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Acquirement and evaluation of murine ventricular extracellular matrix

JIANG Yu-Dong, LI Wen-Si, YU Chong, WANG Lu, SUN Xiao-Xi, XI Jiao-Ya^*

Department of Physiology and Chinese-German Stem Cell Center, School of Basic Medicine, Tongji Medical College, Huazhong University of Science and Technology, the Key Laboratory for Drug Target Researches and Pharmacodynamic Evaluation of Hubei Province, Wuhan 430030, China

Abstract

Cardiac extracellular matrix (ECM), generated from the process of decellularization, has been widely considered as an ideal source of biological scaffolds. However, current ECM preparations are generally difficult to be applied to generate cardiac tissue. Our research was aimed to improve decellularization protocols to prepare cardiac ECM slices. Adult murine ventricular tissues were embedded in low melting agarose and cut into 300 μm slices, and then were divided randomly into three groups: normal cardiac tissue, SDS treated group (0.1% SDS) and SDS+Triton X-100 treated group (0.1% SDS+0.5% Triton X-100). Total RNA content and protein content quantification, HE staining and immunostaining were used to evaluate the removal of cell components and preservation of vital ECM components. Furthermore, murine embryonic stem cell-derived cardiomyocytes (mES-CMs) and mouse embryonic fibroblasts (MEFs) were co-cultured with ECM slices to evaluate biocompatibility. The relative residual RNA and protein contents of ECM slices significantly decreased after decellularization. HE staining showed that SDS+Triton X-100 treatment better destroyed cellular structure and removed nuclei of ECM slices, compared with SDS treatment. Immunostaining showed that collagen IV and laminin were better preserved and presented better similarity to original cardiac tissue in ECM slices acquired by SDS+Triton X-100 treatment. However, collagen IV and laminin were significantly decreased and arranged disorderly in SDS treated group. We observed effective survival (≥ 12 days) of MEFs and mES-CMs on ECM slices acquired by SDS+Triton X-100 treatment, and signs of integration, whereas those signs were not found in SDS treated group. We concluded that, compared with traditional SDS method, new combined protocol (SDS+Triton X-100) generated ECM slices with better component and structural preservation, as well as better biocompatibility.

Key words: myocardium slice; tissue engineering; decellularization; extracellular matrix

Received: 2014-05-11　　Accepted: 2014-07-09

Corresponding author: 席姣娅　　E-mail: zhengyall@hotmail.com

Citing This Article：

JIANG Yu-Dong, LI Wen-Si, YU Chong, WANG Lu, SUN Xiao-Xi, XI Jiao-Ya. Acquirement and evaluation of murine ventricular extracellular matrix. Acta Physiol Sin 2014; 66 (6): 709-717 (in Chinese with English abstract).