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1.
Mol Nutr Food Res ; 68(9): e2300779, 2024 May.
Artigo em Inglês | MEDLINE | ID: mdl-38632845

RESUMO

The intestinal immune system plays a crucial role in obesity and insulin resistance. An altered intestinal immunity is associated with changes to the gut microbiota, barrier function, and tolerance to luminal antigens. Lipid metabolism and its unbalance can also contribute to acute and chronic inflammation in different conditions. In celiac disease (CD), the serum phospholipid profile in infants who developed CD is dramatically different when compared to that of infants at risk of CD not developing the disease. In a mouse model of gluten sensitivity, oral wheat gliadin challenge in connection with inhibition of the metabolism of arachidonic acid, an omega-6 polyunsaturated fatty acid, specifically induces the enteropathy. Recent evidence suggests that gluten may play a role also for development of life-style related diseases in populations on a high fat diet (HFD). However, the mechanisms behind these effects are not yet understood. Exploratory studies in mice feed HFD show that wheat gliadin consumption affects glucose and lipid metabolic homeostasis, alters the gut microbiota, and the immune cell profile in liver.


Assuntos
Doença Celíaca , Dieta Hiperlipídica , Microbioma Gastrointestinal , Gliadina , Obesidade , Animais , Dieta Hiperlipídica/efeitos adversos , Obesidade/metabolismo , Humanos , Microbioma Gastrointestinal/fisiologia , Triticum , Camundongos , Metabolismo dos Lipídeos
2.
Methods Cell Biol ; 179: 157-171, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37625873

RESUMO

A complete understanding of celiac disease (CD) pathogenesis has been hindered to date because of the lack of adequate in vivo models. Herein, we describe two in vivo approaches in HLA-DQ8-transgenic mice to study the intrinsic cytoxicity and immune features of wheat gliadin. By adopting the first method, we explored the mucosal architecture of the small intestine following the intra-gastric administration of wheat gliadin in mice treated with indomethacin, an inhibitor of cyclooxygenases. Mice showed a significant reduction of villus height, increased crypt depth and increased intraepithelial lymphocytes. The second approach involved the mucosal sensitization to gliadin via the intranasal route. This protocol induced a Th1/Th17 phenotype in mesenteric lymph nodes, as described in CD. In conclusion, these methods remain instrumental to analyze in vivo distinct biological features of wheat gliadin and related prolamins. Furthermore, the sensitization protocol could be exploited to test innovative strategies downregulating the gliadin-specific immunity.


Assuntos
Gliadina , Triticum , Camundongos , Animais , Camundongos Transgênicos , Triticum/genética , Antígenos HLA-DQ/genética
3.
Int Rev Immunol ; 42(3): 237-245, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-35225129

RESUMO

Celiac disease (CD) is an autoimmune disease that occurs in genetically predisposed individuals following the ingestion of gluten. Its prevalence is rising worldwide. A gluten-free (GF) diet is mandatory for the management of CD. However, several issues persist regarding the nutritional quality of GF products. Importantly, deep knowledge about the pathogenic mechanisms in CD highlights the central role of CD4+ T cell-mediated immunity in CD. Furthermore, intestinal T regulatory cells are functional in CD, but cytokines such as IL-15, produced under inflammatory conditions, hamper their activity. This paves the way for the development of immunomodulatory strategies to the GF diet. From this perspective, microbiological approaches were considered able to modulate the gluten-specific immune response. Interestingly, gliadin peptide-based immunotherapy to abolish the inflammatory CD4+T cell-mediated response has been explored in CD patients. Furthermore, different biotechnological approaches based on the use of chemically/enzymatically modified gluten molecules have been proved effective in different models of CD. However, the choice of the right age in infants to introduce the antigen and thus induce tolerance still remains an important issue to solve. Addressing all these points should help to design an effective intervention strategy for preventing CD.


