RGD接枝壳聚糖纳米短纤维增强型磷酸钙骨水泥的生物学性能研究

目的: 分析壳聚糖纳米短纤维（CSNF）和RGD对磷酸钙骨水泥（CPC）生物力学和生物相容性的影响。方法: 采用静电纺丝法制备壳聚糖纳米纤维膜，通过高速剪切形成纳米短纤维，并对CSNF进行RGD基团接枝修饰。采用Biocement D法制备钙磷摩尔比为1.5：1的CPC。通过红外光谱、X射线衍射、扫描电镜对CPC、CSNF、RGD接枝CSNF（CSNF-RGD）、CSNF增强型骨水泥（CPC-CSNF）、RGD接枝CPC-CSNF（CPC-CSNF-RGD）进行成分分析和结构观察，利用万能力学试验机检测其生物力学特性，采用免疫荧光染色和MTT法检测成骨细胞（MC3T3）在上述材料上的黏附和增殖情况。结果: 扫描电镜观察发现，CSNF和CSNF-RGD呈现出分散均匀的多孔结构；红外图谱中CSNF在波长为1637和1579 nm处的吸收峰位移至波长1633和1585 nm处，说明RGD成功接枝到CSNF上；X射线衍射图谱显示CPC具有一定的可固化性；应力应变曲线统计分析结果显示，CPC-CSNF和CPC-CSNF-RGD断裂强度分别为（17.74±0.54）MPa和（16.67±0.56）MPa，均高于CPC（均P < 0.05）；实验材料与成骨细胞复合培养240 min后，CPC-CSNF-RGD上细胞数量均明显多于CPC和CPC-CSNF（均P < 0.05）。结论: CSNF和RGD的加入改善了CPC的生物力学性能和生物相容性。
Abstract: Objective: To analysis the biomechanical and biocompatible properties of calcium phosphate cement (CPC) enhanced by chitosan short nanofibers(CSNF) and Arg-Gly-Asp (RGD). Methods: Chitosan nanofibers were prepared by electrospinning, and cut into short fibers by high speed dispersion. CPC with calcium phosphorus ratio of 1.5:1 was prepared by Biocement D method. The composition and structure of CPC, CSNF, RGD modified CSNF (CSNF-RGD), CSNF enhanced CPC (CPC-CSNF), RGD modified CPC-CSNF (CPC-CSNF-RGD) were observed by infrared spectrum, X-ray diffraction (XRD) and scan electron microscopy (SEM). The mechanical properties were measured by universal mechanical testing instrument. The adhesion and proliferation of MC3T3 cells were assessed using immunofluorescence staining and MTT method. Results: The distribution of CSNF in the scaffold was homogeneous, and the porous structure between the nanofibers was observed by SEM. The infrared spectrum showed the characteristic peaks at 1633 nm and 1585 nm, indicating that RGD was successfully grafted on chitosan nanofibers. The XRD pattern showed that the bone cement had a certain curability. The stain-stress test showed that break strengths were (17.74±0.54) MPa for CPC-CSNF and (16.67±0.56) MPa for CPCP-CSNF-RGD, both were higher than that of CPC(all P < 0.05). The immunofluorescence staining and MTT results: indicated that MC3T3 cells grew better on CPC-CSNF-RGD after 240 min of culture(all P < 0.05). Conclusion: CSNF-RGD can improve the biomechanical property and biocompatibility of CPC, indicating its potential application in bone tissue repair. Key words: Glycine Arginine Aspartic acid Peptides Chitosan Nanocomposites Calcium phosphates Biomechanics Biocompatible materials