不同烧结温度对双相钙磷陶瓷颗粒材料介孔结构及其成骨性能的影响  

Influence of different sintering temperatures on mesoporous structure and ectopic osteogenesis of biphasic calcium phosphate ceramic granule materials

作　　者：张栋 宗宪磊 郭小双 杜宏 宋国栋 靳小雷 ZHANG Dong;ZONG Xianlei;GUO Xiaoshuang;DU Hong;SONG Guodong;JIN Xiaolei(The 16th Department of Plastic Surgery,Plastic Surgery Hospital,Peking Union Medical College,Chinese Academy of Medical Sciences,Beijing,100144,P.R.China)

机构地区：[1]中国医学科学院北京协和医学院整形外科医院整形十六科,北京100144

出　　处：《中国修复重建外科杂志》2021年第1期95-103,共9页Chinese Journal of Reparative and Reconstructive Surgery

基　　金：国家自然科学基金青年科学基金项目(81400565);中国医学科学院北京协和医学院院所基金项目(B2018007)。

摘　　要：目的通过体内外实验检测不同烧结温度下制备的不同介孔直径双相钙磷陶瓷(biphasic calcium phosphate,BCP)颗粒材料成骨能力差异,为筛选具备更好临床应用参数的BCP材料提供依据。方法将羟基磷灰石(hydroxyapatite,HA)及β-磷酸三钙(β-tricalcium phosphate,β-TCP)以8∶2比例混合后,分别在1 050、1 150及1 250℃下烧制3 h制备3种BCP材料(分别设为材料1、2、3),比表面积测试法(Brunauer-Emmett-Teller test,BET)测量材料的颗粒内部孔隙率及介孔直径、体积、面积,X线衍射(X-ray diffraction,XRD)评估材料组成成份,扫描电镜观察材料微观表面形态。体外将第3代SD大鼠BMSCs与各材料共培养7 d(分别设为A、B、C组),扫描电镜观察细胞黏附情况,鬼笔环肽染色观察BMSCs贴附于材料表面后的形态,细胞计数试剂盒8法检测细胞增殖活性。体内建立比格犬异位成骨模型:取9只比格犬,于每只犬双侧竖脊肌内制作9个肌袋,将肌袋随机分为3组(每组3个/只),A、B、C组分别置入材料1、2、3。术后1、2、3个月分别麻醉3只比格犬取材行HE、Masson及番红固绿染色,计算BCP间隙中的成骨面积比;行实时荧光定量PCR(real-time fluorescence quantitative PCR,qRT-PCR)检测成骨相关基因ALP、骨桥蛋白(osteopontin,OPN)、骨钙素(osteocalcin,OC)的表达。结果 BET检测示随烧结温度增加,颗粒内部孔隙率无明显变化,但介孔直径、体积及面积逐渐减小;XRD检测示3种材料均可见HA及β-TCP两种X线衍射波;扫描电镜观察示3种材料表面有广泛分布的微孔,孔间有空隙相连。体外实验示BMSCs在3种材料表面黏附、增殖,B、C组材料的细胞生物相容性优于A组。体内实验结果示,术后2个月开始3种材料颗粒孔隙内即可见明显的骨样组织沉积。各组成骨面积比随时间延长均增加,术后2、3个月A组成骨面积比显著高于B、C组,1个月时显著高于B组(P<0.05)。qRT-PCR检测示,A组成骨相关基因表Objective To detect the difference in the osteogenesis ability of biphasic calcium phosphate(BCP)ceramic granular materials with different mesoporous diameters prepared at different sintering temperatures through in vivo and in vitro experiments,so as to provide evidence for screening BCP materials with better clinical application parameters.Methods Three kinds of BCP(materials 1,2,3)were prepared by mixing hydroxyapatite(HA)andβ-tricalcium phosphate(β-TCP)at a ratio of 8∶2 and sintered at 1050,1150,and 1250℃for 3 hours,respectively.The internal porosity and the diameter,volume,and area of the mesopore were measured by Brunauer-Emmett-Teller test(BET);the composition of the material was evaluated by X-ray diffraction(XRD);the microscopic surface morphology of the material was observed by scanning electron microscopy(SEM).The 3rd generation bone marrow mesenchymal stem cells(BMSCs)from Sprague-Dawley rats were co-cultured with the materials 1,2,and 3 for 7 days in vitro respectively(groups A,B,and C),and the cells adhesion on the materials was observed by SEM and phalloidine staining,respectively.Cell proliferation activity was measured by cell counting kit 8 method.In vivo,9 muscle bags were made in dorsal muscles of 9 beagles,respectively.The muscle bags were randomly divided into 3 groups(3 per beagle in each group)and materials 1,2,and 3 were placed into the muscle bags of groups A,B,and C,respectively.After 1,2,and 3 months of operation,3 beagles were anesthetized and the samples were stained with HE,Masson,and Safranin,and the bone formation area ratio in the BCP gap was calculated.Real-time fluorescence quantitative PCR(qRT-PCR)was performed to detect the expressions of bone-related genes[including alkaline phosphatase(ALP),osteopontin(OPN),and osteocalcin(OC)].Results The BET test showed that with the increase of sintering temperature,the internal porosity of the particles did not change significantly,but the diameter,volume,and area of the mesopores gradually decreased.The XRD detection showed that

关 键 词：双相钙磷陶瓷 异位成骨 介孔直径 骨传导性 BMSCS 

分 类 号：R318.08[医药卫生—生物医学工程]