CD8+T-cell auto-reactivity is dependent on the expression of the immunoproteasome subunit LMP7 in exposed to lipopolysaccharide antigen presenting cells and epithelial target cells

Abstract

Proteasome generates peptides for presentation to CD8+ T-cells by antigen presenting cells (APCs) and peripheral cells. Inflammation alters proteasome to immunoproteasome, producing a different pool of peptides. The effect of immunoproteasome expression in APCs and in peripheral cells on CD8+ T-cell self-tolerance was evaluated. Splenocytes (SP) were used as a source of APCs and CD8+ T-cells. Renal epithelial cells (RC) from the same mouse strain were used as peripheral cells. Lipopolysaccharide (LPS) was used for increasing immunoproteasome expression, whereas transduction with lentiviral particles encoding short-hairpin RNA targeting LMP7 (LnLMP7) were used for decreasing the expression of the LMP7 subunit of the immunoproteasome in SP and/or RC. LMP7 expression was tested with Western blotting, while the cytotoxicity of isolated CD8+ T-cells against RC with LDH release assay. LPS increased LMP7 expression in SP, while did not affect its expression in RC. LnLMP7 decreased LMP7 expression in both LPS-treated and untreated SP and RC. CD8+ T-cell auto-reactivity increased in case of LPS-treated APCs, further enhanced in the event of both APCs and RC treated with LPS. Transduction of APCs or RC with LnLMP7 decreased CD8+ T-cell auto-reactivity, which was further decreased in case of concurrent transduction of both APCs and RC. In conclusion, in controlled in vitro conditions, exposure to LPS of APCs and peripheral cells induces CD8+ T-cell auto-reactivity. Over-expression of the LMP7 subunit of the immunoproteasome in APCs due to LPS exposure, as well as LMP7 expression in peripheral cells, are required for CD8+ T-cell auto-reactivity.