Peter Aichele

Antigen localisation regulates immune responses in a dose‐ and time‐dependent fashion: a geographical view of immune reactivity

Summary: This review summarises experimental evidence to illustrate that induction of immune reactivity depends upon antigen reaching and being available in lymphoid organs in a dose‐ and time‐dependent manner. If antigen reaches lymph organs in a localised staggered manner and with a concentration gradient, a response is induced in the draining lymph node. Antigen‐presenting cells are of critical importance to transport antigen from the periphery to local organised lymphoid tissue. If antigen is all over the lymphoid system, then it deletes all specific cells in the thymus or induces them within a few days: because of their limited life‐span they then die off, leaving the repertoire depleted of this specificity. If antigen does not reach lymphoid organs it is ignored by immune cells. Once a response is induced, activated but not resting T cells will reach antigen outside lymphoid organs, whereas activated B cells differentiate into plasma cells in an inducing environment, mostly in lymphoid tissue including bone marrow, but also in chronic lymphoid‐like infiltrations in peripheral organs. In immunopathology (when the infectious agent is known) or in autoimmunity (when the triggering infectious agent is not known or not recognised) lymphoid tissue may become organised close to the antigen (e.g. in organ‐specific autoimmune diseases) and may thereby maintain an autoantigen‐driven disease‐causing immune response for a long time, The notion that naive T cells get induced or silenced in the periphery may be questioned because induction can only occur in lymphoid organs providing anatomical structures where critical cell‐cell interactions are properly guided and where, therefore, cells are likely to meet sufficiently frequently and in a critical milieu. Since overall immune reactivity critically depends upon the localisation of antigens in a dose‐ and time‐dependent manner, it seems more likely ‐ but this remains to be shown ‐ that activated T cells may get exhausted in non‐lymphoid peripheral tissues, whereas they are usually maintained in lymphoid organs. The critical role of antigen in regulating immune responses also has relevance for our understanding of immunological defence against epithelial and mesenchymal tumours, against many infectious diseases and for understanding autoimmunity and immunological memory. Collectively the data indicate that antigen, impendent upon localisation, dose and time, seems to be the simplest regulator of immune responses.

Early granuloma formation after aerosol Mycobacterium tuberculosis infection is regulated by neutrophils via CXCR3-signaling chemokines.

Among the first cells to invade a site of infection, polymorphonuclear neutrophils (PMN) play an important role in the control of numerous infections. While PMN are considered critical for control of acute infections, their role in chronic infections remains less well understood. Here we report that PMN are essential for accurate early granuloma formation during chronic M. tuberculosis infection without influencing mycobacterial growth restriction. The PMN‐mediated regulation of granuloma formation depended on chemokines signaling through CXCR3, in particular MIG, as indicated by immune histochemical analysis of lung sections from C57BL/6 wild‐type and CXCR3–/– mutant mice and supported by microarray transcriptome analysis. Hence, PMN play a central role in regulating the focal granulomatous response in the lung, and this early granuloma formation can be segregated from long‐term protection against pulmonary M. tuberculosis infection.

Macrophages of the Splenic Marginal Zone Are Essential for Trapping of Blood-Borne Particulate Antigen but Dispensable for Induction of Specific T Cell Responses

Rapid removal of pathogens from the circulation by secondary lymphoid organs is prerequisite for successful control of infection. Blood-borne Ags are trapped mainly in the splenic marginal zone. To identify the cell populations responsible for Ag trapping in the marginal zone, mice were selectively depleted of marginal zone macrophages and marginal metallophilic macrophages. In the absence of these cells, trapping of microspheres and Listeria monocytogenes organisms was lost, and early control of infection was impaired. Depletion of marginal zone macrophages and marginal metallophilic macrophages, however, did not limit Ag presentation because Listeria-specific protective T cell immunity was induced. Therefore, marginal zone macrophages and marginal metallophilic macrophages are crucial for trapping of particulate Ag but dispensable for Ag presentation.

Peptide Antigen Treatment of Naive and Virus-Immune Mice: Antigen-Specific Tolerance Versus Immunopathology

Peptide-specific down-regulation of T cell responses may represent a powerful tool to intervene in autoimmune diseases or graft rejections. It is therefore important to know whether peptide treatment tolerizes both naive and antigen-experienced memory T lymphocytes. Here we show that a major histocompatibility complex class I binding peptide, derived from the glycoprotein (GP33 peptide) of lymphocytic choriomeningitis virus (LCMV), specifically tolerized naive cytotoxic T lymphocytes (CTL) when administered three times intraperitoneally in incomplete Freunds adjuvants. However, in the presence of GP33-specific memory CTL in LCMV-primed mice, the same treatment had a general immunosuppressive effect on unrelated third-party antigen-specific T cell responses and caused severe immunopathological damage to the spleen.

Rapid Neutrophil Response Controls Fast-Replicating Intracellular Bacteria but Not Slow-Replicating Mycobacterium tuberculosis

Being one of the first cells to invade the site of infection, neutrophils play an important role in the control of various bacterial and viral infections. In the present work, the contribution of neutrophils to the control of infection with different intracellular bacteria was investigated. Mice were treated with the neutrophil-depleting monoclonal antibody RB6-8C5, and the time course of infection in treated and untreated mice was compared by using intracellular bacterial species and strains varying in virulence and replication rate. The results indicate that neutrophils are crucial for the control of fast-replicating intracellular bacteria, whereas early neutrophil effector mechanisms are dispensable for the control of the slow-replicating Mycobacterium tuberculosis.

