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The present study, employing a comparative proteomic approach, analyzes the protein profile of pig claustrum (CLA), putamen (PU), and insula (IN). Pig brain is an interesting model whose key translational features are its similarities with cortical and subcortical structures of human brain. A greater difference in protein spot expression was observed in CLA vs PU as compared to CLA vs IN. The deregulated proteins identified in CLA resulted to be deeply implicated in neurodegenerative (i.e., sirtuin 2, protein disulfide-isomerase 3, transketolase) and psychiatric (i.e., copine 3 and myelin basic protein) disorders in humans. Metascape analysis of differentially expressed proteins in CLA vs PU comparison suggested activation of the α-synuclein pathway and L1 recycling pathway corroborating the involvement of these anatomical structures in neurodegenerative diseases. The expression of calcium/calmodulin-dependent protein kinase and dihydropyrimidinase like 2, which are linked to these pathways, was validated using western blot analysis. Moreover, the protein data set of CLA vs PU comparison was analyzed by Ingenuity Pathways Analysis to obtain a prediction of most significant canonical pathways, upstream regulators, human diseases, and biological functions. Interestingly, inhibition of presenilin 1 (PSEN1) upstream regulator and activation of endocannabinoid neuronal synapse pathway were observed. In conclusion, this is the first study presenting an extensive proteomic analysis of pig CLA in comparison with adjacent areas, IN and PUT. These results reinforce the common origin of CLA and IN and suggest an interesting involvement of CLA in endocannabinoid circuitry, neurodegenerative, and psychiatric disorders in humans.

Aging is a multifactorial phenomenon characterized by degenerative processes closely connected to oxidative damage and chronic inflammation. Recently, many studies have shown that natural bioactive compounds are useful in delaying the aging process. In this work, we studied the effects of an in vivo supplementation of the stilbenoid pterostilbene on lifespan extension in Drosophila melanogaster. We found that the average lifespan of flies of both sexes was increased by pterostilbene supplementation with a higher effect in females. The expression of longevity related genes (Sir2, Foxo, and Notch) was increased in both sexes but with different patterns. Pterostilbene counteracted oxidative stress induced by ethanol and paraquat and up-regulated the antioxidant enzymes Ho e Trxr-1 in male but not in female flies. On the other hand, pterostilbene decreased the inflammatory mediators dome and egr only in female flies. Proteomic analysis revealed that pterostilbene modulates 113 proteins in male flies and only 9 in females. Only one of these proteins was modulated by pterostilbene in both sexes: vacuolar H[+] ATPase 68 kDa subunit 2 (Vha68-2) that was strongly down-regulated. These findings suggest a potential role of pterostilbene in increasing lifespan both in male and female flies by mechanisms that seem to be different in the two sexes, highlighting the need to conduct nutraceutical supplementation studies on males and females separately in order to give more reliable results.

Nutrition has relevant consequences for human health and increasing pieces of evidence indicate that medicinal mushrooms have several beneficial effects. One of the main issues in Western countries is represented by the challenges of aging and age-related diseases, such as neurodegenerative disorders. Among these, Parkinson's disease (PD) affects 10 million people worldwide and is associated with α-synuclein misfolding, also found in other pathologies collectively called synucleinopathies. Here, we show that aqueous extracts of two edible mushrooms

Aging and age-related neurodegeneration are among the major challenges in modern medicine because of the progressive increase in the number of elderly in the world population. Nutrition, which has important long-term consequences for health, is an important way to prevent diseases and achieve healthy aging. The beneficial effects of Vigna unguiculata on metabolic disorders have been widely documented. Here, we show that an aqueous extract of V. unguiculata beans delays senescence both in Saccharomyces cerevisiae and Drosophila melanogaster, in a Snf1/AMPK-dependent manner. Consistently, an increased expression of FOXO, SIRT1, NOTCH and heme oxygenase (HO) genes, already known to be required for the longevity extension in D. melanogaster, is also shown. Preventing α-synuclein self-assembly is one of the most promising approaches for the treatment of Parkinson's disease (PD), for which aging is a risk factor. In vitro aggregation of α-synuclein, its toxicity and membrane localization in yeast and neuroblastoma cells are strongly decreased in the presence of bean extract. In a Caenorhabditis elegans model of PD, V. unguiculata extract substantially reduces the number of the age-dependent degeneration of the cephalic dopaminergic neurons. Our findings support the role of V. unguiculata beans as a functional food in age-related disorders.

