The NLCs developed can be used as a promising carrier for safer and efficient management of tuberculosis.

eThe establishment of mycobacterial infection is characterized by the formation of granulomas, which are well-organized aggregates of immune cells, namely, infected macrophages. The granuloma's main function is to constrain and prevent dissemination of the mycobacteria while focusing the immune response to a limited area. In some cases these lesions can grow progressively into large granulomas which can undergo central necrosis, thereby leading to their caseation. Macrophages are the most abundant cells present in the granuloma and are known to adapt under hypoxic conditions in order to avoid cell death. Our laboratory has developed a granuloma necrosis model that mimics the human pathology of Mycobacterium tuberculosis, using C57BL/6 mice infected intravenously with a low dose of a highly virulent strain of Mycobacterium avium. In this work, a mouse strain deleted of the hypoxia inducible factor 1␣ (HIF-1␣) under the Cre-lox system regulated by the lysozyme M gene promoter was used to determine the relevance of HIF-1␣ in the caseation of granulomas. The genetic ablation of HIF-1␣ in the myeloid lineage causes the earlier emergence of granuloma necrosis and clearly induces an impairment of the resistance against M. avium infection coincident with the emergence of necrosis. The data provide evidence that granulomas become hypoxic before undergoing necrosis through the analysis of vascularization and quantification of HIF-1␣ in a necrotizing mouse model. Our results show that interfering with macrophage adaptation to hypoxia, such as through HIF-1␣ inactivation, accelerates granuloma necrosis.T he main pathological feature of human tuberculosis is the induction by Mycobacterium tuberculosis of granulomas which undergo central necrosis (caseous necrosis) (1). The mechanisms underlying the latter phenomenon are not clear, but it is generally accepted that caseation plays an essential role in the pathogenesis and dissemination of the infection. Caseous necrosis and cavitation during infection by M. tuberculosis is more prominent in rabbits and guinea pigs than in mice (2). Nevertheless, human-like pathology, as central granuloma necrosis and associated hypoxia, can be obtained in M. tuberculosis-infected I/StSnEgYCit, C3HeB/ FeJ, or IL-13-overexpressing C57BL/6 mouse strains (3-5). Confluent necrotic granulomas also are observed in mice deficient in gamma interferon (IFN-␥) or the type 1 tumor necrosis factor (TNF) receptor (2, 6, 7). However, the fact that immunodeficient human hosts, namely, AIDS patients, tend to present forms of tuberculosis with granulomas lacking necrotic features suggests that necrosis depends on the immune response (8).We have shown that mice infected with a small inoculum of M. avium ATCC 25291 establish a chronic and progressive infection characterized by the formation of discrete granulomas, which gradually increase in size and undergo central necrosis within a few months (9). This requires CD4 ϩ T cells as well as an intact interleukin-12 (IL-12)/IFN-␥ cytokine axis (9) but is independent...

Summary The transcription factor hypoxia‐inducible factor‐1 alpha (HIF‐1α) is a key regulator of the response and function of myeloid cells in hypoxic and inflammatory microenvironments. To define the role of HIF‐1α in tuberculosis, the progression of aerosol Mycobacterium tuberculosis infection was analysed in mice deficient in HIF‐1α in the myeloid lineage (mHIF‐1α−/−). We show that myeloid HIF‐1α is not required for the containment of the infection, as both wild‐type (WT) and mHIF‐1α−/− mice mounted normal Th1 responses and maintained control of bacterial growth throughout infection. However, during chronic infection mHIF‐1α−/− mice developed extensive lymphocytic inflammatory involvement of the interstitial lung tissue and died earlier than WT mice. These data support the hypothesis that HIF‐1α activity coordinates the response of myeloid cells during M. tuberculosis infection to prevent excessive leucocyte recruitment and immunopathological consequences to the host.

IFN-γ is known to be predominantly produced by lymphoid cells such as certain subsets of T cells, NK cells, and other group 1 innate lymphoid cells. In this study, we used IFN-γ reporter mouse models to search for additional cells capable of secreting this cytokine. We identified a novel and rare population of nonconventional IFN-γ-producing cells of hematopoietic origin that were characterized by the expression of Thy1.2 and the lack of lymphoid, myeloid, and NK lineage markers. The expression of IFN-γ by this population was higher in the liver and lower in the spleen. Furthermore, these cells were present in mice lacking both the and the common γ-chain (γc) genes (Rag2γc), indicating their innate nature and their γc cytokine independence. Rag2γc mice are as resistant to as Rag2 mice, whereas Rag2 mice lacking IFN-γ are more susceptible than either or Rag2γc These lineage-negative CD45/Thy1.2 cells are found within the mycobacterially induced granulomatous structure in the livers of infected Rag2γc animals and are adjacent to macrophages that expressed inducible NO synthase, suggesting a potential protective role for these IFN-γ-producing cells. Accordingly, Thy1.2-specific mAb administration to infected Rag2γc animals increased growth in the liver. Overall, our results demonstrate that a population of Thy1.2 non-NK innate-like cells present in the liver expresses IFN-γ and can confer protection against infection in immunocompromised mice.

