The role of Killer immunoglobulin-like receptors (KIRs) in the genetic susceptibility to non-celiac wheat sensitivity (NCWS).

OBJECTIVES: Non-celiac wheat sensitivity (NCWS) is an emerging clinical condition characterized by gastrointestinal and extraintestinal symptoms following the ingestion of gluten-containing foods in patients without celiac disease (CD) or wheat allergy. Despite the great interest for NCWS, the genetic risk factors still need to be fully clarified. In this study, we first assessed the possible contribution of KIR genes and KIR haplotypes on the genetic predisposition to NCWS. METHODS: Fifty patients with NCWS, 50 patients with CD, and 50 healthy controls (HC) were included in this study. KIR genes and KIR genotyping were investigated in all subjects by polymerase chain reaction with the sequence oligonucleotide probe (PCR-SSOP) method using Luminex technology. RESULTS: We found a statistically different distribution of some KIR genes among NCWS, CD, and HC. Specifically, NCWS showed a decreased frequency of KIR2DL1, -2DL3, -2DL5, -2DS2, -2DS3, -2DS4, -2DS5, and -3DS1 genes, and an increased frequency of -3DL1 gene respect to both CD and HC. No difference was detected in the KIR haplotype expression. At the multivariate analysis, KIR2DL5, -2DS4, and -2DS5 were independent predictors of NCWS. CONCLUSIONS: Our findings suggest a role of KIR genes in NCWS susceptibility, with KIR2DL5, -2DS4, and -2DS5 having a protective effect. Further large-scale multicentric studies are required to validate these preliminary findings.

Evaluation of gut microbiota of iranian patients with celiac disease, non-celiac wheat sensitivity, and irritable bowel syndrome: are there any similarities?

BACKGROUND AND AIMS: Individuals with celiac disease (CD), non-celiac wheat sensitivity (NCWS), and irritable bowel syndrome (IBS), show overlapping clinical symptoms and experience gut dysbiosis. A limited number of studies so far compared the gut microbiota among these intestinal conditions. This study aimed to investigate the similarities in the gut microbiota among patients with CD, NCWS, and IBS in comparison to healthy controls (HC). MATERIALS AND METHODS: In this prospective study, in total 72 adult subjects, including CD (n = 15), NCWS (n = 12), IBS (n = 30), and HC (n = 15) were recruited. Fecal samples were collected from each individual. A quantitative real-time PCR (qPCR) test using 16S ribosomal RNA was conducted on stool samples to assess the relative abundance of Firmicutes, Bacteroidetes, Bifidobacterium spp., and Lactobacillus spp. RESULTS: In all groups, Firmicutes and Lactobacillus spp. had the highest and lowest relative abundance respectively. The phylum Firmicutes had a higher relative abundance in CD patients than other groups. On the other hand, the phylum Bacteroidetes had the highest relative abundance among healthy subjects but the lowest in patients with NCWS. The relative abundance of Bifidobacterium spp. was lower in subjects with CD (P = 0.035) and IBS (P = 0.001) compared to the HCs. Also, the alteration of Firmicutes to Bacteroidetes ratio (F/B ratio) was statistically significant in NCWS and CD patients compared to the HCs (P = 0.05). CONCLUSION: The principal coordinate analysis (PCoA), as a powerful multivariate analysis, suggested that the investigated gut microbial profile of patients with IBS and NCWS share more similarities to the HCs. In contrast, patients with CD had the most dissimilarity compared to the other groups in the context of the studied gut microbiota.

LC-MS/MS quantitation of α-amylase/trypsin inhibitor CM3 and glutathione during wheat sourdough breadmaking.

