In Vitro Insights into the Dietary Role of Glucoraphanin and Its Metabolite Sulforaphane in Celiac Disease.

Sulforaphane is considered the bioactive metabolite of glucoraphanin after dietary consumption of broccoli sprouts. Although both molecules pass through the gut lumen to the large intestine in stable form, their biological impact on the first intestinal tract is poorly described. In celiac patients, the function of the small intestine is affected by celiac disease (CD), whose severe outcomes are controlled by gluten-free dietary protocols. Nevertheless, pathological signs of inflammation and oxidative stress may persist. The aim of this study was to compare the biological activity of sulforaphane with its precursor glucoraphanin in a cellular model of gliadin-induced inflammation. Human intestinal epithelial cells (CaCo-2) were stimulated with a pro-inflammatory combination of cytokines (IFN-γ, IL-1ß) and in-vitro-digested gliadin, while oxidative stress was induced by H2O2. LC-MS/MS analysis confirmed that sulforaphane from broccoli sprouts was stable after simulated gastrointestinal digestion. It inhibited the release of all chemokines selected as inflammatory read-outs, with a more potent effect against MCP-1 (IC50 = 7.81 µM). On the contrary, glucoraphanin (50 µM) was inactive. The molecules were unable to counteract the oxidative damage to DNA (γ-H2AX) and catalase levels; however, the activity of NF-κB and Nrf-2 was modulated by both molecules. The impact on epithelial permeability (TEER) was also evaluated in a Transwell® model. In the context of a pro-inflammatory combination including gliadin, TEER values were recovered by neither sulforaphane nor glucoraphanin. Conversely, in the context of co-culture with activated macrophages (THP-1), sulforaphane inhibited the release of MCP-1 (IC50 = 20.60 µM) and IL-1ß (IC50 = 1.50 µM) only, but both molecules restored epithelial integrity at 50 µM. Our work suggests that glucoraphanin should not merely be considered as just an inert precursor at the small intestine level, thus suggesting a potential interest in the framework of CD. Its biological activity might imply, at least in part, molecular mechanisms different from sulforaphane.

Pigmented corn as a gluten-free source of polyphenols with anti-inflammatory and antioxidant properties in CaCo-2 cells.

A high number of varieties from corn (Zea mays L.) have been consumed for long time all over the world, however pigmented varieties are recently gaining renewed attention due to their beneficial effects and polyphenolic content. The natural lack of gluten makes corn suitable for consumption by celiac population, who need to control their inflammatory state through an appropriate gluten-free diet. The biological effects of polyphenols from pigmented corn are poorly investigated in the context of celiac disease. In this work, we analyzed through HPLC-DAD the phenolic composition of two Italian purple and red varieties ("Scagliolo Rosso" and "Rostrato di Rovetta", respectively) comparing their effects in human intestinal epithelial cells (CaCo-2 cells). The possible impact of gastro-intestinal digestion following oral consumption was assessed as well. The phenolic profile showed the presence of phenolic acids in both varieties, while anthocyanins were identified in Scagliolo Rosso only. After simulated digestion, the level of polyphenols did not significantly change and paralleled with an increased scavenging activity. In CaCo-2 cells, stimulated by a proinflammatory cocktail containing gliadin-derived peptides (IL-1ß, IFN-γ, digested gliadin), pigmented corn extracts inhibited the release of CXCL-10 and sICAM-1, with mechanisms partially ascribed to NF-κB impairment. At the same concentration (200 µg/mL), ROS production and catalase depletion were reverted through Nrf-2-independent mechanisms. Our data suggest that polyphenols from pigmented corns might help in controlling the inflammatory and oxidative state of people with celiac disease at intestinal level, at concentrations potentially achievable through a gluten-free diet.

Phenolic Profile and In Vitro Antioxidant Activity of Different Corn and Rice Varieties.

Celiac disease (CD) is an autoimmune disease. To date, the only universally recognized treatment for CD is the gluten-free diet (GFD). Despite the GFD, a state of inflammation and oxidative stress could remain at the intestinal level of celiac patients. Several components of the diet, such as phenolic compounds with known antioxidant properties, could play a protective role in the inflammatory state of patients with CD. The objective of this study was the characterization of the phenolic profile and the antioxidant capacity of pigmented cereals (rice and corn) from the Italian market and farms. Different in vitro methods were applied: Folin-Ciocalteu assay, pH differential method, DPPH assay, TEAC assay, and High-Performance Thin Layer Chromatography technique. According to the results, pigmented varieties are possible valuable sources of phenolic compounds and anthocyanins with high antioxidant activity. They could be used as alternative ingredients for the formulation of gluten-free products.

Evaluation of the Potential Anti-Inflammatory Activity of Black Rice in the Framework of Celiac Disease.

Inflammation and oxidative stress are two mechanisms involved in the pathogenesis of celiac disease (CD). Since the direct effect of gliadin on the intestinal epithelia is less studied, the aims of this study were the development of a specific cellular model based on the use of gliadin as a pro-inflammatory stimulus and the evaluation of the potential antioxidant and anti-inflammatory properties of extracts from different black rice in the framework of CD. The rice extracts were in vitro digested, characterized in terms of phenolic compounds and antioxidant capacity, and tested on Caco-2 cells to investigate their inhibitory effect on Reactive Oxygen Species, the NF-κB transcription and the CXC chemokines (sICAM-1, IL-8, and CXCL-10). In addition, the role of the extracts in modulating the activation of epithelial cells in CD was confirmed by applying the K562(S) agglutination test. The black rice extracts showed inhibitory effects on the production of the oxidative and the inflammatory mediators considered, with particular reference to lymphocyte-attracting CXCL-10 both before and after digestion. The presence of anthocyanins and their digestion metabolites may account for the observed anti-inflammatory activity after in vitro digestion. This work provided preliminary data supporting the use of black rice as a healthy food or ingredient of food supplements for celiacs.

Ancient and Modern Cereals as Ingredients of the Gluten-Free Diet: Are They Safe Enough for Celiac Consumers?

Celiac disease is an autoimmune disorder that occurs in genetically predisposed individuals after consuming prolamins from some cereals. Although the products available for celiac subjects have increased significantly in quality and quantity over the last few decades, research still focuses on identifying new ingredients to improve the nutritional, sensorial and functional qualities of gluten-free products. In terms of toxicity for people with celiac disease, there is a wide variability between ancient and modern grains. The most contradictory results are related to the role of oats in the gluten-free diet. In order to clarify the role of minor cereals (such as oat) and ancient grains in the diets of celiac patients, this review discusses recent in vitro and in vivo studies performed on those cereals for which the toxicity for celiac subjects is still controversial. According to in vivo studies, selected oat varieties could be tolerated by celiac patients. On the other hands, although some wheat-ancient grains (Triticum monococcum, Triticum aestivum ssp. spelta and Kamut®) showed a reduced in vitro toxicity, to date, these grains are still considered toxic for celiac patients. Contradictory results underline the importance of studying the safety of "unusual" cereals in more detail.