Minka Breloer

Heat shock proteins: linking danger and pathogen recognition

Besides their central function in protein folding and transport within the cell, heat shock proteins (HSP) have been shown to modulate innate and adaptive immune response: (1) HSP mediate uptake and MHC presentation of HSP-associated peptides by antigen-presenting cells (APC). (2) HSP function as endogenous danger signals indicating cell stress and tissue damage to the immune system. (3) HSP bind pathogen-associated molecular pattern (PAMP) molecules and modulate PAMP-induced Toll-like receptor (TLR) signaling. Thus, HSP contribute to both, recognition of “danger” defined as uncontrolled tissue destruction and recognition of dangerous “nonself”. In this review these different aspects of immune stimulation by HSP will be discussed.

ISOLATION OF PROCESSED, H-2KB-BINDING OVALBUMIN-DERIVED PEPTIDES ASSOCIATED WITH THE STRESS PROTEINS HSP70 AND GP96

Stress‐induced proteins or heat shock proteins (HSP) of 96 kDa mass (gp96) and 70 kDa mass (HSP70) have been shown previously to elicit specific immunity to tumors from which they are isolated. This immunity is dependent on CD8+ cytotoxic T cells which are readily primed in vivo by immunization with HSP. The immunization capacity of HSP relies on their ability to bind antigenic peptides. Here we show that HSP70 and gp96 preparations purified from the ovalbumin (OVA)‐transfected cell line E.G7 are associated with processed H‐2Kb ‐binding peptides which contain the major H‐2Kb ‐associated epitope SIINFEKL (OVA257 –264). Our data show for the first time in the well‐defined OVA antigen system that not only endoplasmic reticulum‐resident HSP, like gp96, are associated with processed antigenic peptides but that also the cytosolic HSP70 protein forms complexes with major finally processed MHC‐binding epitopes.

Synergistic and Differential Modulation of Immune Responses by Hsp60 and Lipopolysaccharide

Activation of professional antigen-presenting cells (APC) is a crucial step in the initiation of an efficient immune response. In this study we show that Hsp60 mediates immune stimulation by different mechanisms, dependent and independent of lipopolysaccharide (LPS). We have demonstrated earlier that both, Hsp60 and LPS, increase antigen-specific interferon (IFN) γ release in T cells. Here we show that in contrast to LPS Hsp60 induces IFNα production in professional APC. Neutralization of IFNα as well as the absence of functional IFNαβ receptor on APC and T cells interfered with Hsp60-mediated IFNγ secretion in antigen-dependent T cell activation, strongly suggesting that IFNα represents one factor contributing to Hsp60-specific immune stimulation. On the other hand, we show that Hsp60 bound to the cell surface of APC colocalizes with the LPS co-receptor CD14 and LPS binding sites. Hsp60 specifically binds bacterial LPS and both molecules synergistically enhanced IL-12p40 production in APC and IFNγ release in antigen-dependent T cell activation. This effect was Hsp60-specific and dependent on LPS-binding by Hsp60. Furthermore, we show that Hsp60 exclusively binds to macrophages and DC but not to T or B lymphocytes and that both, T cell stimulation by Hsp60 as well as Hsp60/LPS complexes, strictly depends on the presence of professional APC and is not mediated by B cells. Taken together, our data support an extension of the concept of Hsp60 as an endogenous danger signal: besides its function as a classical danger signal indicating unplanned tissue destruction to the innate immune system, in the incident of bacterial infection extracellular Hsp60 may bind LPS and facilitate microbe recognition by lowering the threshold of pathogen-associated molecular pattern (PAMP) detection and enhancing Toll-like receptor (TLR) signaling.

CD83 regulates lymphocyte maturation, activation and homeostasis

The transmembrane CD83 molecule, a conserved member of the immunoglobulin superfamily, is known as one of the most characteristic cell surface markers for fully matured dendritic cells (DCs) in the peripheral circulation. An essential role for CD83 on murine DCs has not been found; however, evidence shows that its function primarily lies in the regulation of T- and B-lymphocyte maturation and in the regulation of their peripheral responses. Here, we review evidence for a role of CD83 in central lymphocyte maturation and novel, sometimes contradictory findings, regarding the function of CD83 in peripheral immune responses.

Toll like receptor 4 mediates cell death in a mouse MPTP model of Parkinson disease

In mammalians, toll-like receptors (TLR) signal-transduction pathways induce the expression of a variety of immune-response genes, including inflammatory cytokines. It is therefore plausible to assume that TLRs are mediators in glial cells triggering the release of cytokines that ultimately kill DA neurons in the substantia nigra in Parkinson disease (PD). Accordingly, recent data indicate that TLR4 is up-regulated by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) treatment in a mouse model of PD. Here, we wished to evaluate the role of TLR4 in the acute mouse MPTP model of PD: TLR4-deficient mice and wild-type littermates control mice were used for the acute administration way of MPTP or a corresponding volume of saline. We demonstrate that TLR4-deficient mice are less vulnerable to MPTP intoxication than wild-type mice and display a decreased number of Iba1+ and MHC II+ activated microglial cells after MPTP application, suggesting that the TLR4 pathway is involved in experimental PD.

