Article Figures & Data
Figures
- Figure 1.
A, p53-deficient H1299 cells were infected with Ad-p53 at 0, 10, 50, and 100 MOI. Expression of exogenous p53 mRNA and p53-targeted gene mRNA (p21, MDM2, Noxa, and p53AIP1) 24 h after Ad-p53 infection in vitro was measured by the real-time quantitative PCR assay. All expression levels are normalized to that of β-globin (HKG) in each well and are expressed as the fold increase for each sample relative to MOCK, which was set at 1. This experiment was repeated twice. Data from one representative experiment are shown. B, expression of exogenous p53 mRNA and p53-targeted gene mRNA (p21, MDM2, Noxa, and p53AIP1) was measured at different time points after Ad-p53 infection (50 MOI) in vitro by the real-time quantitative PCR assay. p53-deficient H1299 cells were infected at 50 MOI of Ad-p53 and harvested at 6, 12, 24, and 30 h after infection. All expression levels are normalized to β-globin (HKG) in each well and are defined as the fold increase for each sample relative to that at 0 h (0 h = 1). This experiment was repeated twice. Data from one representative experiment are shown. C, correlation between phosphorylation at Ser46 of p53 and induction of apoptosis. H1299 and H226Br human NSCLC cells were infected with Ad-p53 at the indicated MOI. Upper panel, phase-contrast photomicrographs. Cell morphology was evaluated 24 h after infection. Note a rapid loss of viability due to massive cell death, as evidenced by floating, highly light refractile cells, in Ad-p53-infected H1299 cells. Lower panel, Western blot analysis with anti-p53 and anti-p53-Ser46 antibodies. Total cell extracts were prepared from MOCK-infected cells or from cells infected with control d1312 or Ad-p53 (indicated MOI) and then analyzed on Western blots.
- Figure 2.
A and B, expression of exogenous p53, p21, MDM2, Noxa, and p53AIP1 mRNA at different time points following intratumoral administration of Ad-p53 (1 × 109 pfu/50 μl) or Ad-LacZ (1 × 109 pfu/50 μl) in nude mice transplanted with H1299 (A) or A549 (B) tumors was measured by real-time quantitative PCR assay. All expression levels are normalized to β-globin (HKG) in each well. Values are relative to day 0 (day 0 = 1). These experiments were repeated twice. Data from one representative experiment are shown. C and D, detection of apoptotic cells in tumor tissue by TUNEL assay at different time points following intratumoral administration of Ad-p53 (1 × 109 pfu/50 μl; C) or Ad-LacZ (1 × 109 pfu/50 μl; D). a, day 0; b, day 1; c, day 2; d, day 3; e, day 7; f, day 14. This experiment was repeated twice. Data from one representative experiment are shown.
- Figure 3.
A, H1299/p53R-GFP cells were infected with Ad-p53 at 0 (a), 10 (b), 50 (c), and 100 (d) MOI, and p53-responsive GFP expression was assessed with fluorescence microscopy and FACS 36 h after infection. B, H1299/p53R-GFP cells were infected with Ad-p53 at 50 MOI, and p53-responsive GFP expression was assessed with fluorescence microscopy at different time points after infection. a, 0 h; b, 12 h; c, 24 h; d, 36 h; e, 48 h; f, 72 h.
- Figure 4.
Exogenous p53 transcriptional activity as intratumoral GFP expression was imaged in real time by 3-CCD camera. The right tumor (arrow) on the mouse's back was injected with Ad-p53 (1 × 109 pfu/50 μl). The left tumor was injected with Ad-LacZ (1 × 109 pfu/50 μl; data not shown). A representative mouse was observed with 3-CCD camera on days 0 [naked (A) and 3-CCD (B)], 1 (C), 2 (D), 3 (E), and 7 (F) after injection.
Volume 3, Issue 1
