TLR9 regulates the mycobacteria-elicited pulmonary granulomatous immune response in mice through DC-derived Notch ligand delta-like 4
Toshihiro Ito, Matthew Schaller, Cory M. Hogaboam, Theodore J. Standiford, Matyas Sandor, Nicholas W. Lukacs, Stephen W. Chensue, Steven L. Kunkel
Toshihiro Ito, Matthew Schaller, Cory M. Hogaboam, Theodore J. Standiford, Matyas Sandor, Nicholas W. Lukacs, Stephen W. Chensue, Steven L. Kunkel
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Research Article Infectious disease

TLR9 regulates the mycobacteria-elicited pulmonary granulomatous immune response in mice through DC-derived Notch ligand delta-like 4

Abstract

TLR9 activation is important for the maintenance of mycobacteria-elicited pulmonary granulomatous responses, hallmarks of protective immune responses following mycobacterial infection. However, the mechanism or mechanisms underlying this effect of TLR9 are not clear. Here, we show that Tlr9-deficient mice challenged with a Mycobacterium antigen display an altered Th17 cytokine profile, decreased accumulation of granuloma-associated myeloid DCs, and profoundly impaired delta-like 4 (dll4) Notch ligand expression. Mechanistic analysis revealed that WT bone marrow–derived DCs but not macrophages promoted the differentiation of Th17 cells from bacillus Calmette-Guérin–challenged (BCG-challenged) lung CD4+ T cells. Both lung and bone marrow DCs isolated from Tlr9-deficient mice inoculated with Mycobacterium antigen expressed lower levels of dll4 Notch ligand than the same cells isolated from WT mice. Passively immunizing WT mice with neutralizing antibodies specific for dll4 during granuloma formation resulted in larger granulomas and lower levels of Th17-related cytokines. In addition, dll4 specifically regulated Th17 activation in vitro. Together, these results suggest dll4 plays an important role in promoting Th17 effector activity during a mycobacterial challenge. Furthermore, TLR9 seems to be required for optimal dll4 expression and the regulation of Mycobacterium antigen–elicited granuloma formation in mice.

Authors

Toshihiro Ito, Matthew Schaller, Cory M. Hogaboam, Theodore J. Standiford, Matyas Sandor, Nicholas W. Lukacs, Stephen W. Chensue, Steven L. Kunkel

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Figure 7

Passive immunization using Abs against dll4 increases granuloma size, abrogates the migration of DCs, and alters the expression of lung Th17-related cytokines and of CCL20.

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Passive immunization using Abs against dll4 increases granuloma size, ab...
(A) Histological appearance of pulmonary granulomas from WT mice treated with either control Ab or Abs directed against dll4 at day 4 after granuloma development. H&E staining. Original magnification, ×40; ×400. Experiments were performed 3 times with similar results. (B) Morphometric analysis of pulmonary granuloma size from mice treated with either control Abs or dll4 Abs after 4 days of lung granuloma development. *P < 0.01 compared with control. (C) FACS profiles for mDCs (CD11b+CD11c+) recovered from granulomatous lung cells isolated from day 4 lesions. †P < 0.03 compared with control. (D) Protein level of IL-17 was measured in whole lungs using a Luminex system. (E) Quantitative real-time PCR was performed on lung mRNA for the cytokines IFN-γ, IL-4, IL-17A, IL-17F, IL-6, and IL-21. *P < 0.01 compared with control. (F) Quantitative real-time PCR was performed on lung mRNA for the indicated chemokines. ‡P < 0.02 compared with control. Data shown are mean ± SEM from 5 mice and are from a representative experiment of 3 independent experiments. Each time point represents 5 mice per group.