AIMS: This study aimed to quantify α-amylase/trypsin inhibitor (ATI) CM3 and glutathione (GSH) during wheat sourdough breadmaking. METHODS AND RESULTS: Breads were made with two wheat cultivars and fermented with Fructilactobacillus sanfranciscensis, F. sanfranciscensis ΔgshR or Latilactobacillus sakei; chemically acidified and straight doughs served as controls. Samples were analysed after mixing, after proofing and after baking. GSH and CM3 were quantified by multi-reaction-monitoring-based methods on an LC-QTRAP mass spectrometer. Undigested ATI extracts were further examined by SDS-PAGE. CONCLUSIONS: GSH abundance was similar after mixing and after proofing but increased after baking (p < 0.001), regardless of fermentation. In breads baked with cv. Brennan, the samples fermented with lactobacilli had higher GSH abundance (p < 0.001) than in the controls. CM3 relative abundance remained similar after mixing and after proofing but decreased after baking (p < 0.001) across all treatments. This trend was supported by the SDS-PAGE analysis in which ATI band intensities decreased after baking (p < 0.001) in all experimental conditions. The overall effect of baking exerted a greater effect on the abundances of GSH and CM3 than fermentation conditions. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first report to quantify ATI over the course of breadmaking by LC-MS/MS in sourdough and straight dough processes.

[Food sensitivities of the digestive tract-Part 2: Food intolerances]. / Nahrungsmittelunverträglichkeiten des Verdauungstraktes ­ Teil 2: Nahrungsmittelintoleranzen.

Adverse reactions to food affect about one third of the population. They are based on very different mechanisms and are triggered by specific foods. They are divided into food intolerances, which manifest mainly in the gastrointestinal tract and food allergies, which can also cause extraintestinal symptoms and have an immunological genesis. In adults, food intolerances are significantly more common than food allergies with a prevalence of approximately 10-20%. The most important food intolerances are sugar intolerances, such as lactose and fructose intolerance but intolerances to wheat also play an increasing role. The diagnostics of food intolerances require extensive exclusion diagnostics, whereby in particular irritable bowel syndrome and intestinal dysbiosis must be differentiated. The therapy of food intolerance is primarily based on a targeted elimination diet. In this advanced education article the most important food intolerances are discussed.

Hybrid QconCAT-Based Targeted Absolute and Data-Independent Acquisition-Based Label-Free Quantification Enables In-Depth Proteomic Characterization of Wheat Amylase/Trypsin Inhibitor Extracts.

Wheat amylase/trypsin inhibitors (ATIs) have gained significant relevance as inducers of intestinal and extra-intestinal inflammation. In this study, we present a novel hybrid data-independent acquisition (DIA) liquid chromatography-mass spectrometry (LC-MS) approach, combining QconCAT technology with short microflow LC gradients and DIA and apply the method toward the quantitative proteome analysis of ATI extracts. The presented method is fast, robust, and reproducible and provides precise QconCAT-based absolute quantification of major ATI proteins while simultaneously quantifying the proteome by label-free quantification (LFQ). We analyzed extracts of 60 varieties of common wheat grown in replication and evaluated the reproducibility and precision of the workflow for the quantification of ATIs. Applying the method to analyze different wheat species (i.e., common wheat, spelt, durum wheat, emmer, and einkorn) and comparing the results to published data, we validated inter-laboratory and cross-methodology reproducibility of ATI quantification, which is essential in the context of large-scale breeding projects. Additionally, we applied our workflow to assess environmental effects on ATI expression, analyzing ATI content and proteome of same varieties grown at different locations. Finally, we explored the potential of combining QconCAT-based absolute quantification with DIA-based LFQ proteome analysis for the generation of new hypotheses or assay development.

Gynecological Disorders in Patients with Non-celiac Wheat Sensitivity.

BACKGROUND: Non-celiac wheat sensitivity (NCWS) most frequently presents clinically with irritable bowel syndrome (IBS)-like symptoms, although many extra-intestinal manifestations have also been attributed to it. No studies to date have evaluated the presence and frequency of gynecological symptoms in NCWS. AIM: To evaluate the frequency of gynecological disorders in patients with NCWS. PATIENTS AND METHODS: Sixty-eight women with NCWS were included in the study. A questionnaire investigating gynecological symptoms and recurrent cystitis was administered, and patients reporting symptoms were then examined by specialists. Three control groups were selected: 52 patients with IBS not related to NCWS, 56 patients with celiac disease (CD), and 71 healthy controls. RESULTS: 59% of the patients with NCWS showed gynecological symptoms, a higher frequency than in healthy controls (P = 0.04), IBS controls (P = 0.01) and CD controls (P = 0.02). Menstrual cycle alterations were more frequent in patients with NCWS than in healthy controls (26.5% vs 11.3%; P = 0.03); the patients with NCWS suffered from recurrent vaginitis (16%) and dyspareunia (6%) significantly more frequently than healthy controls. Twenty-nine percent of patients with NCWS reported recurrent cystitis, a finding higher than in the control groups (vs healthy P = 0.0001, vs IBS P = 0.001, vs CD controls P = 0.04). Microbiological examinations were negative in most of the patients with NCWS and recurrent vaginitis or cystitis. During the 1-year follow-up, 46% of patients with menstrual disorders and 36% with recurrent vaginitis reported resolution of symptoms on a wheat-free diet. CONCLUSIONS: Patients with NCWS showed a significantly higher frequency of gynecological symptoms and recurrent cystitis than patients with IBS.

