Fig. 5.

Apoptosis via flow cytometry and TUNEL labeling of qHSCs without and with superoxide challenge. A: The cells were stained with annexin-Vcy3 and 7-AAD following 24 h incubation with 1 mM hypoxanthine (HX) ± 2 mU/ml xanthine oxidase (XO), and flow cytometry was performed as described in Experimental Procedures. Normal cells (annexin-Vcy3- and 7-AAD-negative, E3); cells in early apoptosis (annexin-Vcy3-positive and 7-AAD-negative, E4); and cells in late apoptosis/necrosis (annexin-Vcy3- and 7-AAD-positive, E2). The values represent number of cells (%) in the specified phase ± SD.B: TUNEL assay was performed on the qHSCs treated with 1 mM HX± 2mU/mlXOfor 24 h as described in Experimental Procedures. A representative experiment performed in duplicates (two repeats) shows apoptotic nuclei (red). Note that there is no significant difference in the level of apoptosis between the control and superoxide-treated cells. [Color figure can be viewed in the online issue, which is available at www.interscience.wiley.com.]