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ARTEMIS Report Summary

Project ID: 330134
Funded under: FP7-PEOPLE
Country: Denmark

Periodic Report Summary 1 - ARTEMIS (Autoimmune Reaction or Tolerance - Endosymbiont Mitochondria and the Immune System)

The ARTEMIS project revolves around the balance between autoimmune reaction and tolerance, with a special focus on the molecular discrimination of self (host-derived nucleic acids and mitochondrial components) and non-self (bacterial and viral components).
Mitochondria, the powerhouses of our cells, are remnants of a eubacterial endosymbiont. Because mitochondria retain many hallmarks of their bacterial origin, they have recently been implicated in the etiology of the sterile systemic inflammatory response syndrome (SIRS), a frequent cause of death in intensive care units (Zhang et al., 2010). However, we found that mitochondria are immunologically inert under normal homeostatic conditions, as they are cleared in a complement-dependent manner in the absence of overt inflammation (Brinkmann et al., 2013). Taken together, these contrasting observations exemplify the fundamental question of how the immune system distinguishes self from non-self.
Mitochondria are centrally involved in initiation and execution of programmed cell death. This process may both release endogenous danger-associated molecular patterns (DAMPs) and pathogen-associated molecular pattern (PAMP) mimetics, as well as produce reactive oxygen species (ROS), which in turn can generate neoepitopes. Autoantibodies to mitochondrial components, e.g. cytochrome C, have been observed in patients with lupus and related autoimmune diseases (Mamula, Jemmerson, & Hardin, 1990). Such autoantibodies are thought to arise in response to oxidative damage caused by free radical generation in cells at sites including the mitochondria, endoplasmic reticulum and nuclear membranes (Casciola-Rosen, Anhalt, & Rosen, 1994). Accordingly, we investigated the release, transport and uptake of such apoptotic cell-derived antigenic material using intravital two-photon imaging (Degn and Carroll, manuscript in preparation). Either labeled apoptotic cells or organelles were exogenously injected, or more elegantly, such material was generated in situ in a fluorescent reporter strain (BFP-SSB, Figure 1) by UV irradiation (Figure 2). We were able to visualize these components in situ but they appear to be efficiently cleared, and do not drain in intact form to skin-draining lymph nodes. An embedded OTII epitope in the BFP-SSB reporter (please refer to Figure 1) was used to specifically address T cell responses in this scenario. Adoptive transfer of transgenic OTII T cells demonstrated efficient surveillance of such neo-autoantigen in skin-draining lymph nodes, indicating drainage of degraded antigen. Yet following transient activation, responding T cell populations contracted, indicating a regulatory response (Figure 3).
An exciting observation springing from these initial investigations was that a restricted autoreactive B cell repertoire is sufficient to drive autoreactive germinal center reactions, recruiting endogenous proto-autoreactive B cells. This process may be a fundamental one underlying the concepts of epitope spreading and repertoire conversion. Autoantibodies are common in autoimmune conditions, and class-switched affinity-matured antibodies to nucleic acid antigens are a hallmark of systemic lupus erythematosus (SLE) (Rahman & Isenberg, 2008). Anti-ds-DNA is more specific for SLE (>75%) compared with healthy controls (~0.5%) and typically comes up a few years before SLE is diagnosed (Arbuckle et al., 2003). SLE patients examined closer to or after onset of disease demonstrate drift in their serological reactivities towards a variety of nuclear, nucleolar, and protein-DNA complexes which may reflect functional epitope spreading and which may propagate disease states. Epitope spreading is driven in part by chronic immune responses and functionally refers to diversification of VJ segment utilization by germinal center (GC) B cells which leads to new reactivities. Presence and levels of titers representing these reactivities are used to guide clinical management without mechanistic understanding of either their role in disease or their widely varying penetrance and magnitude (Arbuckle et al., 2003; Cornaby et al., 2015; Vanderlugt & Miller, 2002). GC are the primary sites of clonal expansion, class-switching and affinity maturation of B cells, directing the production of high-affinity antibodies. Yet the dynamics of self-reactive B cell engagement and persistence in chronic autoreactive GC remain poorly understood. We have found that a self-reactive transgenic B cell clone (derived from the 564 model, (Berland et al., 2006)) is sufficient to break tolerance and initiate autoreactive GC composed predominantly of wild type-derived cells (Degn et al., manuscript in preparation). Autoreactive GC depend on T help, are self- sustained, long-lived, and may be cyclically reseeded by self-reactive cells. Employing a novel Aicda-CreERT2 Confetti reporter system ((Tas et al., 2016), Figure 4), we find that they evolve towards pauciclonality at a decreased rate compared with GC elicited by foreign antigen. Using photoactivatable (PA)-GFP reporters ((Victora et al., 2010), Figure 4), we have examined single GCs. Clonally they converge on stereotypic autoreactive sequence elements, which are mirrored serologically by functional epitope spreading. Our findings have fundamental implications for the understanding of important pathophysiological processes such as epitope spreading and repertoire conversion.

