Regulation and Enhancement of Angiogenesis in Dense Fibrin Matrices

描述（由申请人提供）：血管生成，即从现有的血管系统中萌发新的毛细血管，是一个复杂的生物过程，对许多疾病的治疗和组织工程的成功至关重要。在组织工程中促进血管生成的许多有前途的策略中，包括通过精确的空间和时间控制来控制促血管生成分子的递送。另一种方法是传递一种合适的细胞类型，这种细胞类型可以提供更多的促血管生成线索的生理性混合物，从而加速宿主血管的募集。在与两位高级共同研究者的合作下，PI已经开始在一个强大的、基于纤维蛋白的毛细血管形态发生的体外3d模型中探索后一种策略。使用该模型系统，支持形成稳定的（> - 3周）具有明确管腔的毛细血管样结构，我们已经证明毛细血管形态发生被增加的纤维蛋白基质密度显著抑制。此外，我们已经证明，这种纤维蛋白密度阻滞部分是由于内皮细胞无法上调基质金属蛋白酶（MMPs）的一个亚群，而基质金属蛋白酶是局部重塑基质和维持毛细血管侵袭所必需的。然而，将骨髓来源的间充质干细胞（MSCs）添加到三维组织结构中可以显著增强毛细血管形态发生，通过上调这一MMPs亚群的表达和/或活性，部分克服了由基质密度增加引起的抑制。在这些初步发现的基础上，这项新的研究者生物工程研究拨款提案试图在体外和体内测试MSCs可以刺激致密三维纤维蛋白基质内的血管生成的假设。这些目标将通过以下三个具体目标来实现：量化MSCs嵌入密集纤维蛋白基质的能力，以增强体外毛细血管样网络的形成。2)。探讨mmp的一个子集（mmp -2和-9以及膜型MT1-MMP）在MSCs血管生成增强中的作用。3)。在scid小鼠模型中确定MSCs刺激皮下血管形成的能力。这些研究的成功完成将有助于我们对ECM和MMPs在血管生成中的作用的基本理解，并最终影响合成ECM类似物的设计和干细胞在组织工程中的应用。

DESCRIPTION (provided by applicant): Angiogenesis, the sprouting of new capillary blood vessels from existing vasculature, is a complex biological process of critical importance to the treatment of numerous pathologies and the success of tissue engineering. Amongst the many promising strategies to promote angiogenesis in tissue engineering includes the controlled delivery of pro-angiogenic molecules with precise spatial and temporal control. An alternative approach is to deliver an appropriate cell type that can provide a more physiologic mixture of pro- angiogenic cues that will accelerate the recruitment of host vessels. In conjunction with two senior co- investigators, the PI has begun to explore this latter strategy in a robust, fibrin-based 3-D in vitro model of capillary morphogenesis. Using this model system, which supports the formation of stable (> 3 weeks) capillary-like structures with well-defined lumens, we have shown that capillary morphogenesis is significantly inhibited by increasing fibrin matrix density. Furthermore, we have shown that this fibrin density block is partially due to the inability of endothelial cells to upregulate a subset of matrix metalloproteinases (MMPs) required to locally remodel the matrix and sustain capillary invasion. However, the addition of bone marrow-derived mesenchymal stem cells (MSCs) to the 3-D tissue construct significantly enhances capillary morphogenesis, partially overcoming the inhibition caused by increased matrix density by upregulating the expression and/or activity of this subset of MMPs. Building on these preliminary findings, this new investigator Bioengineering Research Grant proposal seeks to test the hypothesis that MSCs can stimulate angiogenesis within dense 3-D fibrin matrices both in vitro and in vivo. These objectives will be achieved via three specific aims as follows: 1.) Quantify the ability of MSCs embedded throughout dense fibrin matrices to enhance capillary-like network formation in vitro. 2.) Probe the roles played by a subset of MMPs (MMPs-2 and -9 and the membrane-type MT1-MMP) in the angiogenic enhancement by MSCs. 3.) Determine the ability of MSCs to stimulate vascularization in vivo in a subcutaneous site in a SCID-mouse model. Successful completion of these studies will contribute to our fundamental understanding of the role of the ECM and MMPs in angiogenesis, and will ultimately impact the design of synthetic ECM analogs and the use of stem cells in tissue engineering applications.