Tritordeum breads are well tolerated with preference over gluten-free breads in non-celiac wheat-sensitive patients and its consumption induce changes in gut bacteria.

BACKGROUND: The ingestion of wheat and other cereals are related to several gut disorders. The specific components responsible for non-celiac wheat-sensitivity (NCWS) may include gluten and other compounds. Tritordeum is a new cereal derived from crossing durum wheat with a wild barley species, which differs from bread wheat in its gluten composition. In the present work, we examined the response of NCWS patients to tritordeum bread Gastrointestinal symptoms as well as tritordeum acceptability, gluten immunogenic peptides excretion, and the composition and structure of the intestinal microbiota were evaluated. RESULTS: Gastrointestinal symptoms of the subjects showed no significant change between the gluten-free bread and the tritordeum bread. Participating subjects rated tritordeum bread higher than the gluten-free bread. Analysis of the bacterial gut microbiota indicated that tritordeum consumption does not alter the global structure and composition of the intestinal microbiota, and only a few changes in some butyrate-producing bacteria were observed. CONCLUSIONS: All the results derived from acceptability, biochemical and microbiological tests suggest that tritordeum may be tolerated by a sub-set of NCWS sufferers who do not require strict exclusion of gluten from their diet. © 2020 Society of Chemical Industry.

Deciphering the immunogenic potential of wheat flour: a reference map of the salt-soluble proteome from the U.S. wheat Butte 86.

BACKGROUND: Within the complex wheat flour proteome, the gluten proteins have attracted most of the attention because of their importance in determining the functional properties of wheat flour doughs and their roles in human health conditions such as celiac disease and food allergies. However, certain non-gluten proteins also trigger immunological responses but may be present in flour in low amounts or obscured by the more abundant gluten proteins in two-dimensional gels of total protein preparations. METHODS: Non-gluten proteins were preferentially extracted from the flour with a dilute salt solution and separated by two-dimensional gel electrophoresis. Proteins in 173 gel spots were identified by tandem mass spectrometry after cleavage with trypsin or chymotrypsin. Transgenic wheat lines in which specific groups of gluten proteins were suppressed by RNA interference were used to estimate the amount of carry-over of gluten proteins in the salt-soluble protein fraction. RESULTS: Fifty-seven different types of non-gluten proteins were identified, including 14 types that are known or suspected immunogenic proteins. The predominant proteins in 18 gel spots were gluten proteins. Some of these also contained non-gluten proteins. Analysis of the salt-soluble proteins from a transgenic line in which omega-1,2 gliadins were eliminated by RNA interference indicated that certain omega-1,2 gliadins were present in large amounts in the salt-soluble fraction and obscured relatively small amounts of beta-amylase and protein disulfide isomerase. In comparison, analysis of a transgenic line in which alpha gliadins were absent revealed that glyceraldehyde-3 phosphate dehydrogenase was a moderately abundant protein that co-migrated with several alpha gliadins. CONCLUSIONS: In this study, we constructed a proteomic map of the non-gluten protein fraction of wheat flour from the US wheat Butte 86 that complements a proteomic map of the total flour proteins developed previously for the same cultivar. Knowing the identities of low abundance proteins in the flour as well as proteins that are hidden by some of the major gluten proteins on two-dimensional gels is critical for studies aimed at assessing the immunogenic potential of wheat flour and determining which wheat proteins that should be targeted in future gene editing experiments to reduce the immunogenic potential of wheat flour.

