Figure 1.

Isolation and Characterization of Heart-Resident Nestin⁺ Cells from Nestin-GFP Reporter Mice

(A) Heart-derived CD45⁻Ter119⁻CD31⁻ cells were flow cytometrically isolated from the hearts of 7-day-old Nestin-GFP transgenic mice, and Nestin⁺ and Nestin⁻ subpopulations were divided based on GFP expression. Cells from 7-day-old non-transgenic C57BL/6 mice were isolated as a control. (B) Representative images showing the clonal sphere growth of single Nestin-GFP⁺ cells. Cells in the upper and lower columns were observed under bright-field and fluorescence field microscopy, respectively. Scale bars, 50 μm. (C) The expressions of cell surface markers on Nestin-GFP⁺ cells were detected by flow cytometry. (D) Representative stained images show that mouse heart-derived Nestin⁺ cells could differentiate into osteocytes (Alizarin red), adipocytes (Oil red O), and chondrocytes (toluidine blue); these findings were confirmed by qPCR analysis of the differentiation-associated genes Runx2 and SARC (osteogenesis), FabP4 and PPAR-γ (adipogenesis), and Collagen II and Collagen X (chondrogenesis). The “con” refers to undifferentiated Nestin⁺ cells, which represents Nestin⁺ cells that were cultured in the normal growth medium. Scale bars, 100 μm. Data are shown as mean ± SEM; n = 5. **p < 0.01, ***p < 0.001.