Autoimmune Intestinal Pathology Induced by hsp60-Specific CD8 T Cells

Due to their ubiquitous distribution and high degree of structural similarity, heat shock proteins (hsp) are potential target antigens in autoimmune diseases. Here, we describe induction of intestinal inflammation following transfer of hsp60-reactive CD8 T cells into mice. Inflammatory reactions were MHC class I dependent and developed primarily in the small intestine. IFN gamma and TNF alpha, as well as gut-derived hsp60, were elevated at sites of T cell infiltration. Intestinal lesions were drastically reduced in mice lacking receptors for TNF alpha. Pathology also developed in germ-free mice, indicating recognition of host-derived hsp60 by CD8 T cells. This report demonstrates that CD8 T cells with defined antigen specificity cause intestinal inflammation, emphasizing a link between infection and autoimmune disease.

Modified immunohistological staining allows detection of Ziehl-Neelsen-negative Mycobacterium tuberculosis organisms and their precise localization in human tissue

The diagnosis of mycobacterial infection depends on the Ziehl–Neelsen (ZN) stain, which detects mycobacteria because of their characteristic acid‐fast cell wall composition and structure. The histological diagnosis of tuberculosis (TB) comprises various aspects: (1) sensitive detection of mycobacteria; (2) precise localization of mycobacteria in the context of granulomatous lesions; (3) ‘staging’ of disease according to mycobacterial spread and granulomatous tissue integrity. Thus, detection of minute numbers of acid‐fast bacteria in tissue specimens is critical. The conventional ZN stain fails to identify mycobacteria in numbers less than 104 per ml. Hence many infections evade diagnosis. PCR is highly sensitive, but allows neither localization within tissues nor staging of mycobacterial disease, and positive findings frequently do not correlate with disease. In this study, an anti‐Mycobacterium bovis bacille Calmette–Guérin polyclonal antiserum (pAbBCG) was used to improve immunostaining, which was compared to the ZN stain in histological samples. Screening of tissue samples including lungs, pleural lesions, lymph nodes, bone marrow, and skin for mycobacterial infection revealed that pAbBCG staining detects infected macrophages harbouring intracellular mycobacteria or mycobacterial material as well as free mycobacteria that are present at low abundance and not detected by the ZN stain. The positive pAbBCG staining results were confirmed either by PCR analysis of microdissected stained tissue or by culture from tissue. This immunostaining approach allows precise localization of the pathogen in infected tissue. Copyright

Microchimerism maintains deletion of the donor cell–specific CD8 + T cell repertoire

Rare cases of stable allograft acceptance after discontinuation of immunosuppression are often accompanied by macrochimerism (> 1% donor cells in blood) or microchimerism (< 1% donor cells in blood). Here, we have investigated whether persistence of donor cells is the cause or the consequence of long-lasting CTL unresponsiveness. We found that engraftment of splenocytes bearing a single foreign MHC class I-restricted epitope resulted in lifelong donor cell microchimerism and specific CTL unresponsiveness. This status was reversed in a strictly time- and thymus-dependent fashion when the engrafted cells were experimentally removed. The results presented herein show that microchimerism actively maintains CTL unresponsiveness toward a minor histocompatibility antigen by deleting the specific repertoire and thus excluding dominant, T cell extrinsic mechanisms of CTL unresponsiveness independent of systemically persisting donor cell antigen.

Neutrophilia in LFA-1-Deficient Mice Confers Resistance to Listeriosis: Possible Contribution of Granulocyte-Colony-Stimulating Factor and IL-17

LFA-1 (CD11a/CD18) plays a crucial role in various inflammatory responses. In this study, we show that LFA-1−/− mice are far more resistant to Listeria monocytogenes infection than LFA-1+/− mice. Consistent with this, we found the following: 1) the numbers of granulocytes infiltrating the liver were markedly higher in LFA-1−/− mice than in LFA-1+/− mice, 2) increased antilisterial resistance in LFA-1−/− mice was abrogated by depletion of granulocytes, and 3) the numbers of granulocytes in peripheral blood, and the serum levels of both G-CSF and IL-17 were higher in LFA-1−/− mice than in LFA-1+/− mice. Neither spontaneous apoptosis nor survival of granulocytes from LFA-1−/− mice were affected by physiological concentrations of G-CSF. Our data suggest regulatory effects of LFA-1 on G-CSF and IL-17 secretion, and as a corollary on neutrophilia. Consequently, we conclude that increased resistance of LFA-1−/− mice to listeriosis is due to neutrophilia facilitating liver infiltration by granulocytes promptly after L. monocytogenes infection, although it is LFA-1 independent.

Antigen persistence and time of T-cell tolerization determine the efficacyof tolerization protocols for prevention of skin graft rejection

We studied antigen-specific T-cell tolerization therapy using skin transplantation across a defined minor histocompatibility antigen difference. Specific tolerization protocols using short-lived peptide or long-lived spleen cells presenting the peptide as antigen prevented graft rejection without immunosuppression when started before or as long as 10 days after transplantation. Peptide-induced T-cell tolerance was transient, and antigen presentation by the graft was not sufficient to maintain tolerance. In contrast, transfer of antigen-expressing lymphoid cells induced long-lasting tolerance correlating with donor cell chimerism. These findings show that antigen-specific tolerization can induce graft acceptance even when begun after transplantation and that long-term graft survival depends on persistence of the tolerizing antigen.