Control of Stem Cell Behavior by Exposure to Tissue-Specific ECM

摘要t 脂肪组织源性间充质干细胞(AD-MSCs)有可能成为一种重要的工具 对于骨组织工程来说，由于它们在体内的频率相对较高，可以提供非常大的 再生疗法所需的细胞数量。不幸的是，许多研究表明AD-MSCs 与其他类型的干细胞相比，其成骨能力要低得多。我们小组已经开发出一种新的 体外产生的细胞外基质(ECM)，概括了天然骨髓(BM-)的特定属性 和脂肪(AD-)微环境。在以前的研究中，我们证明了BM-ECM显著 与标准培养塑料相比，骨髓(BM)来源的MSCs促进成骨分化。在……里面 在目前的建议中，我们评估了BM-ECM增加AD-MSCs成骨能力的潜力， 通过这样做，显著提高了它们在骨骼修复和再生方面的临床相关性。 我们的初步数据表明，BM和AD来源的ECM能够指导BM-MSC的分化 向成骨细胞和脂肪细胞方向发展。这些ECM的特点是在 生化成分，建筑，甚至机械性能。我们证明了BM-和AD- 在两种组织特异性ECM上培养的MSCs在每种类型的MSC上都更容易增殖 维持在细胞外基质上--源自相同的相应组织。此外，我们观察到BM-和AD- 骨髓间充质干细胞在BM-ECM上呈低环状铺展，在AD-ECM上呈高度环状铺展 传播形态。这些观察结果与最近的报告一致，这些报告表明， 骨髓间充质干细胞的形态最终与涉及谱系决定的信号线索有关。此外，最近 研究表明，焦点黏附复合体的形成可以通过以下方式调节干细胞的命运 经典的Wnt/β-连环蛋白途径。我们将调查是否在组织上形成独特的焦点黏附复合体- 特定的ECM和重建的微环境(S)负责引导MSC分化到 成骨细胞和脂肪细胞谱系。 作为这些观察的结果，我们假设BM-ECM的骨特异性微环境 可以对AD-MSCs进行重新培养，以增强其成骨能力。以下是具体目标 建议检验这一假设：1)确定BM-和AD-MSC分化的差异，评估 通过体外和体内成骨和成脂实验，在组织培养塑料上进行培养。 和组织特异性细胞外基质；2)确定促进MSC的分子机制(S) 在磷酸三钙和组织特异性细胞外基质上培养，向成骨细胞和脂肪细胞分化。

Abstract t Adipose tissue-derived mesenchymal stem cells (AD-MSCs) have the potential to serve as a critical tool for bone tissue engineering as their relatively high frequency in the body can provide the exceedingly large numbers of cells required for regenerative therapies. Unfortunately, many studies have shown that AD-MSCs have considerably less osteogenic potential than other types of stem cells. Our group has developed an in vitro-produced extracellular matrix (ECM) that recapitulates specific attributes of the native bone marrow (BM-) and adipose (AD-) microenvironment. In previous studies, we demonstrated that BM-ECM significantly promotes osteogenic differentiation by bone marrow (BM)-derived MSCs relative to standard culture plastic. In the current proposal, we evaluate the potential of BM-ECM to increase the osteogenic capacity of AD-MSCs, and by doing so, dramatically improve their clinical relevance for skeletal repair and regeneration. Our preliminary data show that BM- and AD-derived ECMs are capable of guiding BM-MSC differentiation towards osteoblast and adipocyte lineages. These ECMs are characterized by significant differences in biochemical composition, architecture and even mechanical properties. We demonstrated that BM- and AD- MSCs cultured on both types of tissue-specific ECM proliferated more readily when each type of MSC was maintained on ECM-derived from the same respective tissue. Additionally, we observed that BM- and AD- MSCs both exhibited low-circularity spreading on BM-ECM, while on AD-ECM, they displayed a high-circularity spreading morphology. These observations are consistent with recent reports demonstrating that changes in MSC morphology are ultimately related to signaling cues involved in lineage-decisions. Furthermore, recent studies have suggested that focal adhesion complex formation can regulate stem cell fate decisions through the canonical Wnt/β-catenin pathway. We will investigate if unique focal adhesion complexes form on tissue- specific ECMs and reconstitute the microenvironment(s) responsible for directing MSC differentiation to either the osteoblast and adipocyte lineage. As a result of these observations, we hypothesize that the bone-specific microenvironment of BM-ECM can "re-train" AD-MSCs in order to enhance their osteogenic potential. The following specific aims are proposed to test this hypothesis: 1) To determine differences in BM- and AD-MSC differentiation, assessed with assays of osteogenesis and adipogenesis in vitro and in vivo, with culture on tissue culture plastic (TCP) and tissue-specific ECM and 2) To determine the molecular mechanism(s) responsible for promoting MSC differentiation to the osteoblast and adipocyte lineages with culture on TCP and tissue-specific ECM.