无机离子仿生骨膜调控免疫微环境促进骨修复  

Inorganic ion bionic periosteum regulates immune microenvironment to promote bone repair

作　　者：张力宸 陈亮[1] 顾勇[1] Zhang Lichen;Chen Liang;Gu Yong(First Affiliated Hospital of Soochow University,Suzhou 215031,Jiangsu Province,China)

机构地区：[1]苏州大学附属第一医院,江苏省苏州市215031

出　　处：《中国组织工程研究》2023年第3期346-353,共8页Chinese Journal of Tissue Engineering Research

基　　金：基金资助:国家自然科学基金项目(81972078,81772312),项目负责人:陈亮;江苏省杰出青年基金(BK20211504),项目负责人:顾勇。

摘　　要：背景:介孔生物活性玻璃纳米颗粒与甲基丙烯酸明胶合成的仿生骨膜可在骨缺损局部释放钙硅磷离子,促进成骨。其中硅离子还有调控巨噬细胞极化表型的作用,因此仿生骨膜对骨缺损局部炎症微环境的影响仍需探索。目的:通过体、内外实验探讨无机离子仿生骨膜调控炎症微环境与促进骨修复之间的联系。方法:①体外实验:将介孔生物活性玻璃纳米颗粒与甲基丙烯酸明胶共混后光交联得到无机离子仿生骨膜,通过电感耦合等离子体光谱检测仿生骨膜浸泡于人体模拟体液中7 d内的Si^(4+)释放。将骨髓间充质干细胞分别接种于甲基丙烯酸明胶水凝胶与仿生骨膜复合水凝胶中,以单纯培养的细胞为对照,进行活死荧光染色与碱性磷酸酶染色。将骨巨噬细胞分别接种于甲基丙烯酸明胶水凝胶与仿生骨膜复合水凝胶中,以单纯培养的细胞为对照,进行一氧化氮合成酶(巨噬细胞M1表型)和CD206(巨噬细胞M2表型)免疫荧光染色。②体内实验:建立大鼠颅骨缺损模型,将甲基丙烯酸明胶水凝胶和仿生骨膜复合水凝胶分别植入骨缺损处,术后7 d,通过RT-PCR检测骨缺损局部巨噬细胞表型;术后8周,通过骨缺损处苏木精-伊红染色检测仿生骨膜的成骨性能。结果与结论:①体外实验:仿生骨膜24 h内释放Si^(4+)达到最大浓度,在之后6 d内缓慢释放。共培养3 d后的活死荧光染色显示,各组干细胞生长良好,有较好的细胞活性,无明显死亡。共培养7 d后的免疫荧光染色显示,仿生骨膜组巨噬细胞多为M2型,甲基丙烯酸明胶水凝胶组和对照组多为M1型。共培养7 d后的碱性磷酸酶染色显示,仿生骨膜组染色强于甲基丙烯酸明胶水凝胶组和对照组。②体内实验:RT-PCR检测结果显示,仿生骨膜组骨缺损局部巨噬细胞大多呈M2型,空白组和GelMA水凝胶组大多呈M1型。苏木精-伊红染色显示,仿生骨膜组骨缺损处新生骨BACKGROUND:The biomimetic periosteum synthesized by mesoporous bioactive glass nanoparticles and methacrylic acid gelatin can release calcium,silicon and phosphorus ions in the local area of bone defect and promote osteogenesis.However,silicon ions can also regulate the polarization of macrophages,so the effect of bionic periosteum on local inflammatory microenvironment of bone defect still needs to be explored.OBJECTIVE:To explore the relationship between the regulation of inflammatory microenvironment and the promotion of bone repair by inorganic ion biomimetic periosteum in vivo and in vitro.METHODS:(1)In vitro:The bionic periosteum was obtained by photocrosslinking mesoporous bioactive glass nanoparticles and methacrylic acid gelatin.Inductively coupled plasma spectroscopy was used to observe the release of Si^(4+) from biomimetic periosteum within 7 days.Bone marrow mesenchymal stem cells were inoculated into methacrylic acid gelatin hydrogel and mesoporous bioactive glass nanoparticles/methacrylic acid gelatin composite hydrogel separately.With the simple cultured cells as control,the biocompatibility of the cells was detected by Live/Dead fluorescence staining.Bone macrophages were inoculated in methacrylic acid gelatin hydrogel and mesoporous bioactive glass nanoparticles/methacrylic acid gelatin composite hydrogel separately.With simple cultured cells as control,the effect of bionic periosteum on macrophage phenotype in vitro was detected by nitric oxide synthase(macrophage M1 phenotype)and CD206(macrophage M2 phenotype)immunofluorescence staining.(2)In vivo:The rat model of skull defect was established,and methacrylic acid gelatin hydrogel and mesoporous bioactive glass nanoparticles/methacrylic acid gelatin hydrogel were implanted into the bone defect.The local macrophage phenotype of bone defect was detected by RT-PCR at 7 days after operation,and the osteogenic property of biomimetic periosteum was detected by hematoxylin-eosin staining at 8 weeks after operation.RESULTS AND CONCLUSION:(1)In vitro:Th

关 键 词：无机离子 仿生骨膜 炎症 巨噬细胞 成骨 免疫微环境 

分 类 号：R459.9[医药卫生—治疗学] R318.08[医药卫生—临床医学] R683.4