Wheat Gliadin Research Articles

CDNA microarray analysis of gene expression in coeliac disease jejunal biopsy samples

In coeliac disease in the jejunum of a genetically susceptible person wheat gliadin and related prolamins from rye and barley trigger an immunological reaction, which induces small-bowel mucosal transformations, villous atrophy and crypt hyperplasia. Though CD4+ specific T cells, intraepithelial lymphocytes, infiltrating plasma cells and autoantibodies are known to have an effect on the coeliac disease, the pathogenic mechanisms leading to the tissue injury remain to be elucidated. Our aim was to find novel gene transcripts, which might have a role in coeliac disease pathogenesis. The gene expression in duodenal biopsy samples from untreated coeliac patients ( n=4), patients on gluten-free diet ( n=4) and healthy controls ( n=4) was studied by cDNA microarray analysis. The method allows monitoring of the expression of thousands of genes simultaneously. Compared to healthy controls, the expression of 156 and 60 genes was changed in untreated and treated coeliac disease, respectively. Between treated and untreated coeliac disease, 98 genes had altered expression. Of the 5184 genes or expressed sequence tags, altogether 263 were affected. Many of these genes are directly or indirectly connected to T-cell activation, B-cell maturation or epithelial cell differentiation. By the microarray method, numerous genes were found to evince altered mRNA expression in coeliac disease. The method holds promise in exploring the pathogenetic mechanisms in the small bowel and may reveal new target genes for the therapy of coeliac disease.

Celiac Disease: Celiac Sprue, Gluten-sensitive Enteropathy

Celiac Disease (CD) or gluten-sensitive enteropathy is a T-cell mediated disease occurring in genetically susceptible individuals induced by the ingestion of one of several proteins found in wheat (gliadins), barley (hordeins) and rye (secalinin). Symptoms classically include episodic diarrhea, abdominal pain and distention and weight loss. Approximately one half of adults develop clinically significant diarrhea. Interestingly, only one half of all patients with CD have symptoms referable to the gastrointestinal tract.

IgE-binding Abilities of Pentapeptides, QQPFP and PQQPF, in Wheat Gliadin

Wheat gliadin is known mainly as the causative substance for enteropathy accompanied by diarrhea (celiac disease) and food-dependent exercise-induced anaphylaxis. However, little is known in regards to the allergenicity of gliadin in atopic dermatitis. In this study, the allergenicity of gliadin was demonstrated using sera of wheat-allergic patients with atopic dermatitis. Since there are many repeated sequences in gliadin, peptides containing each motif were synthesized to evaluate their IgE-binding abilities. As a result, QQPFP and PQQPF were found to be epitopes (minimum structures for the IgE-binding). Gliadin might crosslink patient's IgE-antibodies via these epitopes and trigger subsequent allergic reactions.

Effects of Wheat Protein Fractions on Flour Tortilla Quality

ABSTRACTCommercial wheat protein fractions (10) were evaluated during processing for quality of tortillas prepared using pastry, tortilla, and bread flours. Protein fractions that separately modify dough resistance and extensibility were evaluated in tortillas to determine whether the proteins could increase diameter, opacity, and shelf stability. Tortillas were prepared using laboratory‐scale, commercial equipment with fixed processing parameters. Dough and tortilla properties were evaluated using analytical methods, a texture analyzer, and subjective methods. Tortillas were stored in plastic bags at 22°C for up to 20 days. Adjustments in water absorption and level of reducing agent were made to normalize differences in functionality of 3% added proteins on dough properties. Tortilla weight, moisture, pH, opacity, and specific volume were not affected by added proteins, except for glutenin and vital wheat gluten treatments, which had decreased opacity in tortillas prepared from pastry flour. Increased insoluble polymeric protein content corresponded to decreased tortilla diameter and improved shelf stability. Treatments yielding tortillas with improved shelf stability and similar tortilla properties were produced when commercially processed vital wheat gluten products, FP600, FP6000, FP5000, or gliadin were added to pastry or tortilla flour. These wheat protein fractions improved processing and tortilla quality of wheat flours, especially pastry flour, by modifying protein content and quality.

