CTLs are targeted to kill β cells in patients with type 1 diabetes through recognition of a glucose-regulated preproinsulin epitope
Ania Skowera, … , Bart O. Roep, Mark Peakman
Ania Skowera, … , Bart O. Roep, Mark Peakman
Published September 18, 2008
Citation Information: J Clin Invest. 2008;118(10):3390-3402. https://doi.org/10.1172/JCI35449.
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Research Article

CTLs are targeted to kill β cells in patients with type 1 diabetes through recognition of a glucose-regulated preproinsulin epitope

Abstract

The final pathway of β cell destruction leading to insulin deficiency, hyperglycemia, and clinical type 1 diabetes is unknown. Here we show that circulating CTLs can kill β cells via recognition of a glucose-regulated epitope. First, we identified 2 naturally processed epitopes from the human preproinsulin signal peptide by elution from HLA-A2 (specifically, the protein encoded by the A*0201 allele) molecules. Processing of these was unconventional, requiring neither the proteasome nor transporter associated with processing (TAP). However, both epitopes were major targets for circulating effector CD8+ T cells from HLA-A2+ patients with type 1 diabetes. Moreover, cloned preproinsulin signal peptide–specific CD8+ T cells killed human β cells in vitro. Critically, at high glucose concentration, β cell presentation of preproinsulin signal epitope increased, as did CTL killing. This study provides direct evidence that autoreactive CTLs are present in the circulation of patients with type 1 diabetes and that they can kill human β cells. These results also identify a mechanism of self-antigen presentation that is under pathophysiological regulation and could expose insulin-producing β cells to increasing cytotoxicity at the later stages of the development of clinical diabetes. Our findings suggest that autoreactive CTLs are important targets for immune-based interventions in type 1 diabetes and argue for early, aggressive insulin therapy to preserve remaining β cells.

Authors

Ania Skowera, Richard J. Ellis, Ruben Varela-Calviño, Sefina Arif, Guo Cai Huang, Cassie Van-Krinks, Anna Zaremba, Chloe Rackham, Jennifer S. Allen, Timothy I.M. Tree, Min Zhao, Colin M. Dayan, Andrew K. Sewell, Wendy Unger, Jan W. Drijfhout, Ferry Ossendorp, Bart O. Roep, Mark Peakman

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Figure 7

High ambient glucose increases β cell PPI synthesis and activation of PPI SP epitope PPI15–24–specific CTLs and the efficiency of human β cell killing.

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High ambient glucose increases β cell PPI synthesis and activation of PP...
(A) Agarose gel showing resolution of products of semiquantitative RT-PCR amplification using insulin- and glucagon-specific primers from total RNA extracted from human islet cells cultured for 14 hours in the presence of 5.6, 11, and 20 mM glucose. Results show increasing insulin mRNA synthesis with increasing glucose concentration. (B) Expressed relative to the density of the β-actin gene PCR product, insulin expression is greater than 4-fold higher at 20 mM compared with 5.6 mM glucose. There is an expected reduction (~2-fold) in glucagon expression at 20 mM compared with 5.6 mM glucose due to the known suppression of glucagon secretion by high glucose and insulin concentrations (55–57). (C) Coculture of islet cells at different glucose concentrations with 1E6 PPI15–24–specific T cell clone results in a significant increase (1-way ANOVA, P < 0.01) in TNF-α production by the clone, with a significant linear trend as glucose concentration increases (P < 0.005), indicating an increase in epitope presentation at higher glucose concentration. (D) A similar analysis of islets precultured at different glucose concentrations and prepulsed with PPI15–24 peptide shows that high glucose alone has no effect on T cell clone activation. Symbols indicate different effector/target ratios of 3:1 (circles), 6:1 (diamonds), 12:1 (inverted triangles), and 25:1 (triangles).