Abstract
Fibroblast Growth Factor-2 (FGF2) protects mice from radiation- and LPS-induced lung injury. Here we investigated the effect of FGF2 on DNA damage caused by bleomycin (BLM) in mouse lung endothelial cells (MLEC). MLEC were treated with 0.1-10 ng FGF2/ml for 1 to 4h, and then with 100 or 500 ug BLM/ml for 15-45 min. Cells were either fixed with formaldehyde and permeabilized in 70% ethanol, or fixed with 95% ethanol/1% acetic acid and stored in PBS. In formaldehyde-fixed cells, DNA strand breaks containing 3′OH were labeled with fluorescein-12-dUTP by nick translation with DNA polymerase. Ethanol/acetic acid-fixed cells were immunostained with antibody to the phosphorylated histone (H) 2A variant, H2Ax (phospho-H2Ax), which accumulates at sites of double strand breakage, and a Cy3-secondary antibody. Cells were finally stained with H33342 to mark DNA. Fluorescence microscopic image intensities of fluorescein for 3′OH labeling, or Cy3 for phospho-H2Ax, were analyzed. FGF2 inhibited both measures of strand breakage. The results suggest that FGF2 can protect lung EC from BLM-induced single and/or double strand DNA breakage. Support: NIH HL68054.
Published Version
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