The following supporting information can be downloaded at https://www.mdpi.com/article/10.3390/nu14204368/s1. Table S1. Analytes identified by GC/MS on the basis of match with NIST2014 library and the corresponding target ions used to quantify them. Table S2. Yeast strains used in this study. Figure S1 Calibration curve and linear regression curve for ET. Figure S2 Evaluation of interference of fungal extracts on ThT assay. Figure S3. High fungal extract concentrations are toxic for adult flies., Nutrition has relevant consequences for human health and increasing pieces of evidence indicate that medicinal mushrooms have several beneficial effects. One of the main issues in Western countries is represented by the challenges of aging and age-related diseases, such as neurodegenerative disorders. Among these, Parkinson’s disease (PD) affects 10 million people worldwide and is associated with α-synuclein misfolding, also found in other pathologies collectively called synucleinopathies. Here, we show that aqueous extracts of two edible mushrooms, Grifola frondosa and Hericium erinaceus, represent a valuable source of β-glucans and exert anti-aging effects in yeast. Their beneficial effects are mediated through the inhibition of the Ras/PKA pathway, with increased expression of heat shock proteins, along with a consistent increase of both mean and maximal lifespans. These fungal extracts also reduce the toxicity of α-synuclein heterologously expressed in yeast cells, resulting in reduced ROS levels, lower α-synuclein membrane localization, and protein aggregation. The neuroprotective activity of G. frondosa extract was also confirmed in a PD model of Drosophila melanogaster. Taken together, our data suggest the use of G. frondosa and H. erinaceus as functional food to prevent aging and age-related disorders, further supporting the neuro-healthy properties of these medicinal mushroom extracts., We acknowledge financial support from the Italian Ministry of University and Research (MUR) through grant “Dipartimenti di Eccellenza 2017” to University of Milano-Bicocca, Department of Biotechnology and Biosciences. This research was also supported by the Italian Ministry of University and Research (MUR): Dipartimenti di Eccellenza Program (2018–2022) Dept. of Biology and Biotechnology “L. Spallanzani”, and Fondo di Ricerca e Giovani (FRG, University of Pavia). M.L. was supported by a fellowship from Fondazione Umberto Veronesi. R.M. was supported by a fellowship from the Italian Ministry of University and Research (MUR).

This study investigated the effects of dietary linseed and physiological state on blood concentrations of some metabolic hormones and indicators, in transition dairy ewes. From 21 d before lambing to 60 d post-partum, ewes were provided with one of three isoenergetic and iso‑nitrogenous pelleted concentrates, without (CTR, n = 21), or with 100 (EL-10, n = 22) or 200 g/kg (EL-20, n = 22) of extruded linseed. Animals were fed alfalfa hay ad libitum and had access to mixed pasture. Plasma thyroid hormones and insulin concentrations were not affected by diet and significantly changed by time. The last week of pregnancy T3 (2.27 ± 0.15 ng/mL) and T4 (102.63 ± 2.85 ng/mL) were lower than at 4 weeks before lambing (T3: 3.27 ± 0.27 ng/mL; T4: 125.89 ± 2.63 ng/mL). After lambing, T3 rose to be highest at 4 and 8 weeks of lactation (3.41 ± 0.22). Mean insulin peaked 2 weeks after lambing (0.31 ± 0.02 ng/mL) vs. late pregnancy (0.22 ± 0.01 ng/mL) and progressing lactation (0.18 ± 0.01 ng/mL). Plasma Leptin concentration (2.43 ± 0.03 ng/mL) was not affected by diet nor time. All the blood metabolites and enzymes investigated showed significant time x treatment interaction. Differences of several haematological parameters were found in EL vs. CTR (cholesterol, triglycerides, total protein, bilirubin, ALP, ALT), however, in most cases the values were fairly within the reference physiological ranges. Metabolic hormones are confirmed to be mainly linked to the different physiological states, energy intake and variations of energy balance, without clear effects by different sources of energy and quality of dietary lipids.

