Casting a wider net: Immunosurveillance by nonclassical MHC molecules.

Most studies of T lymphocytes focus on recognition of classical major histocompatibility complex (MHC) class I or II molecules presenting oligopeptides, yet there are numerous variations and exceptions of biological significance based on recognition of a wide variety of nonclassical MHC molecules. These include αβ and γδ T cells that recognize different class Ib molecules (CD1, MR-1, HLA-E, G, F, et al.) that are nearly monomorphic within a given species.

"The Impact of Mycobacterium tuberculosis Immune Evasion on Protective Immunity: Implications for TB Vaccine Design" - Meeting report.

Tuberculosis (TB) is the major cause of death from infectious diseases around the world, particularly in HIV infected individuals. TB vaccine design and development have been focused on improving Bacille Calmette-Guérin (BCG) and evaluating recombinant and viral vector expressed Mycobacterium tuberculosis (Mtb) proteins, for boosting BCG-primed immunity, but these approaches have not yet yielded significant improvements over the modest effects of BCG in protecting against infection or disease. On March 7-8, 2016, the National Institute of Allergy and Infectious Diseases (NIAID) convened a workshop on "The Impact of Mtb Immune Evasion on Protective Immunity: Implications for TB Vaccine Design" with the goal of defining immune mechanisms that could be targeted through novel research approaches, to inform vaccine design and immune therapeutic interventions for prevention of TB.

Reduced frequency of a CD14+ CD16+ monocyte subset with high Toll-like receptor 4 expression in cord blood compared to adult blood contributes to lipopolysaccharide hyporesponsiveness in newborns.

The human innate immune response to pathogens is not fully effective and mature until well into childhood, as exemplified by various responses to Toll-like receptor (TLR) agonists in newborns compared to adults. To better understand the mechanistic basis for this age-related difference in innate immunity, we compared tumor necrosis factor alpha (TNF-α) production by monocytes from cord blood (CB) and adult blood (AB) in response to LAM (lipoarabinomannan from Mycobacterium tuberculosis, a TLR2 ligand) and LPS (lipopolysaccharide from Escherichia coli, a TLR4 ligand). LPS or LAM-induced TNF-α production was 5 to 18 times higher in AB than in CB monocytes, whereas interleukin-1α (IL-1α) stimulated similar levels of TNF-α in both groups, suggesting that decreased responses to LPS or LAM in CB are unlikely to be due to differences in the MyD88-dependent signaling pathway.

Human phagosome processing of Mycobacterium tuberculosis antigens is modulated by interferon-γ and interleukin-10.

Intracellular pathogens, such as Mycobacterium tuberculosis, reside in the phagosomes of macrophages where antigenic processing is initiated. Mycobacterial antigen-MHC class II complexes are formed within the phagosome and are then trafficked to the cell surface. Interferon-γ (IFN-γ) and interleukin-10 (IL-10) influence the outcome of M.

Decline of influenza-specific CD8+ T cell repertoire in healthy geriatric donors.

Background: While influenza vaccination results in protective antibodies against primary infections, clearance of infection is primarily mediated through CD8+ T cells. Studying the CD8+ T cell response to influenza epitopes is crucial in understanding the disease associated morbidity and mortality especially in at risk populations such as the elderly. We compared the CD8+ T cell response to immunodominant and subdominant influenza epitopes in HLA-A2+ control, adult donors, aged 21-42, and in geriatric donors, aged 65 and older.

Mycobacterium tuberculosis synergizes with ATP to induce release of microvesicles and exosomes containing major histocompatibility complex class II molecules capable of antigen presentation.

Major histocompatibility complex class II (MHC-II) molecules are released by murine macrophages upon lipopolysaccharide (LPS) stimulation and ATP signaling through the P2X7 receptor. These studies show that infection of macrophages with Mycobacterium tuberculosis or M. bovis strain BCG enhances MHC-II release in synergy with ATP.

Preserved MHC class II antigen processing in monocytes from HIV-infected individuals.

Background: MHC-II restricted CD4+ T cells are dependent on antigen presenting cells (APC) for their activation. APC dysfunction in HIV-infected individuals could accelerate or exacerbate CD4+ T cell dysfunction and may contribute to increased levels of immunodeficiency seen in some patients regardless of their CD4+ T cell numbers. Here we test the hypothesis that APC from HIV-infected individuals have diminished antigen processing and presentation capacity.

Influenza-induced production of interferon-alpha is defective in geriatric individuals.

Background: The majority of deaths (90%) attributed to influenza are in person's age 65 or older. Little is known about whether defects in innate immune responses in geriatric individuals contribute to their susceptibility to influenza.

Objective: Our aim was to analyze interferon-alpha (IFN-alpha) production in peripheral blood mononuclear cells (PBMCs) isolated from young and geriatric adult donors, stimulated with influenza A or Toll-like receptor (TLR) ligands.

Presenting exogenous antigen to T cells.

Antigen processing and presentation experiments can be done with a wide variety of antigen-presenting cells (APCs). Most experiments will use one of the "professional" APC types: dendritic cells (DCs), macrophages, and B lymphocytes. Other types of cells may be used for antigen presentation in some circumstances.

Choosing and preparing antigen-presenting cells.

The first issue in selecting a system for antigen-presentation experiments is to define the appropriate type of antigen-presenting cell (APC) to study. For some experiments, crude preparations such as splenocytes or peripheral blood mononuclear cells (PBMCs) may suffice to provide APC function for stimulating T cells. This unit develops approaches for preparation of more defined APC populations, including dendritic cells (DCs), macrophages, and B lymphocytes, the three types of "professional" APC.