Influence of an Orally Effective SOD on Hyperbaric Oxygen-related Cell Damage

In a prospective, double-blind, randomised placebo-controlled study, we tested the hypothesis that a new formulation consisting of wheat gliadin chemically combined with a vegetal (thus orally effective) preparation of superoxide dismutase (SOD) allows to prevent hyperbaric oxygen (HBO)-induced oxidative cell stress. Twenty healthy volunteers were exposed to 100% oxygen breathing at 2.5 ATA for a total of 60 min. DNA strand breaks (tail moments) were determined using the alkaline version of the comet assay. Whole blood concentrations of reduced (GSH) and oxidised (GSSG) glutathione and F2-isoprostanes, SOD, glutathione peroxidase (GPx) and catalase (Cat) activities and red cell malondialdehyde (MDA) content were determined. After HBO exposure the tail moment [Formula: See Text] and isoprostane levels [Formula: See Text] were significantly lower in the group that received the vegetal formulation. Neither SOD and Cat nor GSH and GSSG were significantly affected by this preparation or HBO exposure. By contrast, blood GPx activity, which tended to be lower in the SOD-group already before the HBO exposure [Formula: See Text] was significantly lower afterwards [Formula: See Text] We conclude that an orally effective SOD-wheat gliadin mixture is able to protect against DNA damage, which coincided with reduced blood isoprostane levels, and may therefore be used as an antioxidant.

IL12B and IRF1 gene polymorphisms and susceptibility to celiac disease.

Celiac disease (CD) is a common gastro-intestinal disorder resulting from permanent intolerance to wheat gliadins and related proteins in rye and barley. In addition to the strong genetic association with HLA-DQ2 and HLA-DQ8, a genetic region on chromosomes 5 (CELIAC2) has been identified that harbours a susceptibility gene for CD. The gene(s) responsible for this association, however, remains to be identified. In the present study we evaluated polymorphisms in the genes encoding interleukin-12 p40 (IL12B) and interferon regulatory factor 1 (IRF1). Both genes are located in the celiac2 region, and have key roles in inducing interferon (IFN)-gamma secreting T helper 1 (Th1) cells, one of the immunological hallmarks of CD. The frequencies of a TaqI gene polymorphism in the 3' UTR of IL12B and a HinfI gene polymorphism in the 3' UTR of IRF1 were studied in 258 Dutch CD patients and 237 ethnically matched healthy controls. The transmission of the polymorphic variants from parents to affected child was determined in 123 families with at least one affected child. The frequencies of the IL12B TaqI gene polymorphism and the IRF1 HinfI gene polymorphism did not differ significantly between patients and controls. In addition, in the family study, no deviation from the expected transmission from parents to affected child of any of the polymorphic variants was found. The IL12B TaqI and the IRF1 HinfI gene polymorphisms do not appear to be involved in susceptibility to CD. Further studies on the factors that drive the Th1 immunopathology in CD are required.

High microbial production and characterization of strictly periodic polymers modelled on the repetitive domain of wheat gliadins

Primary structures of wheat prolamins contain repetitive domains involved in the mechanical properties of gluten. In order to experience the ability of recombinant strictly periodic polypeptides, modelled on a consensus sequence of wheat gliadins (PQQPY) 8 and (PQQPY) 17 (SPR8 and SPR17 polypeptides, respectively), to be formulated in film solutions, their heterologous expression conditions, in batch culture and low cell densities, were optimized to match the high requirements of this process. A convenient and general purification procedure was also devised. Moreover, FTIR-ATR characterizations indicated that these periodic polypeptides prepared as hydrated doughy state and dried have the tendency to form a protein network through intermolecular β-sheets, strongly maintained by hydrogen bonds. Accordingly, these recombinant polypeptides are assumed to be a suitable candidate for potential application.

Identification of transglutaminase-mediated deamidation sites in a recombinant alpha-gliadin by advanced mass-spectrometric methodologies.

Celiac disease is a permanent immune-mediated food intolerance triggered by ingestion of wheat gliadins in genetically susceptible individuals. It has been reported that tissue transglutaminase plays an important role in the onset of celiac disease by converting specific glutamine residues within gliadin fragments into glutamic acid residues. This process increases binding affinity of gliadin peptides to HLA-DQ2/DQ8 molecules, thus enhancing the immune response. The aim of the present study was to achieve a detailed structural characterization of modifications induced by transglutaminase on gliadin peptides. Therefore, structural analyses were carried out on a recombinant alpha-gliadin and on a panel of 26 synthetic peptides, overlapping the complete protein sequence. Modified glutamine residues were identified by means of advanced mass-spectrometric methodologies on the basis of MALDI-TOF-MS and tandem mass spectrometry. Results led to the identification of 19 of 94 glutamine residues present in the recombinant alpha-gliadin, which were converted into glutamic acid residues by a transglutaminase-mediated reaction. This allowed us to achieve a global view of the modifications induced by the enzyme on this protein. Furthermore, results gathered could likely be utilized as relevant information for a better understanding of processes leading to T-cell recognition of gliadin peptides involved in celiac disease.

