Design of injectable hydrogels to improve cell transplantation for peripheral arterial disease

 描述（由申请人提供）：将移植细胞无效递送至所需部位是阻碍基于细胞的再生医学疗法临床应用的主要障碍。令人沮丧的低移植细胞存活率（~1-32%）是由注射期间的机械力（其破坏细胞膜）和缺乏三维基质以支持原位细胞存活引起的。外周动脉疾病（PAD）发生在50岁以上的美国人中，每20人中就有一人。目前的治疗方案往往是无效的，通常导致截肢的情况下，严重的肢体缺血。一种有前景的疗法是将患者来源的人脂肪来源的基质细胞（hASC）移植到缺血部位以通过分泌促血管生成旁分泌因子来促进再灌注。在这里，我们提出了一种物理交联的双网络水凝胶，其刚度范围从10到3000 Pa，以解决移植后细胞死亡的问题，调节细胞功能以优化hASC的血管生成潜力，并改善PAD小鼠模型中的血液再灌注。在具体目标1中，我将合成和表征经历两种不同物理交联机制的可注射水凝胶。第一交联步骤通过基于肽的分子识别离体发生以将细胞包封在物理水凝胶内，而第二交联步骤通过热相变原位发生以形成增强网络。该水凝胶将通过改变增强网络密度而配制成具有10至3000 Pa的宽范围刚度，以在注射器注射期间提供hASC保护并改善移植后的细胞活力。在具体目标2中，我将探索刚度对体外hASCs分泌促血管生成生长因子的影响，并最大限度地提高封装在这一系列新型水凝胶中的hASCs的血管生成潜力。在具体目标3中，我将选择性能最好的水凝胶制剂并将hASC递送至PAD的鼠后肢缺血模型。将评估血液灌注和新血管形成的恢复，以验证使用我们的新型水凝胶的基于hASC的疗法，寻求加速PAD的细胞疗法的临床实现。

 DESCRIPTION (provided by applicant): Ineffective delivery of transplanted cells to the desired site is a major hurdle hampering the clinical application of cell-based, regenerative medicine therapies. Dismally low transplanted cell viability (~1-32%) is caused both by the mechanical forces during injection, which damage the cell membrane, and the lack of a three- dimensional matrix to support cell survival in situ. Peripheral arterial disease (PAD), occurs to one in 20 Americans over the age of 50. Current treatment options are often ineffective, commonly leading to amputation in cases of critical limb ischemia. A promising therapy is the transplantation of patient-derived human adipose- derived stromal cells (hASCs) into the ischemic site to promote reperfusion through secretion of pro-angiogenic paracrine factors. Here we propose a physically-crosslinked, double-network hydrogel with a wide range of stiffness from 10 to 3000 Pa to address the problem of post-transplantation cell death, to regulate cell functionality to optimize the angiogenic potential of hASCs, and to improve blood reperfusion in a murine model for PAD. In Specific Aim 1, I will synthesize and characterize an injectable hydrogel that undergoes two different physical crosslinking mechanisms. The first crosslinking step occurs ex vivo through peptide-based molecular recognition to encapsulate cells within a physical hydrogel, while the second crosslinking step occurs in situ through a thermal phase transition to form a reinforcing network. This hydrogel will be formulated with a wide range of stiffness from 10 to 3000 Pa by varying the reinforcing network densityto provide hASCs protection during syringe injection and to improve the cell viability post-transplantation. In Specific Aim 2, I will explore the effect of stiffness on the secretion of pro-angiogenic growth factors by hASCs in vitro and maximize the angiogenic potential of hASCs encapsulated within this series of novel hydrogels. In Specific Aim 3, I will choose the best-performing hydrogel formulation and deliver hASCs to a murine hind limb ischemia model of PAD. Recovery of blood perfusion and neovascularization will be assessed to validate the hASC-based therapy using our novel hydrogels, seeking to accelerate the clinical realization of cell therapy for PAD.