聚磷酸盐改性胶原促成骨性能研究  

Bone regeneration ability of polyphosphate crosslinked collagen scaffold

作　　者：顾俊婷 焦凯 马雨轩 王凯燕 陈吉华 牛丽娜 GU Junting;JIAO Kai;MA Yuxuan;WANG Kaiyan;CHEN Jihua;NIU Lina(State Key Laboratory of Military Stomatology&National Clinical Research Center for Oral Diseases&Shaanxi Key Laboratory of Stomatology,Department of Prosthodontics,Stomatological Hospital,Air Force Medical University,Xi'an 710032,China)

机构地区：[1]军事口腔医学国家重点实验室,口腔疾病国家临床医学研究中心,陕西省口腔医学重点实验室,空军军医大学口腔医院修复科,陕西西安710032

出　　处：《空军军医大学学报》2022年第3期274-279,共6页Journal of Air Force Medical University

基　　金：国家自然科学基金(81870805,81870787,81720108011);陕西省重点科技创新团队(2020TD-03);陕西高校青年创新团队(2019-2022年)。

摘　　要：目的通过化学交联法构建聚磷酸盐改性胶原(P-CS),探究其生物安全性与促成骨性能。方法在前期研究基础上,通过化学交联体系将聚磷酸盐交联到胶原表面,以此构建出P-CS。利用甲基绿染色对P-CS上的聚磷酸盐进行表征,并进一步利用扫描电镜观察P-CS表面形貌。利用CCK-8、活/死染色和急性全身毒性实验评价P-CS的生物相容性。随后在体外进行成骨性能检测。将大鼠骨髓间充质干细胞(BMSCs)与P-CS共培养并成骨诱导14 d后,观察P-CS对BMSCs成骨分化的影响。通过PCR检测骨形态形成蛋白(BMP-2)、骨桥蛋白(OPN)和Ⅰ型胶原mRNA表达水平;通过免疫荧光染色检测BMP-2、OPN表达水平;通过碱性磷酸酶染色及定量分析检测BMSCs成骨分化状态;通过茜素红染色及定量分析表征细胞体外矿化能力。最后在大鼠牙槽骨缺损模型中观察P-CS植入7 d后的骨修复效果。结果P-CS保持了胶原的特征性结构且生物安全性良好。体外促成骨研究显示,经14 d成骨诱导后,P-CS可促进BMSCs成骨分化;大鼠牙槽骨缺损修复实验中,P-CS植入7 d后,Goldner染色显示新骨形成显著增多。结论P-CS具有良好的生物安全性以及促成骨性能。Objective To investigate the biosafety and osteo-inductive property of polyphosphate crosslinked collagen scaffold(P-CS).Methods Based on our preliminary studies,the polyphosphate was covalently crosslinked on collagen.The modified collagen was called P-CS.The modification was further verified by methyl green staining,and the surface morphology of P-CS was characterized by scanning electron microscopy.Besides,the biocompatibility of P-CS was determined by CCK-8 assay,live/dead staining,and acute systemic toxicity testing.Additionally,the rat bone marrow stromal cells(BMSCs)were used to investigate the osteo-inductive property of P-CS in vitro.The cells were co-cultured with P-CS,and then subjected to osteogenic induction for 14 d to observe the effect of P-CS on the osteogenic differentiation of BMSCs.PCR was used to detect the mRNA expression levels of bone morphogenetic protein-2(BMP-2),osteopontin(OPN),and collagen type-Ⅰ.The expression changes of BMP-2 and OPN were further demonstrated by immunofluorescence staining.The osteogenic differentiation of BMSCs was studied by alkaline phosphatase staining and quantitative analysis.Alizarin red staining combined with quantitative analysis was used to evaluate the mineralization ability of BMSCs in vitro.After 7 d of P-CS implantation,the alveolar bone defect model of rats was used to investigate the bone regeneration ability of P-CS in vivo.Results P-CS maintained the characteristic structure of collagen and showed good biosafety.After 14 d of osteogenic induction,P-CS enhanced osteogenic differentiation of BMSCs in vitro.After bone regeneration for 7 d,Goldner staining showed that P-CS induced more new bone formation in the alveolar bone defect area of rat models.Conclusion P-CS has favorable biosafety and bone regeneration ability.

关 键 词：聚磷酸盐 胶原支架 生物安全性 成骨分化 骨缺损修复 

分 类 号：R683[医药卫生—骨科学]