Coxsackievirus infection in the pathogenesis of type 1 diabetes: Linking environmental triggers and autoimmune mechanisms.

Federica Vecchio

05 September 2023

Thesis defence

Pratical info

14h30 - 23h
Conference room Rosalind Franklin
research professional
Reduced mobility access

Under the supervision of Sylvaine You, team T-cell Tolerance, Biomarkers and Therapies in type 1 diabetes

Infections by Enteroviruses such as Coxsackievirus B (CVB) are candidate triggers of type 1 diabetes (T1D). Both a temporal correlation between CVB infection that precedes seroconversion of autoantibodies to β-cell antigens, and a spatial association of CVB with insulitis in pancreatic tissues from T1D patients have been observed. CVB has a tropism for pancreatic β-cells, resulting in their lysis. However, very little is known about the precise impact of CVB on β-cells, including the presentation of viral epitopes by HLA class I (HLA-I) molecules and their recognition by cytotoxic CD8+ T cells. Considering the CVB vaccination trials under development, it is essential to understand the mechanisms and relative weight of β-cell destruction primarily mediated by CVB vs. that secondarily induced by anti-CVB CD8+ T cells, as the latter could enhance β-cell autoimmunity during vaccination.

First, we identified by mass spectrometry the peptides presented by the HLA-A2 and HLA-A3 molecules expressed on the surface of a human ECN90 β-cell line infected with different CVB serotypes. We observed a decreased HLA-I expression after infection, associated with the presentation of only few viral peptides derived from structural and non-structural proteins (n=16). By using a combinatorial analysis with different HLA-A2 or HLA-A3 multimers carrying the CVB peptides identified, and phenotypic markers, we showed that only a fraction (30%) of these peptides is recognized by circulating CD8+ T cells from CVB-seropositive healthy donors, and that only another subfraction is targeted by effectors/memory CD8+ T cells (<50% in most individuals). Interestingly, a CVB epitope homologous to an epitope of the β-cell protein Glutamic Acid Decarboxylase (GAD), a major autoantigen of T1D, was identified and recognized by the same CD8+ T cells, demonstrating cross-reactivity. Immunodominant peptides were also recognized by circulating CD8+ T cells from T1D patients, displaying very low frequencies similar to those observed in non-diabetic donors; and by CD8+ T cells from spleen and pancreatic lymph nodes. The latter expressed the PD-1 marker, suggesting an exhausted phenotype, and shared expanded, yet private, antigen T-cell receptors (TCRs) with peripheral tissues. To facilitate functional analysis of anti-CVB CD8+ T-cell responses, CD8+ T-cell transductants were generated by expressing TCRs recognizing immunodominant CVB peptides in primary CD8+ T cells and then used in real-time imaging cytotoxic assays. We observed that infected β-cells were more effectively killed directly by CVB and only to a lesser extent by anti-CVB CD8+ T cells. In addition, analysis of the morphology of infected β-cells showed the formation of filopodia (thin cytoplasmic protrusions), which allowed the infection to spread to other β-cells.

Thus, my work shows that CVB infection generates a limited CD8+ T-cell response and memory in terms of epitope coverage and frequency, which could predispose to the persistent infections that have been associated with the seroconversion of autoantibodies to β-cell antigens. The predominant destruction of β-cells by CVB, promoted by the spread of infection by filipodia, suggests that the triggering of anti-β-cell autoimmunity may involve a virus-mediated release of autoantigens rather than antiviral CD8+ T-cell responses, in the context of limited immune protection. These findings lend rationale to boost such protection through CVB vaccines and provide biomarkers to follow response to infection and vaccination along the natural history of the disease.