(P30-026-23) Buckwheat Bioactive Mitigates Usual Innate Immune Evasion Mechanisms of Human Pathogenic Microorganisms in Whole Blood Model

Objectives: D-fagomine (Fag) is a bioactive present in buckwheat grain. It Is known to inhibit glycosidase enzymes and also the adhesion of some bacteria. During preliminary “ex vivo” human colonic mucosal trials involving a pathogenic E coli strain (AIEC), Fag was found to modulate TNF-α in an apparent double sigmoid curve in the 0 to 5000 mg/L range of concentrations, with the max activity in a middle/low range of 50 mg/L. The aim of this study was to confirm the unexpected effect observed at such a low range of dose, carrying out simpler and more reproducible experimental trials using human whole blood (WB), and also testing other microorganisms with the ability to evade the immune response.

Methods: WB was diluted and treated with Fag (0 or 50 mg/L) and the correspondent inoculum to get 10⁸ CFU/mL of the bacteria E. coli (AIEC), the dimorphic fungus Blastomyces sp (BLS), or the Control (CTR). Collected WB was diluted (1:4) with culture medium (RPMI + 10% FBS). 0,9 ml aliquots were placed in each culture well and incubated at 37ºC in a 95% O2 + 5% CO2 atmosphere during 30 min. Thereafter, inoculum and/or Fag and/or culture medium were added carefully to the wells (50 μL each) to complete additional 100 μL in each well. After 4 h of culture supernatants were collected for measurement of the release of TNF-α which was quantified using ELISA. Results are expressed by the Mean and SEM, all of them in pg/mL (n=3).

Results: The levels of TNF-α produced by immune cells in front of microorganisms with immune evasion mechanisms were higher in the presence of Fag compared to the positive control group. AIEC/Fag⁺: 451.0 ±5.4; AIEC/Fag^-: 355.0 ±14.2; BLS/Fag⁺: 225.2 ±44.4; BLS/Fag^-: 98.7 ±14.3. The levels of TNF-α in the control group without microorganisms are very low and not modified by the presence of Fag. CTR/Fag⁺: 14.0 ±3.1; CTR/Fag^-: 11.5 ±1.3.

Conclusions: These results indicate that Fag at 50 mg/L activates the human innate immune cells preventing evasion mechanisms. The apparent mechanism would seem to be related to complement activation. As far as the proposed range of doses of bioactive are very near to what is possible to get from available regular food, it should be possible a fast clinical translation of the results in this study looking to develop innovative functional food in the immune area.

Funding Sources: None