More fuel to the fire: some patients with non-celiac self-reported wheat sensitivity exhibit adaptive immunological responses in duodenal mucosa.

BACKGROUND: In contrast to the well-characterized Celiac Disease (CD), the clinical scenarios encompassed by the non-celiac self-reported wheat sensitivity (NCSRWS) might be related to different antigens that trigger distinct immune-inflammatory reactions. Although an increased number of intestinal intraepithelial lymphocytes is observed at the inception of both diseases, the subsequent immunopathogenic pathways seem to be different. We aimed to describe the cytokine profile observed in the duodenal mucosa of patients with NCSRWS. METHODS: In a blind, cross-sectional study, we included duodenal biopsies from 15 consecutive untreated patients with active CD, 9 individuals with NCSRWS and 10 subjects with dyspepsia without CD and food intolerances. Immunohistochemistry and flow-cytometry were used to determine the presence of pro-inflammatory cytokine expressing monocytes and monocyte-derived dendritic cells involved in innate immune activation, cytokine-driven polarization and maintenance of Th1 and Th17/Th 22, and anti-inflammatory/profibrogenic cytokines. RESULTS: The percentage of cells expressing all tested cytokines in the lamina propria and the epithelium was higher in CD patients than in the control group. Cytokines that induce and maintain Th1 and Th17 polarization were higher in CD than in NCSRWS and controls, also were higher in NCSRWS compared to controls. Similar differences were detected in the expression of IL-4 and TGF-1, while IL-10-expressing cells were lower in NCSRWS patients than in controls and CD subjects. CONCLUSIONS: NCSRWS patients exhibit components of both, innate and adaptive immune mechanisms but to a lesser extent compared to CD.

Functional Dyspepsia and Food: Immune Overlap with Food Sensitivity Disorders.

PURPOSE OF REVIEW: Functional dyspepsia (FD) is a chronic functional gastrointestinal disorder characterised by upper gastrointestinal symptoms. Here, we aimed to examine the evidence for immune responses to food in FD and overlap with food hypersensitivity conditions. RECENT FINDINGS: A feature of FD in a subset of patients is an increase in mucosal eosinophils, mast cells, intraepithelial cytotoxic T cells and systemic gut-homing T cells in the duodenum, suggesting that immune dysfunction is characteristic of this disease. Rates of self-reported non-celiac wheat/gluten sensitivity (NCW/GS) are higher in FD patients. FD patients commonly report worsening symptoms following consumption of wheat, fermentable oligosaccharides, disaccharides, monosaccharides, or polyols (FODMAPs), high-fat foods and spicy foods containing capsaicin. Particularly, wheat proteins and fructan in wheat may drive symptoms. Immune mechanisms that drive responses to food in FD are still poorly characterised but share key effector cells to common food hypersensitivities including non-IgE-mediated food allergy and eosinophilic oesophagitis.

Reduction of Allergenic Potential in Bread Wheat RNAi Transgenic Lines Silenced for CM3, CM16 and 0.28 ATI Genes.

Although wheat is used worldwide as a staple food, it can give rise to adverse reactions, for which the triggering factors have not been identified yet. These reactions can be caused mainly by kernel proteins, both gluten and non-gluten proteins. Among these latter proteins, α-amylase/trypsin inhibitors (ATI) are involved in baker's asthma and realistically in Non Celiac Wheat Sensitivity (NCWS). In this paper, we report characterization of three transgenic lines obtained from the bread wheat cultivar Bobwhite silenced by RNAi in the three ATI genes CM3, CM16 and 0.28. We have obtained transgenic lines showing an effective decrease in the activity of target genes that, although showing a higher trypsin inhibition as a pleiotropic effect, generate a lower reaction when tested with sera of patients allergic to wheat, accounting for the important role of the three target proteins in wheat allergies. Finally, these lines show unintended differences in high molecular weight glutenin subunits (HMW-GS) accumulation, involved in technological performances, but do not show differences in terms of yield. The development of new genotypes accumulating a lower amount of proteins potentially or effectively involved in allergies to wheat and NCWS, not only offers the possibility to use them as a basis for the production of varieties with a lower impact on adverse reaction, but also to test if these proteins are actually implicated in those pathologies for which the triggering factor has not been established yet.