Assuntos
Doença Celíaca , Lactente , Humanos , Glutens , Tolerância Imunológica , Imunomodulação , Imunoterapia
4.
Food Funct ; 13(17): 8941-8950, 2022 Aug 30.
Artigo em Inglês | MEDLINE | ID: mdl-35929785

RESUMO

A lifelong gluten-free diet (GFD) is currently the only available therapy for coeliac disease (CD). However, GFD compliance is difficult and alternative strategies are envisaged in the near future. We previously found that wheat gliadin following transamidation by microbial transglutaminase (mTG) does not induce IFN-γ secretion by intestinal T cells from CD patients. Fully transamidated gliadin with lysine ethyl ester can be recovered in a soluble protein fraction (spf) generated by the enzymatic treatment of wheat flour. Herein, we analysed the performance of transamidation by mTG on a pilot-scale (1L) by evaluating the reaction kinetics and its biological effect on the intestinal immune response in HLA/DQ8 transgenic mice, a model of gluten sensitivity. At 1 h, all gliadin fractions showed a faster electrophoretic mobility by acid-polyacrylamide gel electrophoresis (A-PAGE) following transamidation in comparison with their native counterparts. In parallel, the yield of residual native gliadin dropped (30% at 180 min), confirming our previous findings on a lab scale. Mucosal sensitisation of mice with gliadin via the intranasal route induced a Th1 phenotype in mesenteric lymph nodes (MLNs). Importantly, IFN-γ secretion was significantly reduced when gliadin-specific MLN cells were challenged in vitro with spf (P < 0.001). Multiplex analysis revealed that the adaptive immune response evoked by spf involved a distinct cell population characterised by secretion of IL-2, IL-3 and IL-5. Notably, spf stimulated in vitro a reduced or null secretion of all of the examined pro-inflammatory markers mainly associated to innate immunity. In conclusion, our data revealed the ability of transamidated gliadin to modulate both innate and adaptive mechanisms involved in the inflammatory response induced by wheat gliadin in the small intestine of DQ8 mice.


Assuntos
Doença Celíaca , Gliadina , Animais , Doença Celíaca/metabolismo , Farinha , Gliadina/metabolismo , Glutens/metabolismo , Antígenos HLA-DQ/imunologia , Intestino Delgado/metabolismo , Camundongos , Camundongos Transgênicos , Transglutaminases/metabolismo , Triticum/metabolismo
5.
J Pediatr Gastroenterol Nutr ; 74(6): 728-733, 2022 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-35442226

RESUMO

ABSTRACT: Coeliac disease (CD) is caused by immunological intolerance to wheat gluten and related proteins of rye and barley. Consequently, gluten-free (GF) products have been developed but technological implementation is required to improve their intrinsic rheological properties. One alternative for increasing the functional properties of GF foodstuff is the incorporation of microbial transglutaminase (mTG), which allows for the cross-linking of proteins that can substitute for the gluten network in the bakery industry. mTG has been, however, suggested to mimic tissue transglutaminase and to be immunogenic in CD patients. Recently, both mTG and gliadin were found to be transported to the endoplasmic reticulum of enterocytes, suggesting cross-presentation and potential interaction with immune cells in CD. Although pathogenetic activity of mTG has not been found to date, these data naturally raise concerns among clinicians and patients about the use of mTG as a food additive. On the contrary, different studies have shown that treatment with mTG was effective in reducing the inflammatory immune response of gluten in CD. In this article, we take advantage of recent advances in gut physiology and CD pathogenesis to revise the literature data on mTG. An updated and unbiased overview of the role of mTG in this pathology allowed us to definitively highlight the beneficial use of this food additive by CD patients.


