论文部分内容阅读
目的细胞外基质是脂肪组织工程材料的研究热点之一。通过探讨骨骼肌无细胞基质的制作方法及生物相容性,为其在脂肪组织工程中的应用奠定基础。方法取健康成年小香猪新鲜骨骼肌组织,横切成厚2～3 mm的组织块,采用低渗-去垢剂法脱细胞处理。处理后采用HE染色、Masson三色染色、免疫组织化学染色及扫描电镜检测骨骼肌无细胞基质是否有细胞成分残留,并观察其基本结构;应用MTT法检测骨骼肌无细胞基质细胞毒性。取乳腺癌患者自愿捐赠脂肪组织,分离培养人脂肪干细胞(human adipose-derived stem cells,hADSCs),从形态学、流式细胞学和成脂、成骨分化能力方面进行鉴定。将骨骼肌无细胞基质与第3代hADSCs共培养,于培养后第1、3、5、7天通过细胞活性检测材料上细胞黏附、扩散和增殖情况,了解其与细胞之间的相互作用。结果 HE、Masson、免疫组织化学染色及扫描电镜观察显示骨骼肌无细胞基质肌纤维去除完全,无细胞核残留,基质结构保留完整;大量连接成网状的胶原纤维呈多孔隙样结构,规则排列。MTT检测示骨骼肌无细胞基质细胞毒性为1级,细胞相容性好。细胞活性检测示hADSCs在骨骼肌无细胞基质上能很好地伸展,且能与周围基质黏附,进入基质内部并相互交织。结论经脱细胞处理的骨骼肌无细胞基质具有良好生物相容性,可能作为脂肪组织工程的支架材料。
Objective Extracellular matrix is one of the hot topics in the research of adipose tissue engineering materials. By discussing the production method and biocompatibility of acellular matrix of skeletal muscle, it laid the foundation for its application in adipose tissue engineering. Methods Fresh skeletal muscle tissue from healthy adult piglets was cut transversely into 2 ~ 3 mm thick tissue blocks and decellularized by hypotonic-detergent method. After treatment, HE staining, Masson trichrome staining, immunohistochemical staining and scanning electron microscopy were used to detect whether any cell components remained in the acellular matrix of skeletal muscle, and the basic structure was observed. MTT assay was used to detect the cytotoxicity of acellular matrix stromal cells. Adipose tissue was donated voluntarily to breast cancer patients. Human adipose-derived stem cells (hADSCs) were isolated and cultured, and their morphological, flow cytological and adipogenic, osteogenic and differentiation abilities were identified. The skeletal muscle cell-free matrix was co-cultured with the third generation hADSCs. Cell adhesion, proliferation and proliferation were detected on the 1st, 3rd, 5th and 7th day after culturing, and the interaction between them was also observed. Results HE, Masson, immunohistochemical staining and scanning electron microscopy showed that the acellular acellular matrix muscle fibers of skeletal muscle were completely removed without residual of cells and the matrix structure remained intact. A large number of collagen fibers connected to the network were porous and arranged regularly. MTT test showed skeletal cell-free matrix cytotoxicity grade 1, good cell compatibility. The cell viability assay showed that hADSCs could stretch well on the acellular matrix of skeletal muscle and could adhere to the surrounding matrix and enter the matrix and interweave with each other. Conclusion The decellularized skeletal muscle cell-free matrix has good biocompatibility and may serve as a scaffold for adipose tissue engineering.