Aging is a multifactorial phenomenon characterized by degenerative processes closely connected to oxidative damage and chronic inflammation. Recently, many studies have shown that natural bioactive compounds are useful in delaying the aging process. In this work, we studied the effects of an

Allium subhirsutum, known as hairy garlic, is a bulbous plant widespread in the Mediterranean area and locally used as a food and spice. In the present study, the chemical profile of the ethanolic extracts from bulbs (BE) and aerial parts (APE) were analyzed by HPLC-ESI-MSn, and antioxidant properties were evaluated by DPPH, ABTS and TEAC assays. The traditional use in the diet, and the well documented biological activity of Allium species suggest a potential as a new nutraceutical. For this reason, the potential usefulness of this food can be considered in the treatment and prevention of degenerative Alzheimer disease. For this reason, acetylcholinesterase inhibitory property was investigated. Furthermore, due to the observed presence of sulfur-containing and phenolic constituents, the cytotoxicity on tumor cells line was investigated. Results revealed significant AChE inhibitory activity for BE and APE. Both extracts exhibited also moderate antioxidant properties in the in vitro assays. Finally, limited cytotoxic activity was observed towards Human colon carcinoma and adenocarcinoma cell line, with differences between the individual parts tested. HPLC-ESI-MSn analysis showed that hairy garlic is a good source of sulphur compounds, flavonoids and phenylpropanoids derivatives, thus being a valid alternative to the common garlic (A. sativum). This work opens new opportunities for the application of A. subhirsutum as a health-promoting food.

Pulicaria undulata is used as a traditional herbal remedy in Egypt. We used gas chromatography-mass spectrometry for analysis of essential oil of this plant growing wild in Egypt and 64 compounds were identified. The oil was rich in oxygenated monoterpenes (64.0%) and aromatic derivatives (18.8%). The major components were carvacrol (46.5%), xanthoxylin (18.1%) and carvotanacetone (8.7%). The oil of the Egyptian plant showed significant differences from the oil results reported on this species derived from different accessions. Antioxidant activity was performed by FRAP, DPPH and ABTS assays, and the oil demonstrated a powerful antioxidant properties. Furthermore, cytotoxicity was assessed using MTT assay against three cell lines (A375, T98G, HCT116) and the oil showed moderate results with IC50 of 18.53, 40.64 and 22.23 μg/ml; respectively. The oil showed a good anti-acetylcholinesterase activity (IC50 = 139.2 μg/ml) using Ellman method. In conclusion, the studied oil exhibited a peculiar fingerprint and promising biological activities.

Increasing experimental data suggests that the regular intake of polyphenols represents a potential way to improve the quality of life in aging. Pterostilbene is a derivative of the polyphenols resveratrol that has higher bioavailability and neuroprotective activity than resveratrol itself. The present study was carried out to investigate the anti-aging and anti-inflammatory effects of a lifelong dietary supplementation of pterostilbene in Drosophila melanogaster focusing on the expression of genes related to lifespan and senescence regulation (SIRT1; FOXO; p16; NOTCH) or to inflammatory response (DOME and Eiger1), after 2 weeks (T1) or 2 months (T2) of pterostilbene supplementation. To test the anti-neuroinflammatory effect of pterostilbene, the expression of IL1 and iNOS has been investigated by RT-PCR in BV2 microglial cells stimulated with LPS. Pterostilbene supplementation significantly increased the mean life span of both male and female fruit flies. Consistently, pterostilbene supplementation significantly increased NOTCH and SIRT1 expression in female flies at T1 and T2, whereas only at T2 in males. FOXO and p16 were up-regulated at T1 in females and at T2 in males. Eiger and DOME were reduced only in females at T1 and T2, respectively. The anti-inflammatory effect was also observed in BV-2 cells were PTS significantly reduced the up-regulation of IL1 and iNOS induced by LPS. These data confirmed the the anti-inflammatory activity of pterostilbene and suggest that it is influenced by gender.

