RNA was extracted using the RNeasy Mini kit (Qiagen, Hilden, Germany). shows that clonal TCR signatures are shared between patients. Increased clonal growth in muscle tissue is significantly correlated with increased CK levels (p=0.03), while it tends to correlate with decreased muscle mass strength (p=0.08). Conclusion Network analysis of clones in muscle mass of IIM patients shows shared clusters of sequences across patients. Muscle-restricted CDR3 TCR clones show specific structural features in their T cell receptor. Our results indicate that clonal TCR growth in muscle tissue might be associated with disease activity. Collectively, these findings support a role for specific clonal T cell responses in muscle tissue in the pathogenesis of the IIM subtypes analyzed. Keywords: myositis, T cells, AIRR seq, T cell receptor (TCR), adaptive immunity Introduction Subacute proximal muscle mass weakness is usually a characteristic feature of idiopathic inflammatory myopathies (IIM). The main subtypes are dermatomyositis (DM), antisynthetase syndrome (ASyS), immune-mediated necrotizing myopathy (IMNM), non-specific/overlap myositis (NM/OM), polymyositis (PM) and inclusion body myositis (IBM) (1, 2). Despite therapy, most patients (70%) have a chronic or polyphasic disease course and develop significant residual disability and reduced quality of life (3). Consequently, there is a clear need for a better understanding of its disease pathogenesis to identify novel therapeutic targets. For decades, lymphocytes have been implicated in the mechanisms responsible for the development of IIMs. For B-lymphocytes this view is supported by the presence of plasma cells in muscle biopsies (4, 5), the presence of myositis specific and myositis related antibodies in blood in 60 to 70% of the IIM KBTBD6 patients (6), the effectiveness of B cell directed therapies (7, 8), and the presence of dominant Z-LEHD-FMK B-cell receptor clones in muscle tissue and peripheral blood which correlates with better response to intravenous immunoglobulins (9). Besides B cells, also T cells of different lineages have been implicated in the pathogenesis of IIM patients. Cytotoxic T cells, which often produce toxic granules such as granzymes and perforins, are present in muscle tissues of PM, juvenile DM and IBM patients (10C12) In addition, expanded T cell clones with cytotoxic properties have been reported in DM and IMNM patients (13). Several reports describe skewing of various T cells subsets in peripheral blood, including subsets which provide help to B cells (14C16). The latter is not unexpected, given the key role these cells play in the generation of serological responses. Few studies have explored the T cell response at the clonal level simultaneously in peripheral blood samples and muscle biopsies. Previous studies using flow cytometry, immunochemistry and CDR3 spectra typing contributed to unraveling T cell involvement in myositis (11, 17) but are less sensitive and of low clonal resolution in detecting relevant repertoire changes (18). In this prospective study on the effects of IVIG in IIM we explored T cell receptor beta (TCR) repertoires in paired peripheral blood samples and muscle biopsies taken prior to IVIG treatment. In this way we aimed to answer the question whether the peripheral blood T cell repertoire is reflective of the repertoire in muscle tissues prior to treatment. Our results confirmed T cell clonal expansion both in muscle tissues and in peripheral blood of IIM patients However, we also identified (expanded) tissue restricted TCR clones which could not be retrieved in peripheral blood. These tissue restricted TCR clones showed features which were quite distinct from those seen in peripheral blood, and shared between patients. Finally, TCR repertoire features in muscle tissues (but not in peripheral blood) correlated significantly with disease activity in these patients. Our data support a key role for abnormal T cell responses in disease development and progression in IIM. Materials and methods Ethical statement All patients signed Z-LEHD-FMK informed consent prior to inclusion in the Z-LEHD-FMK study. This study was conducted with approval of the local medical research ethics committee of the Academic Medical Centre, Amsterdam, and in accordance with the declaration of Helsinki. Patients and study design Peripheral blood and muscle tissues used in this.