Interestingly, animals ZK38 and ZL43 which experienced no weight loss, the lowest PET hot signals and the largest CD4 T cell reactions in the airways also experienced the largest CD4 T cell reactions in individual granulomas. Mtb-specific CD4 T cells simultaneously appeared in the airways and blood ~21C28 days post-exposure, indicating that recently primed effectors are quickly recruited into the lungs after entering blood circulation. Mtb-specific CD4 T cells in granulomas display a tissue-parenchymal CXCR3+CX3CR1?PD-1hiCTLA-4+ phenotype. However, most granuloma CD4 T cells are found within the outer lymphocyte cuff, and few localize to the myeloid cell core comprising the bacilli. Using the intravascular stain approach, we find essentially all Mtb-specific CD4 T cells in granulomas have extravasated across the vascular endothelium into the parenchyma. Consequently, it is unlikely that lung-homing defects launched by terminal differentiation limit the migration of CD4 Mizoribine T cells into granulomas following primary Mtb illness of macaques. However, intralesional placing defects within the granuloma may present a major barrier to T cell-mediated immunity during tuberculosis. INTRODUCTION CD4 T cells are critical for control of illness.1C4 In order to mediate safety, CD4 T cells must recognize MHC class II on the surface of infected macrophages within the lung cells and deliver signals that instruct macrophages to restrict growth of their ingested bacilli through direct cell-to-cell relationships.5 Therefore, the ability of Mtb-specific CD4 T cells to migrate into the site of infection and interact with infected antigen showing cells is key to protection during Mtb infection. In mice, IL-12 and T-bet dependent CD4 T cell differentiation produces unique subsets of Mtb-specific Th1 cells with varying levels of protecting capacity after main illness. CXCR3+ Th1 cells that communicate intermediate levels of T-bet are able to efficiently migrate out of the blood vasculature into the lung, increase and may adoptively transfer safety to infected T cell deficient recipient mice.6C9 In contrast, T cells that undergo extensive Th1 differentiation become CX3CR1+KLRGl+T-bethigh terminal effector cells that cannot increase, poorly exit the pulmonary vasculature into the tissue parenchyma and don’t adoptively transfer protection. Although T-bet manifestation in CD4 T cells is required for IFN production and sponsor safety,9, 10 T-bet haploinsufficient mice do not generate KLRG1+ CD4 T cells and are more resistant to Mtb illness compared to WT mice.11 Based on these observations in the murine model of Mtb infection, there is a hypothesis the differentiation state of CD4 T cells is a major determinant of their protective capacity against Mtb infection, and vaccination should aim to selectively promote the generation of less-differentiated CD4 T cells.12 Indeed, it has been found in mice that vaccine strategies that generate memory space T cell populations that can resist terminal effector cell formation upon Mtb challenge are more protective.13C15 It is not known, however, if the generation of Ag-specific non-protective terminal effector cells happens in other species following Mtb infection or to what extent defects in CD4 T cell migration into the lung due to terminal differentiation limits the overall protective quality of the Mtb-specific effector cell population. After CD4 T cells extravasate across lung blood vascular endothelium, there is a subsequent phase of migration within the EMR2 cells as CD4 T cells locate Mtb infected macrophages. In mice, there is relatively little structure to the organization of immune cells that cluster around sites of bacterial replication, and true human-like granulomas do not form, at least not in Mizoribine the most commonly used inbred mouse strains. Consequently, the mouse model is not ideal for the study of intralesional leukocyte placing and trafficking, and little is known about this aspect of T cell function during tuberculosis in higher mammals. In contrast to mice, Mtb infected nonhuman primates form complex, human-like granulomas with reproducibly identifiable and clearly demarcated cellular strata with unique inflammatory microenvironments.16, 17 Although tuberculosis granulomas can be classified into many subtypes with very different outcomes, in general, CD3+ cells are abundant in a cuff circumscribing a central macrophage rich region where the bacteria reside, and very few lymphocytes are immediately proximal to infected macrophages.18C20 Therefore, both transendothelial diapedesis as well as the intalesional placement of effector CD4 T cell are potential points of failure in T cell trafficking in the setting of tuberculosis. Here we examine the differentiation state of Mtb-specific effector CD4 T cells generated after low-dose Mtb illness of rhesus macaques. We Mizoribine find that rhesus macaque CD4 T cells do not undergo terminal differentiation during the clonal growth phase and display markers of lung tissue-parenchymal cells following pulmonary Mtb illness. Using the intravascular staining technique, we display that CX3CR1+ CD4 T cells found in uninfected Mizoribine rhesus macaques are localized to the lung-associated blood vasculature, indicating that the poor lung-homing of.