Figure 3.

HAECs attenuate DNA damage in LSK cells. (A) LSK cells were FACS-sorted from 580 cGy-irradiated mouse bone marrow and cultured in the presence or absence of HAECs. After 24 h, LSK cells and their progeny were re-sorted based on Sca-1 and CD45 expression, and DNA damage was assessed using γH2AX immunofluorescence and a Comet assay. (B) LSK cells recovered after 24 h, with the + EC group normalized to 100% (n = 9 independent experiments). (C) Re-sorted LSK cells were assessed for DNA double strand breaks using γH2AX immunofluorescence. Representative images of irradiated LSK cells cultured in the absence (top) or presence (bottom) of HAECs (scale bar = 5 μm). (D) Quantification of γH2AX-positive cells (≥5 foci/cell) in LSK cells immediately after irradiation (white bar) and after 24 h culture in the presence (grey bar) or absence (black bar) of HAECs. At least 100 cells were scored per group (combined results from 4 independent experiments). (E) DNA damage in re-sorted LSK cells was also determined using a Comet assay. A representative experiment is shown where >100 LSK cells/group were scored for olive tail moment (n = 3 independent experiments).