FRI0521 Cd28null T Cells Kill Autologous Muscle Cells from Polymyositis Patients in Vitro by Perforin-Dependent Mechanisms
Authors: P. Venalis, J. Pandya, L. Al-Khalili, M.S. Hossain, V. Stache, I.E. Lundberg, V. Malmström, A.E.R. Fasth
Journal: Annals of the Rheumatic Diseases (2014)

<h3>Background</h3> Inflammatory infiltrates of muscles in polymyositis are dominated by CD4+CD28null and CD8+CD28null T cells<sup>1</sup>. In contrast to conventional CD28+ T cells, these cells have a restricted T cell-receptor repertoire, rapidly release large amounts of IFN-γ and TNF, and can kill tissue cells by granzyme B and perforin upon activation<sup>1,2,3,4</sup>. Although their presence in muscles, their effect on muscle cells has not been studied. <h3>Objectives</h3> To in a fully autologous system investigate the myotoxic effect of CD28null T cells on muscle cells from patients with polymyositis. <h3>Methods</h3> Muscle stem cells were extracted from biopsies from 5 patients with polymyositis, and differentiated into myotubes. T cell subsets from the same patients were isolated from peripheral blood by flow-cytometry. Co-cultures of autologous muscle cells and unstimulated or pre-activated (anti-CD3-antibodies) T cells were performed for 24 hours. Dead myotubes were quantified by calcein release or by flow-cytometry. Blocking experiments were performed by adding indicated antibodies to the cultures. <h3>Results</h3> CD4+CD28null and CD8+CD28null T cells caused spontaneous death of 13 60% (n=2 patients) of the muscle cells. No or low muscle cell death was detected in cultures with conventional CD4+ and CD8+ T cells. By blocking perforin the myotoxicity was reduced by a median of 52% with CD4+CD28null T cells and by 56% in cultures with CD8+CD28null T-cells (n=3). TNF or IFN-γ did not induce death of muscle cells in the absence of T cells, but did upregulate MHC class I and II on muscle cells (n=3-5). Blockade of IFN-γ and TNF in co-cultures with T cells reduced the level of dead muscle cells by 46% (CD4+CD28null) and 53% (CD8+CD28null) (n=3). The reduced myotoxicity is likely attributed to reduced MHC expression, as MHC blockade resulted in a comparable reduction in myotoxicity, 61% (CD4+CD28null) and 55% (CD8+CD28null) (n=5). <h3>Conclusions</h3> The myotoxic effect by CD28null T cells in polymyositis is mediated by directed perforin-dependent killing and regulated by IFN-γ-induced MHC expression by muscle cells. Together this suggests that CD28null T cells are key effector cells directly contributing to the muscle cell damage in polymyositis, hence represent target candidate for future therapies. <h3>References</h3> T cell infiltrates in the muscles of patients with dermatomyositis and polymyositis are dominated by CD28null T cells. Fasth AE, Dastmalchi M, Rahbar A, Salomonsson S, Pandya JM, Lindroos E, Nennesmo I, Malmberg KJ, Söderberg-Nauclér C, Trollmo C, Lundberg IE, Malmström V. J Immunol. 2009. Skewed distribution of proinflammatory CD4+CD28null T cells in rheumatoid arthritis. Fasth AE, Snir O, Johansson AA, Nordmark B, Rahbar A, Af Klint E, Björkström NK, Ulfgren AK, van Vollenhoven RF, Malmström V, Trollmo C. Arthritis Res Ther. 2007. Expanded T cell receptor Vβ-restricted T cells from patients with sporadic inclusion body myositis are proinflammatory and cytotoxic CD28null T cells. Pandya JM, Fasth AE, Zong M, Arnardottir S, Dani L, Lindroos E, Malmström V, Lundberg IE. Arthritis Rheum. 2010. T-cell-mediated lysis of endothelial cells in acute coronary syndromes. Nakajima T, Schulte S, Warrington KJ, Kopecky SL, Frye RL, Goronzy JJ, Weyand CM. Circulation. 2002. <h3>Disclosure of Interest</h3> P. Venalis: None declared, J. Pandya: None declared, L. Al-Khalili: None declared, M. Hossain: None declared, V. Stache: None declared, I. Lundberg: None declared, V. Malmström: None declared, A. Fasth Employee of: Associated researcher at Karolinska Institutet, but also employeed by Novartis. The research at Karolinska Institutet is totally independent from the work at Novartis. <h3>DOI</h3> 10.1136/annrheumdis-2014-eular.5189