组织工程化心肌构建研究中形态学检测体系的建立

目的：建立并完善再造工程化心肌组织研究中形态学检测项目平台。方法：取新生大鼠天然心肌组织固定，制作天然心肌石蜡组织切片；用胰酶消化法分离新生大鼠心肌细胞进行细胞爬片培养；以液态胶原为支架材料与来源于新生大鼠的心肌细胞在体外构建工程化心肌组织条带并进行力学拉伸刺激，进而对其进行固定处理，制作石蜡组织切片。之后将天然心肌组织、心肌细胞爬片和工程化心肌组织标本分别进行HE及心肌特异性标志肌钙蛋白T（cTnT）免疫组化染色，并对三者的形态学检测结果进行观察、分析。结果：天然心肌组织的HE及cTnT结果反映了三维立体正常心肌组织在体情况下组织结构、细胞形态、数量比例等组织形态学特点；心肌细胞爬片培养简便直观地反映了体外二维条件下的心肌细胞形态、数量及其比例的改变；工程化心肌组织检测则反映了三维立体再造心肌的组织结构、细胞形态、数量及比例改变。结论：设立天然心肌、心肌细胞爬片及工程化心肌组织三者的HE、cTnT免疫组化检测项目并对其结果进行对照分析，从二维与三维、在体与体外等方面，建立并完善了再造工程化心肌组织研究中形态学检测对照体系，提高了实验效率，节约了实验成本。

A morphological analysis system for cardiac tissue engineering studies

Objective：To develop a necessary analysis system of morphological assay for cardiac tissue engineering studies. Methods： Paraffin sections of normal cardiac tissue from neonatal rats were prepared. Cardiac myocytes of neonatal rats were isolated by trypsin. Some cells were cultured on the slides as the sample of cultured cardiac myocytes. Some were mixed with liquid collagen type Ⅰ and then cast to construct tissue engineered cardiac strips. Paraffin sections of normal cardiac tissue,the sample of cultured cardiac myocytes and tissue engineered cardiac strips were stained with HE and immunohistochemical reagent respectively. The results were observed by microscope. Results： The results of HE and immunohistochemistry staining of the normal cardiac tissue showed histological characters of tissue structure, cell shape and cell ratio, which reflected the normal three-dimensional cardiac tissue. The sample of cultured cardiac myocytes also showed these characters in two-dimensional condition in vitro. The morphological results of tissue engineered cardiac strips showed these characters of reconstructed three-dimensional cardiac tissue. All the above three types of samples showed evident difference in tissue structure, cell shape and cell ratio. Conclusion： A morphological analysis system for cardiac tissue engineering studies is developed. It can provide scientific aids for an experimental system for cardiac tissue engineering studies ,two dimensional or three dimensional, in vivo an in vitro.