Figure 1.

Bach1 overexpression promotes ROS production and apoptosis in the ischemic limbs of mice. Eight-week-old male C57BL/6 mice were subjected to hindlimb ischemia (HLI). GFP, Bach1 adenovirus, or saline (NS) was injected into the adductor and gastrocnemius muscles immediately after surgery. (a) Bach1 inhibited blood flow restoration after HLI. Blood flows of ischemic and nonischemic limb were measured with a laser Doppler imaging system at 0, 7, and 14 days after surgery. Ratios of blood flow from ischemic hindlimb to nonischemic hindlimb are shown (n = 12; ^∗ P < 0.05, ^∗∗ P < 0.01 versus Ad-GFP). ((b) and (c)) ROS level in nonischemic and ischemic muscle tissue on day 7 was determined by an in situ detection of superoxide with dihydroethidium (DHE) fluorescence. Bars, 100 µm (b). DHE-positive cells were quantified as DHE-positive cells/mm². Data from four sections of each mouse muscle tissue are shown in graphics and n = 6 for each group ((c), ^∗ P < 0.05 versus Ad-GFP). (d) Seven days after HLI, cleaved caspase 3 and cyclin D1 protein levels were evaluated in HLI and non-HLI limbs via Western blot (n = 6; ^∗ P < 0.05, ^∗∗ P < 0.01 versus Ad-GFP).