Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2020 Apr 27;221(10):1636-1646.
doi: 10.1093/infdis/jiz663.

Mycobacterium tuberculosis HN878 Infection Induces Human-Like B-Cell Follicles in Mice

José Alberto Choreño-Parra  1   2 , Suhas Bobba  1 , Javier Rangel-Moreno  3 , Mushtaq Ahmed  1 , Smriti Mehra  4   5   6 , Bruce Rosa  7 , John Martin  7 , Makedonka Mitreva  7 , Deepak Kaushal  4   8   9 , Joaquín Zúñiga  10 , Shabaana A Khader  1
Affiliations

Mycobacterium tuberculosis HN878 Infection Induces Human-Like B-Cell Follicles in Mice

José Alberto Choreño-Parra et al. J Infect Dis. .

Abstract

Specific spatial organization of granulomas within the lungs is crucial for protective anti-tuberculosis (TB) immune responses. However, only large animal models such as macaques are thought to reproduce the morphological hallmarks of human TB granulomas. In this study, we show that infection of mice with clinical "hypervirulent" Mycobacterium tuberculosis (Mtb) HN878 induces human-like granulomas composed of bacilli-loaded macrophages surrounded by lymphocytes and organized localization of germinal centers and B-cell follicles. Infection with laboratory-adapted Mtb H37Rv resulted in granulomas that are characterized by unorganized clusters of macrophages scattered between lymphocytes. An in-depth exploration of the functions of B cells within these follicles suggested diverse roles and the activation of signaling pathways associated with antigen presentation and immune cell recruitment. These findings support the use of clinical Mtb HN878 strain for infection in mice as an appropriate model to study immune parameters associated with human TB granulomas.

Keywords: Mycobacterium tuberculosis HN878; B-cell follicles; human granulomatous diseases; lung granuloma; pulmonary tuberculosis.

PubMed Disclaimer

Figures

Figure 1.
Figure 1.
Mycobacterium tuberculosis (Mtb) HN878 infection induces human-like TB granulomas in lungs of mice. (A and B) C57BL/6 mice were aerosol infected with ~100 colony-forming units of Mtb H37Rv (Rv) or Mtb HN878 (HN) and sacrificed at 50, 100, and 150 days postinfection (dpi). Formalin-fixed and paraffin-embedded (FFPE) lung sections were stained with hematoxylin and eosin, and samples were histologically assessed. (A) The Rv infection is representative at 50 dpi. (B) The HN infection is representative of 50 dpi. (C) Human biopsy specimen obtained from pulmonary tuberculosis (TB) patient (left panel) and nonhuman primates with pulmonary TB that were infected with Mtb Erdman (right panel, 13 weeks postinfection). (D) The number of human-like granulomas per mouse lung lobe was determined histologically in FFPE lung sections. Area occupied by human-like granulomas quantified using the morphometric tool of the Zeiss Axioplan microscope. Percentage from lung area occupied by human-like granulomas per mouse lung lobe was determined. The data shown represent mean (±standard deviation) values from 2 to 3 independent experiments per time point (n = 4–5 mice per group in each experiment). Student t test was used to determine differences per time point. *, P < .05.
Figure 2.
Figure 2.
HN878 infection induces robust B-cell lymphoid follicle formation within human-like granulomas. C57BL/6 mice were infected with Mycobacterium tuberculosis (Mtb) H37Rv (Rv) or Mtb HN878 (HN) as described in Figure 1. (A) B-cell lymphoid follicles were stained using antibodies to CD3 and B220, and Mtb localization with inducible nitric oxide synthase (iNOS)-expressing macrophages was also stained. (B) Expression of germinal center follicles within granulomas was assessed by B220/PCNA/PNA staining. (C) At 50 days postinfection, the total number of B-cell follicles, the size of each individual B-cell follicle, and the percentage of the lung area occupied by B-cell follicles per lobe were quantified using the morphometric tool of the Zeiss Axioplan microscope. The data represented are mean (±standard deviation) values from 2 to 3 independent experiments (n = 5 mice in each HN878 experiment, n = 3–4 mice in H37Rv experiments). Student t test was used to determine differences between the groups. *, P < .05; ***, P < .0001.
Figure 3.
Figure 3.
HN878 infection induces differential expression of cytokines and chemokines around lung granulomas. C57BL/6 mice were infected with Mycobacterium tuberculosis (Mtb) H37Rv (Rv) or Mtb HN878 (HN) as described in Figure 1. (A) The expression of CXCL13, (B) STAT-1, (C) CCL2, and (D) interferon-β (IFN-B) was determined by in situ hybridization of messenger ribonucleic acid on formalin-fixed and paraffin-embedded slides.
Figure 4.
Figure 4.
B-cell deficient mice show increased susceptibility to HN878 infection. C57BL/6 (B6) and Ighm−/− mice were aerosol infected with ~100 colony-forming units (CFUs) of Mycobacterium tuberculosis (Mtb) H37RV or Mtb HN878. (A) Bacterial burden in the lung of B6 and Ighm−/− HN-infected animals was determined at different days postinfection (dpi) after plating and counting CFUs and used as a measure of susceptibility to HN infection between B6 and Ighm−/− mice. (B) The percentage of inflamed lung area was determined histologically in formalin-fixed and paraffin-embedded lung sections. Area occupied inflammation was quantified using the Nanozoomer software. Percentage from lung area occupied by inflammation per mouse lung was determined. (C) Bacterial burden in the lung of B6 and Ighm−/− Rv-infected animals was determined at 50 dpi after plating and counting CFUs and used as a measure of susceptibility to Rv infection between B6 and Ighm−/− mice. (D) At 50 dpi, the number of T cells around granulomas in B6 and Ighm−/− mice infected with Rv or HN was quantified in a ×200 field of view using the morphometric tool of the Zeiss Axioplan microscope after staining using antibodies to CD3 and CXCR5. (A–D) The data shown represent mean (±standard deviation) values from 1 to 3 independent experiments per time point (n = 4–5 mice per group in each experiment). Student t test was used to determine differences per time point. *, P < .05; **, P < .005; ***, P < .0005.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Global Tuberculosis Report 2019. World Health Organization 2019; https://www.who.int/tb/publications/global_report/en/
    1. Tsai MC, Chakravarty S, Zhu G, et al. . Characterization of the tuberculous granuloma in murine and human lungs: cellular composition and relative tissue oxygen tension. Cell Microbiol 2006; 8:218–32. - PubMed
    1. Silva Miranda M, Breiman A, Allain S, Deknuydt F, Altare F. The tuberculous granuloma: an unsuccessful host defence mechanism providing a safety shelter for the bacteria? Clin Dev Immunol 2012; 2012:139127. - PMC - PubMed
    1. Lenaerts A, Barry CE 3rd, Dartois V. Heterogeneity in tuberculosis pathology, microenvironments and therapeutic responses. Immunol Rev 2015; 264:288–307. - PMC - PubMed
    1. Lin PL, Ford CB, Coleman MT, et al. . Sterilization of granulomas is common in active and latent tuberculosis despite within-host variability in bacterial killing. Nat Med 2014; 20:75–9. - PMC - PubMed

Publication types

Substances