Assuntos
Doença Celíaca , Transglutaminases , Doença Celíaca/patologia , Aditivos Alimentares , Gliadina , Glutens , Humanos
6.
Prog Lipid Res ; 80: 101066, 2020 11.
Artigo em Inglês | MEDLINE | ID: mdl-32979455

RESUMO

Polyunsaturated fatty acids (PUFA) are fundamental building materials for cells and play crucial function as signaling molecules. When PUFA are used as substrates for non-enzymatic or enzymatic reactions and gut microbiota metabolism, they can generate electrophilic derivatives (called Reactive Lipid Species, RLS) that promptly form adducts with nucleophilic molecules. RLS participate in several signaling pathways, including the activation of the nuclear factor erythroid 2-related factor 2 (Nrf2) pathway, which is the key mechanism in the maintenance of redox, metabolic and protein homeostasis, as well as the regulation of inflammation. Recent studies have provided insights on the localization of enzymes that synthesise reactive oxygen or nitrogen species (ROS or RNS respectively) in plasma membrane compartments (raft/caveolae) which also harbour PUFA esters, from which free acid forms can be released by phospholipase A2 activity (PLA2), and the complex of Nrf2 with the inhibitory protein Kelch-like ECH-associated Protein 1(Keap1). Additional investigations have indicated that dietary PUFA insertion into specific plasma membrane microdomains may alter the lipid environment and thereby influence caveolar composition and cell signaling. Given that PUFA-originated RLS attack such a complex and promote the release of active Nrf2, it cannot be excluded that all the biochemical machinery for Nrf2 activation is present in caveolae, where it triggers the Nrf2-mediated adaptive response for rescuing or maintaining cellular redox homeostasis. Here, we specifically aimed to summarize current information with regard to the roles of dietary PUFA and RLS in Nrf2-mediated redox homeostasis, namely 1) their role as Nrf2 activators, 2) the significance of the in vivo conversion of PUFA into RLS and 3) the caveolar involvement in cell signaling for redox homeostasis.


Assuntos
Gorduras Insaturadas na Dieta/farmacologia , Ácidos Graxos Insaturados/metabolismo , Ácidos Graxos Insaturados/farmacologia , Fator 2 Relacionado a NF-E2/metabolismo , Animais , Microbioma Gastrointestinal , Homeostase , Humanos , Proteína 1 Associada a ECH Semelhante a Kelch/metabolismo , Metabolismo dos Lipídeos , Oxirredução , Proteínas Quinases/metabolismo
7.
Food Chem Toxicol ; 140: 111331, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32304812

RESUMO

The resemblance of physiological traits of cell lines with their target/original tissue is an important prerequisite for the choice of the in vitro model. Although cytoprotective defenses, activated by the nuclear factor erythroid 2-related factor2 (Nrf2), have a preeminent importance in intestinal protection, nevertheless their levels inin vitro models have been never compared with those of their original tissue. Basal level of Nrf2-mediated defenses in murine enterocyte cells (Mode-K) and in human colon adenocarcinoma cells -at differentiated (DCaco2) or confluent stage (CCaco2)- were compared with those found in mouse or human duodenum. The pro-oxidant and cytotoxic effects of peptic-tryptic digest of gluten prepared from wheat bread (PT-b), einkorn (PT-e) or durum wheat (PT-d) were evaluated in Mode-k and DCaco2 by combining enzymatic, immune-enzymatic and real-time PCR assay. The results presented reveal that Mode-k cells resemble cytoprotective defenses of human/murine duodenum and are more susceptible to pro-oxidant, cytotoxic and pro-inflammatory effect of gliadin digest (in comparison with Caco2). Prolamins digests from the considered wheat exhibit different inhibitory effect on Nrf2-mediated defenses (PT-b > PT-e > PT-d). These data indicate, for the first time, that Mode-k are a reliable model for investigating wheat prolamins toxicity and for evaluating the signaling pathway of gluten-associated disease.


Assuntos
Enterócitos/efeitos dos fármacos , Glutens/toxicidade , Animais , Células CACO-2 , Diferenciação Celular , Linhagem Celular , Duodeno/efeitos dos fármacos , Enterócitos/citologia , Gliadina/toxicidade , Humanos , Técnicas In Vitro , Camundongos , Modelos Biológicos , Triticum/química
8.
Nutr Neurosci ; 23(4): 309-320, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30039750