In the present study the use of Selenium-fortified olive leaves as potential dietary source of Se in rabbit nutrition was evaluated. Sixty New Zealand White rabbits (35 days of age) were randomly assigned to the following dietary treatments: standard diet (C), and C supplemented with either 10% olive leaves (OL) or 10% Selenium-fortified olive leaves (SeOL; 100 mg/L of foliar spray sodium selenate solution). At 70 days of age, 10 rabbits per group were slaughtered and the oxidative status, mineral profile and Selenium speciation of Longissimus dorsi meat was analyzed. Meat of the SeOL group exhibited better oxidative status (lower TBARS, higher GPx and α-tocopherol values) and a 5-fold higher Se content compared to that of the other treatments. The main Se form was SeMet (7-fold higher in the SeOL group), followed by SeCys2. The present trial demonstrates the possibility of using agro-industrial by-products as ingredients in rabbit feeds, thereby enriching meat bioactive compound content.

The effects of an oregano and/or rosemary (Origanum vulgare L. and/or Rosmarinus officinalis L.) dietary supplementation to the diet of fattening pigs were investigated. Thirty-two grower-finisher pigs (45 kg) were divided into four dietary groups identified as: control diet (CTR); CTR+ 0.2% oregano (O); CTR + 0.2% rosemary(R), and CTR+ 0.1% oregano + 0.1% rosemary (OR). During the finishing period, all groups received a further supplementation of 0.5% of conjugated linoleic acids (CLA). Blood samples were collected after an adaptation period of 15 days to the new diet (T1) and at the end of the finishing period (T2) to evaluate antioxidant status (total antioxidant power and reactive oxygen metabolites) and immune responses (lymphocytic phenotyping and IgG levels). Pork meat samples were evaluated for glutathione peroxidase activity (GSHPx), total phenolic content and preference rating. A significant increase in B lymphocytes (CD79+) and a higher IgG level was observed in the R and O groups (P

A small CD3+ T-cell population, that lacks both CD4 and CD8 molecules, defined as double negative (DN), is expanded in the peripheral blood of patients with systemic lupus erythematosus, produces IL-17 and accumulates in the kidney during lupus nephritis. Since IL-17 production is enhanced in salivary gland infiltrates of patients with primary Sjögren’s syndrome (pSS), we aimed to investigate whether DN T cells may be involved in the pathogenesis of salivary gland damage. Fifteen patients with SS and 15 normal controls (NC) were enrolled. Peripheral blood mononuclear cells were stimulated with anti-CD3 antibody and cultured in presence or absence of dexamethasone (Dex). Phenotypic characterization was performed by flow cytometry in freshly isolated cells and after culture. Minor salivary glands (MSG) from pSS were processed for immunofluorescence staining. Total circulating DN T cells were increased in pSS compared to NC (4.7±0.4% vs 2.6±0.4%). NC and pSS freshly isolated DN T cells produce consistent amounts of IL-17 (67.7±5.6 in NC vs 69.2±3.3 in pSS). Notably, DN T cells were found in the pSS-MSG infiltrate. Dex was able to down-regulate IL-17 in vitro production in NC (29±2.6% vs 15.2±1.9% vs 13±1.6%) and pSS (49±4.8% vs 16±3.8% vs 10.2±0.8%) conventional Th17 cells and in DN T cells of NC (80±2.8% vs 3.8±2.1% vs 4.2±1.8%), but not of pSS (81±1.5% vs 85.4±0.8% vs 86.2±1.7%). DN T cells are expanded in pSS PB, produce IL-17 and infiltrate pSS MSG. In pSS, conventional Th17 cells are inhibited by Dex, but DN T cells appear to be resistant to this effect. Taken together, these data suggest a key role of this T-cell subset in the perpetuation of chronic sialoadenitis and eventually in pSS prognosis.