Isolation and characterization of angiotensin I‐converting enzyme inhibitory peptides from wheat gliadin hydrolysate

Angiotensin I-converting enzyme (ACE) inhibitory peptide was isolated from wheat gliadin hydrolysate prepared with acid protease. Consecutive purification methods were used for peptide isolation including ion-exchange chromatography, size-exclusion chromatography, and reverse-phase high-performance liquid chromatography. The amino acid sequence of this peptide was identified as Ile-Ala-Pro, and the ACE inhibitory activity (IC50 value) was 2.7 microM. The hypotensive activity of Ile-Ala-Pro on spontaneously hypertensive rats was investigated. This peptide inhibited the hypertensive activity of angiotensin I with intravenous injection, and decreased the blood pressure significantly with intraperitoneal administration.

Fast ω-gliadin is a major allergen in wheat-dependent exercise-induced anaphylaxis

Background: Wheat-dependent exercise-induced anaphylaxis is an anaphylaxy induced by physical exercise after ingestion of wheat. An immediate-type hypersensitivity to water/salt-insoluble fraction of wheat proteins (gluten) has been considered to underlie in this disease. Objective: The aim of the study is to determine the major allergen in Japanese patients with wheat-dependent exercise-induced anaphylaxis by using a panel of purified wheat gliadins and glutenins. Methods: Water/salt-insoluble wheat proteins, α-gliadin, β-gliadin, γ-gliadin, fast ω-gliadin, slow ω-gliadin, high molecular weight glutenin and low molecular weight glutenin, were purified, and five patients with wheat-dependent exercise-induced anaphylaxis, whose diagnose had been determined by positive-challenge test, were evaluated for skin prick test, dot-blotting test and CAP–RAST inhibition test by using these purified wheat proteins. Results: The fast ω-gliadin was the most potent allergen among these water/salt-insoluble proteins when evaluated by skin prick test and dot-blotting test. Fast and slow ω-gliadin, and γ-gliadin caused dose-dependent inhibition of the serum IgE-binding to solid-phase gluten in the patients. The incubation with fast ω-gliadin of the patient's serum caused dose-dependent inhibition in the IgE-binding to γ-gliadin as well as slow ω-gliadin, indicating a cross-reactivity of these proteins in IgE-binding. Conclusion: We concluded that fast ω-gliadin is a major allergen among these water/salt-insoluble proteins for wheat-dependent exercise-induced anaphylaxis in Japanese patients, and IgE against fast ω-gliadin cross-reacts to γ-gliadin and slow ω-gliadin.

Comparative investigations of gluten proteins from different wheat species

The only commercially available immunoassay for gliadin determination in gluten-free food which has been ring-tested and in use for many years, is a test kit based on monoclonal antibodies against ω-gliadins. Various studies of the literature have shown that different gliadin standards resulted in different calibration curves, and it has been proposed that the affinity of ω-gliadins to the monoclonal antibodies varied among wheat varieties. To clarify this fundamental problem, total gliadins and the ω-gliadins from a winter wheat ("Rektor"), a spring wheat ("CWRS"), a wheat rye hybrid ("Herzog") and varieties of spelt, durum wheat, emmer and einkorn, were isolated and analyzed by means of an enzyme-linked immunoabsorbent assay (ELISA) kit based on antibodies against ω-gliadins. Additionally, single α- and γ-gliadins of Rektor wheat were studied. The results demonstrated that the calibration curves derived for total gliadins differed, in parts, strongly from that of the kit gliadin standard; only the curves for the durum wheat and spelt gliadins were in congruence with the kit gliadin. Single ω-gliadins revealed strong differences between and within wheat species and ELISA equivalents had a range from 10 to 220% according to kit gliadin. The affinity of ω-gliadins was not correlated with the calibration curves of total gliadins. Some of the α-gliadins of Rektor wheat showed ELISA equivalents similar to those of ω-gliadins. Because the proportions of α-gliadins in total gliadins were significantly higher than those of ω-gliadins, the unspecific binding of α-gliadins contributed much more to the total affinity of gliadins than the specific binding of ω-gliadins. γ-Gliadins, however, did not show any detectable affinity.

