透明软骨材料经脱细胞处理后细胞外基质成分变化的比较

Alteration of extracellular matrix in decellularized articular cartilage treated by different technologies

目的比较Triton X-100和十二烷基硫酸钠(SDS)试剂对人关节软骨脱细胞处理后细胞外基质成分的变化,指导筛选适宜的制备。方法首先制备人关节软骨微粒,再采用两种不同试剂分别作用于软骨细胞微粒,清洗和消化步骤相同,脱细胞完成后分别进行DNA、总氨基酸及总胶原和葡萄糖胺聚糖(GAG)含量检测,分为Triton X-100组和SDS组,并与对照组(正常软骨组织)进行基质成分的比较分析。结果 Triton X-100组、SDS组和对照组经脱细胞处理后其GAG含量分别为(84.71±3.72)、(122.63±6.05)、(232.78±75.33)μg/mg,羟脯氨酸含量分别为(24.92±1.74)、(24.23±1.68)、(24.54±2.99)μg/mg,DNA含量分别为(0.242±0.065)、(0.225±0.057)、(5.679±1.873)μg/mg。结论Triton X-100和SDS两种试剂均能将软骨微粒中软骨细胞成分有效去除,胶原成分保留量在两组中无明显差异;但是应用SDS行脱细胞处理后能保留更多的GAG成分,更有利于维持软骨细胞功能,是较好的材料制备技术。

Objective To compare the changes of extracellular matrix in decellularized articular carti- lage treated by Triton X-100 and sodium dodecyl sulfate（SDS） to optimize the suitable preparation. Methods First, cartilage panicles were prepared, then the two reagents was used to treat the ceils of cartilage particles respectively, with the same experimental procedures of rinsing and digestion. After decellularization, the contents of DNA, amino acid, collagen and glycosaminoglycan （GAG） were measured, and the results were compared with those of the normal cartilaginous tissue. Results After decellularization, the GAG contents were 84.71 ±3.72 and 122.63 ± 6.05 μg/mg respectively in the extracellular matrix of articular cartilages treated by Triton X-100 and SDS, while that was 232.78 ＋ 75.33 μg/mg in the extraeellular matrix of normal cartilaginous cells. Hydroxyproline content was 24.92 ± 1.74 and 24.23 ＋ 1.68 μg/mg respectively after the decellularization, and that in the normal matrix was 24.54 ＋ 2.99 μg/mg. DNA content was 0. 242 ＋ 0.065 and 0. 225 ± 0. 057 μg/mg respectively, and that in the normal one was 5. 679 ± 1. 873 μg/mg. Conclusion Cell components are removed effectively from cartilage particles by both Triton X-100 and SDS without obvious difference of collagen remains. While SDS reserves more GAG components and is more helpful to maintain the cartilage cells＇ function. So SDS decellularization is a better preparative techniaue for tissue engineering.