Diversity of Human and Macaque Airway Immune Cells at Baseline and during Tuberculosis Infection

Abstract

Immune cells of the distal airways serve as "first responders" of host immunity to the airborne pathogen Mycobacterium tuberculosis (Mtb). Mtb infection of cynomolgus macaques recapitulates the range of human outcomes from clinically silent latent tuberculosis infection (LTBI) to active tuberculosis of various degrees of severity. To further advance the application of this model to human studies, we compared profiles of bronchoalveolar lavage (BAL) cells of humans and cynomolgus macaques before and after Mtb infection. A simple gating strategy effectively defined BAL T-cell and phagocyte populations in both species. BAL from Mtb-naive humans and macaques showed similar differential cell counts. BAL T cells of macaques were composed of fewer CD4⁺cells but more CD8⁺ and CD4⁺CD8⁺ double-positive cells than were BAL T cells of humans. The most common mononuclear phagocyte population in BAL of both species displayed coexpression of HLA-DR, CD206, CD11b, and CD11c; however, multiple phagocyte subsets displaying only some of these markers were observed as well. Macaques with LTBI displayed a marked BAL lymphocytosis that was not observed in humans with LTBI. In macaques, the prevalence of specific mononuclear phagocyte subsets in baseline BAL correlated with ultimate outcomes of Mtb infection (i.e., LTBI versus active disease). Overall, these findings demonstrate the comparability of studies of pulmonary immunity to Mtb in humans and macaques. They also indicate a previously undescribed complexity of airway mononuclear phagocyte populations that suggests further lines of investigation relevant to understanding the mechanisms of both protection from and susceptibility to the development of active tuberculosis within the lung.

Keywords: alveolar macrophages; bronchoalveolar lavage; human subjects; macaques; tuberculosis.

Figure 1.

Total cell counts observed in bronchoalveolar lavage (BAL) of Mycobacterium tuberculosis (Mtb)–naive macaques (n = 21), macaques with latent tuberculosis infection (LTBI) (n = 9), Mtb-naive human subjects (n = 10), and humans with LTBI (n = 11). More cells were obtained from BAL of humans than of macaques (****P < 0.001, two-way analysis of variance with Sidak’s multiple comparison), consistent with differences in the sizes of the hosts as well as the volume of fluid instilled. Differences in BAL cell yields were not significant (N.S.) among Mtb-naive versus LTBI groups within either species. Squares and circles represent cell counts obtained from individual macaques or human subjects, respectively; horizontal lines indicate median values for each group.

Figure 2.

BAL differential cell counts of Mtb-naive humans and macaques are similar but differ substantially during LTBI. As detailed in the text, differential cell counts of (A) Mtb-naive humans and (B) macaques are both dominated by mononuclear phagocytes (dark gray) with limited percentages of lymphocytes (black) and fewer neutrophils (white) or eosinophils (light gray). (C) Human subjects with LTBI continue to display this same profile. (D) In contrast, lymphocytes comprise 33% of BAL cells in macaques with LTBI, representing a marked increase from naive animals; BAL neutrophils are increased in macaques with LTBI as well.

Figure 3.

Heterogeneous populations of BAL phagocytes exist in Mtb-naive and LTBI populations in both humans and macaques. The prevalence of BAL phagocytes defined by coexpression of receptors CD206, HLA-DR, CD11b, and CD11c is illustrated for human subjects (n = 10 Mtb-naive and n = 11 LTBI) and cynomolgus macaques (n = 16 Mtb-naive and n = 8 LTBI). As detailed in the text, the “4⁺” population is less common in LTBI BAL than in Mtb-naive BAL from both species (but approaches significance only for human subjects). The prevalence of the “3⁺/CD11c⁻” population is decreased significantly in macaques with LTBI compared with Mtb-naive animals, whereas no significant differences in the prevalence of any of the “3⁺” combinations were observed between Mtb-naive human subjects and humans subjects with LTBI. (^§P = 0.06, *P < 0.05, Mann–Whitney.) Results for individual subjects as well as median values are displayed for each group. CD, cluster of differentiation; HLA-DR, human leukocyte antigen-antigen D related.

Figure 4.

Prevalence of BAL phagocyte subsets in Mtb-naive cynomolgus macaques correlates with subsequent outcome of Mtb infection. In preinfection BAL phagocytes of macaques that ultimately developed active tuberculosis (n = 11), the “4⁺” subpopulation coexpressing CD206, CD11b, CD11c, and HLA-DR was less common compared with those of macaques that would develop LTBI. In contrast, the “3⁺/CD11c⁻” population was more prevalent in animals that would later develop LTBI (n = 5) compared with active tuberculosis. (P = 0.07, *P < 0.05, Mann–Whitney). Results for individual macaques as well as median values are displayed.