Since the pioneering discoveries, by the Nobel laureates Jules Hoffmann and Bruce Beutler, that Toll and Toll-like receptors can sense pathogenic microorganisms and initiate, in vertebrates and invertebrates, innate immune responses against microbial infections, many other families of pattern recognition receptors (PRRs) have been described. One of such receptor clusters is composed by, if not all, at least several members of the scavenger receptor cysteine-rich (SRCR) superfamily. Many SRCR proteins are plasma membrane receptors of immune cells; however, a small subset consists of secreted receptors that are therefore in circulation. We here describe the first characterization of biological and functional roles of the circulating human protein SSC4D, one of the least scrutinized members of the family. Within leukocyte populations, SSC4D was found to be expressed by monocytes/macrophages, neutrophils, and B cells, but its production was particularly evident in epithelial cells of several organs and tissues, namely, in the kidney, thyroid, lung, placenta, intestinal tract, and liver. Similar to other SRCR proteins, SSC4D shows the capacity of physically binding to different species of bacteria, and this opsonization can increase the phagocytic capacity of monocytes. Importantly, we have uncovered the capacity of SSC4D of binding to several protozoan parasites, a singular feature seldom described for PRRs in general and here demonstrated for the first time for an SRCR family member. Overall, our study is pioneer in assigning a PRR role to SSC4D.

Granuloma formation is a hallmark of several infectious diseases, including those caused by Mycobacterium sp. These structures are composed of accumulations of inflammatory cells, and it has been shown that cytokines such as IFN-g and TNF-a are required for granuloma assembly during M. avium infections in mice. Macrophages (MFs) insensitive to IFN-g (MIIG) mice have MFs, monocytes, and dendritic cells that are unresponsive to IFN-g. We observed that although IFN-g 2/2 mice present an exacerbated infection, the same is not true for MIIG animals, where the same levels of protection as the wild-type animals were observed in the liver and partial protection in the spleen. Unlike IFN-g 2/2 mice, MIIG mice still develop well-defined granulomas, suggesting that IFN-g-mediated MF activation is not required for granuloma assembly. This work also shows that MIIG animals exhibit increased cell recruitment with higher CD4 + T cells numbers as well as increased IFN-g and TNF-a expression, suggesting that TNF-a may have a role in protection and may compensate the lack of MF response to IFN-g in the MIIG model. TNF-a-deficient MIIG mice (MIIG.TNF-a 2/2) exhibited increased bacterial burdens when compared with MIIG mice. These results suggest that in the absence of IFN-g signaling in MFs, TNF-a has a protective role against M. avium.

The global burden of sepsis, with an estimated 49 million cases and 11 million deaths in 2017, often passes unnoticed to the general public even though it is the direct cause of nearly 20% of all deaths worldwide. This unawareness is perhaps due to misconceptions, or miscoding in the reporting of the ultimate causes of death, as in many diseases it is not the actual infectious agent that causes the biggest harm. Rather, it is the uncontrolled inflammation leading to septic shock that is the most menacing manifestation associated with many infections, and becomes deadly serious once it has passed the stage where anti-microbial drugs no longer have any effect to inactivate or destroy the pathogen. Here we show that the combined anti-bacterial and anti-inflammatory properties of the scavenger receptor cysteine-rich (SRCR) protein CD5L contribute to a remarkable therapeutic effect of the protein to fight sepsis, such that when exogenously administered in C57BL/6 mice with induced lethal-grade sepsis, it can be a very effective curative agent to treat this condition. The resistance conferred by CD5L to polybacterial-induced sepsis using the cecal ligation and puncture (CLP) model is consistent with the reported observations that CD5L physically binds and inactivates diverse species and strains of bacteria. Accordingly, our CD5L-knockout mice are significantly more susceptible to experimentally-induced mid-grade CLP than wild-type animals. We show that CD5L is centered on promoting neutrophil recruitment and activation, overall contributing to reducing the bacteria burden of the animals. However, the dramatic susceptibility of CD5L-deficient animals is not necessarily correlated only with pathogen load, as these mice are also extremely susceptible to sterile sepsis induced by nonlethal doses of LPS. Notwithstanding the observed capacity of CD5L to directly bind to a broad range of pathogens, typical of many PRRs, our evidence suggests that the anti-inflammatory properties of the protein are at least as important as its pathogen-binding potential, and can, and should, be explored to treat the deadly inflammation storm that is sepsis.

T-cell membrane scaffold proteins play important roles in T cell biology, functioning as multi-functional signaling hubs. CD6 assembles a large intracellular signalosome but, unlike typical membrane-attached scaffolds like LAT or PAG, it has a sizeable ectodomain that binds a well-characterized ligand, CD166. It is unclear whether CD6 has net inhibitory or costimulatory functions or how its ectodomain influences these activities. To explore these questions, we dissected the signaling functions of the extracellular and cytoplasmic regions of CD6. We found that CD6 was delivered to the immunological synapse and suppressed T cell responsiveness in vitro wholly dependently of its cytoplasmic domain, indicating that CD6 very potently imposes tonic inhibition, acting as a structural and signaling inhibitory hub. However, the cell-intrinsic suppression of autoimmunity by CD6 in vivo was also impacted by extracellular interactions, demonstrated by the increased susceptibility of mice to experimental autoimmune encephalomyelitis after removal of the ligand binding region of the ectodomain of CD6. Our work identifies CD6 as a new class of ‘on/off switching’ scaffold-receptor that constrains immune responsiveness at two speeds. First, it sets signaling thresholds via tonic inhibition, functioning as a cytoplasmic membrane-bound scaffold and, second, by cycling between signaling-enabling and signalinginhibiting ectodomain isoforms it functions as an immune checkpoint.