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. 2023 Dec 19;14:8423. doi: 10.1038/s41467-023-44152-8

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© The Author(s) 2023

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 7 — a–f Wildtype C57BL/6 mice at four weeks post-infection treated with 100 μg α-OX40 agonist or isotype control antibody injections twice weekly for two weeks. Antibiotic mice also treated from four weeks post-infection to harvest with isoniazid and rifampin in water continuously. Mice harvested at 0, 13, 42, and 76 days post-treatment for flow cytometry of lung bacterial load. b Lung bacterial load of α-OX40 antibody or isotype control antibody, with or without antibiotics. Pretreatment wildtype lungs harvested at 0 days post-treatment. Data representative of two repeat experiments. Two-way ANOVA results: antibiotic treatment contributed 43% of variance p < 0.0001, antibody treatment contributed 30% of variance p < 0.0001, interaction contributed 13% of variance p = 0.0015; p values shown in figure calculated with two-tailed t testing adjusted for multiple comparisons. Mice per group: 4 for pre-treatment, 5 for no treatment, 5 for no antibiotics + Isotype, 9 for no antibiotics + OX40, 5 for antibiotics + Isotype, 9 for antibiotics + OX40. c, d CFUs and number of CD4 T cells in the lungs of α-OX40 antibody or isotype control antibody treated mice. Gated on live CD4⁺ CD44⁺ cells. Cell numbers assessed using counting beads. P values calculated with an unpaired two tailed t-test, adjusted for multiple comparisons. Mice per group: 5 for isotype and 10 for OX40 at day 13, 5 each for isotype and OX40 at day 42, and 6 each for isotype and OX40 at day. e, f Frequency of terminally differentiated CD4 T cells in the lungs of α-OX40 antibody or isotype control antibody, with or without antibiotics. Gated on live CD4⁺ CD44⁺ CX3CR1⁺ KLRG1⁺ cells. Two-way ANOVA results: antibiotic treatment contributed 39% of variance p = 0.0005, antibody treatment contributed 27% of variance p = 0.0005, interaction contributed 1% of variance p = 0.43; p values shown in figure calculated with two-tailed t testing adjusted for multiple comparisons, 5 mice per group. Error bars indicate standard deviation.