Adverse Reactions to Wheat or Wheat Components.

Wheat is an important staple food globally, providing a significant contribution to daily energy, fiber, and micronutrient intake. Observational evidence for health impacts of consuming more whole grains, among which wheat is a major contributor, points to significant risk reduction for diabetes, cardiovascular disease, and colon cancer. However, specific wheat components may also elicit adverse physical reactions in susceptible individuals such as celiac disease (CD) and wheat allergy (WA). Recently, broad coverage in the popular and social media has suggested that wheat consumption leads to a wide range of adverse health effects. This has motivated many consumers to avoid or reduce their consumption of foods that contain wheat/gluten, despite the absence of diagnosed CD or WA, raising questions about underlying mechanisms and possible nocebo effects. However, recent studies did show that some individuals may suffer from adverse reactions in absence of CD and WA. This condition is called non-celiac gluten sensitivity (NCGS) or non-celiac wheat sensitivity (NCWS). In addition to gluten, wheat and derived products contain many other components which may trigger symptoms, including inhibitors of α-amylase and trypsin (ATIs), lectins, and rapidly fermentable carbohydrates (FODMAPs). Furthermore, the way in which foods are being processed, such as the use of yeast or sourdough fermentation, fermentation time and baking conditions, may also affect the presence and bioactivity of these components. The present review systematically describes the characteristics of wheat-related intolerances, including their etiology, prevalence, the components responsible, diagnosis, and strategies to reduce adverse reactions.

Gluten-free living in China: The characteristics, food choices and difficulties in following a gluten-free diet - An online survey.

BACKGROUND: Individuals adhering to a gluten-free diet (GFD) have been scarcely researched within a Chinese population. This present study was conducted to assess the characteristics, food choices, shopping preferences, and emotions and attitudes of individuals following a GFD. METHODS: Individuals following a GFD were invited to complete an online questionnaire about their demography and geography characteristics, gluten-free food choices, shopping preferences, GFD management and subjective difficulty towards the diet. Distribution of the characteristics was described, and univariate and multivariate logistic regressions performed to explore the relationship between the above-mentioned variables and subjective difficulty in following a GFD. RESULT: 209 individuals following a GFD completed the questionnaire, most of whom were young, single and well-educated females from developed provinces or municipalities in China. Multiple regression showed that age, education level, advice on starting a GFD, duration before discovering a gluten intolerance, food choices and ways of GFD management were significantly associated with the subjective difficulty in following a GFD. CONCLUSION: Our findings offer a basic characteristics pattern of the population on a GFD in mainland China. Nearly one-third of GFD followers found the diet challenging to be followed. We suggest that sufficient celiac disease and gluten-induced disorder education be conducted among healthcare practitioners. Early diagnosis of gluten-induced disorders and defining an Asian-adapted GFD, as well as an increase in public awareness, may help adherence to a strict GFD in China.

Sieving out non-celiac gluten sensitivity amongst patients with irritable bowel syndrome.

OBJECTIVES: It is challenging to make diagnosis of non-celiac gluten sensitivity/non-celiac wheat sensitivity (NCGS/NCWS) in clinical practice, since there is no biomarker and diagnosis is based on response to gluten-free-diet (GFD). We used anti-gliadin antibody (AGA) for screening patients with IBS for gluten-sensitivity. METHODS: 492 Adult-patients with IBS underwent screening for celiac disease and gluten-sensitivity using IgA anti-tissue transglutaminase antibody and IgA-AGA and IgG-AGA, respectively. Patients with positive AGA (IgA and/or IgG) were invited to follow GFD, those willing were put on GFD for 6-weeks. Responsive patients were given gluten re-challenge. Diagnosis of NCGS was confirmed if they had recurrence of symptoms. RESULTS: Of 492 patients with IBS, AGA was positive in 61(12.4 %), hence suspected to have gluten-sensitivity. Of 31 who agreed to participate and followed GFD for 6-weeks, 17 (54.8 %) had complete (>30 % improvement) and 10(32.2 %) had partial (>20 % improvement) response. All 17 complete-responders were given gluten re-challenge for 6-weeks, symptoms recurred in all and hence were confirmed to have NCGS/NCWS. Significant decrease in AGA levels occurred almost in all GFD-responders. CONCLUSIONS: 12.4 % IBS patients have biological evidence of gluten/wheat-sensitivity. Almost 87 % patients with IBS having AGA responded to GFD. The value of AGA may further be explored as a biomarker for screening for the presence of NCGS, before recommending this test for the clinical practice.

