N-Acetylcysteine improves intestinal function and attenuates intestinal autophagy in piglets challenged with ß-conglycinin.

ß-Conglycinin (ß-CG), an anti-nutritional factor, is a major allergen in soybeans to induce intestinal dysfunction and diarrhea in neonatal animals, including piglets and human infants. This study with a piglet model determined the effects of N-acetylcysteine (NAC) on intestinal function and autophagy in response to ß-CG challenge. Twenty-four 12-day-old piglets (3.44 ± 0.28 kg), which had been weaned at 7 days of age and adapted for 5 days after weaning, were randomly allocated to the control, ß-CG, and ß-CG + NAC groups. Piglets in the control group were fed a liquid diet containing 10% casein, whereas those in the ß-CG and ß-CG + NAC groups were fed the basal liquid diets containing 9.5% casein and 0.5% ß-CG for 2 days. Thereafter, pigs in the ß-CG + NAC group were orally administrated with 50 mg (kg BW)-1 NAC for 3 days, while pigs in the other two groups were orally administrated with the same volume of sterile saline. NAC numerically reduced diarrhea incidence (- 46.2%) and the concentrations of hydrogen peroxide and malondialdehyde, but increased claudin-1 and intestinal fatty-acid binding protein (iFABP) protein abundances and activities of catalase and glutathione peroxidase in the jejunum of ß-CG-challenged piglets. Although ß-CG challenge decreased the villus height, villus height/crypt depth ratio, and mRNA levels of claudin-1 and occludin, no significant differences were observed in these indices between the control and ß-CG + NAC groups, suggesting the positive effects of NAC supplementation on intestinal mucosal barrier function. Moreover, NAC increased the concentrations of citrulline and D-xylose in the plasma, as well as the expression of genes for aquaporin (AQP) 3, AQP4, peptide transporter 1 (PepT1), sodium/glucose co-transporter-1 (SGLT-1), potassium inwardly-rectifying channel, subfamily J, member 13 (KCNJ13), and solute carrier family 1 member 1 (SLC1A1) in the jejunum, demonstrating that NAC augmented intestinal metabolic activity and absorptive function. Remarkably, NAC decreased Atg5 protein abundance and the LC3II/LC3I ratio (an indicator of autophagy) in the jejunum of ß-CG-challenged piglets. Taken together, NAC supplementation improved intestinal function and attenuated intestinal autophagy in ß-CG-challenged piglets.

Digestion and Transport across the Intestinal Epithelium Affects the Allergenicity of Ara h 1 and 3 but Not of Ara h 2 and 6.

SCOPE: No accepted and validated methods are currently available which can accurately predict protein allergenicity. In this study, the role of digestion and transport on protein allergenicity is investigated. METHODS AND RESULTS: Peanut allergens (Ara h 1, 2, 3, and 6) and a milk allergen (ß-lactoglobulin) are transported across pig intestinal epithelium using the InTESTine model and afterward basophil activation is measured to assess the (remaining) functional properties. Additionally, allergens are digested by pepsin prior to epithelial transport and their allergenicity is assessed in a human mast cell activation assay. Remarkably, transported Ara h 1 and 3 are not able to activate basophils, in contrast to Ara h 2 and 6. Digestion prior to transport results in a significant increase in mast cell activation of Ara h 1 and 3 dependent on the length of digestion time. Activation of mast cells by Ara h 2 and 6 is unaffected by digestion prior to transport. CONCLUSIONS: Digestion and transport influences the allergenicity of Ara h 1 and 3, but not of Ara h 2 and 6. The influence of digestion and transport on protein allergenicity may explain why current in vitro assays are not predictive for allergenicity.

Detection of ricin contamination in liquid egg by electrochemiluminescence immunosorbent assay.

UNLABELLED: A monoclonal antibody-based electrochemical luminescence method was developed for detecting and quantifying ricin in liquid egg, with a limit of detection of 0.2 ng/mL. Because this highly toxic protein, present in the seeds of Ricinus communis (castor), has been used for intentional poisoning in the past, it is important to have sensitive and reliable analytical methodology to detect ricin in food matrices such as liquid egg. The detection of this quantity of pure or crude ricin spiked into commercial samples of liquid egg provides approximately 50000-fold greater sensitivity than required to detect a toxic dose of ricin (>1 mg) in a 100 g sample. PRACTICAL APPLICATION: Because ricin has been used for intentional poisoning, there is a need for analytical methodology to detect ricin in food matrices to assure a safe food supply. Using monoclonal antibodies to ricin developed in our laboratory, we explored an assay readout system known as electrochemiluminescence. This technique afforded sensitive and specific analysis of ricin intentionally added to liquid egg and could potentially be used to monitor egg-based vaccine production.

