Genistein antagonizes gliadin-induced CFTR malfunction in models of celiac disease.

In celiac disease (CD), an intolerance to dietary gluten/gliadin, antigenic gliadin peptides trigger an HLA-DQ2/DQ8-restricted adaptive Th1 immune response. Epithelial stress, induced by other non-antigenic gliadin peptides, is required for gliadin to become fully immunogenic. We found that cystic-fibrosis-transmembrane-conductance-regulator (CFTR) acts as membrane receptor for gliadin-derived peptide P31-43, as it binds to CFTR and impairs its channel function. P31-43-induced CFTR malfunction generates epithelial stress and intestinal inflammation. Maintaining CFTR in an active open conformation by the CFTR potentiators VX-770 (Ivacaftor) or Vrx-532, prevents P31-43 binding to CFTR and controls gliadin-induced manifestations. Here, we evaluated the possibility that the over-the-counter nutraceutical genistein, known to potentiate CFTR function, would allow to control gliadin-induced alterations. We demonstrated that pre-treatment with genistein prevented P31-43-induced CFTR malfunction and an epithelial stress response in Caco-2 cells. These effects were abrogated when the CFTR gene was knocked out by CRISP/Cas9 technology, indicating that genistein protects intestinal epithelial cells by potentiating CFTR function. Notably, genistein protected gliadin-sensitive mice from intestinal CFTR malfunction and gliadin-induced inflammation as it prevented gliadin-induced IFN-γ production by celiac peripheral-blood-mononuclear-cells (PBMC) cultured ex-vivo in the presence of P31-43-challenged Caco-2 cells. Our results indicate that natural compounds capable to increase CFTR channel gating might be used for the treatment of CD.

Fetuin B links vitamin D deficiency and pediatric obesity: Direct negative regulation by vitamin D.

Vitamin D (VD) deficiency (VDD) correlates to obesity, with VD a recognized mediator of metabolic diseases. From a previous proteomic study identifying adiponectin as a link between VDD and pediatric obesity, herein we analysed another protein (SSP2301) increased with VDD. A focused 2D-electrophoretic analysis identified 4 corresponding plasma proteins, with one predicted to be fetuin B (FETUB). FETUB was studied due to its emerging role in metabolic diseases and cytogenetic location (3q27.3) with adiponectin. Results were confirmed in obese children, where plasma FETUB was higher with VDD. A direct effect by 1α,25-(OH)2D3 on hepatocellular FETUB synthesis was observed, with a time and dose dependent reduction. Further, we demonstrated the VD-receptor (VDR) is key, with FETUB "released" with VDR silencing. Finally, VD supplementation (6weeks) to juvenile mice fed a standard diet, reduced plasma FETUB. Only at 22weeks did liver FETUB correspond to plasma FETUB, highlighting the contribution of other VD-responsive tissues. Overall, FETUB is a key protein linking VDD to pediatric obesity. With an emerging role in metabolic diseases, we demonstrate that VD/VDR directly regulate FETUB.

Thymosin α1 represents a potential potent single-molecule-based therapy for cystic fibrosis.

Cystic fibrosis (CF) is caused by mutations in the gene encoding the cystic fibrosis transmembrane conductance regulator (CFTR) that compromise its chloride channel activity. The most common mutation, p.Phe508del, results in the production of a misfolded CFTR protein, which has residual channel activity but is prematurely degraded. Because of the inherent complexity of the pathogenetic mechanisms involved in CF, which include impaired chloride permeability and persistent lung inflammation, a multidrug approach is required for efficacious CF therapy. To date, no individual drug with pleiotropic beneficial effects is available for CF. Here we report on the ability of thymosin alpha 1 (Tα1)-a naturally occurring polypeptide with an excellent safety profile in the clinic when used as an adjuvant or an immunotherapeutic agent-to rectify the multiple tissue defects in mice with CF as well as in cells from subjects with the p.Phe508del mutation. Tα1 displayed two combined properties that favorably opposed CF symptomatology: it reduced inflammation and increased CFTR maturation, stability and activity. By virtue of this two-pronged action, Tα1 has strong potential to be an efficacious single-molecule-based therapeutic agent for CF.

Reactive nitrogen species modulate the effects of rhein, an active component of senna laxatives, on human epithelium in vitro.

BACKGROUND: Senna laxatives are used worldwide. However, their misuse can lead to chronic mucosal inflammation with the accumulation of pigment-laden leukocytes and may cause colon cells to undergo apoptosis. This study explores the mechanisms by which rhein, an active component of senna, acts on a human intestinal cell line to induce ion secretion, apoptosis, and indirect chemotaxis of polymorphonuclear leukocytes. METHODS: Human colonic adenocarcinoma (CaCo-2) monolayer cells, in the presence or in the absence of rhein, were used to monitor the production of reactive nitrogen species using the Griess reaction. Modified Ussing chambers were used to study electrolyte secretion. The capacity to recruit human polymorphonuclear leukocytes was evaluated using masked well chemotaxis chambers. Rhein-induced apoptosis was investigated by counting apoptotic nuclei stained with Hoechst 33258 dye. RESULTS: Rhein caused a dose-dependent increase in short-circuit current that was abolished in chloride-free bathing buffer or by preincubating with 100 micromol/L NG-nitro-L-arginine (L-NAME) methyl ester. The concentration that maximally stimulated intestinal secretion, 50 micromol/L rhein, induced nitrate production. Supernatants obtained from CaCo-2 cultures after incubation with 50 micromol/L rhein stimulated a time-dependent polymorphonuclear leukocytes chemotaxis that was significantly decreased with 100 micromol/L L-NAME, whereas rhein per se was not active. Neutralizing antibodies anti-interleukin-8 (IL-8) and anti-ENA78 also inhibited chemotaxis. Overnight rhein incubation produced an increased number of apoptotic cells in the culture supernatant that was significantly decreased by preincubation with 100 micromol/L L-NAME. Light-degraded rhein had no effects on CaCo-2 monolayers. CONCLUSIONS: The integrity of rhein is crucial to generating nitric oxide, which mediates, with different time courses, ion secretion, chemotaxis, and apoptosis of human-derived cells.