Ethnopharmacological relevance The top flowering aerial parts of the Hypericum species are traditionally used to prepare ointments to heal cuts and burns. Sometimes even the fruits are used for these purposes. Hypericum androsaemum L., commonly known as tutsan or shrubby St. John's Wort, is a Mediterranean medicinal plant which has been traditionally used to prepare an ointment for treating cuts and wounds. Aim of the study To evaluate the extracts obtained from H. androsaemum red berries as functional ingredients for skin care formulations. Materials and methods The methanolic extract was obtained by Soxhlet extraction while the aqueous extract was prepared by decoction; their composition was determined by HPLC analysis. Their biological activities were measured in terms of proliferation and migration of human fibroblasts, inhibition of collagenase activity, and immunomodulatory effects on human peripheral blood mononuclear cells (PBMCs). In addition, we evaluated their photostability by UV spectroscopy and their protective effects against APPH-induced hemolysis in red blood cells (RBC). Results The polar extracts contained significant amounts of shikimic (108,143.7–115,901.3 mg/kg) and chlorogenic acids (45,781.1–57,002.7 mg/kg). The main components of these extracts made an important contribution to a significant increase in human fibroblast migration. Both extracts were also active as collagenase inhibitors, with the aqueous one showing a greater inhibitory capacity (IC 50 value of 88.1 µg/mL), similar to that of chlorogenic acid. The kinetic parameters determined for the enzymatic reaction revealed for both aqueous extract and chlorogenic acid an uncompetitive mechanism of inhibition. The methanolic extract showed important effects on PBMCs by modulating IL-6. Both extracts proved to be photostable in the UVA/B range and protected RBC against peroxidation at low concentrations. Conclusions H. androsaemum red berries were proven to contain phytochemicals that improve skin regeneration, hence potentially employable in skin care formulations.

Diabetes mellitus (DM) is a common disorder in middle-aged to older dogs. Treatment options, similar to those for humans, include insulin injection, dietary changes and exercise. Since some diabetic dogs may develop humoral immune responses to exogenous insulin resulting in treatment failure, the use of alternative medicine could represent an interesting new therapeutic strategy for DM in addition to the traditional insulin therapy. The main objective of this report was to evaluate a new therapeutic strategy for DM, based on the association of insulin injections with an orally administered extract of Boswellia serrata to improve glycemic control in a diabetic dog. A nine year old female neutered mixed breed dog diagnosed with DM was treated with increasing doses of a porcine insulin zinc suspension starting from 0.2 U/kg up to 0.6 U/kg q 12 hours. Twenty weeks after the start of insulin therapy the duration of effect and glucose nadir were ideal and clinical symptoms had disappeared but hyperglycemia, although less severe, persisted. Supplementation with a dry extract from the gum- resin of Boswellia serrata was initiated, at a dosage of 15 mg/kg q 12 hours, to improve insulin sensitivity or possible remaining endogenous insulin secretion. B. serrata supplementation led to good glycemic control. No side-effect or adverse reaction were observed during the study. The present case report provides the first evidence in veterinary medicine of a positive effect of dietary supplementation with boswellic acids associated with traditional insulin therapy on glycemic control in a diabetic dog. A major pitfall of the study is the lack of a control. More extensive clinical trials are required to provide definitive evidence of B. serrata efficacy. Key words: diabetes mellitus; dog; boswellic acids; glycemic control  VPLIV DODAJANJA BOSVELIJE ( Boswellia serrata ) OB ZDRAVLJENJU Z INZULINOM NA UREJANJE RAVNI GLUKOZE V KRVI PRI PSU S SLADKORNO BOLEZNIJO Sladkorna bolezen oz. diabetes mellitus (DM) je pogosta motnja pri psih v srednjih letih in starejAiih psih. MoA¾nosti zdravljenja, ki so podobne kot pri ljudeh, vkljuAujejo dodajanje inzulina, spremembe v prehrani in telesno aktivnost. Ker lahko nekateri psi s sladkorno boleznijo razvijejo imunski odziv na dodajanje inzulina, kar lahko vodi do neuspeAinega zdravljenja, lahko uporaba alternativne medicine predstavlja zanimivo novo metodo zdravljenja DM poleg tradicionalnega zdravljenja z inzulinom. Glavni cilj raziskave je bil ovrednotiti novo metodo zdravljenja DM, ki temelji na hkratnem dodajanju inzulina in peroralnem dodajanju izvleAka bosvelije ( Boswellia serrata ) za izboljAianje urejanja ravni glukoze v krvi pri psu s sladkorno boleznijo. Devet let stara sterilizirana samica meAiane pasme, ki je bila diagnosticirana s sladkorno boleznijo, je bila zdravljena z naraAiAajoAimi odmerki suspenzije praAiiAjega inzulina in cinka (od 0,2 E/kg do 0,6 U/kg), ki jih je dobivala vsakih 12 ur. Dvajset tednov po zaAetku zdravljenja z inzulinom so bili uAinki in najniA¾ja vrednost izmerjene glukoze v normalnih mejah vrednosti, kliniAni simptomi so izginili, le hiperglikemija je bila Aie vedno prisotna, vendar pa je bila manj resna. Dodatek suhega izvleAka iz gumijeve smole bosvelije Boswellia serrata v odmerku 15 mg/kg vsakih 12 ur je poveAal obAutljivost na inzulin oz. morebitne preostanke endogenega izloAanja inzulina. Dodatek B. serrata je pozitivno vplival na nadzorovanje sladkorne bolezni. Med zdravljenjem niso bili opaA¾eni stranski ali nezaA¾eleni uAinki. PoroAilo o opisanem primeru vsebuje prve dokaze o pozitivnem uAinku prehranskega dopolnila z bosveliAnimi kislinami v veterinarski medicini, povezanimi z obiAajnim zdravljenjem z inzulinom, pri nadzoru glukoze v krvi pri psu s sladkorno boleznijo. Najbolj neugoden del te Aitudije je odsotnost kontrole. Za zagotovitev dokonAnega dokaza o uAinkovitosti B. serrata so potrebni obseA¾nejAii kliniAni poskusi. KljuAne besede: diabetes mellitus ; pes; bosveliAna kislina; nadzor sladkorne bolezn

