Oral intake of xanthohumol attenuates lipoteichoic acid-induced inflammatory response in human PBMCs

Abstract

Purpose: The aim of the study was to determine if xanthohumol, a prenylated chalcone found in Hop (Humulus lupulus), has anti-inflammatory effects in healthy humans if applied in low doses achievable through dietary intake.

Methods: In a placebo-controlled single-blinded cross-over design study, 14 healthy young men and women either consumed a beverage containing 0.125 mg xanthohumol or a placebo. Peripheral blood mononuclear cells (PBMCs) were isolated before and 1 h after the intake of the beverages. Subsequently, PBMCs were stimulated with or without lipoteichoic acid (LTA) for 24 and 48 h. Concentrations of interleukin-1β (IL-1β), interleukin-6 (IL-6) and soluble cluster of differentiation (sCD14) protein were determined in cell culture supernatant. Furthermore, hTLR2 transfected HEK293 cells were stimulated with LTA in the presence or absence of xanthohumol and sCD14.

Results: The stimulation of PBMCs with LTA for 24 and 48 h resulted in a significant induction of IL-1β, IL-6, and sCD14 protein release in PBMCs of both, fasted subjects and subjects after the ingestion of the placebo. In contrast, after ingesting xanthohumol, LTA-dependent induction of IL-1β, IL-6, and sCD14 protein release from PBMCs was not significantly higher than in unstimulated cells after 48 h. In hTLR2 transfected HEK293 cells xanthohumol significantly suppressed the LTA-dependent activation of cells, an effect attenuated when cells were co-incubated with sCD14.

Conclusion: The results of our study suggest that an ingestion of low doses of xanthohumol can suppress the LTA-dependent stimulation of PBMCs through mechanisms involving the interaction of CD14 with TLR2. Study registered at ClinicalTrials.gov (NCT04847193, 22.03.2022).

Keywords: Hop; Inflammation; LTA; TLR2; Xanthohumol.

Fig. 1

Study design and cytokine concentrations in supernatant of stimulated PBMCs. Graphical illustration of the general study design a and on the study days b as well as protein concentrations of IL-1β c and IL-6 d in cell culture supernatant of cells stimulated with 0 or 10 µg/ml LTA isolated from healthy study participants receiving either a placebo or xanthohumol. IL interleukin, LTA lipoteichoic acid, PBMC peripheral blood mononuclear cell, XN xanthohumol. Data are expressed as means ± SEM. *p < 0.05. Figure was created with Microsoft PowerPoint (a, b) and GraphPad Prism 7 (c, d)

Fig. 2

Fluorescence imaging of PBMCs isolated from study participants as well as hTLR2 receptor activities of HEK293 cells stimulated with 0–8 μg/ml xanthohumol for 12 h. Fluorescence imaging of PBMCs (a) as well as lymphocytes and monocytes (b) obtained from study participants before and after oral ingestion of xanthohumol (magnification: 630×) and hTLR2 receptor activities from HEK293 cells co-incubated with lipoteichoic acid (10 μg/ml) and increasing concentrations of xanthohumol (0–8 μg/ml) (c). hTLR2 human toll-like receptor 2, LTA lipoteichoic acid, PBMC peripheral blood mononuclear cell, XN xanthohumol. Data are expressed as means ± SEM. *p < 0.05. Figure was created with Leica LAS X (a, b) and GraphPad Prism 7 (c)

Fig. 3

Protein concentration of TLR2 in cell lysate and sCD14 in cell culture supernatant of LTA-stimulated PBMCs as well as effect of sCD14 on LTA-stimulated HEK293 cells co-incubated with xanthohumol. Representative blots (a) and densitometric analysis of TLR2 western blot (b) in cell lysate as well as representative blots (c) and densitometric analysis of sCD14 western blot (d) in cell culture supernatant of LTA-stimulated (10 μg/ml) PBMCs obtained from study participants either receiving a placebo or xanthohumol. hTLR2 receptor activities of HEK293 cells incubated with xanthohumol (3 μg/ml) and sCD14 (200 ng/ml) (e). LTA lipoteichoic acid, hTLR2 human toll-like receptor 2, sCD14 soluble cluster of differentiation 14, PBMC peripheral blood mononuclear cell, XN xanthohumol. Data are expressed as means ± SEM. *p < 0.05. Figure was created with Bio-Rad Image Lab 4.0 (a, c) and GraphPad Prism 7 (b, d, e)

Fig. 4

Schematic summary of the study results. CD14 cluster of differentiation 14, LTA lipoteichoic acid, TLR2 toll-like receptor 2. Figure was created with Microsoft PowerPoint