Inflammation and the microbiome

Flavobiotics that change the microbiome, regenerate the intestinal barrier and activate the immune system

Mikrobióm+®

Mikrobióm+® is a top-class flavobiotic from citrus fruit that contains specifically active flavonoids. Mikrobióm+® draws directly on the potential of the gut microbiome and improves the function of the intestinal barrier. Mikrobióm+® is specially designed to give positive support to the activity of the immune system.

In 2019 a clinical study was carried out with a daily dose of 500 mg of Mikrobióm+® to evaluate the benefit for the composition of the gut microbiome. The results showed a significant change in the composition of short-chain fatty acids (SCFAs). This is a direct effect of the change in the composition of the gut microbiome. In addition, a strong trend towards a reduction in faecal calprotectin was shown, which indicated immunomodulation. Both results strengthen the hypothesis that citrus polyphenol compounds are able to modulate the composition and function of the gut microbiota, thereby supporting gut and host health through their anti-inflammatory effects.

Clinical study

Introduction

Developed countries are successfully fighting infectious diseases, but the prevalence of metabolic, cardiovascular and autoimmune diseases is rising (1). The rise in inflammatory and autoimmune diseases is linked to an altered “Western” diet, which is an important factor influencing the intestinal ecosystem, and this in turn may be very important in the development of human diseases. Recent data suggest that various polyphenol compounds, derived for example from citrus fruit, are able to modulate the structure and function of the gut microbiome, thereby favourably influencing gut health (2).

Mikrobióm+® is extracted from citrus fruit with excellent stability and a high concentration of two key flavonoids: hesperidin and naringin.

The benefits of Mikrobióm+® were evaluated in a clinical study, which showed a dual action of the ingredient:

1. The active flavonoids act directly in the intestinal lumen by protecting and markedly improving the function of the intestinal barrier.

2. They activate the immune system and reduce gut inflammation, with a positive change in the composition of the microbiome: as a result, the butyrate level increased (P = 0.031) and the calprotectin level fell, which suggests a lower pathogen load.

Aims

The primary aim of the clinical study was to examine the effect of 12 weeks of Mikrobióm+® supplementation on the intestinal ecosystem and on calprotectin levels in overweight individuals with an unbalanced lipid profile. Because the participants were overweight, they probably showed dysbiosis at the microbiome level and low-grade chronic inflammation in the gut.

Methods

The 12-week randomised, parallel, double-blind, placebo-controlled study was designed to investigate the effects of a daily dose of 500 mg of Mikrobióm+® on the composition of the gut microbiome and on calprotectin levels. The study was carried out in a group of 50 overweight subjects, and therefore with an altered microbiota (Table 1). Each subject attended three test days. After overnight fasting, a stool sample was taken from the subjects on the first and last test day. Blood pressure was measured on all three test days.

Results

1) Short-chain fatty acids

After 12 weeks of daily intake of Mikrobióm+®, the share of butyrate in total SCFAs increased significantly (P = 0.031) (Figure 1).

In addition, the butyrate-to-acetate ratio increased significantly in the Mikrobióm+® group compared with the placebo group (P = 0.020).

2) Changes in the microbiome

After the 12-week intervention, an increase in the butyrate-producing cluster Clostridium cluster XIVa was observed between the Mikrobióm+® and placebo groups, with a change from baseline of 43.9% in the intervention group compared with 15.9% in the placebo group. (Figure 2).

3) Calprotectin levels

After 12 weeks of daily supplementation with Mikrobióm+®, a strong reduction in faecal calprotectin was seen compared with the placebo group (P = 0.058) (Figure 3). This reduction reflects a reduction in the pathogen load in the gut.

In vitro studies

Aims

The aim of the in vitro study was to observe the immediate effect of Mikrobióm+® at lower doses in the TIM-2 model. The TIM-2 model is a well-validated in vitro model of the colon.

In this controlled experiment, two different doses (250 mg and 350 mg) of Mikrobióm+® were administered over 72 hours to the TIM-2 intestinal model, which contained a representative sample of human microbiota from 6 healthy older volunteers.

Results

1) Effect on short-chain fatty acids

After daily intake of Mikrobióm+® for 72 hours, an increase in SCFAs over time was observed. The 350 mg dose caused the greatest increase (Figure 4).

2) Difference in butyrate, acetate and propionate

After 12 weeks of daily intake of Mikrobióm+®, an increase in butyrate, acetate and propionate was observed over time, with acetate showing the largest quantitative increase compared with the control.

Butyrate and propionate regulate the immune system and are also an important modulator of satiety.

Acetate plays an important role in suppressing appetite by stimulating the release of the satiety hormones PYY and GLP-1.

