English corner: Zonulin Pathway as a Therapeutic Target

Zonulin as a Potential Therapeutic Target in Microbiota-Gut-Brain Axis Disorders
Main ideas
- Zonulin is a protein modulator of intestinal permeability.
- Zonulin opens the tight junctions between intestinal cells, temporarily causing leaky gut.
- This pathway is part of the immune system and may represent a defensive mechanism to get rid of allergens.
- In the presence of gliadin (wheat) restoration of the brush border is compromised. Inflammation and autoimmune reaction are amplified.
- Lectins too influence the integrity of tight junctions.
- In the presence of some polyphenols (EGCG) most allergenic proteins are blocked, except alpha-gliadin.
- Trophic amino acids, such as threonine, arginine and glutamine preserves gut mucosa integrity.
- Hydrolyzing or sequestrating agglutinin molecules may represent a sealing pathway, a therapeutic strategy to remain under the radar of the immune system.
- The best option would be to limit drastically bread and pasta, when not yet marked IBS.

Article
What is zonulin?
Zonulin modulates the permeability of tight junctions between cells of the wall of the digestive tract. It was discovered in 2000 by Alessio Fasano and his team at the University of Maryland School of medicine. 
Dr. Fasano found that cells in the human intestine produce a protein that is almost identical to the zonula occludens toxin. This toxin, one of the bacterial toxins associated with Vibrio Cholera, rapidly and reversibly opens the tight junctions between intestinal cells, temporarily causing leaky gut, and subsequently watery, and debilitating diarrhea.
Zonulin as regulator or tight junctions
“The intestinal epithelium with its barrier function is in charge of a tightly controlled antigen trafficking from the intestinal lumen to the submucosa”. Fasano & Sturgeon 2016 (1)
Figure: Tight junctions & allergens
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= = = > Tight junctions are very close to each other. Normal situation. Red arrows: large molecules aren’t allowed to pass the brush barrier.
Zonulin is a protein modulator of intercellular tight junctions. It has been shown to induce a significant and reversible increase in gastroduodenal and small intestinal permeability and is involved in tolerance/immune response balance.
The zonulin action mechanism (2)
When zonulin is secreted, it activates two receptors present at the level of enterocytes (PAR2 and EGFR). It then triggers a cascade of reactions inside the cell, which changes its architecture and causes the tight junction to dismay. Like a relaxing mesh, the permeability of the intestine increases. The action of zonulin is reversible and only temporary. Due to its direct relationship with the cohesion of tight junctions, zonulin is considered a biomarker of intestinal permeability. Its dosage can be done in the blood or in the stool. (3)
Intestinal Mucosal Barrier: Integrity restored during the night
In a physiologically healthy gut, the intestinal barrier is characterized by selective permeability (semi-permeability), allowing the adsorption of water and nutrients.
To modulate the intestinal physiology, trophic amino acids, such as threonine, arginine, and glutamine, play a very important role on the intestinal mucosa and may support increased epithelial turnover rates to improve intestinal recovery following an insult. 
L-Glutamine supplementation decreases intestinal permeability and preserves gut mucosa integrity.
Glutamine and Arginine – These amino acids keep the gut lining healthy and well equipped to heal in the face of constant challenge. => Bone broth (glycine & arginine) + glutamine supplement.
What causes zonulin secretion?
Among the factors leading to the production of zonulin, two ones predominate: the presence of a large quantity of bacteria in the small intestine, and the consumption of gluten.
This pathway may represent a defensive mechanism, which flushes out microorganisms (or undesirable / allergisable molecules) so contributing to the innate immune response of the host against bacterial colonization of the small intestine.
Note: Lectins too influence the integrity of tight junctions. Lectins have the capacity to bind to saccharides (glycoconjugaison).
Stimuli that cause zonulin release in the gut 
Beside bacterial exposure, we have seen that gliadin, a storage protein present in wheat and that often triggers celiac disease in genetically susceptible individuals, but possibly differently according to gene susceptibility and weakness of the individual ground. Differently expressed, the pathology seems nothing to do with the digestive system but however it has the same starting etiology: a defensive mechanism that tries to flash out the invaders, or the so-called wrongly identified ones, due to a misleading / excessive reaction of the immune system.
Back to zonulin
In the presence of gliadin, whose alpha-gliadin is the most allergenic type, contained in wheat, recovery and restoration of the sealing of the intestinal wall (brush border) is compromised. Inflammation and autoimmune reaction are amplified.
Gliadin sequestration
Hydrolyzing or sequestrating agglutinin molecules
So, we know that the defense system is upregulated in presence of wheat gliadin but some polyphenols have demonstrated the capacity to modulate the immune reaction. However it doesn’t say if agglutinin molecules are precipitated / sequestrated like some ECGG from green tea could do it with gliadin from wheat (sequestration).
Int J Mol Sci. 2021 Jan. doi: 10.3390/ijms22020595
Gliadin Sequestration as a Novel Therapy for Celiac Disease: A Prospective Application for Polyphenols.
Aspirin (salicylic acid) has shown the capacity to alleviate side-effects. They are downregulated: less immune reaction, thus lowering inflammation and digestive issues. Note that it's probably a temporary effect if you don't change the way you manage. It calms down but it doesn't deal with the cause. The problem will soon upraise by another way. If your stomach is irritated, I won’t take mere aspirin but white willow bark extract.

