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Extracellular matrix regulation of differentiation via modulation of ILK: application to 3D bioprinting of cardiac tissue

通过调节 ILK 进行细胞外基质分化调节：在心脏组织 3D 生物打印中的应用

基本信息

批准号：
10001078
负责人：
Michael McAlpine
金额：
$ 45.21万
依托单位：
UNIVERSITY OF MINNESOTA
依托单位国家：
美国
项目类别：
财政年份：
2017
资助国家：
美国
起止时间：
2017-04-01 至 2022-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10001078
关键词：
3-Dimensional 3D Print Accounting Activities of Daily Living Acute Address Adult Arteries Behavior Biochemistry Biological Biological Models Bioreactors Blood Vessels Cardiac Cardiac Myocytes Cardiac development Cardiovascular Diseases Cardiovascular Physiology Cause of Death Cell Differentiation process Cells Cessation of life Chemicals Chronic Collagen Type I Core Facility Couples Cyclin-Dependent Kinase Inhibitor 3 Dependence Deposition Developed Countries Dimensions Disease model Drug Screening Elements Endothelial Cells Endothelium Engineering Event Extracellular Matrix Extracellular Matrix Proteins Fibronectins Focal Adhesion Kinase 1 Focal Adhesions Formulation Gene Expression Generations Genes Genetic Transcription Goals Heart Heart Transplantation Human In Vitro Individual Injury Integrins Laboratories Ligation Link Literature Mass Spectrum Analysis Mediating Mesoderm Molecular Motivation Mus Myocardial Myocardial tissue Myocarditis Myocardium Natural regeneration Nutrient Outcome Oxygen Pathway interactions Perfusion Pluripotent Stem Cells Printing Proteins RNA Interference Regulation Research Signal Pathway Signal Transduction Spatial Design Technology Testing Therapeutic Thick Time Tissue Engineering Tissues Toxicity Tests Transcriptional Activation United States arteriole base beta catenin bioprinting cell behavior cell type design glycogen synthase kinase 3 beta inhibitor in vivo induced pluripotent stem cell innovation insight integrin-linked kinase kinase inhibitor matrigel mechanical force meetings muscle form optical imaging organ regeneration programs stem cell differentiation stem cells tissue repair tool transcription factor trend

项目摘要

PROJECT SUMMARY The primary objective of this proposal is to couple a) mechanistic insight relating differentiation outcomes to ECM engagement via intracellular signaling events triggered at the focal adhesion (FA), with b) 3D printing of ECM and ECM-associated proteins as a means to direct cell distribution with maturation and thereby enable fabrication of thick, functional cardiac tissue. The proposal is significant as it has the potential to generate replacement tissues and even heart grafts for individuals suffering from acute and chronic injury to the heart. It is the first of its kind to address the conundrum of the apparent uniformity of the focal adhesion relative to the myriad of different ECM/integrin combinations and the corresponding variety of cell behaviors that emerge from ECM engagement. It does so by proposing that elements of the FA, namely integrin linked kinase (ILK) and associated phosphatase, act as sensitive rheostats that can be co-opted to yield desired behavior. Here, the desired behavior is cardiac differentiation, and the innovation is the utilization of optimized ECM formulations as bioinks to create 3D cardiac tissue mimics (3DCTM) capable of directing cell distribution of multiple cell types with differentiation. This concept is feasible as the Ogle laboratory has long-studied the biochemistry of the ECM and stem cell behaviors associated with ECM engagement, and the McAlpine laboratory has focused on 3D printing of functional materials for a range of applications, from biological to electronic and the merger of these materials. These groups will also interface with expertise of the Kamp lab with respect to their recent generation of induced cardiac progenitor cells (iCPCs) to populate the 3DCTM, the Provenzano lab to assist with molecular mechanisms associated with FA formation, the Garry lab to assist with bioreactor implementation, the Zhang lab to add cardiovascular physiology expertise, and the Talkachova lab to assist with optical imaging to assess function of the 3DCTM and core facilities for mass spectrometry of ECM components and gene editing tools for modulating ILK activity. Together, this expertise will be funneled toward meeting the primary objective of the proposal via the following aims: 1) determine whether activation of integrin-linked kinase (ILK) of focal adhesions or costameres couples integrin activation to β-catenin activation via GSK3β to enable expression of genes associated with cardiomyocyte specification, 2) use 3D ECM-based model systems to identify ECM formulations supportive of endothelial differentiation and assess ILK dependence, and 3) use ECM-based 3D printing to modulate differentiation of cardiac cell types spatially in a cardiac tissue mimic. The motivation for this concept was based on extensive literature search, and results from our own experimentation; the approach was designed to insure that the interpretations of the results are subject to minimal bias and the hypotheses posed are truly tested.

