Article Figures & Data
Figures
- FIGURE 1.
Absence of TRAIL expression in TRAIL gene-targeted mice. Liver MNC were isolated from WT or TRAIL−/− B6 mice and were stained with FITC-conjugated anti-CD3 mAb, PE-conjugated anti-TRAIL mAb, and biotin-conjugated anti-NK1.1 mAb followed by streptavidin-PerCP. A and C, NK1.1/CD3 staining of WT and TRAIL−/− mice, respectively, with R2 representing the gated NK cell population. B, TRAIL expression on WT liver NK cells (R2 gated). D, Absence of TRAIL on TRAIL−/− liver NK cells (R2 gated). Solid lines represent staining with isotype-matched control mAb; dotted lines represent staining with anti-mouse TRAIL mAb. These analyses have been performed on more than three occasions and these profiles are representative.
- FIGURE 2.
TRAIL gene-targeted mice do not exert TRAIL-mediated cytotoxicity. Liver and spleen MNC were isolated from untreated BALB/c WT, BALB/c pfp−/−, BALB/c IFN-γ−/−, or BALB/c TRAIL−/− mice (A and B), and liver MNC were isolated from α-GalCer (2 μg i.p. on days −5 and −1)-treated BALB/c WT or BALB/c TRAIL−/− mice (C). Their cytotoxic activities were tested against TRAIL-sensitive 4T1 (A and C) and Renca (B and C) tumor cells in the presence or absence of 50 nM CMA, 10 μg/ml anti-TRAIL mAb, or 10 μg/ml control rat IgG2a (control) by an 8-h 51Cr release assay at three different E:T ratios (highest, 100:1 for spleen MNC; highest, 50:1 for liver MNC, shown) as indicated. Data are representative of those across the E:T ratio range (100 to 5:1, spleen; 50 to 5:1, liver) and are recorded as the mean ± SE of triplicate samples. Similar results were obtained in two independent experiments.
- FIGURE 3.
TRAIL contributes to NK cell suppression of experimental Renca metastases to the liver. Groups of WT or TRAIL−/− BALB/c mice were inoculated i.s. with Renca tumor cells ranging from 1 × 104–3 × 105 (A), i.s. with 3 × 105 Renca tumor cells (B), or i.v. with 3 × 105 Renca tumor cells (C) on day 0. As indicated, some groups of mice were treated with 200 μg of anti-asGM1 Ab i.p. on days −1, 0, and 7 (A–C), 0.25 mg of anti-TRAIL mAb i.p. on days 0, 1, and 7 (A–C), and 2 μg of α-GalCer i.p. on days 0, 4, and 8 (B and C). The livers (A and B) or lungs (C) were removed from mice on day 14, and the metastatic nodules were quantified as described in Materials and Methods. Data are indicated as the mean ± SE of five mice in each group, with the significance compared with untreated (A) or α-GalCer-treated (B and C) WT mice as defined by the Mann-Whitney U test. ∗, p < 0.01; ∗∗, p < 0.05. α-GalCer was also statistically effective alone compared with no treatment in the livers and lungs of WT mice (p < 0.01). We have previously shown no significant effects of rabbit and mouse control Igs in these models (14 ).
- FIGURE 4.
TRAIL contributes to the suppression of primary 4T1 tumor growth and spontaneous metastasis. A–C, Groups of female BALB/c WT or TRAIL−/− mice were inoculated on day 0 into the abdominal mammary fat pad with 5 × 103 (A), 1 × 104 (B), or 2.5 × 104 (C) 4T1 mammary carcinoma cells. Some groups of mice were treated with 200 μg of anti-asGM1 Ab i.p. on days −1, 0, 7, and 14, or 0.25 mg of anti-TRAIL mAb i.p. on days 0, 1, 4, 7, 10, 14, and 21. Tumor size was measured over the course of 30 days. Data are shown as the mean ± SE of five tumor-bearing mice in each group. D–F, Groups of female BALB/c WT or TRAIL−/− mice were inoculated on day 0 into the abdominal mammary fat pad with 2.5 × 104 4T1 mammary carcinoma cells and were i.p. administered with 2 μg of α-GalCer on days 0, 4, 8, 12, and 16. Some groups of mice were treated with 200 μg of anti-asGM1 Ab i.p. on days −1, 0, 7, and 14 or 0.25 mg of anti-TRAIL mAb i.p. on days 0, 1, 4, 7, 10, 14, and 21. Primary tumor growth in the mammary gland was measured over the course of 30 days (D). Lung metastases (E) and liver metastases (F) were measured as described in Materials and Methods. Data are indicated as the mean ± SE of five mice in each group, with the significance compared with α-GalCer-treated WT mice as defined by the Mann-Whitney U test. ∗, p < 0.01. α-GalCer was also statistically effective alone compared with no treatment in the livers and lungs of WT mice (p < 0.01).
- FIGURE 5.
TRAIL suppresses MCA induction of fibrosarcomas. Groups of 10–30 B6 WT (▪) or TRAIL−/− (□) mice were inoculated s.c. in the hind flank with 400, 100, 25, or 5 μg of MCA in 0.1 ml of corn oil. Development of sarcomas was monitored over the course of 80–160 days, and the percentage of mice with sarcoma was recorded. ∗, Statistically increased sarcoma incidence above that observed in WT (Fisher’s exact test, p < 0.05).
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