RESUMO

Objectives: A strong rise of the fructose content in the human diet occurred in the last decade, as corn syrup is widely used as a sweetener for beverages and processed food. Since young people make a widespread consumption of added sugars, we evaluated the effects of a two weeks fructose-rich diet on brain redox homeostasis, autophagy and synaptic plasticity in the cortex of young and adults rats, in order to highlight the early risks to which brain is exposed.Methods and Results: Short-term fructose feeding was associated with an imbalance of redox homeostasis, as lower amount of Nuclear factor (erythroid derived 2)-like 2, lower activity of Glucose 6-phosphate dehydrogenase and Glutathione reductase, together with lower Glutathione/Oxidized Glutathione ratio, were found in fructose-fed young and adult rats. Fructose-rich diet was also associated with the activation of autophagy, as higher levels of Beclin, LC3 II and P62 were detected in cortex of fructose-fed rats. A diet associated decrease of synaptophysin, synapsin I, and synaptotagmin I, was found in fructose-fed young and adult rats. Interestingly, BDNF amount was significantly lower only in fructose-fed adult rats, while the level of its receptor TrkB decreased in both groups of treated rats. A further marker of brain functioning, Acetylcholinesterase activity, was found increased only in fructose-fed young animals.Conclusion: Overall, our findings suggest that young rats may severely suffer from the deleterious influence of fructose on brain health as the adults and provide experimental data suggesting the need of targeted nutritional strategies to reduce its amount in foods.


Assuntos
Autofagia/efeitos dos fármacos , Córtex Cerebral/efeitos dos fármacos , Córtex Cerebral/metabolismo , Frutose/administração & dosagem , Fator 2 Relacionado a NF-E2/metabolismo , Sinapses/efeitos dos fármacos , Sinapses/metabolismo , Animais , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Masculino , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Estresse Oxidativo , Ratos Sprague-Dawley , Receptor trkB/metabolismo , Transdução de Sinais/efeitos dos fármacos
9.
Mol Nutr Food Res ; 63(21): e1900243, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31398773

RESUMO

Inflammation and oxidative stress play an important role in the pathogenesis of depressive disorders and nuclear erythroid related factor 2 (Nrf2), a regulator of RedOx homeostasis and inflammation, is a promising target for depression prevention/treatment. As fish oil (FO) and conjugated linoleic acid (CLA) are known Nrf2 inducers, their protective ability is comparatively evaluated in a murine model of depression (MRL/MpJ-Faslpr ). Oxidative stress, fatty acids content, and critical factors reflecting brain functioning-namely brain-derived neurotrophic factor (BDNF), synaptic markers, and cholinergic signaling-are preliminarily evaluated in the frontal cortex of 8-week (Young) and in 22-week old animals (Old), which are used as model of depression. These markers are measured in Old mice at the end of a 5-week pretreatment with FO or CLA (728 or 650 mg kg-1 , respectively). Old mice exhibit disrupted Redox homeostasis, compensatory Nrf2 hyperactivation, lower docosaheaxaenoic acid (DHA), and lower BDNF and synaptic function proteins compared to Young mice. FO and CLA treatment relieves almost all the pathophysiological hallmarks at a level comparable to Young mice. Presented data provide the first evidence for the comparable efficacy of FO or CLA supplementation in preventing depression signs in Old MRL/lpr mice, likely through their ability of improving Nrf2-mediated antioxidant defenses.


Assuntos
Encéfalo/efeitos dos fármacos , Depressão/dietoterapia , Óleos de Peixe/farmacologia , Ácidos Linoleicos Conjugados/farmacologia , Fator 2 Relacionado a NF-E2/metabolismo , Envelhecimento , Animais , Antidepressivos/farmacologia , Autoimunidade/efeitos dos fármacos , Biomarcadores/metabolismo , Encéfalo/metabolismo , Encéfalo/patologia , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Depressão/metabolismo , Depressão/patologia , Suplementos Nutricionais , Ácidos Docosa-Hexaenoicos/metabolismo , Elongases de Ácidos Graxos/genética , Ácidos Graxos/metabolismo , Inflamação/dietoterapia , Fígado/efeitos dos fármacos , Fígado/metabolismo , Masculino , Camundongos Endogâmicos MRL lpr , Estresse Oxidativo/efeitos dos fármacos , Estearoil-CoA Dessaturase/genética , Fator de Necrose Tumoral alfa/metabolismo
10.
Mol Nutr Food Res ; 63(22): e1900350, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31410984

