Qualitative characterization of unrefined durum wheat air-classified fractions

Durum wheat milling is a key process step to improve the quality and safety of final prod-ucts. The aim of this study was to characterize three bran-enriched milling fractions (i.e., F250, G230 and G250), obtained from three durum wheat grain samples, by using an innovative micronization and air-classification technology. Milling fractions were characterized for main standard quality parameters and for alveographic properties, starch composition and content, phenolic acids, antioxidant activity and ATIs. Results showed that yield recovery, ash content and particle size distributions were influenced either by the operating conditions (230 or 250) or by the grain samples. While total starch content was lower in the micronized sample and air-classified fractions, the P/L ratio increased in air-classified fractions as compared to semolina. Six main individual phenolic acids were identified through HPLC-DAD analysis (i.e., ferulic acid, vanillic acid, p-coumaric acid, sinapic acid, syringic and p-hydroxybenzoic acids). Compared to semolina, higher contents of all individual phenolic components were found in all bran-enriched fractions. The highest rise of TPAs occurred in the F250 fraction, which was maintained in the derived pasta. Moreover, bran-enriched fractions showed significant reductions of ATIs content versus semolina. Overall, our data suggest the potential health benefits of F250, G230 and G250 and support their use to make durum-based foods

Effects of L-histidine on hydrogen peroxide-induced DNA damage and cytotoxicity in cultured mammalian cells.

L-Histidine markedly increased the growth- and DNA synthesis-inhibitory effects elicited by hydrogen peroxide in cultured Chinese hamster ovary cells. DNA single-strand breakage was also higher in the presence of the amino acid and, in addition, these breaks were characterized by a slower rate of repair, compared with that of the breaks generated by the oxidant alone. In the presence of L-histidine, hydrogen peroxide also produced DNA double-strand breakage, a lesion that cannot be detected in cells treated with even exceedingly high concentrations of the oxidant alone. Data reported herein suggest that the L-histidine-mediated increase of the cytotoxic response of cultured Chinese hamster ovary cells to hydrogen peroxide may be at least partially dependent on the formation of DNA double-strand break

Functional assessment of cancer therapy questionnaire for breast cancer (FACT-B+4): Italian version validation

BACKGROUND: Improvements in breast cancer diagnosis and treatment led to an increased incidence of survivors' rate. The healthcare system has to face new problems related not only to the treatment of the disease, but also to the management of the quality of life after the diagnosis. The aim of this study was to validate the Italian version of the Functional Assessment of Cancer Therapy - Breast (FACT-B+4) questionnaire and to evaluate its reliability. METHODS: The questionnaire was administered twice, with an interval of three days between each administration, to a cohort of women of the Breast Surgical Unit, PoliclincoUmberto I. Cronbach's alpha was used as a measure of the internal consistency of the Italian version. RESULTS: The Italian version of the tool was administered to 55 subjects. The Cronbach's alpha for most scores registered values >0.7, both at baseline and at the follow-up analysis, therefore the subscale showed good internal consistency. CONCLUSIONS: The Italian version of FACT-B+4 demonstrated acceptable reliability properties in the Breast Unit patients. The use of this questionnaire seemed to be effective and in line with the results derived from the English and Spanishversions. Internal consistency and validity had similar performance results

Isolation and preliminary characterization of a Chinese hamster ovary cell line with high-degree resistance to hydrogen peroxide.

We have isolated and conducted preliminary characterization of a cell line derived from the Chinese hamster ovary cell line AA8, which we have designated AG8 and which is highly resistant to the cytotoxic effects of H2O2 (approximately 17-fold when the H2O2 treatment was at 37 degrees; approximately 11-fold when the H2O2 treatment was at 4 degrees). AG8 cells were moderately (but significantly; P Be+) fast neutrons. As regards their biochemical status, AG8 and AA8 cells contain similar non-protein sulfhydryl levels per milligram of protein. Catalase activity (assessed by both spectrophotometry and polarography) was significantly higher in AG8 than in AA8 cells irrespective of whether enzyme activity was expressed per 10(6) cells (approximately 3.6-fold increase) or per milligram of protein (approximately 1.6-fold increase). AG8 cells also exhibited significantly greater glutathione reductase activity than wild-type cells when the data were expressed per 10(6) cells (approximately 2.9-fold) or per milligram of protein (approximately 1.3-fold). Glutathione peroxidase activity was immeasurably low in both cell lines. The susceptibility of the two cell lines to H2O2-mediated generation of DNA single-strand breaks (as measured by alkaline elution) indicated a slightly (approximately 1.5-fold) decreased yield in the resistant AG8 cell line. The two cell lines repaired these breaks with similar kinetics. In contrast, no measurable induction of DNA double-strand breaks (as measured by pulsed-field gel electrophoresis) was apparent in either cell line after survival-curve range concentrations of H2O2. On the basis of these data, it appears that the AG8 phenotype involves two previously identified resistance mechanisms, namely an adaptive component that may or may not involve increased antioxidant capacity, and a second component that does involve increased antioxidant (primarily catalase) capacity

CRISPR-Cas9 Multiplex Editing of the α-Amylase/Trypsin Inhibitor Genes to Reduce Allergen Proteins in Durum Wheat

