CAJ | 학술논문

目的评价三相蚕丝组织工程韧带的理化性质及生物相容性。方法编织蚕丝网状支架，提取丝素蛋白，利用硫酸软骨素、透明质酸钠、羟基磷灰石( HA )及丝素蛋白修饰三相支架，测定支架材料形貌及表面元素。支架材料接种骨髓间充质干细胞( bone marrow mesenchymal stem cell，BMSCs )后利用扫描电镜( SEM )、激光共聚焦显微镜，观察细胞的黏附及增殖情况。结果蚕丝去除丝胶后，表面变光滑，脱丝胶前蚕丝纤维的直径为(9.89±1.98)μm，脱丝胶后大小为(9.14±1.75)μm，表面经脱丝胶处理后蚕丝纤维直径没有显著改变(P＝0.221)。表面元素分析表明 B 区中的 S 和 Na 元素及 C 区中的 Ca 元素比 A 区有显著提高，并且材料的孔径在60～80μm。A、B、C 区的细胞在培养后4 h 与培养后2 h 相比，细胞黏附率分别为：A 区(83.2±3.4)%，B 区(81.8±2.8)%，C 区(71.1±5.4)%，差异有统计学意义(P＝0.0001)，表明细胞培养后4 h 大部分细胞在材料上有较好的黏附。扫描电镜观察，接种后1天，BMSCs 在材料上黏附良好，接种后7天细胞爬满材料表面并可见细胞伪足深入材料内部生长。活/死细胞染色利用激光共聚焦显微镜观察培养的7天比1天的细胞增殖明显，且未见明显死亡，细胞在材料的多个层次可见。结论构建的三相蚕丝组织工程韧带具有良好的生物相容性，利于 BMSCs 的黏附增殖。
목적평개삼상잠사조직공정인대적이화성질급생물상용성。방법편직잠사망상지가，제취사소단백，이용류산연골소、투명질산납、간기린회석( HA )급사소단백수식삼상지가，측정지가재료형모급표면원소。지가재료접충골수간충질간세포( bone marrow mesenchymal stem cell，BMSCs )후이용소묘전경( SEM )、격광공취초현미경，관찰세포적점부급증식정황。결과잠사거제사효후，표면변광활，탈사효전잠사섬유적직경위(9.89±1.98)μm，탈사효후대소위(9.14±1.75)μm，표면경탈사효처리후잠사섬유직경몰유현저개변(P＝0.221)。표면원소분석표명 B 구중적 S 화 Na 원소급 C 구중적 Ca 원소비 A 구유현저제고，병차재료적공경재60～80μm。A、B、C 구적세포재배양후4 h 여배양후2 h 상비，세포점부솔분별위：A 구(83.2±3.4)%，B 구(81.8±2.8)%，C 구(71.1±5.4)%，차이유통계학의의(P＝0.0001)，표명세포배양후4 h 대부분세포재재료상유교호적점부。소묘전경관찰，접충후1천，BMSCs 재재료상점부량호，접충후7천세포파만재료표면병가견세포위족심입재료내부생장。활/사세포염색이용격광공취초현미경관찰배양적7천비1천적세포증식명현，차미견명현사망，세포재재료적다개층차가견。결론구건적삼상잠사조직공정인대구유량호적생물상용성，리우 BMSCs 적점부증식。
Objective To evaluate physicochemical properties and biocompatibility of the three-phase silk tissue-engineered ligament.Methods The silk mesh was woven, and silk ifbroin was extracted. The three-phase scaffold was modified with chondroitin sulfate, hyaluronic acid sodium, hydroxyapatite ( HA ) and silk fibroin. The surface morphology and element of the scaffold were determined. After bone marrow mesenchymal stem cells ( BMSCs ) seeding, the cell’s proliferation and adhesion were observed by scanning electron microscope ( SEM ) and laser confocal microscopy.Results After the removal of sericin, silk surface became smooth. The silk ifber diameter was ( 9.89±1.98 ) μm. After the removal of sericin, the diameter was ( 9.14±1.75 ) μm. No signiifcant differences existed in the diameter of silk ifber (P=0.221 ). A surface element analysis showed the Ca element in area C, S and Na elements in area B were all higher than those in area A. The pore size of the material was 60-80 μm. The cell adhesion rates of area A, B and C 4hrs later were ( 83.2±3.4 ) %, ( 81.8±2.8 ) % and ( 71.1±5.4 ) %, which were signiifcantly different from those 2hrs later (P=0.0001 ). It indicated that most of the cells were adherent to the material after 4hrs’ cells culture. One day after the seeding, the BMSCs in the material adhered well by SEM. Seven days later, the cells covered the material surface and the pseudopodia grown into internal were visible. Using life / dead cell staining, the cells cultured 7d were of higher proliferation than 1d by laser confocal microscopy. The cells were visible in multiple levels of the material. Conclusions The constructed three-phase silk tissue-engineered ligament has good biocompatibility, which is conducive to the proliferation of BMSCs. It lays the foundation for the vivo experiment in the next.