微量蠕动泵法制备大鼠全肾脏脱细胞基质及其成份鉴定

Preparation of whole - kidney acellular matrix using peristaltic pump perfusion and identifica- tion the composition of matrix

目的通过微量蠕动泵灌注法制备全肾脏脱细胞基质支架，并鉴定肾脏脱细胞基质是否保留了天然细胞外基质。方法成年雄性SD大鼠（200-250g），使用微量蠕动泵按序将脱细胞洗涤液灌流，脱细胞处理后，HE染色和DAPI染色观察支架微观结构；通过免疫组化方法对脱细胞基质成份进行鉴定。结果HE染色和DAPI染色结果证实肾脏脱细胞基质支架中未见细胞残留；免疫组织化学染色结果证实保存了细胞外基质蛋白的天然表达类型，包括I型、IV型、纤维连接蛋白、层粘连蛋白。结论使用微量蠕动泵灌流法成功制备了完整的全肾脏脱细胞基质支架，这种支架保留了天然细胞外基质的重要蛋白，是一种理想的天然支架用于肾脏组织工程中。

Objective To make a whole - kidney acellular matrix scaffold through the method of peristaltic pump perfusion and to identify whether the kidney aeellular matrix reserves extracellular matrix proteins or not. Methods The male Sprague - Dawley （SD） rats, 200 ~ 250 g. Intravenous eathers were injected into the renal artery to establish the channels for perfusion, followed by the perfu- sion of deeellularization solution by using peristaltic pumps. After the deeellularization, the acellular matrix was observed under microscope after hematoxylin and eosin（HE） staining and fluorescence mi- croscope after DAPI staining. Immunohistochemistry was performed to identify the composition of kid- ney acellualr matrix. Results The result of HE staining and DAPI staining demonstrate the whole re- moval of cellular material in kidney acelluar matrix. Immunohistoehemistry confirmed the preservation of the natural expression type of extraeellular matrix proteins including collagen types I and IV, fi- hroneetin and laminin. Conclusions Peristaltic pump perfusion is a successful method to make the kidney acellular matrix, and this scaffold retains the critical proteins of natural extraeellular matrix. Therefore the kidney acellular matrix is an ideal natural scaffold for renal tissue engineering.