Identification of RNAi hypoallergic bread wheat lines for wheat-dependent exercise-induced anaphylaxis patients.

Wheat-dependent exercise-induced anaphylaxis (WDEIA) is one of the most severe forms of wheat allergy. It occurs in patients when they exercise after ingesting wheat-containing foods. Nowadays, the only possible alternative for WDEIA patients is to avoid such foods. This study investigated the potential of six RNA of interference (RNAi) wheat lines with low-prolamin content as alternatives for WDEIA patients. For that purpose, a high performance-liquid chromatography (HPLC) analysis was performed to evaluate differences in gluten protein fractions among these lines. Next, western blots were conducted to measure the immunoglobulin E (IgE) reactivity to wheat proteins in sera from five WDEIA patients. Additionally, monoclonal antibodies (moAb) recognition sites and the IgE binding sites were searched in all peptides identified by LC-MS/MS after protein digestion. The results showed a 61.4%-81.2% reduction in the gliadin content of the RNAi lines, accompanied by an increase in their high-molecular weight (HMW) glutenin content compared to the wild type bread wheat line (WT). In all cases, the reduction in gliadin content correlated with a decrease in IgE reactivity observed in the sera of WDEIA patients, highlighting the E82 and H320 lines. These two RNAi lines exhibited a ≤90% reduction in IgE reactivity. This reduction could be attributed to an absence of IgE binding sites associated with α- and ω5-gliadins, which were present in the WT. Overall, these lines offer a potential alternative for foodstuff for individuals with WDEIA.

Duodenal lymphogram as a complementary tool in the diagnosis of celiac disease in adults.

INTRODUCTION: celiac disease (CD) patients have a specific pattern of lymphocytic infiltrate in the duodenal mucosa. Flow cytometry is a complementary tool for the diagnosis of CD, which allows the quantification and characterization of intraepithelial lymphocytes (IELs) by what is commonly called a lymphogram. Here we describe our experience with this technique in the diagnosis of CD in adult patients. METHODS: lymphograms from 157 patients performed in our center between 2009 and 2017 were retrospectively analyzed. Fourteen patients had a previous diagnosis of CD and followed a gluten-free diet (GFD), 21 had a new diagnosis of CD and the remaining were considered as non-celiac. The association of the lymphogram results (total IELs, CD3- lymphocytes and TcRγδ lymphocytes) with the CD diagnosis, compliance with the GFD, time since diagnosis and IgA anti-TG2 titer were determined. RESULTS: the area under the ROC curve of TcRγδ lymphocytes for CD patients varied between 0.86 and 0.86. The percentage of TcRγδ lymphocytes in GFD-treated patients was lower; 12 (8.5) vs 20.5 (8.7), p = 0.0153. However, it remained high compared to non-CD; 12 (8.5) vs 6.7 (6), p = 0.135. The time since diagnosis and IgA anti-TG2 titer correlated with the lymphogram results. Helicobacter pylori infection and treatment with angiotensin receptor antagonist 2 (ARA2) were associated with differences in the lymphogram results in patients without CD. CONCLUSIONS: the duodenal lymphogram is a reliable complementary tool in adults for the diagnosis of CD. However, compliance and duration of the GFD and other factors may condition its diagnostic capacity.

Expression and potential role of apolipoprotein D on the death-survival balance of human colorectal cancer cells under oxidative stress conditions.

PURPOSE: Inverse correlations of apolipoprotein D (ApoD) expression with tumor growth have been shown, therefore proposing ApoD as a good prognostic marker for diverse cancer types, including colorectal cancer (CRC). Besides, ApoD expression is boosted upon oxidative stress (OS) in many pathological situations. This study aims at understanding the role of ApoD in the progression of human CRC. METHODS: Samples of CRC and distant normal tissue (n = 51) were assayed for levels of lipid peroxidation, expression profile of OS-dependent genes, and protein expression. Three single-nucleotide polymorphisms in the ApoD gene were analyzed (n = 139), with no significant associations found. Finally, we assayed the effect of ApoD in proliferation and apoptosis in the CRC HT-29 cell line. RESULTS: In CRC, lipid peroxides increase while ApoD messenger RNA and protein decrease through tumor progression, with a prominent decrease in stage I. In normal mucosa, ApoD protein is present in lamina propia and enteroendocrine cells. In CRC, ApoD expression is heterogeneous, with low expression in stromal cells commonly associated with high expression in the dysplastic epithelium. ApoD promoter is basally methylated in HT-29 cells but retains the ability to respond to OS. Exogenous addition of ApoD to HT-29 cells does not modify proliferation or apoptosis levels in control conditions, but it promotes apoptosis upon paraquat-induced OS. CONCLUSION: Our results show ApoD as a gene responding to OS in the tumor microenvironment. Besides using ApoD as marker of initial stages of tumor progression, it can become a therapeutic tool promoting death of proliferating tumor cells suffering OS.

Cytokine gene polymorphisms in retinal detachment patients with and without proliferative vitreoretinopathy: a preliminary study.

PURPOSE: Cytokines and other growth factors such as interleukins play an important role in the pathogenesis of proliferative vitreoretinopathy (PVR). Interindividual variations in cytokine production seem to correlate with some cytokine gene polymorphisms. The purpose of this study was to analyse the distribution of these cytokine gene variants in patients with rhegmatogenous retinal detachment (RD) with and without PVR. METHODS: Single nucleotide polymorphisms were analysed for five cytokines: tumour necrosis factor-alpha (TNF-alpha), transforming growth factor-beta1 (TGF-beta1), interferon-gamma (IFN-gamma), interleukin-6 (IL-6) and interleukin-10 (IL-10). Patients were divided into two surgically treated groups of RD patients: group RD had 27 patients with RD, and group PVR had 31 patients with RD complicated by PVR. A control group was composed of 46 ethnically matched healthy individuals. RESULTS: The genotype distribution of the TGF-beta1 codon 10 polymorphism differed between PVR and RD patients (p = 0.018) and between PVR patients and controls in codon 25 (p = 0.011). There was a higher frequency of TGF-beta1 codon 10 allele T in PVR patients compared with RD patients (p = 0.023). No statistically significant differences between groups were observed for the other polymorphisms examined. CONCLUSION: An association between the TGF-beta1 genetic profile and the development of PVR was detected in this study. Further studies are necessary to confirm this finding and to establish its clinical relevance.