Identification of Albizia lebbeck seed coat chitin-binding vicilins (7S globulins) with high toxicity to the larvae of the bruchid Callosobruchus maculatus

Seed coat is a specialized maternal tissue that interfaces the embryo and the external environment during embryogenesis, dormancy and germination. In addition, it is the first defensive barrier against penetration by pathogens and herbivores. Here we show that Albizia lebbeck seed coat dramatically compromises the oviposition, eclosion and development of the bruchid Callosobruchus maculatus. Dietary supplementation of bruchid larvae with A. lebbeck seed coat flour causes severe weight loss and reduces survival. By means of protein purification, mass spectrometry and bioinformatic analyses, we show that chitin-binding vicilins are the main source of A. lebbeck tegumental toxicity to C. maculatus. At concentrations as low as 0.1 percent, A. lebbeck vicilins reduce larval mass from 8.1 ± 1.7 (mass of control larvae) to 1.8 ± 0.5 mg, which corresponds to a decrease of 78 percent. Seed coat toxicity constitutes an efficient defense mechanism, hindering insect predation and preventing embryo damage. We hypothesize that A. lebbeck vicilins are good candidates for the genetic transformation of crop legumes to enhance resistance to bruchid predation.

Identification of Albizia lebbeck seed coat chitin-binding vicilins (7S globulins) with high toxicity to the larvae of the bruchid Callosobruchus maculatus.

Seed coat is a specialized maternal tissue that interfaces the embryo and the external environment during embryogenesis, dormancy and germination. In addition, it is the first defensive barrier against penetration by pathogens and herbivores. Here we show that Albizia lebbeck seed coat dramatically compromises the oviposition, eclosion and development of the bruchid Callosobruchus maculatus. Dietary supplementation of bruchid larvae with A. lebbeck seed coat flour causes severe weight loss and reduces survival. By means of protein purification, mass spectrometry and bioinformatic analyses, we show that chitin-binding vicilins are the main source of A. lebbeck tegumental toxicity to C. maculatus. At concentrations as low as 0.1%, A. lebbeck vicilins reduce larval mass from 8.1 ± 1.7 (mass of control larvae) to 1.8 ± 0.5 mg, which corresponds to a decrease of 78%. Seed coat toxicity constitutes an efficient defense mechanism, hindering insect predation and preventing embryo damage. We hypothesize that A. lebbeck vicilins are good candidates for the genetic transformation of crop legumes to enhance resistance to bruchid predation.

A 13-week sub-chronic dietary toxicity study of a cruciferin-rich canola protein isolate in rats.

The objective was to evaluate the safety of a cruciferin-rich canola protein isolate (Puratein) when fed as a protein source at various dietary levels to rats for 13-weeks. The study included four groups (20 animals/sex/group) of young Sprague Dawley rats. They were fed ad libitum with an AIN-93 G based protein-free diet added respectively with 5%, 10% and 20% (w/w) Puratein (test article) or 20% (w/w) vitamin-free casein (control article). Protein levels were adjusted in all groups at 18% using vitamin-free casein. Body weights, food consumption, locomotor activity and behavioral and clinical pathology parameters were recorded at various points of the study, followed by macroscopic examination, determination of organ weights and microscopic tissue examination. There were no test article-related effects on body weight, food consumption, clinical observations, functional observational battery, motor activity, clinical pathology, or ophthalmic examinations. A slightly higher thyroid/parathyroid weight (g/100g BW) noted in the 20% Puratein group was not correlated with histopathological changes. The no-observed-effect-level (NOEL) was 10%, whereas the no-observed-adverse-effect-level (NOAEL) was the highest fed level of 20%, equivalent to 11.24 g/kg BW/day for males and 14.11 g/kg BW/day for females. The cruciferin-rich canola protein isolate (Puratein) was considered safe under the conditions tested.

A catalogue of Triticum monococcum genes encoding toxic and immunogenic peptides for celiac disease patients.

The celiac disease (CD) is an inflammatory condition characterized by injury to the lining of the small-intestine on exposure to the gluten of wheat, barley and rye. The involvement of gluten in the CD syndrome has been studied in detail in bread wheat, where a set of "toxic" and "immunogenic" peptides has been defined. For wheat diploid species, information on CD epitopes is poor. In the present paper, we have adopted a genomic approach in order to understand the potential CD danger represented by storage proteins in diploid wheat and sequenced a sufficiently large number of cDNA clones related to storage protein genes of Triticum monococcum. Four bona fide toxic peptides and 13 immunogenic peptides were found. All the classes of storage proteins were shown to contain harmful sequences. The major conclusion is that einkorn has the full potential to induce the CD syndrome, as already evident for polyploid wheats. In addition, a complete overview of the storage protein gene arsenal in T. monococcum is provided, including a full-length HMW x-type sequence and two partial HMW y-type sequences.