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Evidence of Rare T Cell population expansion and T Cell Clonality in inclusion body myositis

Thomas, Emily (2019) Evidence of Rare T Cell population expansion and T Cell Clonality in inclusion body myositis. Honours thesis, Murdoch University.

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Background and objectives: Inclusion body myositis (IBM) is an inflammatory myopathy that frequently affects patients over 45, causing atrophy and weakness in proximal and distal muscles. The muscle histology is characterized by the infiltration of T cells, that direct responses against muscle leading to their destruction. T cells responding to single or common antigens will consist of monoclonal or oligoclonal populations respectively. Previous studies have sequenced the receptor of infiltrating T cell populations and found evidence of clonal (TCR) restriction. Furthermore, expansion of rare T cells which includes CD8+ T cell large granular lymphocytes (TLGL) and gamma-delta (γδ) T cells, are increasing in IBM patients. T-LGL expansion is a precursor of T-LGL leukaemia and presents an additional burden for IBM patients. γδ-T cells constitute an unconventional T cell subset whose functions include anti-microbial and anti-tumor immunity. We aim to confirm if 1) if T-LGL leukaemia is associated with IBM and if we can distinguish these T-LGL on the basis of phenotype 2) if γδ-T cells with alterations in their repertoire are expanded in IBM patients and 3) determine the clonal diversity of T cells in peripheral blood and muscle.

Methods: Twenty-five IBM patients (Male =10, Female=15, mean age=73) and seventeen healthy controls (Male =10, Female=7, mean age=65) were recruited through the institute for immunology and infectious disease (IIID), Murdoch University. Blood samples were collected and analysed using flow cytometry. Additionally, muscle biopsies and PBMCs were taken from two IBM patients were analysed via bulk Next Generation TCR sequencing.

Results: T-LGLs are late-differentiated cells that display a characteristic CD8+ CD57+, CD5-, KLRG1+, and variable expression of CD94+, and CD56+ phenotype. 8/25 (32%) of IBM patients showed expansion of T-LGL compared to 17% of healthy controls (H.C). The proliferation of CD5CD57 populations was assessed using ki67. Overall, these populations’ proliferation potential was reduced; however IBM + T-LGL patients show a slightly higher frequency of ki67 than IBM T-LGL- and HC.

Increased frequency of γδ T cells (>4% of total T cells) is observed in 27% of IBM patients compared to 7% of H.C. The proportion of Vδ1 T cells is slightly greater than Vδ2 T cells, yet is seen in both IBM and H.C The Vδ1 subset in IBM and H.C display higher proportion of TEMRA cells that show reduced expression of Ki67 yet show higher frequency of CD27- and variable expression of CX3CR1 and CD57. The Vδ2 cell has a higher proportion of TEM and has slightly higher expression of Ki67 and displays the highest frequencies of CX3CR1 yet CD57 frequency was variable. Frequency of Vδ2 CD27+ was highest in IBM patients compared to healthy controls.

Lastly, the TCR sequence analysis of peripheral blood CD4 and CD8 and muscle homogenate reveals common TCRβV and TCRβJ rearrangements and CDR3 sequences and/or terminal motifs across the three samples and between the two IBM donors.

Conclusions: We found evidence of two populations of rare T cells expanding in IBM patients; however, we also see a considerable portion of healthy controls with the same phenotypic changes proposing that these changes are due to the effects of ageing. TCR sequences analysis indicates an oligoclonal repertoire with a few common sequences found between IBM patients suggesting an immunogenic response directed towards common antigen(s).

Item Type: Thesis (Honours)
Murdoch Affiliation(s): School of Veterinary and Life Sciences
Supervisor(s): Coudert, Jerome
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