CAJ | 학술논문

目的研究用于制备新型生物活性接骨螺钉的纳米羟基磷灰石/聚酰胺66/玻璃纤维(nano-hydroxyapatite/polyamide 66/glass fiber,n-HA/PA66/GF)对成骨细胞生物学行为的影响,为生物螺钉后期临床应用提供实验依据. 方法制备圆片状n-HA/PA66/GF及其浸提液;取小鼠成骨细胞与n-HA/PA66/GF浸提液共培养,采用直接接触实验观察细胞生长变化情况;MTT实验检测细胞增殖情况;流式细胞仪检测细胞凋亡率;ELISA法测定细胞骨钙素含量;Transwell小室实验观察细胞迁移能力;扫描电镜观察细胞黏附和生长情况;细胞免疫荧光检查细胞骨架及肌动蛋白纤维表达. 结果直接接触实验表明,n-HA/PA66/GF对成骨细胞无明显细胞毒性.MTT检测n-HA/PA66/GF浸提液与成骨细胞共培养2,4,6d后,吸光度(A)值分别为0.96-0.14,1.54 ±0.15,2.39 ±0.27.结果显示,成骨细胞数量随着培养时间延长而增多(P＜0.05).流式细胞检测结果证实,n-HA/PA66/GF能使更多成骨细胞进入S期而对细胞凋亡率无明显影响,成骨细胞骨钙素分泌随着共培养时间延长而增多.Transwell实验显示,含浸提液和FBS的α-MEM培养基小室穿过半透膜的细胞数分别为(8.73±3.26)个/视野、(9.47±3.29)个/视野,差异无统计学意义(P＞0.05).扫描电镜观察n-HA/PA66/GF表面的成骨细胞形态规整,紧紧黏附在材料表面,逐渐呈现复层生长.免疫荧光结果表明,材料浸提液对细胞骨架及肌动蛋白纤维的分布无明显影响. 结论新型生物螺钉n-HA/PA66/GF具有良好的成骨细胞相容性,对细胞生长、增殖、分泌、黏附、周期和骨钙素分泌等均有一定调节作用.
목적 연구용우제비신형생물활성접골라정적납미간기린회석/취선알66/파리섬유(nano-hydroxyapatite/polyamide 66/glass fiber,n-HA/PA66/GF)대성골세포생물학행위적영향,위생물라정후기림상응용제공실험의거. 방법 제비원편상n-HA/PA66/GF급기침제액;취소서성골세포여n-HA/PA66/GF침제액공배양,채용직접접촉실험관찰세포생장변화정황;MTT실험검측세포증식정황;류식세포의검측세포조망솔;ELISA법측정세포골개소함량;Transwell소실실험관찰세포천이능력;소묘전경관찰세포점부화생장정황;세포면역형광검사세포골가급기동단백섬유표체. 결과 직접접촉실험표명,n-HA/PA66/GF대성골세포무명현세포독성.MTT검측n-HA/PA66/GF침제액여성골세포공배양2,4,6d후,흡광도(A)치분별위0.96-0.14,1.54 ±0.15,2.39 ±0.27.결과현시,성골세포수량수착배양시간연장이증다(P＜0.05).류식세포검측결과증실,n-HA/PA66/GF능사경다성골세포진입S기이대세포조망솔무명현영향,성골세포골개소분비수착공배양시간연장이증다.Transwell실험현시,함침제액화FBS적α-MEM배양기소실천과반투막적세포수분별위(8.73±3.26)개/시야、(9.47±3.29)개/시야,차이무통계학의의(P＞0.05).소묘전경관찰n-HA/PA66/GF표면적성골세포형태규정,긴긴점부재재료표면,축점정현복층생장.면역형광결과표명,재료침제액대세포골가급기동단백섬유적분포무명현영향. 결론 신형생물라정n-HA/PA66/GF구유량호적성골세포상용성,대세포생장、증식、분비、점부、주기화골개소분비등균유일정조절작용.
Objective To detect the effect of a new bioactive bone screws composed of nanohydroxyapatite/polyamid 66/glass fiber (n-HA/PA66/GF) biomaterial on biological behavior of osteoblasts with a view to a later clinical application of the screws.Methods Mice osteoblasts were co-cultured with n-HA/PA66/GF biomaterial or its extract.Cell growth was detected under a microscope,cell proliferation using MTT assay,apoptosis by flow cytometry,osteocalcin by ELISA method,cell migration using Transwell invasion assay,cell adhesion and growth by scanning electron microscope,and cytoskeleton and actin filament distribution using immunofluorescene.Results Direct contact test demonstrated n-HA/PA66/GF material had no obvious cytotoxicity to osteoblasts.Extract of n-HA/PA66/GF material stimulated osteoblast proliferative activity over time with absorbance value of 0.96 ± 0.14,1.54 ± 0.15,and 2.39 ±0.27 respectively after 2,4,and 6 hours of coculture (P ＜ 0.05).The n-HA/PA66/GF material induced more osteoblasts to entering S period without obvious influence on apoptosis rate and promoted osteocalcin secretion.Migrated cells in medium supplied with n-HA/PA66/GF material or FBS was 8.73 ± 3.26 and 9.47 ± 3.29 in each visual field,but there was no significant difference (P ＞ 0.05).Osteoblasts had a regular form on the surface of n-HA/PA66/GF material,closely adhered to the surface,and reproduced and aggregated with each other to form stratified cell layers.However,nHA/PA66/GF material exhibited no obvious influence on distribution of cytoskeleton and actin filament.Conclusion The new n-HA/PA66/GF screws has excellent cytocompatibility with positive regulatory effect on cell growth,proliferation,secretion,adhesion,cycle,and osteocalcin secretion.