Fig. 6: iMoP – a circulating monocyte precursor in M.tb infection.
From: Monocyte progenitors give rise to multinucleated giant cells
a, c Frequency of cMoP and iMoP in bone marrow (BM) and blood of mice infected via aerosol with M.tb (H37Rv, 200 CFU) analyzed by FACS 7, 14 and 21 days post infection (p.i.). Graphs show mean ± SD of n = 8–9 mice (uninfected controls), n = 8–10 mice (day 14, day 21) in two experiments and n = 4–5 mice (day 7) in one experiment. Detailed numbers of mice (n) for each condition/time point are presented in Supplementary Table 5. *p = 0.0125, **p = 0.0017 (iMoP in BM), **p = 0.0022 (iMoP in blood, D14), **p = 0.0036 (iMoP in blood, D21), ****p < 0.0001 (refers to all other time points of the respective mouse line), (two-way ANOVA, Tukey’s MCT). Two outliers were removed from iMoP in blood at day 14 C57BL/6 (2.49) and 129S2 (8.50), calculated by Grubbs test. b Flow cytometry gating of iMoP from BM of Cx3cr1gfp/+ mice. d Hemacolor staining of murine BM precursors immediately after cell sorting. Scale bar: 50 µm. Gamma-correction was adjusted (0.53) in all images. e MGC formation of cMoP, iMoP (red) and MC stimulated with BCG (MOI 20), heat-fixed BCG (106/ml) or heat-fixed M.tb (10 µg/ml) for 6 days. Bars show mean ± SEM of n = 5 (M.tb, cMoP-BCG) or n = 6 (control, BCG hf, iMoP-BCG, MC-BCG) biologically independent samples examined over three experiments. ns = not significant (p = 0.7241), ***p = 0.0001 (M.tb: cMoP – iMoP), ***p = 0.0009 (M.tb: MC – iMoP), ***p = 0.0004 (BCG hf: MC – iMoP), ****p < 0.0001 (two-way ANOVA, Tukey’s MCT). f TNFα production of BM precursors from Cx3cr1gfp/+ mice after stimulation with heat-fixed BCG (106/ml) for 5 h determined by FACS compared to M-CSF controls. Depicted are mean ± SEM of n = 6 independent biological samples examined over two experiments. ****p < 0.0001 (two-way ANOVA, Sidak’s MCT). g M.tb aerosol infection (H37Rv, 200 CFU) of 129S2 mice, followed by intraperitonal injection of a CCR2-specific antibody (MC-21, 20 µg daily) or isotype control (MC-67), starting on day 14 p.i.. FACS analysis of progenitor frequency in blood and BM at day 19 p.i.. Graphs show mean ± SD of n = 10 mice per group examined over two experiments. ****p < 0.0001 (student’s unpaired t-test, two-tailed).