Intranasal administration of a recombinant α-gliadin down-regulates the immune response to wheat gliadin in DQ8 transgenic mice

The mucosal lesion in coeliac disease (CD) is an immune-mediated injury triggered by gliadin and associated with HLA-DQ2 and HLA-DQ8. In view of this, an approach that re-induces tolerance to this antigen should be considered as possible alternative to a strict gluten-free diet in treating CD. However, T-cell activation to multiple antigens, as a consequence of the chemical complexity shown by the antigen gliadin, could hamper the efforts to identify single component(s) useful for tolerance induction. To address this issue, a recombinant α-gliadin was tested in tolerance experiments in HLA-DQ8 transgenic mice. As tissue transglutaminase (tTG) treatment of gliadin, previously reported to enhance its stimulatory activity in CD, did not increase its immunogenicity when parenterally administered in these mice, untreated gliadin was used as immunogen. A decrease in systemic T cell responses to the recombinant α-gliadin was found after nasal administration of antigen, reflected by lymphocytes proliferation assay. Interestingly, while the immunisation protocol induced transcription of both Th1 (IFN-γ) and Th2 (IL-4 and IL-10) cytokines, the tolerisation protocol down-regulated significantly only the IFN-γ mRNA expression. More important, the recombinant α-gliadin induced a similar down-regulatory effect in mice immunised with a commercial preparation of wheat gliadin, that is a mixture of many different gliadin components. As the Th1 phenotype and the HLA-DQ8 molecule play a role in the pathogenesis of CD, our data underlined the potential usefulness of this recombinant protein for the immunomodulation of this disease.

Wheat gliadin induces apoptosis of intestinal cells via an autocrine mechanism involving Fas–Fas ligand pathway

Wheat gliadin and other cereal prolamins have been said to be involved in the pathogenic damage of the small intestine in celiac disease via the apoptosis of epithelial cells. In the present work we investigated the mechanisms underlying the pro-apoptotic activity exerted by gliadin-derived peptides in Caco-2 intestinal cells, a cell line which retains many morphological and enzymatic features typical of normal human enterocytes. We found that digested peptides from wheat gliadins (i) induce apoptosis by the CD95/Fas apoptotic pathway, (ii) induce increased Fas and FasL mRNA levels, (iii) determine increased FasL release in the medium, and (iv) that gliadin digest-induced apoptosis can be blocked by Fas cascade blocking agents, i.e. targeted neutralizing antibodies. This favors the hypothesis that gliadin could activate an autocrine/paracrine Fas-mediated cell death pathway. Finally, we found that (v) a small peptide (1157 Da) from durum wheat, previously proposed for clinical practice, exerted a powerful protective activity against gliadin digest cytotoxicity.

Recent advances in coeliac disease

Coeliac disease, or gluten-sensitive enteropathy, affects 1 in 100-200 people in the UK. The condition, which is exacerbated by wheat, rye, barley and possibly oats, can be treated with a gluten-free diet in which these cereals are omitted. Serological screening, particularly of high-risk groups, with both IgA and IgG based systems can be used to identify cases. Diagnosis depends on the use of a small intestinal biopsy, which reveals the classical changes of loss of the normal villous architecture. Evidence suggests that gluten-sensitive T cells are involved in the pathogenesis of the disease. Use of in vitro systems has suggested an immuno dominant epitope within wheat gliadin, which has been shown to exacerbate the condition in vivo. This information can be used to devise strategies to develop immuno-modulatory peptides and cereals with the baking and nutritional qualities of wheat, rye and barley, but which do not exacerbate the condition.

Wheat gliadin promotes the interleukin-4-induced IgE production by normal human peripheral mononuclear cells through a redox-dependent mechanism.