项目概要该提案的主要目标是将 a) 将差异化结果与 ECM 通过在粘着斑 (FA) 处触发的细胞内信号事件参与，b) 3D 打印 ECM 和 ECM 相关蛋白作为指导细胞成熟分布的手段，从而使制造厚实的功能性心脏组织。该提案意义重大，因为它有可能产生为遭受急性和慢性心脏损伤的患者提供替代组织甚至心脏移植物。它是同类中第一个解决粘着斑相对于无数不同的 ECM/整合素组合以及相应的各种细胞行为 ECM 参与。它通过提出 FA 的元素，即整合素连接激酶 (ILK) 和相关的磷酸酶，充当敏感的变阻器，可以选择产生所需的行为。在这里，期望的行为是心脏分化，创新是利用优化的 ECM 配方作为生物墨水来创建能够指导多个细胞的细胞分布的 3D 心脏组织模拟物 (3DCTM) 具有差异化的类型。这个概念是可行的，因为奥格尔实验室长期研究生物化学与 ECM 参与相关的 ECM 和干细胞行为，麦卡尔平实验室重点关注 3D 打印功能材料的一系列应用，从生物到电子，以及这些材料。这些小组还将就他们最近的研究与坎普实验室的专业知识进行交流。生成诱导心脏祖细胞 (iCPC) 以填充 3DCTM，Provenzano 实验室可协助与 FA 形成相关的分子机制，Garry 实验室协助生物反应器实施中，Zhang 实验室增加心血管生理学专业知识，Talkachova 实验室提供协助通过光学成像评估 3DCTM 的功能和 ECM 质谱分析的核心设施用于调节 ILK 活性的组件和基因编辑工具。这些专业知识将共同用于通过以下目标满足提案的主要目标： 1) 确定是否激活粘着斑或肋节的整合素连接激酶 (ILK) 将整合素激活与 β-连环蛋白激活结合起来通过 GSK3β 表达与心肌细胞规格相关的基因，2) 使用基于 3D ECM 的模型系统来识别支持内皮分化的 ECM 配方并评估 ILK 依赖性，3) 使用基于 ECM 的 3D 打印在空间上调节心肌细胞类型的分化心脏组织模拟物。这个概念的动机是基于广泛的文献搜索和结果来自我们自己的实验；该方法旨在确保结果的解释是受到最小偏差的影响，并且所提出的假设得到了真正的检验。

项目成果

期刊论文数量（11）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Implementing Biological Pacemakers: Design Criteria for Successful.

DOI：
10.1161/circep.121.009957
发表时间：
2021-10
期刊：
Circulation. Arrhythmia and electrophysiology
影响因子：
0
作者：
Komosa ER;Wolfson DW;Bressan M;Cho HC;Ogle BM
通讯作者：
Ogle BM

Laminin 411 mediates endothelial specification via multiple signaling axes that converge on β-catenin.

DOI：
10.1016/j.stemcr.2022.01.005
发表时间：
2022-03-08
期刊：
STEM CELL REPORTS
影响因子：
5.9
作者：
Hall, Mikayla L.;Givens, Sophie;Santosh, Natasha;Iacovino, Michelina;Kyba, Michael;Ogle, Brenda M.
通讯作者：
Ogle, Brenda M.

Kinases of the Focal Adhesion Complex Contribute to Cardiomyocyte Specification.

局灶性粘附复合物的激酶有助于心肌细胞规范。