Figure: Polyphenols capture / surround the gliadin molecules.
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Interactions between polyphenols and proteins have been explored as potential therapeutic strategies for food allergies, showing that immunoglobulin E responses (IgE) can be reduced and digestion of allergenic proteins can be blocked [[Vous devez être inscrit et connecté pour voir ce lien],[Vous devez être inscrit et connecté pour voir ce lien],[Vous devez être inscrit et connecté pour voir ce lien],[Vous devez être inscrit et connecté pour voir ce lien]]. Recent studies suggest that the anti-nutritional characteristics of polyphenols including flavonoids and procyanidins can be used to physically sequester gliadin proteins, disrupting key steps in disease pathogenesis in vitro.

Gluten proteins
Gluten is a protein found primarily in grains such as wheat, barley and rye [[Vous devez être inscrit et connecté pour voir ce lien]]. These proteins are roughly divided in 2 subclasses: glutenins, a cysteine-rich class of fibrous proteins, and gliadins, which are classified as prolamins due to their richness in proline and glutamine residues [[Vous devez être inscrit et connecté pour voir ce lien]]. Prolamins make approximatively half of the total protein found in mature grains, with the exception of oat and rice. Note there are two types of oat and two types of quinoa, producing a different immune reaction.
Figure: Different intensity in immune reaction for cereals and pseudo-cereals
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Source: Paleo Mom

Percentage of prolamins in cereals
Prolamin levels (gluten) in cereals and pseudo-cereals
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[Vous devez être inscrit et connecté pour voir ce lien]  and see my forum for details (for references given in the above board) (6)
NB: 7.5 (+/2.5) This indicates a standard deviation of 2.5 between several analyzes. 7.5 is an average.
Note: Prolamins are considered to be the toxic fraction of gluten in celiac patients. European legislation tolerates maximum 20 mg/kg if the mention "gluten-free" is seen on a product. Tolerance is often established under 10 mg per portion ingested in most coeliac patients. But not all. Depending on sources, the type of cultivar or cultivation conditions (land and weather) can lead to significant variations in prolamin content.
Note: Percentage of alpha-gliadin in wheat prolamins (7)
The wheat endosperm contains 8–15% protein, of which 80% is gliadins and glutenins. α-gliadins account for 15–30% of the total seed storage proteins in the wheat grain[Vous devez être inscrit et connecté pour voir ce lien]. As we know they are major triggers of celiac disease, a food-sensitive autoimmune disorder.
doi: [Vous devez être inscrit et connecté pour voir ce lien]
Concluding remarks
Genetic predisposition, miscommunication between innate and adaptive immunity, exposure to environmental triggers, and loss of intestinal barrier function secondary to the activation of the zonulin pathway by food-derived environmental triggers or changes in gut microbiota, all seem to be key ingredients involved in the pathogenesis of inflammation, autoimmunity, and cancer.
This process is not auto-perpetuating. It can be modulated or even reversed by preventing the continuous interplay between genes and the environment. If you don’t excite the system and remain under the radar. Otherwise, the adaptive system will have memorized the “enemy”. Zero allergen / gluten will be tolerated.
Since zonulin-dependent TJ dysfunction allows such interactions, new therapeutic strategies aimed at re-establishing the intestinal barrier function by downregulating the zonulin pathway could be undertaken. 

Sources and References
1) Zonulin, a regulator of epithelial and endothelial barrier functions, and its involvement in chronic inflammatory diseases. Craig Sturgeon and Alessio Fasano. 2016
=> See the graphic named “Disrupted barrier function between intestine cells from gluten consumption”
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doi: [Vous devez être inscrit et connecté pour voir ce lien]
1. Gliadin from wheat gluten binds to 2. CXCR3 receptors (ligand to induce migration, with an immune response). 3. Zonulin is produced in response and released through epithelial cells of tight junctions (tj). 4. Zonulin widens tight junctions. 5. Indigested molecules (or micro-organisms) penetrate blood flow. 6. An auto-immune reaction occurs. 7. Immune cells, cytokines and inflammation cause GI disease.
Vocabulary: CXCR3 receptors
CXCR3 receptors are ligands, mainly expressed on the surface of activated T cells, B cells, and natural killer cells, and plays an essential role in infection, autoimmune diseases, and tumor immunity by binding to specific receptors on target cell membranes to induce targeted migration and immune responses.
2) La zonuline, un marqueur de la perméabilité intestinale
Par Julien Venesson
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3) Dosage of zonulin can be done in the blood or in the stool
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4) Lectins attacks membrane integrity
Lectins have the capacity to bind to saccharides (glycoconjugaison). Lectins that bind to the surface of cells potently inhibit membrane repair.
Lectins are proteins that bind carbohydrate structures (glycans) and are found within numerous plant and animal tissues and organisms.
5) Polyphenols capture the gliadin molecules
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Recent studies suggest that preventing the digestion and absorption of gluten proteins by sequestering the protein from interaction with the gastrointestinal tract may be an effective, novel therapy for celiac disease.
6) Type de protéines dans les céréales et pseudo céréales
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Taux de prolamines dans différentes céréales et pseudo-céréales.
Fonctions métaboliques & structurelles, fonction de stockage et fonction de défense.
7) Percentage of alpha-gliadin in wheat prolamins
The wheat endosperm contains 8–15% protein, of which 80% is gliadins and glutenins. α-gliadins account for 15–30% of the total seed storage proteins in the wheat grain[Vous devez être inscrit et connecté pour voir ce lien]. As we know they are major triggers of celiac disease, a food-sensitive autoimmune disorder.
doi: [Vous devez être inscrit et connecté pour voir ce lien]