For further information, please contact:
Søren E. Degn, PhD
Department of Biomedicine, Aarhus University, Bartholins Allé 6, Building 1242, 565, 8000 Aarhus C, Denmark
Phone: +4587167274

Arbuckle, M. R., McClain, M. T., Rubertone, M. V., Scofield, R. H., Dennis, G. J., James, J. A., & Harley, J. B. (2003). Development of autoantibodies before the clinical onset of systemic lupus erythematosus. The New England Journal of Medicine, 349(16), 1526–1533.
Berland, R., Fernandez, L., Kari, E., Han, J.-H., Lomakin, I., Akira, S., et al. (2006). Toll-like receptor 7-dependent loss of B cell tolerance in pathogenic autoantibody knockin mice. Immunity, 25(3), 429–440.
Brinkmann, C. R., Jensen, L., Dagnaes-Hansen, F., Holm, I. E., Endo, Y., Fujita, T., et al. (2013). Mitochondria and the lectin pathway of complement. The Journal of Biological Chemistry, 288(12), 8016–8027.
Casciola-Rosen, L. A., Anhalt, G., & Rosen, A. (1994). Autoantigens targeted in systemic lupus erythematosus are clustered in two populations of surface structures on apoptotic keratinocytes. The Journal of Experimental Medicine, 179(4), 1317–1330.
Cornaby, C., Gibbons, L., Mayhew, V., Sloan, C. S., Welling, A., & Poole, B. D. (2015). B cell epitope spreading: mechanisms and contribution to autoimmune diseases. Immunology Letters, 163(1), 56–68.
Mamula, M. J., Jemmerson, R., & Hardin, J. A. (1990). The specificity of human anti-cytochrome c autoantibodies that arise in autoimmune disease. The Journal of Immunology, 144(5), 1835–1840.
Rahman, A., & Isenberg, D. A. (2008). Systemic Lupus Erythematosus. New England Journal of Medicine, 358(9), 929–939.
Tas, J. M. J., Mesin, L., Pasqual, G., Targ, S., Jacobsen, J. T., Mano, Y. M., et al. (2016). Visualizing antibody affinity maturation in germinal centers. Science (New York, NY), 351(6277), 1048–1054.
Vanderlugt, C. L., & Miller, S. D. (2002). Epitope spreading in immune-mediated diseases: implications for immunotherapy. Nature Reviews Immunology, 2(2), 85–95.
Victora, G. D., Schwickert, T. A., Fooksman, D. R., Kamphorst, A. O., Meyer-Hermann, M., Dustin, M. L., & Nussenzweig, M. C. (2010). Germinal center dynamics revealed by multiphoton microscopy with a photoactivatable fluorescent reporter. Cell, 143(4), 592–605.
Zhang, Q., Raoof, M., Chen, Y., Sumi, Y., Sursal, T., Junger, W., et al. (2010). Circulating mitochondrial DAMPs cause inflammatory responses to injury. Nature, 464(7285), 104–107.

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Merete Kamp, (Project economist)
Tel.: +45 8715 2847


Life Sciences
Record Number: 191497 / Last updated on: 2016-11-21
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