CAJ | 학술논문

目的探索新型脱细胞半月板细胞外基质（dMECM）的制备方法，并对其细胞相容性进行研究。方法无菌条件下收集新鲜的猪半月板组织，切碎，采用湿法粉碎、差速离心的方法制备 dMECM生物材料。通过天狼猩红、甲苯胺蓝染色方法分别比较天然半月板与 dMECM中胶原以及糖胺聚糖含量的区别；采用 Hoechst 33258荧光染色法观察脱细胞后 dMECM中 DNA残留情况。将 P3代的兔内侧半月板细胞种植在铺有 dMECM的盖玻片上，体外培养7 d后行扫描电镜检测其细胞相容性。结果本实验制备的 dMECM天狼猩红、甲苯胺蓝染色均呈阳性，并且与天然半月板染色类似；dMECM行 Hoechst 33258染色呈阴性；扫描电镜结果显示种植于 dMECM表面上的半月板细胞黏附紧密，可见大量细胞 ECM的分泌。结论本实验制备的 dMECM可以很好地保留天然半月板 ECM成分，有效地去除 DNA物质，并且具有良好的生物相容性，是未来半月板组织工程领域非常有前景的支架材料。
목적탐색신형탈세포반월판세포외기질（dMECM）적제비방법，병대기세포상용성진행연구。방법무균조건하수집신선적저반월판조직，절쇄，채용습법분쇄、차속리심적방법제비 dMECM생물재료。통과천랑성홍、갑분알람염색방법분별비교천연반월판여 dMECM중효원이급당알취당함량적구별；채용 Hoechst 33258형광염색법관찰탈세포후 dMECM중 DNA잔류정황。장 P3대적토내측반월판세포충식재포유 dMECM적개파편상，체외배양7 d후행소묘전경검측기세포상용성。결과본실험제비적 dMECM천랑성홍、갑분알람염색균정양성，병차여천연반월판염색유사；dMECM행 Hoechst 33258염색정음성；소묘전경결과현시충식우 dMECM표면상적반월판세포점부긴밀，가견대량세포 ECM적분비。결론본실험제비적 dMECM가이흔호지보류천연반월판 ECM성분，유효지거제 DNA물질，병차구유량호적생물상용성，시미래반월판조직공정영역비상유전경적지가재료。
Objective To explore a novel approach to prepare the decellularized meniscal extracellular matrix (dMECM) and study its biocompatibility. Methods Porcine knee meniscus were collected immediately after slaughter, and cut into slices under aseptic condition. The novel dMECM was prepared by using waterproof pulverization and differential centrifugation approach. Picrosirius red and toluidine blue staining were carried out with the purpose of comparing the collagen and glycosaminoglycans (GAGs) content of native meniscus with dMECM. Hoechst 33258 staining was used to detect the presence of DNA debris in dMECM. The rabbit inner meniscal fibrochondrocytes (P3) were seeded in the dMECM modified coverslips, and then the cells/coated surface constructs were observed by scanning electron microscopy (SEM) to evaluate the biocompatibility after 7 d culture. Results As for the histological assessment, Picrosirius red and toluidine blue staining of dMECM were all positive and similar to native meniscus, but Hoechst 33258 staining was negative. SEM assessment showed that cells grown on the modified growth surface of dMECM adhered tightly and secreted numerous ECM. Conclusion dMECM prepared in this experiment can well preserve meniscal ECM components, effectively remove the cellular DNA, and possess a good biocompatibility. It may be a promising candidate biomaterial for meniscal tissue engineering applications in advance.