electrospun PLGA. Thus, PLGA-chitosan-PLGA biomimetic membrane was constructed. RESULTS AND CONCLUSION: Electrospun PLGA membrane prepared with hexafluoroisopropanol as electrospinning solvent had the smallest average diameter and the largest porosity. When the relative molecular weight of chitosan was 50,000 and the mass concentration was 20 g/L, the size of chitosan microspheres was basically the same, and the average diameter was 366.6 μm. In addition, chitosan microsphere had good monodispersity, fullness, and stability. Chitosan microspheres formed under 28 kV voltage and were more in line with the requirements of biomimetic membrane for avulsed tooth. The surface of microspheres prepared by 5% sodium tripolyphosphate had medium-sized pores, which are most conducive to clinical periodontal membrane regeneration. Chitosan microspheres can sustainably release stromal cell derived factor 1 alpha for about 1 month. In this study, we constructed a three-dimensional PLGA-chitosan-PLGA periodontal biomimetic membrane that can adhere to the avulsed tooth root surface and allow sustained-release of growth factors and obtained the optimal parameters of constructing the periodontal biomimetic membrane. Based on the PLGA-chitosan-PLGA periodontal biomimetic membrane, the effect and mechanism of tissue engineering on replantation of avulsed tooth can be further studied.