Strongyloides ratti Infection Induces Expansion of Foxp3+ Regulatory T Cells That Interfere with Immune Response and Parasite Clearance in BALB/c Mice

To escape expulsion by their host’s immune system, pathogenic nematodes exploit regulatory pathways that are intrinsic parts of the mammalian immune system, such as regulatory T cells (Tregs). Using depletion of Treg mice, we showed that Foxp3+ Treg numbers increased rapidly during infection with the nematode Strongyloides ratti. Transient depletion of Tregs during the first days of infection led to dramatically reduced worm burden and larval output, without aggravation of immune pathology. The transient absence of Tregs during primary infection did not interfere with the generation of protective memory. Depletion of Tregs at later time points of infection (i.e., day 4) did not improve resistance, suggesting that Tregs exert their counterregulatory function during the priming of S. ratti-specific immune responses. Improved resistance upon early Treg depletion was accompanied by accelerated and prolonged mast cell activation and increased production of types 1 and 2 cytokines. In contrast, the blockade of the regulatory receptor CTLA-4 specifically increased nematode-specific type 2 cytokine production. Despite this improved immune response, resistance to the infection was only marginally improved. Taken together, we provide evidence that Treg expansion during S. ratti infection suppresses the protective immune response to this pathogenic nematode and, thus, represents a mechanism of immune evasion.

CD83 Expression in CD4+ T Cells Modulates Inflammation and Autoimmunity

The transmembrane protein CD83 has been initially described as a maturation marker for dendritic cells. Moreover, there is increasing evidence that CD83 also regulates B cell function, thymic T cell maturation, and peripheral T cell activation. Herein, we show that CD83 expression confers immunosuppressive function to CD4+ T cells. CD83 mRNA is differentially expressed in naturally occurring CD4+CD25+ regulatory T cells, and upon activation these cells rapidly express large amounts of surface CD83. Transduction of naive CD4+CD25− T cells with CD83 encoding retroviruses induces a regulatory phenotype in vitro, which is accompanied by the induction of Foxp3. Functional analysis of CD83-transduced T cells in vivo demonstrates that these CD83+Foxp3+ T cells are able to interfere with the effector phase of severe contact hypersensitivity reaction of the skin. Moreover, adoptive transfer of these cells prevents the paralysis associated with experimental autoimmune encephalomyelitis, suppresses proinflammatory cytokines IFN-γ and IL-17, and increases antiinflammatory IL-10 in recipient mice. Taken together, our data provide the first evidence that CD83 expression can contribute to the immunosuppressive function of CD4+ T cells in vivo.

Protein kinase CK2 enables regulatory T cells to suppress excessive TH2 responses in vivo

The quality of the adaptive immune response depends on the differentiation of distinct CD4+ helper T cell subsets, and the magnitude of an immune response is controlled by CD4+Foxp3+ regulatory T cells (Treg cells). However, how a tissue- and cell type–specific suppressor program of Treg cells is mechanistically orchestrated has remained largely unexplored. Through the use of Treg cell–specific gene targeting, we found that the suppression of allergic immune responses in the lungs mediated by T helper type 2 (TH2) cells was dependent on the activity of the protein kinase CK2. Genetic ablation of the β-subunit of CK2 specifically in Treg cells resulted in the proliferation of a hitherto-unexplored ILT3+ Treg cell subpopulation that was unable to control the maturation of IRF4+PD-L2+ dendritic cells required for the development of TH2 responses in vivo.

Heat Shock Protein 60 Is Released in Immune-Mediated Glomerulonephritis and Aggravates Disease: In Vivo Evidence for an Immunologic Danger Signal

Heat shock proteins (Hsp) are ubiquitous intracellular proteins that can be released in various forms of cellular stress. Some Hsp, such as Hsp60, have been shown to stimulate directly T cell-mediated immune responses in vitro. Here, it is demonstrated that Hsp60 is released from the kidneys and excreted into the urine of mice with nephrotoxic nephritis (NTN), a model of rapidly progressive glomerulonephritis. For examining the functional relevance of Hsp60 release, this protein was injected into mice with subnephritogenic NTN, in which only transient proteinuria and minimal organ damage occur that do not progress to terminal kidney failure. Injection of Hsp60 strikingly aggravated disease, as evidenced by global glomerular necrosis, tubulointerstitial damage, and complete anuria after 10 to 12 d. This effect was mediated neither by endotoxin contaminations of Hsp60 nor by autologous antibodies. It was strictly T cell dependent but not associated with a systemic Th1/Th2 shift. Thus, Hsp60 is an endogenous mediator stimulating immune effector mechanisms that contribute to the progression of NTN. These findings demonstrate in vivo that Hsp60 fulfills criteria of immunologic danger signals and suggest that such signals may be involved in immune-mediated kidney disease.

CD83 is a regulator of murine B cell function in vivo

The transmembrane glycoprotein CD83 has been described as a specific maturation marker for dendritic cells and several lines of evidence suggest that CD83 regulates thymic T cell maturation as well as peripheral T cell activation. Here we show for the first time that CD83 is involved also in the regulation of B cell function. CD83 is up‐regulated on activated B cells in vivo, specifically in the draining lymph nodes of Leishmania major‐infected mice. The ubiquitous transgenic (Tg) expression of CD83 interferes with Leishmania‐specific T cell‐dependent and with T cell‐independent antibody production. This defect is restricted to the B cell population since the antigen‐specific T cell response of CD83Tg mice to L. major infection is unchanged. The defective immunoglobulin (Ig) response is due to Tg expression of CD83 on the B cells because wild‐type B cells display normal antigen‐specific responses in CD83Tg hosts and CD83Tg B cells do not respond to immunization in a mixed wild‐type/CD83Tg bone marrow chimera. Finally, the treatment of non‐Tg C57BL/6 mice with anti‐CD83 mAb induces a dramatic increase in the antigen‐specific IgG response to immunization, thus demonstrating a regulatory role for naturally induced CD83 on wild‐type B cells.