Wheat and its derived foods are widespread, representing one of the main food sources globally. During the last decades, the incidence of disorders related to wheat has become a global issue for the human population, probably linked to the spread of wheat-derived foods. It has been ascertained that structural and metabolic proteins, like \u3b1-amylase/trypsin inhibitors (ATI), are involved in the onset of wheat allergies (bakers' asthma) and probably Non-Coeliac Wheat Sensitivity (NCWS). The ATI are a group of exogenous protease inhibitors, which are encoded by a multigene family dispersed over several chromosomes in durum and bread wheat. WTAI-CM3 and WTAI-CM16 subunits are considered among the main proteins involved in the onset of bakers' asthma and probably NCWS. A CRISPR-Cas9 multiplexing strategy was used to edit the ATI subunits WTAI-CM3 and WTAI-CM16 in the grain of the Italian durum wheat cultivar Svevo with the aim to produce wheat lines with reduced amount of potential allergens involved in adverse reactions. Using a marker gene-free approach, whereby plants are regenerated without selection agents, homozygous mutant plants without the presence of CRISPR vectors were obtained directly from T0 generation. This study demonstrates the capability of CRISPR technology to knock out immunogenic proteins in a reduced time compared to conventional breeding programmes. The editing of the two target genes was confirmed either at molecular (sequencing and gene expression study) or biochemical (immunologic test) level. Noteworthy, as a pleiotropic effect, is the activation of the ATI 0.28 pseudogene in the edited lines

The suppression of TdMRP3 genes reduces the phytic acid and increases the nutrient accumulation in durum wheat grain

Micronutrient malnutrition affects more than half of the world population. Reduced bioavailability of microelements in the raw materials is considered one of the main causes of mineral deficiency in populations whose diet is largely based on the consumption of staple crops. In this context, the production of low phytic acid (lpa) cereals is a main goal of the breeding programs, as phytic acid (PA) binds essential mineral cations such as iron (Fe), zinc (Zn), manganese (Mn), potassium (K), calcium (Ca) and magnesium (Mg) precipitating in the form of phytate salts poorly digested by monogastric animals, including humans, due to the lack of phytases in the digestive tract. Since PA limits the bioavailability of microelements, it is widely recognized as an anti-nutritional compound. A Targeting Induced Local Lesions IN Genomes (TILLING) approach has been undertaken to silence the genes encoding the TdABCC13 proteins, known as Multidrug-Resistance associated Proteins 3 (TdMRP3), transporters involved in the accumulation of PA inside the vacuole in durum wheat. The TdMRP3 complete null genotypes showed a significant reduction in the content of PA and were able to accumulate a higher amount of essential micronutrients (Fe, Zn, Mn) compared to the control. The number of spikelets and seeds per spike, traits associated with the agronomic performances, were reduced compared to the control, but the negative effect was in part balanced by the increased grain weight. The TdMRP3 mutant lines showed morphological differences in the root apparatus such as a significant decrease in the number of root tips, root length, volume and surface area and an increase in root average diameter compared to the control plants. These materials represent a promising basis for obtaining new commercial durum wheats with higher nutritional value

High resolution melting analysis for the detection of EMS induced mutations in wheat SBEIIa genes

Background Manipulation of the amylose-amylopectin ratio in cereal starch has been identified as a major target for the production of starches with novel functional properties. In wheat, silencing of starch branching enzyme genes by a transgenic approach reportedly caused an increase of amylose content up to 70% of total starch, exhibiting novel and interesting nutritional characteristics. In this work, the functionality of starch branching enzyme IIa (SBEIIa) has been targeted in bread wheat by TILLING. An EMS-mutagenised wheat population has been screened using High Resolution Melting of PCR products to identify functional SNPs in the three homoeologous genes encoding the target enzyme in the hexaploid genome. Results This analysis resulted in the identification of 56, 14 and 53 new allelic variants respectively for SBEIIa-A, SBEIIa-B and SBEIIa-D. The effects of the mutations on protein structure and functionality were evaluated by a bioinformatic approach. Two putative null alleles containing non-sense or splice site mutations were identified for each of the three homoeologous SBEIIa genes; qRT-PCR analysis showed a significant decrease of their gene expression and resulted in increased amylose content. Pyramiding of different single null homoeologous allowed to isolate double null mutants showing an increase of amylose content up to 21% compared to the control. Conclusion TILLING has successfully been used to generate novel alleles for SBEIIa genes known to control amylose content in wheat. Single and double null SBEIIa genotypes have been found to show a significant increase in amylose content

An Integrated Approach to Skeletal Muscle Health in Aging

A decline in muscle mass and function represents one of the most problematic changes associated with aging, and has dramatic effects on autonomy and quality of life. Several factors contribute to the inexorable process of sarcopenia, such as mitochondrial and autophagy dysfunction, and the lack of regeneration capacity of satellite cells. The physiologic decline in muscle mass and in motoneuron functionality associated with aging is exacerbated by the sedentary lifestyle that accompanies elderly people. Regular physical activity is beneficial to most people, but the elderly need well-designed and carefully administered training programs that improve muscle mass and, consequently, both functional ability and quality of life. Aging also causes alteration in the gut microbiota composition associated with sarcopenia, and some advances in research have elucidated that interventions via the gut microbiota-muscle axis have the potential to ameliorate the sarcopenic phenotype. Several mechanisms are involved in vitamin D muscle atrophy protection, as demonstrated by the decreased muscular function related to vitamin D deficiency. Malnutrition, chronic inflammation, vitamin deficiencies, and an imbalance in the muscle-gut axis are just a few of the factors that can lead to sarcopenia. Supplementing the diet with antioxidants, polyunsaturated fatty acids, vitamins, probiotics, prebiotics, proteins, kefir, and short-chain fatty acids could be potential nutritional therapies against sarcopenia. Finally, a personalized integrated strategy to counteract sarcopenia and maintain the health of skeletal muscles is suggested in this review