Fecal calprotectin levels in patients with non-celiac wheat sensitivity: a proof of concept.

Some data suggest the existence of intestinal inflammation in patients with non-celiac wheat sensitivity (NCWS). We aimed to verify whether fecal calprotectin (FCP), a marker of intestinal inflammation, could be used to confirm this inflammatory status and to test its diagnostic performance in differentiating NCWS from irritable bowel syndrome/functional dyspepsia (IBS/FD). We conducted a multicenter study, comparing NCWS patients, diagnosed by a double-blind placebo-controlled wheat challenge, with IBS/FD subjects. In the retrospective phase, FCP values were analyzed to define the prevalence of its positivity and its role as a NCWS diagnostic biomarker. In the prospective phase we tested the effects of a strict 6-month wheat-free diet (WFD) on FCP values. 31.3% (n = 63/201) of NCWS patients had above normal FCP values (NCWS FCP +), whereas all IBS/FD patients proved negative (P = 0.0001). FCP using a cut-off value > 41 µg/g showed a 58.6% sensitivity and a 98.0% specificity (AUC 0.755, 95% C.I. 0.702-0.837) in distinguishing NCWS from IBS/FD patients. Of the 63 NCWS FCP+, 65.1% had negative FCP values after ≥ 6 months of WFD, with a significant reduction in FCP values (P < 0.0001). All NCWS FCP- subjects still preserved negative FCP values after ≥ 6 months of WFD. Our study showed that FCP can be a useful but supplementary diagnostic marker for differentiating between NCWS and IBS/FD. Strict WFD adherence reduced FCP values, normalizing them in 65.1% of NCWS FCP + subjects. These data suggest the existence of two NCWS subgroups: NCWS FCP + characterized by a probable predominantly inflammatory/immunologic pattern and NCWS FCP- featuring non-immuno-mediated etiopathogenetic mechanisms. (Registration number NCT01762579).

The Gliadin Hydrolysis Capacity of B. longum, L. acidophilus, and L. plantarum and Their Protective Effects on Caco-2 Cells against Gliadin-Induced Inflammatory Responses.

BACKGROUND: Non-celiac wheat sensitivity (NCWS) is a poorly understood gluten-related disorder (GRD) and its prominent symptoms can be ameliorated by gluten avoidance. This study aimed to determine the effectiveness of a probiotic mixture in hydrolyzing gliadin peptides (toxic components of gluten) and suppressing gliadin-induced inflammatory responses in Caco-2 cells. METHODS: Wheat dough was fermented with a probiotic mix for 0, 2, 4, and 6 h. The effect of the probiotic mix on gliadin degradation was monitored by SDS-PAGE. The expression levels of IL-6, IL-17A, INF-γ, IL-10, and TGF-ß were evaluated using ELISA and qRT-PCR methods. RESULTS: According to our findings, fermenting wheat dough with a mix of B. longum, L. acidophilus, and L. plantarum for 6 h was effective in gliadin degradation. This process also reduced levels of IL-6 (p = 0.004), IL-17A (p = 0.004), and IFN-γ (p = 0.01) mRNA, as well as decreased IL-6 (p = 0.006) and IFN-γ (p = 0.0009) protein secretion. 4 h fermentation led to a significant decrease in IL-17A (p = 0.001) and IFN-γ (p = 0.003) mRNA, as well as reduced levels of IL-6 (p = 0.002) and IFN-γ (p < 0.0001) protein secretion. This process was also observed to increase the expression levels of IL-10 (p < 0.0001) and TGF-ß (p < 0.0001) mRNA. CONCLUSIONS: 4 h fermentation of wheat flour with the proposed probiotic mix might be a good strategy to develop an affordable gluten-free wheat dough for NCWS and probably other GRD patients.

Anemia in non-celiac wheat sensitivity: Prevalence and associated clinical and laboratory features.

