Early Identification and Characterization of Connective Tissue Progenitors

结缔组织祖细胞的早期识别和表征

基本信息

批准号：
8806193
负责人：
George F Muschler
金额：
$ 20.01万
依托单位：
CLEVELAND CLINIC LERNER COM-CWRU
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-09-15 至 2016-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8806193
关键词：
Address Area Aspirate substance Back Biologic Characteristic Biological Biological Assay Biological Markers Bone Marrow Bone Tissue Cartilage Cell Count Cell Density Cell Proliferation Cell Therapy Cell Transplantation Cell physiology Cell surface Cells Cellular Morphology Cellular biology Characteristics Clinical Collection Colony-Forming Units Assay Connective Tissue Data Correlations Development Device or Instrument Development Devices Diagnostic tests Differentiation Antigens Disease Early identification Embryology Environment Flow Cytometry Fluorescence Future Growth and Development function Health Heterogeneity Human Human Characteristics Image In Situ In Vitro Individual Kinetics Knowledge Label Laboratories Learning Life Location Maintenance Marrow Measurement Measures Methods Metric Morphology Natural regeneration Nature Pathogenesis Pattern Performance Pharmaceutical Preparations Phase Phenotype Population Population Heterogeneity Predictive Value Prevalence Process Proliferating Property Protocols documentation Regenerative Medicine Reproducibility Sampling Sea Source Staining method Stains Stem cells Surface Suspension substance Suspensions System Testing Therapeutic Time Tissue Sample Tissues Variant Video Microscopy acronyms base biological systems bone cell motility cell type cellular imaging design drug development imaging modality improved in vivo insight interest novel diagnostics progenitor public health relevance regenerative therapy repaired response stem stem cell population tissue regeneration

项目摘要

DESCRIPTION: Cell based regenerative therapy requires the activity of stem progenitor cells for tissue regeneration. Connective tissue progenitors (CTPs) are defined as the heterogeneous population of tissue resident stem and progenitor cells that are capable of proliferating and differentiating into one or more connective tissue phenotypes. The number and heterogeneity of CTPs can be estimated using the colony formation assay, in which colony founding CTPs are detected and measured by their ability to proliferate and colony of progeny that express a connective tissue phenotype in vitro. In addition to measuring the number and prevalence of CTPs in a tissue sample, differences in biological potential between various CTPs can be assessed based on variation in cell number, distribution, morphology and expression patterns between colonies that are formed. While very useful, a very important limitation of traditional colony formation assays is that their only direct measurement is made on the progeny of the founding CTP (the colony). Information about the founding CTP can only be inferred from the number of colonies and their features. Colony assays to not reveal the physical or biological properties of the rare individual colony founding CTPs themselves, before proliferation. To address this limitation, we have developed a live cell, videomicroscopy system that enables systematic characterization of colony founding cells (morphology, expression of surface markers) before colony formation. Using multicolor labeling methods, a mixed tissue derived cell population is labeled for cell markers of interest at the time of plating. Time lapse phase contras images are then captured over 6 days, while colonies form, over a large field of view. At the end of the culture period, colonies are characterized with respect to established markers and colony metrics. By reversing the video, each colony can also be traced back to identify the colony founding cell, as one in a sea of other non-progenitors. Using this method we propose to significantly advance our understanding of the nature of human CTPs, their diversity and subtypes, their location in bone tissue and define factors that can be measured non-invasively and used to predict their biological potential, or select CTP best suited for specific therapeutic purposes.

描述：基于细胞的再生疗法需要茎祖细胞的活性以进行组织再生。结缔组织祖细胞（CTP）被定义为能够扩增和区分成一个或多个结缔组织表型的组织居民茎和祖细胞的异质种群。 CTP的数量和异质性可以使用菌落形成测定法估算，其中检测并通过其增殖和菌落在体外表达结缔组织表型的菌落创始CTP进行测量。除了测量组织样本中CTP的数量和患病率外，还可以根据细胞数，分布，形态和表达模式的变化来评估各种CTP之间的生物学潜力的差异。虽然非常有用，但传统菌落形成测定法的一个非常重要的局限性是，它们唯一的直接测量是对基础CTP（殖民地）的后代进行的。有关创建CTP的信息只能从殖民地数量及其特征中推断出来。菌落测定在扩散之前，不要揭示CTPS本身的罕见单个菌落的物理或生物学特性。为了解决这一局限性，我们开发了一个活细胞，视频显微镜系统，该系统能够在菌落形成之前系统地表征菌落基础细胞（形态，表面标记的表达）。使用多色标记方法，在电镀时，将混合组织衍生的细胞群标记为感兴趣的细胞标记。随后在6天内捕获了延时阶段对抗图像，而菌落形成在很大的视野上。在文化时期结束时，殖民地的特征是既定标记和殖民指标。通过逆转视频，也可以追溯到每个菌落以识别菌落创始细胞，作为其他非企业家的海洋。使用这种方法，我们建议显着提高我们对人CTP的性质，它们的多样性和亚型，它们在骨骼组织中的位置以及定义可以非侵入性测量的因素，并用于预测其生物学潜力的因素，或者选择最适合特定治疗目的的CTP。