3) Effect on the composition of the microbiota

Roseburia spp. is a member of the Clostridium IVa cluster and is a butyrate-producing bacterial strain that is associated with weight reduction and reduced insulin resistance (prediabetes). After daily intake of Mikrobióm+® for 12 weeks, a significant increase in Roseburia spp. in the microbiome was observed (Figure 5).

B. eggerthii is a species within the phylum Bacteroidetes, more precisely the Bacteroides fragilis group, which plays a role in the production of acetate and the prevention of intestinal inflammation. As shown in the figure below, after daily intake of Mikrobióm+® there was a significant increase in B. eggerthii in the microbiome (Figure 6).

4) A shift in the microbiome towards one similar to that seen with a plant-based diet

At the phylum level, the intervention with Mikrobióm+® led to a relative increase in Bacteroidetes and a relative decrease in Firmicutes. Such a shift in the Bacteroidetes / Firmicutes ratio is often associated with a plant-based diet, which is usually high in fibre and low in fat. Similarly, the increase in Roseburia spp. reverses the trend that we usually see with an animal-based diet.

Effect on the immune system

More than 80% of immune cells reside in the colon, which is the single largest organ with a direct function in our immune system.

Mikrobióm+® exerts its positive effects in precisely this important part of our immune system in several ways:

1) Mediates a change in the microbiota

In vitro studies show that Mikrobióm+® can significantly modulate the gut microbiota.

It influences the growth of specific bacterial groups, in particular Clostridium cluster XIVa, which are able to provide health benefits through the production of SCFAs (e.g. butyrate, acetate, propionate).

2) Improves the function of the intestinal barrier

In vitro experiments showed that butyrate reduces permeability and improves the function of the intestinal barrier (measured as transepithelial resistance). Specifically, butyrate mediates the formation of tight junction proteins, such as claudins and occludins, via AMPK.

The formation of tight junction proteins is a dynamic process that is crucial for creating and maintaining the function of the intestinal barrier.

3) Reduces gut inflammation

As shown in the first clinical study, calprotectin – a clinical marker of intestinal inflammation – decreased as a result of consuming Mikrobióm+®.

In cooperation with Maastricht University, a mechanistic study was carried out on an innovative 2-dimensional in vitro model of Caco2 cell lines combined with LPS-stimulated macrophages (Figure 8).

The purpose of the study was to find out whether specific flavonoid metabolites in Mikrobióm+® influence mediators of inflammatory reactions, such as TNF-α.

Hesperetin and naringenin, both direct metabolites of Mikrobióm+®, have a direct and significant effect on the release of TNF-α. By reducing the release of TNF-α, it is possible to counteract chronic low-grade inflammation in the gut (Figures 9 and 10).

Conclusion

Mikrobióm+® supports gut and host health through its anti-inflammatory effects, owing to its ability to modulate the composition of the gut microbiome. The significant shift from acetate to the health-beneficial butyrate and the marked increase in the share of butyrate in total SCFAs are the result of the altered composition of the microbiome.

Butyrate has a positive effect on the function of the intestinal barrier and reduces gut inflammation, which is reflected in lower calprotectin levels. That is why Mikrobióm+® is a top-class flavobiotic that strengthens the intestinal barrier and increases immunity.

References

1. Manzel A, Muller DN, Hafler DA, Erdman SE, Linker RA, Kleinewietfeld M. Role of “Western diet” in inflammatory autoimmune diseases. Current allergy and asthma reports. 2014;14(1):404.
2. Romier B, Schneider YJ, Larondelle Y, During A. Dietary polyphenols can modulate the intestinal inflammatory response. Nutr Rev. 2009;67(7):363-78.
3. Guinane CM, Cotter PD. Role of the gut microbiota in health and chronic gastrointestinal disease: understanding a hidden metabolic organ. Therapeutic advances in gastroenterology. 2013;6(4):295-308.
4. Canani RB, Costanzo MD, Leone L, Pedata M, Meli R, Calignano A. Potential beneficial effects of butyrate in intestinal and extraintestinal diseases. World Journal of Gastroenterology : WJG. 2011;17(12):1519-28.
5. Stevens Y, Rymenant EV, Grootaert C, Camp JV, Possemiers S, Masclee A, et al. The Intestinal Fate of Citrus Flavanones and Their Effects on Gastrointestinal Health. Nutrients. 2019;11(7):1464.
6. Wang, Xiaozhen, Xican Li, and Dongfeng Chen. 2011. “Evaluation of Antioxidant Activity of Isoferulic Acid in Vitro.” Natural product communications 6(9):1285–88. Retrieved December 13, 2017 (http://www.ncbi.nlm.nih.gov/pubmed/21941899).