Plasma and fecal zonulin are not altered by a high green leafy vegetable dietary intervention: secondary analysis of a randomized control crossover trial

Abstract

Background Zonulin is observed in animal models to regulate intestinal permeability and influenced by dietary intake, gut microbiota, and inflammation. We conducted a secondary analysis of a randomized controlled crossover trial (NCT03582306) in individuals with a BMI greater than 30 kg/m2 and high habitual red meat intake and low habitual green leafy vegetable (GLV) intake. Methods Participants were provided with frozen GLV during the first or last four weeks (immediate or delayed intervention) of the twelve-week trial. Biological and anthropometric measures were taken at the beginning and at each four-week interval. A subset of 20 participants was selected for this secondary analysis of the intestinal permeability and inflammation-related biomarkers: serum and fecal zonulin; serum lipopolysaccharide binding protein (LBP), Alpha-1-acid glycoprotein 1 (ORM-1), tumor necrosis factor α (TNFα), interleukin-6 (IL-6), and C-reactive protein; 8-hydroxy-2'-deoxyguanosine (8OHdG) and plasma Vitamin K1 as a marker of protocol adherence. Nutrient and food group intake from two-24-h dietary recalls collected at each time point were assessed. Fecal microbiota was measured by 16 s rRNA PCR sequencing. Changes in biological markers, dietary factors, and microbial taxa were assessed with Wilcoxon Sign Ranks Tests. Exploratory analyses of the relationship between changes in outcome variables were conducted with Spearman correlations. Results No changes in serum and fecal zonulin and serum LBP were observed. Plasma Vitamin K (p = 0.005) increased, while plasma 8OHdG (p = 0.023) decreased during the intervention compared to the control. The only dietary factors that changed significantly were increases during intervention in Vitamin K and Dark GLV (p < 0.001 for both) compared to control. Fecal microbiota did not change significantly across all times points; however, change in serum zonulin was associated with change in Proteobacteria (ρ = − 0.867, p = 0.001) in females and Bifidobacterium (ρ = − 0.838, p = 0.009) and Bacteroidaceae (ρ = 0.871, p = 0.005) in men. Conclusions A high GLV dietary intervention increased serum zonulin levels and had no effect on fecal zonulin. Lack of concordance between several inflammation-associated biomarkers and zonulin corroborate recent reports of limited utility of zonulin in obese adults free of lower gastrointestinal disease. Trial Registration information: https://clinicaltrials.gov/ct2/show/NCT03582306 (NCT03582306) registered on 07/11/2018.

Assessment of Intestinal Permeability and Inflammation Bio-Markers in Patients with Rheumatoid Arthritis

“…This issue may not be relevant if these ELISA kits correlate strongly with functional gut permeability as assessed via established tests [ 13 ]. At present, this is the best method for measuring zonulin levels in human samples and is widely used for measuring intestinal permeability [ 61 ].…”

“…Environmental factors such as diet may have an important role in regulating intestinal integrity [ 61 , 62 ]. A study that targeted intestinal permeability by treating mice with butyrate found that a reduction in the severity of the observed arthritis was due to the improvement of intestinal permeability and was associated with a reduction in serum zonulin concentrations [ 8 ].…”

“…A total of 61 patients were enrolled in the study, comprising 20/61 (33%) males and 41/61 (67%) females. The median age was 65 (58)(59)(60)(61)(62)(63)(64)(65)(66)(67)(68)(69)(70)(71) years for females and 57 (53.3-61.5) years for males, and the median disease duration was 3 (1-11.5) years for females and 0.8 (0.5-3.9) years for males. In this sample, 6/61 (9.8%) were current smokers (3 female and 3 male) and 29/61 (47.5%) had never smoked.…”

Impaired Intestinal Permeability Assessed by Confocal Laser Endomicroscopy—A New Potential Therapeutic Target in Inflammatory Bowel Disease

“…It is considered one of the main factors that guarantee the good function of the intestinal barrier due to its effects on epithelial tightness. Increased concentrations of zonulin in blood or feces have been correlated with the important alteration of intestinal permeability in the case of diabetes, IBS, or IBD [ 52 ].…”