Figure 4.

Adenovirus Carrying a Decoy Sequence for miR-15a and -16 (Ad.Luc-Decoy) Sensor Activity and Inhibition Efficiency In Vivo

(A) Luciferase activity measured on an IVIS of HUVECs infected with Ad.Luc control or Ad.Luc-Decoy virus cultured under complete (2% FBS) or serum-deprived (0% FBS) media for 24 h. Results are expressed per cell and relative to Ad.-matching, 2% FBS condition. Results are presented as the mean of n = 4–5 per condition ± SEM. *p < 0.05, **p < 0.01, ***p < 0.001 versus Ad.-matching, 2% FBS condition; #p < 0.05 versus Ad.-Luc, 0% FBS condition, Student’s t test. (B and C) After limb ischemia, bioluminescence activity (expressed as photons per second per centimeter squared per steradian (P⋅s⁻¹⋅cm⁻²⋅sr⁻¹) was measured and analyzed in the ischemic legs using the Xenogen IVIS. D-Luciferin was subcutaneously injected into the ischemic limb, and the consequent bioluminescence from the reaction with the luciferase in either Ad.Luc- or Ad.Luc-Decoy-injected legs was measured from day 1 to day 21 after injection of the viruses. Representative images are presented. Data are shown as mean ± SEM (n ≥ 10 per condition). *p < 0.05 versus time-matched Ad.Luc condition using Student’s t test. (D) Schematic explanation of the mechanisms of miR-15a and -16 inhibition, trapping, and sensing in cells expressing the Luc control construct, lacking complementary-decoy sequence for miR-15a/16 or the Luc-Decoy construct.