3D打印PLA-HA复合材料构建组织工程骨的实验研究  

Experimental reserarch of constructing tissue engineered bone using three-dimensional printed polylactic acid-hydroxyapatite composite scaffolds

作　　者：张海峰[1,2] 杜子婧[1] 毛曦媛 赵丹阳[1] 杜朝[1] 姜闻博[3] 韩冬[1] 

机构地区：[1]上海交通大学医学院附属第九人民医院整复外科,200011 [2]上海市骨科内置物重点实验室,200011 [3]上海交通大学材料科学与工程学院,200240

出　　处：《国际骨科学杂志》2016年第1期57-63,共7页International Journal of Orthopaedics

基　　金：国家自然科学基金(81272132)

摘　　要：目的探讨并观察3D打印聚乳酸(PLA)-羟基磷灰石(HA)复合支架材料在体内成骨的可行性。方法将骨髓基质细胞与3D打印PLA-HA复合材料进行体外复合培养,通过扫描电镜观察细胞在支架材料上的生长黏附情况。选择成年新西兰大白兔单侧胫骨内侧建立体内生物反应器模型,分离隐动、静脉血管束,并剥离胫骨内侧骨膜,将游离的动静脉血管束完全穿越支架材料内部并固定于胫骨骨膜囊内,此种方法构建的组织工程骨为实验组;单纯将复合骨髓基质细胞的3D打印PLA-HA复合材料回植于胫骨内侧骨膜囊内,此种方法构建的组织工程骨为对照组。术后6周取材,分别采用聚合酶链式反应(PCR)检测骨分化相关基因,并进行显微CT三维重建及骨形态计量分析、HE染色组织学观察。结果骨髓基质细胞能在3D打印PLA-HA复合材料上黏附生长。定量PCR检测成骨分化基因OPN及COLⅠ的表达,结果显示实验组OPN及COLⅠ相对表达量分别为8.0333±0.5820和7.5452±0.5608;对照组OPN及COLⅠ相对表达量分别为5.7248±0.9975和4.0173±0.2654,差异有统计学意义(P<0.05)。显微CT扫描显示,实验组新生骨组织体积及骨小梁相对数目较对照组高,且差异有统计学意义(P<0.05)。组织学观察可见实验组有新生骨组织形成,部分新生骨组织为编织骨,骨细胞体积大、数量多,呈编织状排列,骨细胞分化成熟;对照组可见部分新生骨样组织形成,骨细胞分化较成熟。结论以骨髓基质细胞为种子细胞,3D打印PLA-HA复合材料为支架,复合隐动、静脉血管束及自体骨膜,在体内能构建出功能相对完善的组织工程骨;3D打印PLA-HA复合材料可作为骨组织工程中支架材料。Objective To explore the feasibility of constructing the tissue engineered bone using three-dimensional （3D） printed polylactie acid-hydroxyapatite （PLA-HA） composite scaffolds. Methods Bone marrow stromal cells （BMSCs） were seeded on 3D printed PLA-HA composite scaffolds, then they were cultured in vitro. The BMSCs of growing morphology and adhesion were observed with the scanning electron microscopy. An animal model of vivo bioreactor on adult New Zealand rabbits inside the unilateral tibial was established. Then saphenous arteriovenous blood bundles were fully free and dissected, and the tibia[ upper periosteum was isolated. The vascular bundle crossed through the central channel of scaffolds and was fixed with a suture under the periosteum capsule. Using the above way to construct bioreactor in vivo was marked experimental group, and only 3D printed pLAHA scaffolds combined with autologous BMSCs transplanted to tibial periosteum without blood vessel was designated as control group. In the postoperative 6 weeks, quantitative statistics of osteogenic related genes in real-time reverse transcription polymerase chain reaction （PCR） method, 3D reconstructions as well as bone morphology measurementand analysis in micro Computed Tomography （micro CT）, histological examination stained with hematoxylin and eosin （HE） were evaluated, respectively. Results BMSCs had fine adhesion capability in 3D printed PLA-HA composite scaffolds. Osteopontin （OPN） and collagen type I （COL I ） were expressed in both groups by PCR measurement. The relative levels of expression of OPN and COL I in the experimental group were 8.0333± 0. 5820 and 7. 5452 ± 0. 5608, and they were 5. 7248 ± 0. 9975 and 4. 0173 ±0. 2654 in the control group. There existed significant differences between two groups （P〈0.05）. The outcomes from micro-CT analysis revealed that the amount of newly formed bone volume and bone trabecula in experimental group and it in control group had significant differences （P〈0.05）.

关 键 词：3D打印 体内生物反应器 骨组织工程 聚乳酸 羟基磷灰石 

分 类 号：R318.08[医药卫生—生物医学工程]