BACKGROUND: Patients suffering from non-celiac wheat sensitivity (NCWS) frequently report extra-intestinal symptoms, such as anemia. AIMS: We investigated the prevalence and associated clinical features of anemia in NCWS patients. METHODS: Data from 244 NCWS patients, diagnosed by double-blind placebo-controlled wheat challenge, were retrospectively reviewed and compared with 2 control groups (celiac disease (CD) and irritable bowel syndrome (IBS)). Furthermore, 31 NCWS anemic patients were prospectively re-evaluated after at least 12 months on the "strict" wheat-free diet (WFD). RESULTS: Anemia prevalence in NCWS patients was 34.8% (mean hemoglobin 10.4 ± 1.4 g/dl), significantly higher than in IBS (17.4%, P = 0.03), but not in CD ones. The NCWS group, on the whole, had sideropenic-like features with low serum iron and altered iron deposits. Both anemia prevalence and sideropenic-like features were more evident in CD than in NCWS patients, whereas only a few IBS subjects showed such features. Significant differences were found in anemic vs non-anemic NCWS patients as regards to female sex, diagnostic delay, poly/hypermenorrhea, iron deficiency, and higher TSH values. A long-term WFD significantly reduced anemia and improved iron metabolism. CONCLUSION: Microcytic/hypochromic anemia and altered iron metabolism occur frequently in NCWS and can be treated with a long-term strict WFD. NCWS should be included in differential diagnosis of anemic patients with "functional gastrointestinal troubles".

No correlation between amylase/trypsin-inhibitor content and amylase inhibitory activity in hexaploid and tetraploid wheat species.

Wheat amylase/trypsin-inhibitors (ATI) are known triggers for wheat-related disorders. The aims of our study were to determine (1) the inhibitory activity against different α-amylases, (2) the content of albumins and globulins (ALGL) and total ATI and (3) to correlate these parameters in wholegrain flour of hexaploid, tetraploid and diploid wheat species. The amount of ATI within the ALGL fraction varied from 0.8% in einkorn to 20% in spelt. ATI contents measured with reversed-phase high-performance liquid chromatography (RP-HPLC) revealed similar contents (1.2-4.2 mg/g) compared to the results determined by LC-MS/MS (0.2-5.2 mg/g) for all wheat species except einkorn. No correlation was found between ALGL content and inhibitory activity. In general, hexaploid cultivars of spelt and common wheat had the highest inhibitory activities, showing values between 897 and 3564 AIU/g against human salivary α-amylase. Tetraploid wheat species durum and emmer had lower activities (170-1461 AIU/g), although a few emmer cultivars showed similar activities at one location. In einkorn, no inhibitory activity was found. No correlation was observed between the ATI content and the inhibitory activity against the used α-amylases, highlighting that it is very important to look at the parameters separately.

Attenuation of immune activation in patients with multiple sclerosis on a wheat-reduced diet: a pilot crossover trial.

Background: Western lifestyle has been associated with an increase in relapsing-remitting multiple sclerosis (RRMS). In mice, dietary wheat amylase-trypsin inhibitors (ATIs) activate intestinal myeloid cells and augment T cell-mediated systemic inflammation. Objective: The aim of this study was to assess whether a wheat- and thus ATI-reduced diet might exert beneficial effects in RRMS patients with modest disease activity. Methods: In this 6-month, crossover, open-label, bicentric proof-of-concept trial, 16 RRMS patients with stable disease course were randomized to either 3 months of a standard wheat-containing diet with consecutive switch to a > 90% wheat-reduced diet, or vice versa. Results: The primary endpoint was negative, as the frequency of circulating pro-inflammatory T cells did not decrease during the ATI-reduced diet. We did, however, observe decreased frequencies of CD14+ CD16++ monocytes and a concomitant increase in CD14++ CD16- monocytes during the wheat-reduced diet interval. This was accompanied by an improvement in pain-related quality of life in health-related quality of life assessed (SF-36). Conclusion: Our results suggest that the wheat- and thus ATI-reduced diet was associated with changes in monocyte subsets and improved pain-related quality of life in RRMS patients. Thus, a wheat (ATI)-reduced diet might be a complementary approach accompanying immunotherapy for some patients. Registration: German Clinical Trial Register (No. DRKS00027967).