The effects of a diet supplemented with oregano essential oil on performance, oxidative status, pork quality traits and sensorial properties were evaluated. In two studies, 72 pigs in indoor or outdoor conditions were assigned to either a control diet or an identical diet supplemented with 0.2% oregano essential oil. Pigs reared outdoor showed lower live weight, average daily gain and average gain:feed ratio compared to indoor pigs. The oregano supplementation improved the growth performance of the outdoor- but not the indoor-reared animals. The serum oxidative status was influenced by the diet. A higher oxidative stability was observed in the oregano-supplemented groups. As for the rearing conditions, the data suggest that after an initial adapting period, the free-range farming systems could be better tolerated by pigs. Meat derived from pigs reared outdoor showed higher pH and a* values. Lightness was influenced by both the diet and the rearing conditions. The control group reared indoor showed shear force values higher than both supplemented groups, while no differences were detected with the control group reared outdoor. In the consumer test performed under blind conditions, the oregano groups achieved higher consistency scores compared with the control. Under informed conditions, the meat derived from the oregano-supplemented pigs reared outdoor received the highest scores for consistency and overall liking regardless of the rearing system. The same result for the overall liking score was obtained in the expectation test. The data obtained showed that dietary oregano essential oil can be effective in reducing performance losses due to the outdoor-rearing system, increasing the oxidative status of the animal and oxidative stability of the meat, without modifying the meat quality traits and improving consumer perceptions of the meat quality.

Boswellia serrata(BS) is an important traditional medicinal plant that currently represents an interesting topic for pharmaceutical research since it possesses several pharmacological properties (e.g., anti-inflammatory, antimicrobial, and antitumour). The safety and versatility of this dietary supplement should allow for its use in numerous pathological conditions; however the quality of the extracts needs to be standardized to increase the clinical success rate resulting from its use. In the present study, different commercially availableB. serrataextracts were employed to compare their AKBA content and in vitro antioxidant power. Furthermore, their ability to modulate the immune system regulatory properties was investigated. Our results showed that the AKBA content varied from3.83±0.10to0.03±0.004%, with one sample in which it was not detectable. The highest antioxidant power and phenolic content were shown by the same extract, which also exhibited the highest AKBA concentration. Finally, the BS extracts showed the ability to influence the regulatory and effector T-cell compartments. Our results suggest that frankincense should be further investigated for its promising potentiality to modulate not only inflammation/oxidative stress but also immune dysregulation, but attention should be paid to the composition of the commercial extracts.

