去细胞的肿瘤细胞外基质制备及结构研究

摘要

Objective:To investigate the effect of decellular treatment on framework constituents of extracellular matrices (ECMs)and biomechanical properties of tumor tissues,and provide evidence for in vitro tumor engineering. Methods:Human A549 pulmonary adenocarcinoma cells were implanted into severe combined immunodeficient (SCID)mice to establish xenotransplatation models.Tumors were retrieved as mass and sliced into plates.Then they were decellularized by four different procedures.Histological techniques were used to study ECM framework constitu-ents of treated examples,fresh tissues were used as controls.DNA,collagen and glycosaminoglycan (GAG)contents were assayed respectively to the fresh and the four decellular groups (n=10).The porosity and mechanical strengths (n=10)were measured to Tris-Trypsin-Triton and SDS group due to complete cellular removal.Results:Histo-chemical and DNA content analysis proved complete removal of nuclear and other cell components for Tris-Trypsin-Triton and SDS group.Collagen content increased and GAG content decreased for the two acellular groups com-pared with the native group (P<0.05).Tris-Trypsin-Triton and SDS group had increased porosity but decreased elastic modulus compared with the native group (P<0.05).SDS group had larger porosity than the Tris-Trypsin-Triton group (P<0.05).Conclusion:Tris-Trypsin-Triton and SDS group got complete cellular removal,and Tris-Trypsin-Triton group had larger porosity.The two acellular tumor ECMs had decreased elastic modulus and should be further cross-linked before being used as a 3D scaffold for tumor engineering.%目的：探讨去细胞处理对肿瘤细胞外基质骨架成分、结构及组织稳定性的影响，为构建体外三维肿瘤提供依据。方法：以人肺腺癌A549肿瘤细胞种植免疫缺陷鼠而得到的肿瘤组织块为材料，切成薄片后接受不同方法的去细胞处理，组织化学染色了解细胞去除程度。新鲜和去细胞处理的肿瘤组织片（n＝10）分别进行DNA，胶原和氨基聚糖含量检测；对细胞去除彻底的Tris-Trypsin-Triton 和SDS组孔隙率和机械性能进行检测，新鲜肿瘤组织作为对照。结果：Tris-Trypsin-Triton 和SDS 组经去细胞处理后细胞成分完全去除，组织间隙增宽。与新鲜组织比较，各组DNA含量明显降低，尤其Tris-Trypsin-Triton和SDS处理组DNA含量较新鲜组明显减少。经去细胞处理后，Tris-Trypsin-Triton 和SDS处理组肿瘤基质的孔隙率均大于新鲜组（P＜0．05），SDS组孔隙率大于Tris-Trypsin-Triton组（P＜0．05）。经过去细胞处理，Tris-Trypsin-Triton 和SDS处理组弹性模量均有所下降（P＜0．05）。结论：Tris-Trypsin-Triton 和SDS处理较彻底去除肿瘤组织细胞成分，SDS处理组有更高的孔隙率。去细胞处理后肿瘤组织弹性模量有所下降，所得到的肿瘤细胞外基质基本成分需经进一步交联处理可以作为一种肿瘤工程支架材料。