Figure 6.
Figure 6. Preservation of lung structure after pulmonary BCG infection upon constitutive or myeloid-specific Ifnγr1 overexpression. (A) Representative macroscopic photos of lungs isolated from mice 9 weeks after infection (positive control, mouse that received WT cells; negative control, mouse that received Lv.GFP-transduced Ifnγr1−/− cells). (B) Representative light sheet microscopy photos of post caval lung lobes depicting the lung structure (LS; green) and infiltration of DsRed-labeled BCG (red) (positive control, mouse that received WT cells; negative control, nontransplanted Ifnγr1−/− mouse). Scale bars represent 500 μm. (C) Quantification of light sheet microscopy depicting the percentage of infected volume of the total lung volume. Mean and SEM are shown for 1 representative experiment (positive control, 9 mice that received WT cells; negative control, 13 nontransplanted Ifnγr1−/− mice; Lv.SFFV.Ifnγr1, n = 10; Lv.MSP.Ifnγr1, n = 9; one-way ANOVA using Tukey’s multiple comparisons post hoc testing). (D) BCG CFU counts of lung homogenates 9 weeks after infection. Mean and SEM (error bars) are shown for 2 independent experiments (positive control, 6 mice that received WT cells; negative control, 5 mice that received Lv.GFP-transduced Ifnγr1−/− cells and nontransplanted Ifnγr1−/− mice: Lv.SFFV.Ifnγr1, n = 4; Lv.MSP.Ifnγr1, n = 5; one-way ANOVA using Dunnett’s multiple comparisons post hoc testing). (E) Representative hematoxylin and eosin–stained histologic sections of lungs isolated 9 weeks after infection are shown for (i-ii) Lv.MSP.Ifnγr1 mouse and (iii-iv) negative control (mouse that received Lv.GFP-transduced Ifnγr1−/− cells). (i) Macrophages and multinucleated cells within the alveoli indicated by black arrows (original magnification ×200). (ii) Higher magnification of black box indicated in (i). (iii) Granuloma indicated by asterisk and macrophages and neutrophils surrounded by collagen fibers (dotted arrow) (original magnification ×200). (iv) Higher magnification photo of giant cells (black arrows). (F) Representative GFP-stained histologic lung sections isolated 9 weeks after infection are shown for (i-ii) Lv.MSP.Ifnγr1 mouse and (iii-iv) negative control mouse that received Lv.GFP-transduced Ifnγr1−/− cells. (i) GFP+ cells (brown signals) are distributed equally throughout Lv.MSP.Ifnγr1 mouse lung tissue and give rise to macrophages (black arrows; ×200). (ii) Higher magnification (×600) of GFP+ intra-alveolar macrophages. (iii) GFP+ cells are numerous in granulomas of negative control mice (*) and macrophages and neutrophils (black arrows) surrounded by collagen fibers (dotted arrow; ×200). (iv) Higher magnification (×600) photo of clustered GFP+ cells (black arrows). (G) Ziehl-Neelson staining of histologic lung sections indicating intracellular acid-fast bacteria (black arrows) (original magnification ×600 for i-ii). *P ≤ .05; **P ≤ .01; ****P ≤ .0001.

Preservation of lung structure after pulmonary BCG infection upon constitutive or myeloid-specific Ifnγr1 overexpression. (A) Representative macroscopic photos of lungs isolated from mice 9 weeks after infection (positive control, mouse that received WT cells; negative control, mouse that received Lv.GFP-transduced Ifnγr1−/− cells). (B) Representative light sheet microscopy photos of post caval lung lobes depicting the lung structure (LS; green) and infiltration of DsRed-labeled BCG (red) (positive control, mouse that received WT cells; negative control, nontransplanted Ifnγr1−/− mouse). Scale bars represent 500 μm. (C) Quantification of light sheet microscopy depicting the percentage of infected volume of the total lung volume. Mean and SEM are shown for 1 representative experiment (positive control, 9 mice that received WT cells; negative control, 13 nontransplanted Ifnγr1−/− mice; Lv.SFFV.Ifnγr1, n = 10; Lv.MSP.Ifnγr1, n = 9; one-way ANOVA using Tukey’s multiple comparisons post hoc testing). (D) BCG CFU counts of lung homogenates 9 weeks after infection. Mean and SEM (error bars) are shown for 2 independent experiments (positive control, 6 mice that received WT cells; negative control, 5 mice that received Lv.GFP-transduced Ifnγr1−/− cells and nontransplanted Ifnγr1−/− mice: Lv.SFFV.Ifnγr1, n = 4; Lv.MSP.Ifnγr1, n = 5; one-way ANOVA using Dunnett’s multiple comparisons post hoc testing). (E) Representative hematoxylin and eosin–stained histologic sections of lungs isolated 9 weeks after infection are shown for (i-ii) Lv.MSP.Ifnγr1 mouse and (iii-iv) negative control (mouse that received Lv.GFP-transduced Ifnγr1−/− cells). (i) Macrophages and multinucleated cells within the alveoli indicated by black arrows (original magnification ×200). (ii) Higher magnification of black box indicated in (i). (iii) Granuloma indicated by asterisk and macrophages and neutrophils surrounded by collagen fibers (dotted arrow) (original magnification ×200). (iv) Higher magnification photo of giant cells (black arrows). (F) Representative GFP-stained histologic lung sections isolated 9 weeks after infection are shown for (i-ii) Lv.MSP.Ifnγr1 mouse and (iii-iv) negative control mouse that received Lv.GFP-transduced Ifnγr1−/− cells. (i) GFP+ cells (brown signals) are distributed equally throughout Lv.MSP.Ifnγr1 mouse lung tissue and give rise to macrophages (black arrows; ×200). (ii) Higher magnification (×600) of GFP+ intra-alveolar macrophages. (iii) GFP+ cells are numerous in granulomas of negative control mice (*) and macrophages and neutrophils (black arrows) surrounded by collagen fibers (dotted arrow; ×200). (iv) Higher magnification (×600) photo of clustered GFP+ cells (black arrows). (G) Ziehl-Neelson staining of histologic lung sections indicating intracellular acid-fast bacteria (black arrows) (original magnification ×600 for i-ii). *P ≤ .05; **P ≤ .01; ****P ≤ .0001.

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