FIGURE 1 from Overcoming Xenoantigen Immunity to Enable Cellular Tracking and Gene Regulation with Immune-competent “NoGlow” Mice

<p>Tolerance to foreign antigens is animal model specific. <b>A</b> and <b>B,</b> Mice expressing GFP and Luc in the pituitary (GH; <i>n</i> = 5), full-body GFP, and Luc (CAG Luc-GFP; <i>n</i> = 5), in FoxP3⁺ cells (Foxp3-GFP; <i>n</i> = 4), or wild-type (WT) C57Bl/6 (<i>n</i> = 4) were vaccinated with adenovirus (Ad) encoding GFP/Luc (2.6 × 10¹⁰ vp/mouse) and anti-Luciferase (A) or anti-GFP (B) in the serum were evaluated by ELISA after 2 weeks. Only CAG Luc-GFP animals, which express both GFP and Luc, display humoral tolerance to both upon vaccination. <i>P</i> values shown are at 1:50 dilution by two-way ANOVA with Tukey multiple comparisons test. <b>C</b> and <b>D,</b> E0771 GFP-Luc cells were implanted into the MFP of female WT C57Bl/6, GH, or CAG Luc-GFP mice (<i>n</i> = 10 each) and subsequently vaccinated with Ad-GFP/Luc or Ad-LacZ as control on day 3 (<i>n</i> = 5/group each). Growth of individual tumors for the duration of the experiment (C) or average tumor volume until time of first euthanasia (D) comparing control or antigen-specific Ad demonstrate that E0771 GFP-Luc cells grow significantly larger in tolerant CAG Luc-GFP mice. <i>P</i> values shown are at day of first euthanasia by two-way ANOVA with Bonferroni multiple comparisons test. <b>E,</b> Female WT (<i>n</i> = 3) or mice expressing human HER2 under the whey acid protein (WAP) promoter (WAP HER2; <i>n</i> = 5) or MMTV Cre–driven CAG promoter (MMTV CAG HER2; <i>n</i> = 3) were vaccinated with Ad-Human Her2 and splenocytes were analyzed for anti-HER2 responses by ELISPOT after 3 weeks. WAP HER2 mice displayed reduced anti-HER2 responses compared with WT animals; however, MMTV CAG HER2 animals produced a limited anti-HER2 T-cell response compared with both WT and WAP HER2. Groups were compared by two-way ANOVA with Tukey multiple comparisons test. <b>F,</b> Doxycycline-inducible GFP animals were given doxycycline (dox)-containing chow at the indicated time points, vaccinated at 9 weeks old, and euthanized after two weeks. <b>G,</b> Serum anti-GFP responses in animals from F reveal that GFP remains highly immunogenic in mice with late (8 weeks old) versus early (4 weeks old/at birth) expression of GFP via doxycycline. <i>P</i> values shown are at 1:50 dilution by two-way ANOVA with Tukey <i>post hoc</i> analysis. All <i>P</i> values represent mean ± SEM.</p>

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