Yang Xiong-Gang, Zhong Hui, Zhang Gui-Qian, Zhou Jing-Cheng, Hu Yong-Cheng, Lu Sheng
Department of Orthopedic Surgery, The First People's Hospital of Yunnan Province, The Affiliated Hospital of Kunming University of Science and Technology, No. 157 Jinbi Road, Xishan District, Kunming City, 650034, Yunnan Province, China.
The Key Laboratory of Digital Orthopedics of Yunnan Province, No. 157 Jinbi Road, Xishan District, Kunming City, 650034, Yunnan Province, China.
Cell Tissue Bank. 2025 Jun 4;26(3):28. doi: 10.1007/s10561-025-10177-0.
To compare the differences in biological and biomechanical characteristics of cadaver tendon treated by deep freezing, 95wt.% ethanol, tributyl phosphate (TBP), and sodium dodecyl sulfate (SDS) respectively. A total of 50 fresh tendon specimens were randomly divided into five groups, and four groups were treated with deep freezing (- 80 °C), 95wt.% ethanol, 1% TBP, or 1% SDS, respectively. Hematoxylin-eosin (HE), toluidine blue (TB) and 4',6-diamidino-2-phenylindole (DAPI) staining, and transmission/scanning electron microscope observations were performed. Then, cyclic creep and tensile tests were conducted to investigate the biomechanical properties. The content of residual DNA was tested. The HE, TB and DAPI staining showed a significant decrease in tendon cells following treatments, compared with fresh specimens. SDS, TBP, alcohol can almost completely decellularize the tendon, and deep-frozen group remained a few number of tendon cells. The residual DNA content was significantly lowered, with an average percentage of 50.97%, 79.16%, 88.91% and 72.56%, for groups of deep freezing, alcohol, TBP and SDS, respectively. The arrangement of collagen fibers was significantly disrupted, and the gap between fibers was widened, following treatments by alcohol, TBP and SDS. However, the biomechanical properties were generally similar among the five groups, with significantly lowered cyclic creep for ethanol group and lowered Young's modulus for SDS group exclusively. The four treatments can effectively reduce the number of residual cells and DNA content. Among them, cryogenic treatment has almost no damage to tendon histology and biomechanics, while ethanol, SDS and TBP decellularization methods cause different degrees of damage.
为比较经深度冷冻、95wt.%乙醇、磷酸三丁酯（TBP）和十二烷基硫酸钠（SDS）处理的尸体肌腱的生物学和生物力学特性差异。共50个新鲜肌腱标本随机分为五组，其中四组分别用深度冷冻（-80°C）、95wt.%乙醇、1%TBP或1%SDS处理。进行苏木精-伊红（HE）、甲苯胺蓝（TB）和4',6-二脒基-2-苯基吲哚（DAPI）染色以及透射/扫描电子显微镜观察。然后，进行循环蠕变和拉伸试验以研究生物力学性能。检测残余DNA含量。与新鲜标本相比，HE、TB和DAPI染色显示处理后肌腱细胞显著减少。SDS、TBP、酒精可使肌腱几乎完全脱细胞，而深度冷冻组仍残留少量肌腱细胞。残余DNA含量显著降低，深度冷冻、酒精、TBP和SDS组的平均百分比分别为50.97%、79.16%、88.91%和72.56%。酒精、TBP和SDS处理后，胶原纤维排列明显紊乱，纤维间间隙增宽。然而，五组之间的生物力学性能总体相似，仅乙醇组的循环蠕变显著降低，SDS组的杨氏模量降低。这四种处理方法均可有效减少残余细胞数量和DNA含量。其中，低温处理对肌腱组织学和生物力学几乎无损伤，而乙醇、SDS和TBP脱细胞方法会造成不同程度的损伤。