Increased levels of serum IgE have been described in gliadin-intolerant patients; however, biological mechanisms implicated in this immunoglobulin production remained unknown. In this study, we demonstrated that in vitro crude gliadins and gliadin lysates (Glilys) promoted the IL-4-induced IgE production by human peripheral blood mononuclear cells (PBMC), indicating that the biological process related to gliadin intolerance and/or allergy may lead to IgE production in vivo. It was found that crude gliadin and Glilys potentiated, after 13 days of culture in a dose-dependent manner, IL-4-induced IgE production and, to a lesser extent, the IgG production, while they did not affect IgA or IgM productions. This promoting effect of gliadin and Glilys on the IL-4-induced activation of normal human PBMC was also observed on the early release (2 days) of the soluble fraction of CD23, suggesting its possible involvement in IgE potentiation. The promoting effect of crude gliadin and Glilys appeared to be indirect because they did not modify purified B-lymphocytes IgE production after IL-4 and anti-CD40 monoclonal antibody stimulation. In addition, as revealed by luminol-dependent chemiluminescence, we demonstrated that crude gliadin and Glilys promoted a substantial production of free radicals by normal human PBMC, treated or not with IL-4. This redox imbalance associated with an increased IgE production led us to evaluate the effect of pharmacological antioxidants (N-acetyl-cysteine (NAC) and Cu/Zn-superoxide dismutase (SOD1)) on IgE production by human PBMC. The NAC and the intracellularly delivered SOD1 were found to suppress the IL-4+/-crude gliadin or Glilys-induced IgE production by normal human PBMC. Taken together, these data indicated that gliadin specifically enhanced IL-4-induced IgE production by normal human PBMC, probably by the regulation of redox pathways, and that this 'pro-allergenic' effect could be counteracted by natural antioxidants: thiols and/or vectorized SOD1.

Bread wheat gliadin cytotoxicity: a new three‐dimensional cell model

Background: In an attempt to clarify the role of gliadin toxicity in the pathogenesis of gluten intolerance (celiac disease), previous in vitro studies have been based on two‐dimensional human cell cultures. However, the specific morphological and biochemical properties of in vivo tissue are better maintained in three‐dimensional cell cultures (multicellular spheroids, MCS). The aim of this study was to develop a three‐dimensional in vitro model to investigate the effects of gliadin on epithelial cells and broaden our understanding of the early tissue damage occurring in celiac disease. Methods: The three‐dimensionally growing Lovo cell line was exposed to increasing concentrations of peptic‐tryptic‐digested bread wheat gliadin (from 125 to 1000 μg/mL) for 7 days in order to evaluate cell viability (colony‐forming assay), and at the standard concentration of 500 μg/mL for 7 days in order to evaluate MCS diameters, volumes and cell morphology using light and electron microscopy. Results: In comparison with the controls, the cell viability of the gliadin‐treated MCS was significantly reduced (20–80%), but there was no difference in size. Various degrees of cell damage (autophagic vacuoles and intra‐cytoplasmic lipid‐like droplets) were detected by both light and electron microscopy. Conclusion: This is the first study investigating the effects of gliadin on MCS. Lovo MCS seem to be responsive to gliadin exposure, thus confirming previous results obtained using two‐dimensional cell cultures. The data suggest that three‐dimensional cell cultures may be useful in broadening our understanding of some of the early effects of gliadin peptides on epithelial cells.

Bread wheat gliadin cytotoxicity: a new three-dimensional cell model

Background: In an attempt to clarify the role of gliadin toxicity in the pathogenesis of gluten intolerance (celiac disease), previous in vitro studies have been based on two‐dimensional human cell cultures. However, the specific morphological and biochemical properties of in vivo tissue are better maintained in three‐dimensional cell cultures (multicellular spheroids, MCS). The aim of this study was to develop a three‐dimensional in vitro model to investigate the effects of gliadin on epithelial cells and broaden our understanding of the early tissue damage occurring in celiac disease. Methods: The three‐dimensionally growing Lovo cell line was exposed to increasing concentrations of peptic‐tryptic‐digested bread wheat gliadin (from 125 to 1000 μg/mL) for 7 days in order to evaluate cell viability (colony‐forming assay), and at the standard concentration of 500 μg/mL for 7 days in order to evaluate MCS diameters, volumes and cell morphology using light and electron microscopy. Results: In comparison with the controls, the cell viability of the gliadin‐treated MCS was significantly reduced (20–80%), but there was no difference in size. Various degrees of cell damage (autophagic vacuoles and intra‐cytoplasmic lipid‐like droplets) were detected by both light and electron microscopy. Conclusion: This is the first study investigating the effects of gliadin on MCS. Lovo MCS seem to be responsive to gliadin exposure, thus confirming previous results obtained using two‐dimensional cell cultures. The data suggest that three‐dimensional cell cultures may be useful in broadening our understanding of some of the early effects of gliadin peptides on epithelial cells.

The use of extended amino acid motifs for focussing on toxic peptides in coeliac disease.

