Prolyl endopeptidase-degraded low immunoreactive wheat flour attenuates immune responses in Caco-2 intestinal cells and gluten-sensitized BALB/c mice

• Published in: Food and chemical toxicology 2019-07, Vol.129, p.466-475
• Main Authors: Mohan Kumar, B.V., Vijaykrishnaraj, M., Kurrey, Nawneet K., Shinde, Vijay S., Prabhasankar, P.
• Format: Article
• Language: English
• Subjects: AN-PEP; BALB/C mice; Caco-2; Gliadin; Gluten; Immunofluorescence
• ISSN: 0278-6915; 1873-6351
• DOI: 10.1016/j.fct.2019.05.011

Targeted degrading Aspergillus niger-derived prolyl endopeptidase (AN-PEP) is promising in gluten hydrolysis because it specifically cleaves the proline-rich sites in gluten. The current study aims to understand the safety aspects of AN-PEP hydrolysed low immunoreactive wheat flours by testing immune responses using cell line and animal models. In the AN-PEP hydrolysed wheat flour (AN-PEP HWF) gliadin extract, there was no increase in the levels of zonulin-1 (Zo-1) and pro-inflammatory cytokines (IL-6 and IL-8) but a significant increase was noted in the control wheat flour (CWF) gliadin-treated Caco-2 cells. The Zo-1 localization in Caco-2 cells was significantly noted in the reacted positive fluorescence cells that were treated with the control wheat flour. Further, a safety evaluation of HWF was carried out in gluten-sensitized BALB/c mice. Mouse anti-gliadin (IgG, IgA and IgE) antibodies were significantly generated in the CWF treated animals rather than the AN-PEP HWF groups. The serum pro-inflammatory (IL-1β, IL-4, IL-6, IL-15, TNF-α and IFN-γ) markers were observed in significant levels in CWF challenged mice and a similar trend was observed in ex-vivo splenocyte cells. A small intestine histopathological sectioning revealed that there are no abnormalities or structural changes in AN-PEP HWF challenged mice. [Display omitted] •Prolyl endopeptidase enzyme cleaves the immunoreactive peptides in wheat flour.•Zonulin (ZO-1) proteins are not expressed in hydrolysed flour treated Caco-2 cells.•Pro-inflammatory markers are significantly reduced in hydrolysed flour treated animals.•In-vitro and in-vivo analysis of treated cells and animals reveal that hydrolysed flour is safe.•Small intestine villi brush border does not show any adverse morphological changes.