The angiogenic potential of PBMNCs and QQMNCs was assessed using the EPC-CFA, as previously described6 (link),14 (link). The EPC-CFA was designed to differentiate total EPC colony-forming units (tEPC-CFUs) into two types of EPC-CFUs (primitive and definitive). Primitive EPC-CFU (pEPC-CFUs) are a predominantly proliferative population of cells, and definitive EPCs-CFU (dEPC-CFU) are a predominantly vasculogenic population with greater adhesion, migration, differentiation, and tubularization potential.
Briefly, adhesive EPC colonies were counted using the EPC-CFA (MethoCult SFBIT; Stem Cell Technologies Inc., Vancouver, BC, Canada) with semisolid culture medium containing proangiogenic growth factors/cytokines in 35-mm Primaria dishes (BD Falcon; BD Biosciences). Twelve aliquots of cells were seeded at 2 × 105 cells/dish (three dishes per subject) for EPC-CFA. Sixteen to 18 days after culture initiation, the number of adherent colonies per dish was measured using a gridded scoring dish (Stem Cell Technologies) under phase-contrast light microscopy (Eclipse TE300; Nikon, Tokyo, Japan). The experiments were performed in triplicate. Experimental conditions were masked and two investigators counted the number of pEPC-CFUs and dEPC-CFUs.
Briefly, adhesive EPC colonies were counted using the EPC-CFA (MethoCult SFBIT; Stem Cell Technologies Inc., Vancouver, BC, Canada) with semisolid culture medium containing proangiogenic growth factors/cytokines in 35-mm Primaria dishes (BD Falcon; BD Biosciences). Twelve aliquots of cells were seeded at 2 × 105 cells/dish (three dishes per subject) for EPC-CFA. Sixteen to 18 days after culture initiation, the number of adherent colonies per dish was measured using a gridded scoring dish (Stem Cell Technologies) under phase-contrast light microscopy (Eclipse TE300; Nikon, Tokyo, Japan). The experiments were performed in triplicate. Experimental conditions were masked and two investigators counted the number of pEPC-CFUs and dEPC-CFUs.
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