解剖学报

解剖学报 ›› 2016, Vol. 47 ›› Issue (6): 824-828.doi: 10.16098/j.issn.0529-1356.2016.06.018

• 组织学胚胎学发育生物学 • 上一篇    下一篇

猪小肠黏膜下层脱细胞支架的制备及组织学评价

王付燕 杜立群*   

  1. 山东大学齐鲁医院眼科， 济南 250000
  • 收稿日期:2016-03-28 修回日期:2016-06-07 出版日期:2016-12-06 发布日期:2016-12-06
  • 通讯作者: 杜立群 E-mail:dlqzqt@163.com
  • 基金资助:

    山东省自然科学基金面上项目;山东省优秀中青年科学家科研奖励基金

Preparation and histological evaluation of the decellularized scaffold for porcine small intestinal submucosa

WANG Fu-yan DU Li-qun*   

  1. Department of Ophthalmology, Qilu Hospital of Shandong University, Jinan 250000,Chian
  • Received:2016-03-28 Revised:2016-06-07 Online:2016-12-06 Published:2016-12-06
  • Contact: DU Li-qun E-mail:dlqzqt@163.com

PDF

摘要：

目的 评价不同浓度的十二烷基磺酸钠（SDS）处理猪小肠黏膜下层（SIS）的脱细胞效果，筛选最佳脱细胞浓度，为组织工程化角膜上皮的制备提供支架材料。方法 配制0.1%、0.2%、0.3%、0.5% SDS随机分成A、B、C、D 4组，分别脱细胞处理SIS 15min、30min、1h、2h（n=20），同时观察SIS的大体形态变化，HE和4，6-二脒基-2-苯基吲哚(DAPI)染色观察脱细胞前后SIS的生物学特性，并对脱细胞支架进行基因组 DNA 分析(n=5)。 结果 脱细胞SIS肉眼观察呈乳白色、半透明的膜状物，具有一定的弹性、韧性及透光性；HE和DAPI染色光学显微镜观察结果显示，A组及B组处理30min时SIS生物支架无细胞及DNA残留，组织结构保留完整，胶原纤维间孔隙率增加；A组及B组处理30min脱细胞率可达90%以上。 结论 0.1%及0.2%SDS处理30min条件下脱细胞效果较好，能更好地降低SIS的免疫原性，是SDS脱细胞处理SIS比较理想的浓度时间条件，为组织工程化角膜上皮的构建奠定理论基础。

关键词: 组织工程, 支架材料, 脱细胞, 小肠黏膜下层, HE染色, DAPI染色

Abstract:

Objective To evaluate the different concentrations of sodium dodecyl sulfonate（SDS）on efficiency of cell removal from porcine small intestinal submucosa（SIS）and optimize the best concentration to produce an acellular porcine SIS scaffold for use in developing a tissueengineered corneal epithelium. Methods Fresh porcine SIS were decellularized with SDS (concentration of 0.1%, 0.2%, 0.3%, and 0.5% for 15 minutes, 30 minutes, 1 hour, and 2 hours) and randomly divided into 4 groups (A, B, C, D, 20 pieces in each group ) and the decellularization process was observed at the same time.After decellularization，the scaffolds were examined through the HE, 4,6-diamidino-2-phenylindole dihydrochloride (DAPI) staining and the genomic DNA contect analysis to observe the biological characteristics of SIS. Results A cellular SIS was shown ivory, translucent and had certain transmittance. HE and DAPI staining revealed that when treatment with the time of 30 minutes in group A and B SIS were decellularized completely and preserved the overall tissue histoarchitecture, the collagen fiber porosity was increased, the acellular rate was more than 90%. Conclusion The acellular matrix treatment effect of 0.1% and 0.2% SDS for a period of 30min is satisfactory. The immunogenicity of the SIS can be reduced, therefore, it is an ideal acellular concentration for SIS and provides a theoretical basis for the construction of tissue engineering corneal epithelium.

Key words: Tissue engineering, Scaffold, Decellularization, Small intestinal submucosa, HE staining, DAPI staining