Skip to main page content
Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
, 11 (12)

Stimulatory Response of Celiac Disease Peripheral Blood Mononuclear Cells Induced by RNAi Wheat Lines Differing in Grain Protein Composition

Affiliations

Stimulatory Response of Celiac Disease Peripheral Blood Mononuclear Cells Induced by RNAi Wheat Lines Differing in Grain Protein Composition

Susana Sánchez-León et al. Nutrients.

Abstract

Wheat gluten proteins are responsible for the bread-making properties of the dough but also for triggering important gastrointestinal disorders. Celiac disease (CD) affects approximately 1% of the population in Western countries. The only treatment available is the strict avoidance of gluten in the diet. Interference RNA (RNAi) is an excellent approach for the down-regulation of genes coding for immunogenic proteins related to celiac disease, providing an alternative for the development of cereals suitable for CD patients. In the present work, we report a comparative study of the stimulatory capacity of seven low-gluten RNAi lines differing in grain gluten and non-gluten protein composition, relevant for CD and other gluten pathologies. Peripheral blood mononuclear cells (PBMCs) of 35 patients with active CD were included in this study to assess the stimulatory response induced by protein extracts from the RNAi lines. Analysis of the proliferative response and interferon-gamma (INF-γ) release of PBMCs demonstrated impaired stimulation in response to all RNAi lines. The lower response was provided by lines with a very low content of α- and γ-gliadins, and low or almost devoid of DQ2.5 and p31-43 α-gliadin epitopes. The non-gluten protein seems not to play a key role in PBMC stimulation.

Keywords: NCWS; PBMCs; RNAi; celiac disease; low-gluten wheat.

Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
Grain protein composition of RNAi lines and wild-type (BW208) lines: (a) Protein distribution of three main protein groups in the wheat grain; (b) Protein distribution of prolamin fractions. NGPs, non-gluten proteins; HMW, high-molecular-weight glutenin subunits; LMW, low-molecular-weight glutenin subunits. Dunnett multiple comparison of means with BW208 wild-type line; ns = not significant; * p < 0.05; ** p < 0.01; *** p < 0.001.
Figure 2
Figure 2
(a) Average number of peptides per protein and genotype identified by liquid chromatography–tandem mass spectrometry (LC–MS/MS). (b) Number of protein peptides identified corresponding to the different gluten and non-gluten protein fractions. NGPs, non-gluten proteins; ATIs, amylase trypsin inhibitors; HMW, high-molecular-weight glutenin subunits; LMW, low-molecular-weight glutenin subunits. Dunnett multiple comparison of means with BW208 wild-type line; ns = not significant; * p < 0.05; ** p < 0.01; *** p < 0.001.
Figure 3
Figure 3
(a) Number of DQ-restricted epitopes identified in LC–MS/MS peptides. (b) The gluten content — based on monoclonal antibody G12 — was measured by an enzyme-linked immunosorbent assay (ELISA) and expressed in parts per million (ppm). Lines with the same letter are not significant different according to the Tukey test comparison of means (p < 0.05).
Figure 4
Figure 4
(a) Immunogenicity of different RNAi wheat lines. Bars represent the proliferative responses of peripheral blood mononuclear cells (PBMCs) to pepsin and trypsin digested (PT-digested) protein extracts from the wild-type (BW208) and RNAi wheat lines defined as a stimulatory index (SI). Line and dots represent IFN-γ release by PBMCs in response to PT-digested protein extracts from the wild-type (BW208) and RNAi wheat lines. Results represent the mean of 35 patients ± standard deviation (SD). (b) Cell proliferation and (c) IFN-γ production groups according to Tukey HSD multiple range test. Different letters denote significant differences (p < 0.05).
Figure 5
Figure 5
Pearson’s correlation analysis of variables measured by Reverse-Phase High-Performance Liquid Chromatography (RP-HPLC), proteomics, PBMCs and G12 moAb.
Figure 6
Figure 6
PCA analysis of the variables measured by RP-HPLC, proteomics, PBMCs and G12 moAb.

Similar articles

See all similar articles

References

    1. Lebwohl B., Sanders D.S., Green P.H.R. Coeliac disease. Lancet. 2018;391:70–81. doi: 10.1016/S0140-6736(17)31796-8. - DOI - PubMed
    1. Choung R.S., Larson S.A., Khaleghi S., Rubio-Tapia A., Ovsyannikova I.G., King K.S., Larson J.J., Lahr B.D., Poland G.A., Camilleri M.J., et al. Prevalence and Morbidity of Undiagnosed Celiac Disease From a Community-Based Study. Gastroenterology. 2017;152:830–839.e835. doi: 10.1053/j.gastro.2016.11.043. - DOI - PMC - PubMed
    1. Junker Y., Zeissig S., Kim S.-J., Barisani D., Wieser H., Leffler D.A., Zevallos V., Libermann T.A., Dillon S., Freitag T.L., et al. Wheat amylase trypsin inhibitors drive intestinal inflammation via activation of toll-like receptor 4. J. Exp. Med. 2012;209:2395–2408. doi: 10.1084/jem.20102660. - DOI - PMC - PubMed
    1. Tjon J.M., van Bergen J., Koning F. Celiac disease: How complicated can it get? Immunogenetics. 2010;62:641–651. doi: 10.1007/s00251-010-0465-9. - DOI - PMC - PubMed
    1. Brandtzaeg P. The changing immunological paradigm in coeliac disease. Immunol. Lett. 2006;105:127–139. doi: 10.1016/j.imlet.2006.03.004. - DOI - PubMed
Feedback