Figure 5 from T Cells Promote Metastasis by Regulating Extracellular Matrix Remodeling following Chemotherapy
<p>Systemic LOX promotes ECM remodeling following PTX chemotherapy. Bone marrow cells were flushed from Cre<sup>+/−</sup>LOX<sup>flox/flox</sup> or their control counterpart Cre<sup>−/−</sup>LOX<sup>flox/flox</sup> mice. The bone marrow cells wer...
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2025
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| Итог: | <p>Systemic LOX promotes ECM remodeling following PTX chemotherapy. Bone marrow cells were flushed from Cre<sup>+/−</sup>LOX<sup>flox/flox</sup> or their control counterpart Cre<sup>−/−</sup>LOX<sup>flox/flox</sup> mice. The bone marrow cells were then implanted into lethally irradiated wild-type BALB/c mice to generate a chimeric mouse model of inducible LOX depletion specifically in bone marrow cells. The chimeric mice (<i>n</i> = 6 mice/group) were treated with tamoxifen for 5 days, and PTX was administered on day 7. After 72 hours, mice were sacrificed and lungs and bone marrow were harvested. <b>A,</b> A schematic of the experimental procedure is presented. LOX levels in bone marrow cells were assessed by Western blot. <b>B,</b> Lung sections were stained with Sirius red to detect both collagen and elastin. Representative images are shown in two magnifications. Arrows, positive staining. Scale bar, 200 μm. Collagen and elastin levels were quantified by calculating the percentage of red pixels per field (<i>n</i> > 4 fields/lung). <b>C–G,</b> Tumor-free, 8- to 10-week-old SCID mice (<i>n</i> = 4–5 mice/group) were treated with PTX or vehicle control. After 72 hours, lungs were removed. Collagen and elastin in lung sections were detected by Sirius staining. Representative images in two magnifications are shown in <b>C</b> (scale bar, 200 μm), and quantification is shown in <b>D</b>. Lung sections were immunostained using antibodies against Collagen I (<b>E</b>), Collagen IV (<b>F</b>), and LOX (<b>G</b>). Nuclei were stained with DAPI (blue). Representative images (scale bar, 50 μm) and quantifications (<i>n</i> > 4 fields/lung) are shown. <b>H</b> and <b>I,</b> Tumor-free, 8- to 10-week-old SCID mice (<i>n</i> = 5–6 mice/group) were treated with PTX or vehicle control. After 72 hours, EMT6-GFP<sup>+</sup> cells (25 × 10<sup>4</sup> cells/mouse) were injected through the tail vein to generate an <i>ex vivo</i> PuMA. After 15 minutes, lungs were perfused, excised, and sectioned. Lung sections were cultured for 1 week and analyzed by FluorVivo Mag system. Representative images are shown in <b>H</b>. Scale bar, 200 μm. The percentage of GFP<sup>+</sup> cells in single-cell suspensions of lung sections was analyzed by flow cytometry (<b>I</b>). Statistical significance was assessed by one-way ANOVA followed by Tukey posttest when more than two groups were analyzed or unpaired two-tailed <i>t</i> test when only two groups were analyzed. Significant <i>P</i> values are shown as *, <i>P</i> < 0.05; **, <i>P</i> < 0.01; ***, <i>P</i> < 0.001 from control or otherwise indicated in the figure.</p> |
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