P2X7 receptor-stimulated secretion of MHC class II-containing exosomes requires the ASC/NLRP3 inflammasome but is independent of caspase-1.

We recently reported that P2X7 receptor (P2X7R)-induced activation of caspase-1 inflammasomes is accompanied by release of MHC class II (MHC-II) protein into extracellular compartments during brief stimulation of murine macrophages with ATP. Here we demonstrate that MHC-II containing membranes released from macrophages or dendritic cells (DCs) in response to P2X7R stimulation comprise two pools of vesicles with distinct biogenesis: one pool comprises 100- to 600-nm microvesicles derived from direct budding of the plasma membrane, while the second pool is composed of 50- to 80-nm exosomes released from multivesicular bodies. ATP-stimulated release of MHC-II in these membrane fractions is observed within 15 min and results in the export of approximately 15% of the total MHC-II pool within 90 min.

MHC molecules and microbial antigen processing in phagosomes.

Macrophages and dendritic cells are phagocytic antigen presenting cells that internalize bacteria and other particulate antigens into phagosomes. The phagosome must then balance microbicidal and proteolytic degradation functions with the generation of antigenic peptides for presentation by class I and class II MHC molecules to CD8 and CD4 T cells, respectively. Understanding the host and bacterial factors that affect phagosomal antigen processing may help facilitate new strategies to eliminate pathogens.

Class II MHC antigen processing in phagosomes.

Phagocytic antigen-presenting cells (APCs) are involved in innate and adaptive immune responses to bacteria. Adaptive responses to bacteria involve processing of bacterial antigens for presentation by class II major histocompatibility complex (MHC II) molecules and class I MHC (MHC I) molecules to stimulate CD4(+) and CD8(+) T cells, respectively. To examine the role of phagosomes in processing of antigens for presentation by MHC II molecules to CD4(+) T cells, phagosomes have been biochemically and functionally analyzed by a variety of techniques that include flow analysis (flow organellometry), SDS-PAGE/Western blotting, and an antigen-presenting organelle assay.

Class II MHC antigen presentation defect in neonatal monocytes is not correlated with decreased MHC-II expression.

Neonates are at increased risk of infections compared to adults. To dissect the mechanisms that contribute to neonatal immune deficiency, we compared MHC-II antigen processing and presentation by monocytes from umbilical cord blood and unrelated adult controls. Antigen-specific, co-stimulation-independent murine T hybridoma cells were used to detect peptide:HLA-DR complexes.

The immunostimulant RU41740 from Klebsiella pneumoniae activates human cells in whole blood to potentially stimulate innate and adaptive immune responses.

The compound RU41740 from Klebsiella pneumoniae, when used as an immunostimulant, improves responses to bacterial and yeast infections in murine models and in human trials. The aim of this study was to determine in vitro, the capacity of RU41740 to stimulate human leukocytes in whole blood. Blood samples from healthy adult donors were incubated with RU41740 for 4 or 24 h and leukocytes were assessed for levels of activation markers and cytokine production by flow cytometry and ELISA.

Role of phagosomes and major histocompatibility complex class II (MHC-II) compartment in MHC-II antigen processing of Mycobacterium tuberculosis in human macrophages.

Mycobacterium tuberculosis resides in phagosomes inside macrophages. In this study, we analyzed the kinetics and location of M. tuberculosis peptide-major histocompatibility complex class II (MHC-II) complexes in M.

Enhancement of dendritic cell antigen cross-presentation by CpG DNA involves type I IFN and stabilization of class I MHC mRNA.

Dendritic cells (DCs) internalize exogenous Ags and process them for cross-presentation by class I MHC (MHC-I) to CD8+ T cells. This processing can occur by transporter for Ag presentation (TAP)-dependent or TAP-independent mechanisms. We observed that CpG DNA enhanced cross-presentation of Ags by Flt-3L-cultured bone marrow-derived murine DCs by a type I IFN (IFN-alphabeta)-dependent mechanism.

Phagosomal processing of Mycobacterium tuberculosis antigen 85B is modulated independently of mycobacterial viability and phagosome maturation.

Control of Mycobacterium tuberculosis infection requires CD4 T-cell responses and major histocompatibility complex class II (MHC-II) processing of M. tuberculosis antigens (Ags). We have previously demonstrated that macrophages process heat-killed (HK) M.

Localization of peptide/MHC class II complexes in macrophages following antigen processing of viable Streptococcus pyogenes.

The subcellular localization of peptide/MHC complexes was investigated during processing of the surface M5 protein from Streptococcus pyogenes. Bone marrow-derived macrophages were pulsed with viable S. pyogenes for 20 min followed by various periods of chase.

Alternate class I MHC antigen processing is inhibited by Toll-like receptor signaling pathogen-associated molecular patterns: Mycobacterium tuberculosis 19-kDa lipoprotein, CpG DNA, and lipopolysaccharide.

Pathogen-associated molecular patterns (PAMPs) signal through Toll-like receptors (TLRs) to activate immune responses, but prolonged exposure to PAMPs from Mycobacterium tuberculosis (MTB) and other pathogens inhibits class II MHC (MHC-II) expression and Ag processing, which may allow MTB to evade CD4(+) T cell immunity. Alternate class I MHC (MHC-I) processing allows macrophages to present Ags from MTB and other bacteria to CD8(+) T cells, but the effect of PAMPs on this processing pathway is unknown. In our studies, MTB and TLR-signaling PAMPs, MTB 19-kDa lipoprotein, CpG DNA, and LPS, inhibited alternate MHC-I processing of latex-conjugated Ag by IFN-gamma-activated macrophages.