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. 2022 Jun 28;11(7):1283.
doi: 10.3390/antiox11071283.

Immunomodulatory Effects of Aronia Juice Polyphenols-Results of a Randomized Placebo-Controlled Human Intervention Study and Cell Culture Experiments

Sonja Lackner  1 , Tommaso Sconocchia  1   2 , Tobias Ziegler  1   3 , Christina Passegger  1 , Nathalie Meier-Allard  1 , Elke Schwarzenberger  1 , Willibald Wonisch  4 , Theresa Lahousen  5 , Alexandra Kohlhammer-Dohr  5 , Sabrina Mörkl  5 , Martina Derler  1   6 , Herbert Strobl  1 , Sandra Johanna Holasek  1
Affiliations

Immunomodulatory Effects of Aronia Juice Polyphenols-Results of a Randomized Placebo-Controlled Human Intervention Study and Cell Culture Experiments

Sonja Lackner et al. Antioxidants (Basel). .

Abstract

Dietary polyphenols, which are present in Aronia melanocarpa, have been associated with various beneficial effects on human health including antioxidant, antiviral, and anti-inflammatory activities. We aimed to investigate the immunomodulatory effects of aronia juice polyphenols in a randomized placebo-controlled human intervention study and cell culture experiments. A total of 40 females were asked to consume either 200 mL of aronia juice or a placebo drink for six weeks and were investigated again after a washout period of another six weeks. We observed that only half of the participants tolerated the aronia juice well (Vt) and the other half reported complaints (Vc). The placebo (P) was generally tolerated with one exception (p = 0.003). Plasma polyphenol levels increased significantly in Vt after the intervention (p = 0.024) but did neither in P nor in Vc. Regulatory T cell (Treg) frequencies remained constant in Vt and P during the intervention, whereas Tregs decreased in Vc (p = 0.018). In cell culture, inhibiting effects of ferulic acid (p = 0.0005) and catechin (p = 0.0393) on the differentiation of Tregs were observed as well as reduced activation of CD4-T cells in ferulic acid (p = 0.0072) and aronia juice (p = 0.0163) treated cells. Interestingly, a CD4+CD25-FoxP3+ cell population emerged in vitro in response to aronia juice, but not when testing individual polyphenols. In conclusion, our data strengthen possible individual hormetic effects, the importance of the food matrix for bioactivity, and the need for further investigations on possible impacts of specific physiological features such as the gut microbiota in the context of personalized nutrition.

Keywords: Aronia melanocarpa juice; bioavailability; hormesis; immunomodulation; oxidative stress; polyphenols; regulatory T cells; tolerability.

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Conflict of interest statement

The authors declare no conflict of interest, except W.W. who is affiliated with and employed at Omnignostica Ltd. where some of the kits were bought.

Figures

Figure 1
Figure 1
Study design: the participants were investigated at three time points at baseline (I), after the intervention period of six weeks (II), and after another six weeks of washout (III) to observe any potential persistent effects of the intervention. The participants were asked to consume 100 mL of the aronia juice or the polyphenol-free placebo drink, respectively, twice a day for six weeks.
Figure 2
Figure 2
Progression of total plasma polyphenol microtiter (PPm) concentrations over the three investigation time points: (a) PPm increased significantly between baseline (I) and after the intervention (II) in the whole verum (V) group (p = 0.040); however, the significance did not remain after Bonferroni correction (p = 0.119) and PPm of the placebo (P) group did not change during the intervention. PPm decreased in both groups over time. (b) The separation of V into participants who tolerated the juice (Vt) compared to those who reported complaints (Vc) revealed significantly lower PPm in Vt at I (p = 0.010). PPm in Vt increased significantly from I to II (p = 0.024) whereas the PPm plasma concentrations of Vc and P remained constant. After Bonferroni correction, the increase in Vt did not remain significant. (c) The absolute difference of plasma PPm between the measurement time points (Δ) was the highest in Vt from I to II whereas PPm almost remained stable in Vc and P. The absolute increase in Vt was significantly higher compared to P (p = 0.015) and at least a trend towards significance was observed compared to Vc (p = 0.051). A decrease in PPm plasma concentrations was observed in all three groups during the washout phase. In Vt PPm almost returned to baseline concentrations after the washout period. p-values < 0.05 are marked with *, < 0.01 with **, and < 0.001 with ***. Outlayers of V and Vt are highlighted as black circles, of Vc as circles, and of P as black triangles. Abbreviations: V: verum, Vt: verum tolerated, Vc: Verum complaints, P: placebo, PPm: polyphenol microtiter, I: baseline, II: after intervention, III: after washout, Δ: difference between two measurement points.
Figure 3
Figure 3
Progression of regulatory T-cell frequencies. (a) Treg frequencies decreased in V over the course of the study whilst Tregs in P remained almost constant. (b) When separated for tolerability groups Vc had significantly higher Treg % at baseline compared to Vt and P. Treg values decreased in Vc continuously during the study. (c) Tregs remained almost stable in Vt and P whilst a significant reduction in Treg % was observed for Vc. p-values < 0.05 are marked with *, < 0.01 with **. Outlayers of V and Vt are highlighted as black circles, of Vc as circles, and of P as black triangles. Abbreviations: V: verum, Vt: verum tolerated, Vc: Verum complaints, P: placebo, Tregs: regulatory T-cells, I: baseline, II: after intervention, III: after washout, Δ: difference between two measurement points.
Figure 4
Figure 4
Results of the Treg differentiation experiments. The effects of polyphenols typically present in aronia juice and the natural aronia juice on Treg differentiation and T cell activation have been tested in vitro. (a) Ferulic acid (p = 0.0005) and catechin (p = 0.0393) showed significant inhibitory effects on Treg differentiation whereas the aronia juice slightly promoted Treg differentiation (n.s.); (b) Ferulic acid (p = 0.0072) and the aronia juice (p = 0.0163) significantly inhibited CD25 expression. (c–e) Representative flow cytometry plots from the Treg differentiation experiments. Following the treatment of naïve CD4+ T cells with IL-2 and TGF-β1, T cells were analyzed for the expression of CD4, CD25, and FoxP3. The representative plots show the percentages of CD25 and FoxP3 cells by gated CD4+ cells. Cultures treated with DSMO (c) and placebo drink (d) did not exhibit a CD25-FoxP3+ population; Cells treated with aronia juice (e) showed a unique population of CD25 negative cells that expressed FoxP3. p-values < 0.05 are marked with *, < 0.01 with **, and < 0.001 with ***. Abbreviations: Treg: regulatory T cell, iTreg: induced regulatory T cell, Ctrl: Control; DMSO: Dimethylsulfoxid.
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