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. 2019 Jun 24;11(8):e9266. doi: 10.15252/emmm.201809266

Figure 4. Combining Apelin blockage and sunitinib treatment reduces tumor angiogenesis and normalizes tumor blood vessels.

Figure 4

  • A
    Experimental setup for the MRI (non‐invasive magnetic resonance imaging) experiments.
  • B, C
    (B) Representative MRI images; scale bar = 5 mm and (C) quantification (mean ± SEM) of relative tumor blood volume (rTBV) over time after NeuT+ mammary tumors were size‐matched at 20–70 mm3 (0 weeks). Treatments and genotypes are indicated. NeuT;Apln +/+ control (n = 4), NeuT;Apln −/− control (n = 4), NeuT;Apln +/+ sunitinib (n = 5), NeuT;Apln −/− sunitinib (n = 5); **P < 0.01, and ***P < 0.001 compared to untreated NeuT;Apln +/+ mice and # P < 0.05 compared to untreated control NeuT;Apln −/− mice; two‐way ANOVA. Of note, in NeuT;Apln +/+ mice, only two tumors could be analyzed after 6 weeks as some mice had to be sacrificed due to the large tumor sizes following ethical guidelines. Thus, we did not perform any statistical analysis on the 6 weeks time points.
  • D
    Analysis (mean values ± SEM) of CD31+ area (×104 μm2)/field, number of dilated tumor vessels, and percentage of alphaSMA+ area per CD31+ blood vessels in mammary tumors of untreated (control) and sunitinib‐treated NeuT;Apln +/+ and NeuT;Apln −/− mice, assessed 6 weeks after mammary tumors were size‐matched. NeuT;Apln +/+ control (n = 4), NeuT;Apln −/− control (n = 4), NeuT;Apln +/+ sunitinib (n = 5), NeuT;Apln −/− sunitinib (n = 4); *P < 0.05; **P < 0.01; ***P < 0.001; one‐way ANOVA and Kruskal–Wallis test.
  • E
    Representative immunofluorescence and immunohistochemistry images from Fig 5D quantification. Dilated blood vessels are marked by a red asterisk. DAPI (blue) is shown as a counterstain to visualize nuclei. Scale bars = 100 μm (upper panels), 50 μm (middle panels), and 20 μm (lower panels).