RESUMO

SCOPE: Polyphenols exhibit their antioxidant activity downstream the activation of the nuclear factor erythroid 2-related factor 2 pathway (Nrf2), but the connection between lipid metabolism and the Nrf2 pathway is still unknown. Flavonoid-rich concentrated extract from Prunus mahaleb (mahaleb concentrated fruit extract; MCFE) may act on oxido-reductive homeostasis and hepatic lipid metabolism via Nrf2. METHODS & RESULTS: MCFE ability to enhance the activity of Nrf2-mediated antioxidant/detoxifying enzymes is investigated in liver and colon of BALB/c mice. After a 4-week supplementation, macroscopic, histological, and biochemical signs of colitis are examined in mouse colon pulsed with 5% (w/v) dextran sodium sulfate (DSS). Untreated or DSS-supplemented mice are used as negative or positive control. MCFE effect on liver lipid metabolism and its possible link with the Nrf2 pathway is investigated. MCFE intake increases antioxidant defenses in mice colon and its pretreatment blunts pathological signs of colitis, as compared to positive control. In the liver, the increase in antioxidant defenses is associated with enhanced oxidative metabolism and with higher levels of peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α) and of hemeoxygenase-1 (HO-1), in comparison with negative controls. CONCLUSION: Cytoprotective and hypolipidemic effect produced by MCFE intake results, at least in part, by the activation of the Nrf2 pathway.


Assuntos
Colite/prevenção & controle , Mitocôndrias/efeitos dos fármacos , Fator 2 Relacionado a NF-E2/fisiologia , Extratos Vegetais/farmacologia , Prunus , Animais , Sulfato de Dextrana , Feminino , Frutas , Metabolismo dos Lipídeos/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos BALB C , Mitocôndrias/metabolismo , Oxirredução , Estresse Oxidativo/efeitos dos fármacos , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo/fisiologia , Transdução de Sinais/efeitos dos fármacos
11.
Cytokine ; 97: 141-148, 2017 09.
Artigo em Inglês | MEDLINE | ID: mdl-28648868

RESUMO

Lactobacilli and bifidobacteria play a primary role in modulation of gut immunity. By considering that microbiota composition depends on various factors, including diet, we asked whether functional differences could characterize faecal populations of lactobacilli and bifidobacteria isolated from individuals with different dietary habits. 155 healthy volunteers who followed omnivorous, ovo-lacto-vegetarian or vegan diets were recruited at four Italian centres (Turin, Parma, Bologna and Bari). Faecal samples were collected; lactobacilli and bifidobacteria were isolated on selective media and their immunomodulatory activity was tested in mouse dendritic cells (DCs). Pre-incubation with lactobacilli increased LPS-induced expression of the maturation markers CD80 and CD86, whereas pre-incubation with bifidobacteria decreased such expression. Analysis of the cytokine profile indicated that strains of both genera induced down-regulation of IL-12 and up-regulation of IL-10, whereas expression of TNF-α was not modulated. Notably, analysis of anti-inflammatory potential (IL-10/IL-12 ratio) showed that lactobacilli evoked a greater anti-inflammatory effect than did bifidobacteria in the omnivorous group (P<0.05). We also found significantly reduced anti-inflammatory potential in the bacterial strains isolated from Bari's volunteers in comparison with those from the cognate groups from the other centres. In conclusion, lactobacilli and bifidobacteria showed a genus-specific ability of modulating in vitro innate immunity associated with a specific dietary habit. Furthermore, the geographical area had a significant impact on the anti-inflammatory potential of some components of faecal microbiota.


