Inhibition of uPAR and MMP-9 induces apoptosis <i>in vivo</i>.

<p>Daoy cells were implanted intracranially in nude mice, which were treated with control pSV or pUM (150 µg) either alone or in combination with radiation as described in Materials and Methods. <b>A–B)</b> Paraffin-embedded brain tumor sections from mice that received control shRNA or pUM either alone and combination of radiation were analyzed by immunohistochmeical analysis using antibodies for <b>A)</b> uPAR and <b>B)</b> MMP-9 followed by treatment with secondary antibody conjugated with Alexa Fluor 594 and 488 fluorescent tagged. <b>C)</b> Induction of apoptosis in pUM-treated tumors was confirmed by carrying out TdT-mediated nick end labeling (TUNEL) assay. The tissues were analyzed for apoptosis using fluorescence microscopy. <b>D)</b> The relative number of TUNEL-positive cells was counted and represented graphically. Bars represent the means ± s.d. from three independent experiments, * <i>p</i><0.05. <b>E)</b> Immunoflourscence analysis of paraffin-embedded brain sections to detect the levels of NF-κB p65 and Stat3 in pUM and pSV treated mice (with and without radiation). After incubating with the anti Rel-A and Stat3 antibodies for overnight, the tissue sections were washed and incubated in secondary antibody conjugated with Alexa Fluor 594 and 488 fluorescent tagged, respectively. Representative images from three independent experiments were shown in each panel.