Quantifying Proliferative and Surface Marker Heterogeneity in Colony Founding Connective Tissue Progenitors and Their Progeny Using Time‐Lapse Microscopy

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 dura-

Influence of Glucose Concentration on Colony-Forming Efficiency and Biological Performance of Primary Human Tissue–Derived Progenitor Cells

“…83 From work done previous in our lab, we understand that the average lag period from the day cells are placed into in vitro culture until the cells undergo their first cell division is 2 to 3 days. 84 As a result, the cells probably had a chance to undergo at most 3 to 5 rounds of replication. Although, there is no literature evidence comparing the proliferation potential of native CTPs derived from various tissue sources, researchers have reported proliferation potentials of culture expanded populations.…”