Assuntos
Bifidobacterium/imunologia , Células Dendríticas/imunologia , Dieta Vegana , Dieta , Microbioma Gastrointestinal/imunologia , Imunomodulação , Lactobacillus/imunologia , Animais , Antígeno B7-1/genética , Antígeno B7-2/genética , Bifidobacterium/isolamento & purificação , Citocinas/genética , Células Dendríticas/microbiologia , Regulação para Baixo , Fezes/microbiologia , Humanos , Interleucina-10/genética , Interleucina-12/genética , Lactobacillus/isolamento & purificação , Camundongos , Fator de Necrose Tumoral alfa/genética , Regulação para Cima , Vegetarianos
12.
Cytokine ; 90: 80-87, 2017 02.
Artigo em Inglês | MEDLINE | ID: mdl-27863334

RESUMO

Enterocytes are actively involved in the defense against pathogens and they limit penetration of commensal microbes into tissues. They also have an important role in gut immunity as enterocytes confer mucosal dendritic cell specialisation. On the other hand, the microbiota is directly involved in the development and modulation of the intestinal immune system. Particularly, lactobacilli and bifidobacteria play a primary role in shaping the immune response. We further explored this issue by evaluating whether functional differences in Caco-2 cells could characterise faecal populations of lactobacilli (155 samples) and bifidobacteria (110 samples) isolated from three dietary cohorts (omnivores, ovo-lacto-vegetarians and vegans) recruited at four Italian centres (Turin, Parma, Bologna and Bari). According to our findings, tested bacteria were unable to modulate expression of IL-8, IL-10, TGF-ß or thymic stromal lymphopoietin (TSLP) cytokines in unstimulated Caco-2 cells. Conversely, in phorbol 12-myristate 13-acetate and ionomycin (PMA/Io) stimulated Caco-2 cells, lactobacilli from the omnivorous group and all bifidobacteria significantly down-regulated IL-8. Notably, both genera also lowered the TSLP expression in stimulated Caco-2 cells, regardless of the diet regimen. By further examining these data on the basis of geographical origin, we found that lactobacilli from the vegetarian group recruited in Bari, significantly up-regulated this cytokine. In conclusion, we highlighted a peculiar immune-modulatory activity profile for lactobacilli on enterocytes undergoing a stimulatory signal, which was associated with a specific dietary habit. Furthermore, the geographical area had a significant impact on the inflammatory potential of members of the Lactobacillus genus.


Assuntos
Bifidobacterium/imunologia , Citocinas/imunologia , Comportamento Alimentar , Preferências Alimentares , Interleucina-8/imunologia , Lactobacillus/imunologia , Adolescente , Adulto , Bifidobacterium/isolamento & purificação , Células CACO-2 , Feminino , Humanos , Lactobacillus/isolamento & purificação , Masculino , Pessoa de Meia-Idade , Acetato de Tetradecanoilforbol/farmacologia , Linfopoietina do Estroma do Timo
13.
Nutrients ; 8(10)2016 Oct 14.
Artigo em Inglês | MEDLINE | ID: mdl-27754398

RESUMO

The most recent trend in research on probiotic bacteria aims at the exploitation of bioactive bacterial compounds that are responsible for health-promoting effects and suitable for medical applications. Therefore, the main purpose of this study was to ascertain if the immunomodulatory effects of L. paracasei strains on dendritic cells (DCs) were caused by bacterial metabolites released in the culture medium. For that reason, bacterial strains were grown in two media generally used for the culture of DCs, and the effects of culture filtrates on the maturation of DCs and cytokine production were evaluated. Moreover, to reveal potential synergistic effects on the immunomodulation of DCs, an artichoke phenolic extract (APE) was added to the media before bacterial growth. The experiments pointed out an interesting anti-inflammatory activity of a culture filtrate obtained after growing a probiotic L. paracasei strain in one of the media supplemented with APE. Therefore, this culture filtrate-which combines the anti-inflammatory activity and the other well-known health-promoting properties of artichoke phenolic compounds-could represent the basis for future particular exploitations.


