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首页> 外文期刊>Journal of tissue engineering and regenerative medicine >Quantifying proliferative and surface marker heterogeneity in colony‐founding connective tissue progenitors and their progeny using time‐lapse microscopy
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Quantifying proliferative and surface marker heterogeneity in colony‐founding connective tissue progenitors and their progeny using time‐lapse microscopy

机译:用延时显微镜量化菌落创建结缔组织祖细胞的增殖和表面标记异质性及其后代

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Abstract 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 connective tissue phenotypes. The prevalence and variation in clonal progeny of CTPs can be characterized using a colony formation assay. However, colony assays do not directly assess the characteristics of the colony‐founding CTP. We performed large, field‐of‐view, time‐lapse microscopy to manually track colonies back to the founding cells. Image processing and analysis was used to characterize the colonies and their founding cells. We found that the traditional colony‐forming unit (CFU) assay underestimates the number of founding cells as colonies can be formed by more than one founding cell. After 6?days in culture, colonies do not completely express CD73, CD90, and CD105. Heterogeneity in colony cells was characterized by two cell populations, proliferative and spread cells. Regression modelling of duration of lag phase and doubling time by cell marker suggests the presence of CD90 and CD105 in CTP subpopulations with different proliferative capabilities. From mathematical modelling of clonal colonies, we quantitatively characterized proliferation, migration, and cell marker expression rates to identify desirable clones for selection. Direct assessment of colony formation parameters led to more accurate assessment of CFU heterogeneity. Furthermore, these parameters can be used to quantify the diversity and hierarchy of stem and progenitor cells from a cell source or tissue for tissue engineering applications.
机译:摘要结缔组织祖细胞(CTPS)被定义为能够增殖和分化成结缔组织表型的组织植物茎和祖细胞的异质群。 CTP的克隆后代的患病率和变化可以使用菌落形成测定来表征。然而,菌落测定不直接评估殖民地成立CTP的特征。我们进行了大,视野,延时显微镜,以手动跟踪殖民地回到创始细胞。图像处理和分析用于表征菌落及其创始细胞。我们发现传统的菌落形成单元(CFU)测定低估了作为菌落的成立细胞的数量可以由多于一个创始细胞形成。在培养6?天后,菌落不完全表达CD73,CD90和CD105。菌落细胞中的异质性以两种细胞群,增殖和涂抹细胞的特征在于。滞后阶段持续时间和细胞标记倍增时间的回归建模表明CD90和CD105在具有不同增殖能力的CTP亚群中的存在。从克隆菌落的数学建模中,我们定量表征增殖,迁移和细胞标志物表达率以鉴定所需的选择克隆。直接评估菌落形成参数导致对CFU异质性的更准确评估。此外,这些参数可用于量化来自用于组织工程应用的细胞源或组织的茎和祖细胞的分集和层次。

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