SARS-CoV-2-specific T cell responses in patients with multisystem inflammatory syndrome in children

Abstract

Multisystem inflammatory syndrome in children (MIS-C) is a severe complication of SARS-CoV-2 infections that occurs in the pediatric population. We sought to characterize T cell responses in MIS-C compared to COVID-19 and pediatric hyperinflammatory syndromes. MIS-C was distinct from COVID-19 and hyperinflammatory syndromes due to an expansion of T cells expressing TRBV11-2 that was not associated with HLA genotype. Children diagnosed with MIS-C, but who were negative for SARS-CoV-2 by PCR and serology, did not display Vβ skewing. There was no difference in the proportion of T cells that became activated after stimulation with SARS-CoV-2 peptides in children with MIS-C compared to convalescent COVID-19. The frequency of SARS-CoV-2-specific TCRs and the antigens recognized by these TCRs were comparable in MIS-C and COVID-19. Expansion of Vβ11-2⁺ T cells was a specific biomarker of MIS-C patients with laboratory confirmed SARS-CoV-2 infections. Children with MIS-C had robust antigen-specific T cell responses to SARS-CoV-2.

Keywords: Coronavirus disease 2019 (COVID-19); Multisystem inflammatory syndrome in children (MIS-C); Pediatrics; Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2); T cell receptor (TCR) repertoire.

Fig. 1

TCR Repertoire Diversity and Clonality in MIS-C. A-B) Shannon Entropy, a measure of repertoire diversity, estimated at fixed sample coverage. C—D) Median frequency (percent) among the top 100 most frequent clones with T cell clone defined as amino acid sequence in CDR3β of the TCR. Summary data on bar graphs is mean ± standard error. Statistical testing for A-D: Kruskal-Wallis test with Dunn's multiple comparisons test. P value * ≤ 0.05, *** ≤ 0.001. CDR3, complementarity determining region 3; TCR, T cell receptor; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2; MIS-C, multisystem inflammatory syndrome in children; peds, pediatric; ctrls, controls; KD, Kawasaki disease; sJIA, systemic juvenile idiopathic arthritis; MAS, macrophage activation syndrome.

Fig. 2

Expansion of T cells expressing TRBV11–2 in MIS-C. A) The frequency of T cell clonotypes expressing each TRBV gene in patients with MIS-C SARSCoV2^pos. B) The frequency of T cell clonotypes expressing TRBV11–2 in each study group. Statistical testing: two-way ANOVA comparing study group and Vβ gene with the Dunnett's multiple comparisons test. P value **** ≤ 0.0001. C) Principal components analysis (PCA) of TRBV gene usage. D) The frequency of T cell clonotypes expressing TRBV11–2 in MIS-C SARSCoV2^pos and adults in the ImmuneCODE database. Statistical testing: Wilcoxon test. Summary data on bar graphs is mean ± standard error in A-B and median with IQR in D. P value, **** ≤ 0.0001, *** = 0.001, ** = 0.01, * = 0.05. SARS-CoV-2, severe acute respiratory syndrome coronavirus 2; MIS-C, multisystem inflammatory syndrome in children; peds, pediatric; ctrl, control; KD, Kawasaki disease; sJIA, systemic juvenile idiopathic arthritis; MAS, macrophage activation syndrome; var., variance.

Fig. 3

HLA Class I Alleles in MIS-C. A) The frequency of each HLA class I genotype in children with pediatric COVID-19 and MIS-C. B) The of frequency each HLA class I genotype in the National Marrow Donor Program database shown by race and ethnicity as defined by US census categories. Peds, pediatric; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2; MIS-C, multisystem inflammatory syndrome in children.

Fig. 4

Diverse TRBJ Gene Usage in MIS-C: A) Heatmap depicting the mean frequency of T cell clonotypes expressing each TRBJ gene in the given study groups. B—C) Chord diagram depicting the Vβ-Jβ pairing of total T cell clonotypes expressing TRBV11–2 in B) children with COVID-19 (n = 10) and C) MIS-C SARSCoV2^pos patients (n = 16). SARS-CoV-2, severe acute respiratory syndrome coronavirus 2; MIS-C, multisystem inflammatory syndrome in children; peds, pediatric; ctrl, control; KD, Kawasaki disease; sJIA, systemic juvenile idiopathic arthritis; MAS, macrophage activation syndrome.

Fig. 5

SARS-CoV-2-Specific T Cells in MIS-C. Peripheral blood mononuclear cells were incubated with either water (negative control) or peptide pools covering the S1, N, and M viral proteins. SARS-CoV2-antigen-specific T cells were identified by their expression of CD154 (CD40 ligand). A) Flow cytometry dot plots depicting CD40L expression on T cells from a healthy control, child with convalescent COVID-19, and child with MIS-C SARS-COV-2^pos. B) Frequency of CD4⁺CD40L⁺ or CD8⁺CD40L⁺ T cells in the given study group. C) Frequency of CD4⁺CD40L⁺ or CD8⁺CD40L⁺ T cells in children with moderate or severe MIS-C SARS-COV-2^pos. D) Frequency of CD4⁺CD40L⁺ or CD8⁺CD40L⁺ T cells in MIS-C SARS-COV-2^pos patients with and without expansions of T cell clonotypes expressing TRBV11–2. Statistical testing: Kruskal-Wallis test with Dunn's multiple comparisons test for panel B and for Mann Whitney tests for panels C—D. P value * ≤ 0.05, ** ≤ 0.01. Summary data on bar graphs is mean ± standard error. SARS-CoV-2, severe acute respiratory syndrome coronavirus 2; MIS-C, multisystem inflammatory syndrome in children; peds, pediatric; ctrl, control; KD, Kawasaki disease.

Fig. 6

SARS-CoV-2-Specific T Cell Receptors in MIS-C. SARS-CoV-2-specific TCR sequences were obtained from the ImmuneCODE database. Antigen-specific T cells were identified by the expression of activation markers following exposure to SARS-CoV-2 peptides. These T cells were then isolated for TCR sequencing. A) The median frequency of the top 10 most abundant SARS-CoV-2-specific TCRs for each patient in each study group. Statistical analysis Kruskal-Wallis test with Dunn's multiple comparison test. B) The median frequency of the top 10 most abundant SARS-CoV-2-specific TCRs in patients with moderate and severe MIS-C. Statistical analysis: Mann-Whitney test C) The median frequency of the top 10 most abundant SARS-CoV-2-specific TCRs that express TRBV11–2 in each patient and each study group. Statistical analysis Kruskal-Wallis test with Dunn's multiple comparison test. D-E) Viral antigens recognized by SARS-CoV-2-specific TCRs in D) MIS-C SARSCOV2^pos and E) pediatric COVID-19. Statistical analysis by permutation test with no difference between MIS-C SARSCOV2^pos and peds COVID-19. P value, ** ≤ 0.01. Summary data on bar graphs is mean ± standard error. SARS-CoV-2, severe acute respiratory syndrome coronavirus 2; MIS-C, multisystem inflammatory syndrome in children; peds, pediatric; ctrl, control; TCR, T cell receptor.