Small intestinal CD8+TCRγδ+NKG2A+ intraepithelial lymphocytes have attributes of regulatory cells in patients with celiac disease
Govind Bhagat, … , Peter H.R. Green, John S. Manavalan
Govind Bhagat, … , Peter H.R. Green, John S. Manavalan
Published December 6, 2007
Citation Information: J Clin Invest. 2008;118(1):281-293. https://doi.org/10.1172/JCI30989.
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Small intestinal CD8+TCRγδ+NKG2A+ intraepithelial lymphocytes have attributes of regulatory cells in patients with celiac disease

Abstract

Intraepithelial lymphocytes (IELs) bearing the γδ TCR are more abundant in the small intestinal mucosa of patients with celiac disease (CD) compared with healthy individuals. However, their role in disease pathogenesis is not well understood. Here, we investigated the functional attributes of TCRγδ+ IELs isolated from intestinal biopsies of patients with either active celiac disease (ACD) or those on a gluten-free diet (GFD). We found that compared with individuals with ACD, individuals on GFD have a higher frequency of CD8+TCRγδ+ IELs that express the inhibitory NK receptor NKG2A and intracellular TGF-β1. TCR triggering as well as cross-linking of NKG2A increased both TGF-β1 intracellular expression and secretion in vitro. Coculture of sorted TCRγδ+NKG2A+ IELs, IL-15–stimulated TCRαβ+ IELs, and HLA-E+ enterocytes resulted in a decreased percentage of cytotoxic CD8+TCRαβ+ IELs expressing intracellular IFN-γ and granzyme-B and surface NKG2D. This inhibition was partially abrogated by blocking either TGF-β alone or both NKG2A and HLA-E. Thus, our data indicate that suppression was at least partially mediated by TGF-β secretion as a result of engagement of NKG2A with its ligand, HLA-E, on enterocytes and/or TCRαβ+ IELs. These findings demonstrate that human small intestinal CD8+TCRγδ+ IELs may have regulatory potential in celiac disease.

Authors

Govind Bhagat, Afzal J. Naiyer, Jayesh G. Shah, Jason Harper, Bana Jabri, Timothy C. Wang, Peter H.R. Green, John S. Manavalan

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Figure 4

Suppression by CD3+TCRγδ+ NKG2A+ IELs requires interaction with HLA-E.

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Suppression by CD3+TCRγδ+ NKG2A+ IELs requires interaction with HLA-E. ...
(A) Percentage (mean ± SEM) of CD3+TCRαβ IELs expressing intracellular IFN-γ and granzyme B in cocultures of CD3+TCRαβ IELs with ESA+ ECs, in the presence of rhIL-15. Addition of CD3+TCRγδ+ NKG2A+ IELs to these cultures resulted in a statistically significant decrease (*) in the percentage of CD3+TCRαβ+ IELs expressing intracellular IFN-γ (P = 0.002) and granzyme B (P = 0.001). No significant decrease (‡) in the percentage of CD3+TCRαβ+ IELs expressing intracellular IFN-γ (P = 0.1) and granzyme B (P = 0.3) was seen in cultures to which CD3+TCRγδ+ NKG2A– IELs were added. When either 10 μg/ml anti-human NKG2A, 10 μg/ml anti-human HLA-E (MEM-E/06), or a cocktail of anti-human NKG2A and anti-human HLA-E were added to cultures containing CD3+TCRγδ+NKG2A+ IELs, the percentage of CD3+TCRαβ IELs expressing intracellular IFN-γ increased by 38.5% ± 10.2%, 43.0% ± 9.8% and 46.2% ± 9.5%, respectively, as did intracellular granzyme B–expressing CD3+TCRαβ IELs (27.0% ± 7.4%, 38.2% ± 9.1%, and 44.2% ± 10.2%). (B) Representative flow cytometry dot plots showing percentages of CD3+TCRαβ+ IELs that express HLA-E, from healthy controls, patients with ACD, and patients on GFD. (C) Box-and-whisker plots representing frequencies of CD3+TCRαβ+ IELs expressing HLA-E in jejunal biopsy samples from ACD patients (n = 8), GFD patients (n = 12), and controls (n = 7). (D) Representative flow cytometry histogram showing an increase in the percent of HLA-E–expressing CD3+TCRαβ+ IELs sorted from the jejunal biopsy of an ACD patient, after 24 hours treatment with 25 ng/ml of rhIFN-γ. (E) Percentage (mean ± SEM) of CD3+TCRαβ IELs expressing intracellular IFN-γ and granzyme B in cocultures of CD3+TCRαβ IELs treated with 25 ng/ml of rhIL-15 in the bottom chamber of a diffusion chamber system. When CD3+TCRγδ+NKG2A+ IELs were added to the bottom chamber or CD3+TCRγδ+NKG2A+ IELs with sorted HLA-E+CD3+TCRαβ IELs (from an ACD patient) were added to the top chamber, a significant decrease (*) in the percentage of CD3+TCRαβ IELs expressing intracellular IFN-γ (P = 0.004 and P = 0.006, respectively) and granzyme B (P = 0.003 for both) was seen. No significant decrease (‡) in the percentage of CD3+TCRαβ IELs expressing intracellular IFN-γ (P = 0.09) and granzyme B (P = 0.08) was seen when CD3+TCRγδ+NKG2A+ IELs and HLA-E–CD3+TCRαβ IELs were added to the top chamber (P values determined by Wilcoxon’s signed-rank test).