Cereal prolamins of wheat, rye and barley are the major proteins that have been implicated in toxicity in patients with coeliac disease. The gliadins of wheat are the best characterised with the identification of toxic peptides from rye and barley not as well advanced. This study has employed extended motifs, based on the known toxic motifs are derived from the sequence of A-gliadin, to search protein databases for matches with coeliac-toxic cereals. The results obtained have provided pointers to specific regions in rye and barley prolamins, which have received little attention in in vitro and in vivo studies of toxicity in coeliac disease. The results obtained in this study indicate that the size of the extended motif is critical when searching for coeliac-toxic cereals using protein databases. Extended motifs that are common to all three coeliac-toxic cereals and found in active wheat gliadin peptides are QQPYP, PQQPY and QQQPFP.

Human Genome Search in Celiac Disease: Mutated Gliadin T-cell-like Epitope in Two Human Proteins Promotes T-cell Activation

Discovery of a number of novel and known human genes whose protein products bear striking similarity to two or more wheat gliadin domains raised the possibility that human intestinal non-HLA peptides homologous to celiac T-cell epitopes could play a role in non-HLA gene specification in celiac disease. Database searching of the entire human genome identified only 11 gut-expressed proteins with high T-cell epitope homology, particularly to the DQ2-γ-I-gliadin epitope (i.e. TFIIA, FOXJ2 and IgD; mean BestFit quality score=40 versus random value of 24). Others were similar to DQ2-α-I-gliadin (i.e. PAX9; BestFit quality 46 versus 20 for random), or DQ2-α-II-gliadin (PHLDA1, known in mice as the T-cell death-associated gene; BestFit quality 43 versus 30 for random) epitopes. Among proteins previously screened for gliadin homology, noteworthy was achaete scute homologous protein (DQ2-α-I-gliadin; BestFit quality 41 versus 22 for random). With the exception of IgD, all are nuclear factors. Paying particular attention to the position of potential major histocompatibility complex (MHC) anchor residues, several were selected for testing in a DQ2-γ-I-gliadin-restricted T-cell system. All native 10-mer peptides were inactive, even when deamidated, but V96F substitution of deamidated TFIIA amino acid residues 91–100 stimulated IL-2 release at levels exceeding the wheat gliadin positive control. Also active, but only slightly, was L1009F substitution of AIB3 amino acid residues 1004–1013. PlotSimilarity alignment of TFIIAs from eight species revealed subthreshold similarity score in the peptide region, in contrast to the highly conserved amino and carboxy termini. Molecular modeling of TFIIA[V96F] peptide points to an important juxtaposition of an upwardly projecting phenylalanine residue at peptide position 6 that likely contacts a receptor complementarity-determining region, and a downwardly projecting glutamic acid residue that fits into the shallow MHC P7 pocket. These observations tentatively point to a new multi-gene hypothesis for the initiation of celiac disease in which deamidated free human peptides with T-cell epitope homology (particularly those made more homologous by mutation) escape negative selection, as per deamidation of the HEL 48–62 peptide in the hen egg lysozyme model of autoimmunity. Deamidation following peptide release due to injury triggers inflammation, thereafter repeatedly provoked by dietary gliadin immunodominant peptides concentrated in the proximal small intestine.

In vitro cytotoxic effect of bread wheat gliadin on the LoVo human adenocarcinoma cell line

The pathogenesis of celiac disease is not completely understood but, although the initial step of the process is still unclear, an altered immune response seems to play a major role. Previous studies of the biological properties of gliadin have highlighted its cytotoxic effects, and the aim of this study was to develop an in vitro technique to study them. The LoVo (human colon adenocarcinoma) cell line grown in two-dimensional cultures was exposed to different concentrations of digested bread wheat gliadin (62, 125, 250, 500 and 750 μg/ml) for 48 h, after which cell growth and oxidative balance (the content of reduced glutathione (GSH), and peroxidase, transferase and reductase activity) was evaluated. Other food proteins were used as controls. Our data revealed a statistically significant inhibition of cell growth in proportion to the gliadin concentration (from 26 to 100%), combined with a decrease in GSH content (−38% at 500 μg/ml) and reduced enzymatic activity (−30% at 500 μg/ml). The controls did not show any noxious effect. Our results confirm the usefulness of LoVo cells in evaluating gliadin cytotoxicity and that they can be used to investigate the biological properties of gliadin.