Supplementary Material for: Tissue-Derived Signals for Mesenchymal Stem Cell Stimulation: Role of Cardiac and Umbilical Cord Microenvironments

The tissue microenvironment regulates such stem cell behaviors as self-renewal and differentiation. Attempts to mimic components of these microenvironments could provide new strategies for culturing and directing the behavior of stem cells. The aim of the present study was to mimic cardiac and umbilical cord tissue microenvironments in vitro and compare the resulting bone marrow-derived mesenchymal stem cell (BM-MSC) behaviors. We generated tissue microenvironments using conditioned medium (CM) and extracellular matrix (ECM) samples obtained from human heart and umbilical cord tissue explant cultures and by tissue decellularization. Mass spectrometry and immunostaining were used to characterize and determine the specific protein profiles of the ECMs and CMs. We demonstrated that the ECMs and CMs were not cytotoxic to BM-MSCs and could thus be tested via cell culture. The BM-MSCs showed a higher proliferation rate when cultured with umbilical cord-derived CM compared with the other analyzed conditions. Furthermore, the ECMs increased cell adhesion and migration. However, although the conditions tested in this work were able to maintain the viability and affect the proliferation, adhesion and migration of BM-MSCs in vitro, mimicking tissue microenvironments using ECM and CM was not sufficient to induce the cardiomyogenic differentiation of BM-MSCs. The present study provides a thorough characterization of the biological activity of these ECMs and CMs in human BM-MSC cultures.<br>

组织微环境（tissue microenvironment）可调控干细胞的自我更新与分化等行为。模拟该类微环境组分的研究可为干细胞的培养与行为调控提供全新策略。本研究旨在体外模拟心脏与脐带组织微环境，并对比由此得到的骨髓间充质干细胞（bone marrow-derived mesenchymal stem cell，BM-MSC）的相关行为。本研究通过从人类心脏与脐带组织外植体培养物中获取条件培养基（conditioned medium，CM）与细胞外基质（extracellular matrix，ECM）样本，并结合组织脱细胞处理技术，构建了组织微环境模型。研究采用质谱法（mass spectrometry）与免疫染色（immunostaining）技术，对ECM与CM的特异性蛋白质谱进行表征与鉴定。我们证实，ECM与CM对BM-MSCs无细胞毒性，因此可用于细胞培养实验。与其他受试培养条件相比，采用脐带来源CM培养的BM-MSCs呈现出更高的增殖速率。此外，ECM可促进细胞黏附与迁移。尽管本研究中所用的培养条件可在体外维持BM-MSCs的存活，并影响其增殖、黏附与迁移行为，但采用ECM与CM模拟组织微环境，并不足以诱导BM-MSCs发生心肌分化（cardiomyogenic differentiation）。本研究全面表征了上述ECM与CM在人类BM-MSC培养体系中的生物学活性。