Assuntos
Cynara scolymus/química , Citocinas/metabolismo , Células Dendríticas/efeitos dos fármacos , Lacticaseibacillus paracasei/metabolismo , Extratos Vegetais/farmacologia , Polifenóis/farmacologia , Probióticos , Células Dendríticas/metabolismo , Humanos , Lacticaseibacillus paracasei/crescimento & desenvolvimento
14.
Foods ; 5(3)2016 Jun 29.
Artigo em Inglês | MEDLINE | ID: mdl-28231144

RESUMO

Celiac disease (CD) is an immune-mediated disorder caused by the ingestion of wheat gluten. A lifelong, gluten-free diet is required to alleviate symptoms and to normalize the intestinal mucosa. We previously found that transamidation reaction by microbial transglutaminase (mTG) was effective in down-regulating the gliadin-specific immune response in CD patients. In this study, the two-step transamidation protocol was adopted to treat commercial wheat semolina on a pilot scale. The effectiveness of the enzymatic reaction was tested by means of consolidated biochemical and immunological methods on isolated prolamins. We found that water-insoluble gliadin and glutenin yields decreased in wheat semolina to 5.9% ± 0.3% and 11.6% ± 0.1%, respectively, after a two-step transamidation reaction. Using DQ8 transgenic mice as a model of gluten sensitivity, we observed a dramatic reduction in IFN-γ production in spleen cells challenged in vitro with the residual insoluble gliadin from transamidated semolina (N = 6; median values: 850 vs. 102; control vs. transamidated semolina, p < 0.05). The technological properties of treated wheat semolina were then tested by manufacturing classical pasta (spaghetti). Notably, the spaghetti manufactured with transamidated semolina had only minor changes in its features before and after cooking. In conclusion, the two-step transamidation reaction modified the immunogenic epitopes of gliadins also on a pilot-scale level without influencing the main technological properties of semolina. Our data shed further light on a detoxification strategy alternative to the current gluten-free diet and may have important implications for the management of CD patients.

15.
Microb Cell Fact ; 12: 98, 2013 Oct 29.
Artigo em Inglês | MEDLINE | ID: mdl-24168229

RESUMO

BACKGROUND: Mucosal infections are a major global health problem and it is generally accepted that mucosal vaccination strategies, able to block infection at their entry site, would be preferable with respect to other prevention approaches. However, there are still relatively few mucosal vaccines available, mainly because of the lack of efficient delivery systems and of mucosal adjuvants. Recombinant bacterial spores displaying a heterologous antigen have been shown to induce protective immune responses and, therefore, proposed as a mucosal delivery system. A non-recombinant approach has been recently developed and tested to display antigens and enzymes. RESULTS: We report that the binding subunit of the heat-labile toxin (LTB) of Escherichia coli efficiently adsorbed on the surface of Bacillus subtilis spores. When nasally administered to groups of mice, spore-adsorbed LTB was able to induce a specific immune response with the production of serum IgG, fecal sIgA and of IFN-γ in spleen and mesenteric lymph nodes (MLN) of the immunized animals. Dot blotting experiments showed that the non-recombinant approach was more efficient than the recombinant system in displaying LTB and that the efficiency of display could be further increased by using mutant spores with an altered surface. In addition, immunofluorescence microscopy experiments showed that only when displayed on the spore surface by the non-recombinant approach LTB was found in its native, pentameric form. CONCLUSION: Our results indicate that non-recombinant spores displaying LTB pentamers can be administered by the nasal route to induce a Th1-biased, specific immune response. Mutant spores with an altered coat are more efficient than wild type spores in adsorbing the antigen, allowing the use of a reduced number of spores in immunization procedures. Efficiency of display, ability to display the native form of the antigen and to induce a specific immune response propose this non-recombinant delivery system as a powerful mucosal vaccine delivery approach.


Assuntos
Bacillus subtilis/metabolismo , Proteínas de Bactérias/metabolismo , Esporos Bacterianos/metabolismo , Animais , Bacillus subtilis/química , Bacillus subtilis/imunologia , Proteínas de Bactérias/análise , Proteínas de Bactérias/imunologia , Camundongos , Esporos Bacterianos/química , Esporos Bacterianos/imunologia
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