Diverse immune environments in human lung tuberculosis granulomas assessed by quantitative multiplexed immunofluorescence

Authors: Marta Abengozar-Muela¹, María Villalba Esparza^{1 2 3}, David Garcia-Ros^{1 2 4}, Cindy Estefanía Vásquez⁴, José I Echeveste^{1 2}, Miguel Angel Idoate^{1 2}, Maria D Lozano^{1 2 3}, Ignacio Melero^{2 3 5 6}, Carlos E de Andrea^{7 8 9 10}

Online: https://www.nature.com/articles/s41379-020-0600-6

Issue: Modern Pathology. 2020 Jun 26. doi: 10.1038/s41379-020-0600-6. Online ahead of print.

Abstract

The precise nature of the local immune responses in lung tuberculosis (TB) granulomas requires a comprehensive understanding of their environmental complexities. At its most basic level, a granuloma is a compact, organized immune aggregate of macrophages surrounded by myeloid, B and T cells. We established two complementary multiplex immunolabeling panels to simultaneously evaluate the myeloid and lymphocytic contexture of 14 human lung TB granulomas in formalin-fixed paraffin-embedded tissue samples. We observed diverse CD3+ and CD8+ T-cell and CD20+ B lymphocyte compositions of the granuloma immune environment and a relatively homogeneous distribution of all myeloid cells. We also found significant associations between CD8+ T-cell densities and the myeloid marker CD11b and phagocytic cell marker CD68. In addition, significantly more CD68+ macrophages and CD8+ T cells were found in Mycobacterium tuberculosis-infected granulomas, as detected by Ziehl-Neelsen staining. FOXP3 expression was predominately found in a small subset of CD4+ T cells in different granulomas. As the success or failure of each granuloma is determined by the immune response within that granuloma at a local and not a systemic level, we attempted to identify the presence of reactive T cells based on expression of the T-cell activation marker CD137 (4-1BB) and programmed cell death-1 (PD-1). Only a small fraction of the CD4+ and CD8+ T cells expressed PD-1. CD137 expression was found only in a very small fraction of the CD4+ T cells in two granulomas. Our results also showed that multinucleated giant cells showed strong PD-L1 but not CTLA-4 membrane staining. This study offers new insights into the heterogeneity of immune cell infiltration in lung TB granulomas, suggesting that each TB granuloma represents a unique immune environment that might be independently influenced by the local adaptive immune response, bacterial state, and overall host disease status.

AUTHOR AFFILIATIONS

1. Department of Pathology, Clínica Universidad de Navarra, University of Navarra, Pamplona, Spain.
2. Navarra Institute for Health Research (IDISNA), Pamplona, Spain.
3. Centro de Investigación Biomédica en Red de Cáncer (CIBERONC), Madrid, Spain.
4. Department of Anatomy, Physiology and Pathology, University of Navarra, Pamplona, Spain.
5. Department of Immunology and Immunotherapy, Clínica Universidad de Navarra, Pamplona, Spain.
6. Program of Immunology and Immunotherapy, Cima Universidad de Navarra, Pamplona, Spain.
7. Department of Pathology, Clínica Universidad de Navarra, University of Navarra, Pamplona, Spain.
8. Navarra Institute for Health Research (IDISNA), Pamplona, Spain.
9. Centro de Investigación Biomédica en Red de Cáncer (CIBERONC), Madrid, Spain.
10. Department of Anatomy, Physiology and Pathology, University of Navarra, Pamplona, Spain.