In the present work we conducted a comprehensive chemical analysis of blue honeysuckle (Lonicera caerulea) spontaneously growing in eastern Russia. HPLC-DAD-ESI/MS analysis showed cyanidin-3-glucoside as the major constituent among phenolics, while nutritional analysis revealed fibre, protein, calcium and magnesium as the most important macro- and micronutrients, respectively. Fatty acid composition was dominated by polyunsaturated fatty acids, linoleic acid being the most abundant. Furthermore, we evaluated several in vitro biological activities such as antioxidant, antimicrobial, antiproliferative, wound healing and immunomodulatory effects of blue honeysuckle aqueous and ethanolic extracts that are often incorporated in food and nutraceutical preparations. While the fruit extracts were revealed to be potent radical scavengers with significant inhibition of ABTS radical, thus confirming the literature data, their inhibitory effects against microbial pathogens and tumor cell lines were negligible. The fruit aqueous extract did not show toxicity to human fibroblasts, but 24 h treatment with 150-200 μg per mL of extract slightly enhanced the cell migration when tested by scratched wound assay. Worth mentioning was the inhibitory effect displayed by the blue honeysuckle fruit aqueous extract on human lymphocytes.

A 57-day study was performed to determine the effects of two aqueous extracts (AEs) on broiler performance, immune function and intestinal microflora. Four groups of 75 one-day-old female broilers (Ross308) received one of the following treatments: (1) a standard commercial feed (C); (2) C supplemented with 2 g/kg rosemary AE (R); (3) C supplemented with 2 g/kg oregano AE (O); (4) C supplemented with 1 g/kg oregano AE + 1 g/kg rosemary AE (OR). Individual body weight, average daily gain, average daily feed intake and feed conversion efficiency were determined at 1, 11, 22, 36 and 57 days. Sample collections for IgG titration and intestinal microflora examination were performed at 22 and 57 days. The addition of oregano AE alone or in combination with rosemary AEs improved body weight up to 36 days of age (P

Hypericum androsaemum, also known as Tutsan, is a small evergreen shrub common in the Mediterranean basin where it is traditionally used as diuretic and hepatoprotective herbal drug. This plant possesses the peculiarity to produce fleshy and berry-like fruits that ripen from red to shiny black. In the present work, the chemical constituents of methanolic extracts and infusions of red and black fruits were analyzed by HPLC, and correlated with their antioxidant properties which were evaluated by the DPPH, β-Carotene/linoleic acid, and hypochlorous acid tests. In addition, the red pigment of the fruit was isolated by column chromatography and structurally elucidated by NMR. Results showed that H. androsaemum fruits contain high amounts of shikimic and chlorogenic acids, while their color was given by a tetraoxygenated-type xanthone, reported for the first time in Hypericum species. The red berries infusion gave the highest content of total phenolic compounds, DPPH, and hypochlorous acid scavenging activity, and β-carotene bleaching. Cytotoxicity of the berries extracts on three human tumor cell lines (malignant melanoma, breast adenocarcinoma, and colon carcinoma) was evaluated by MTT assay, and relevant inhibition on colon carcinoma cells (IC50 value of 8.4 μg/mL) was found. Finally, the effects of red berries extract on the immune system were evaluated by peripheral blood mononuclear cell (PBMC) proliferation assay that revealed a strong stimulation on lymphocytes at low doses (0.4–6 μg/mL).

Chemotypes, chemically distinct entities within a plant species based on genetic variation, are exceptionally frequent for secondary compounds such as essential oils and can influence the quality/activity of plant materials. This is the case with hyssop ( Hyssopus officinalis ) which is present in Italy with two main subspecies, officinalis and aristatus . While many phytochemical investigations have examined the former, there is a dearth of studies on the latter. In this work, we studied the morphology and histochemistry of H. officinalis subsp. aristatus growing in Abruzzo (central Italy), and conducted comprehensive phytochemical analysis of it. Furthermore, the cytotoxic, antioxidant, antimicrobial and immunostimulant effects of the essential oil were investigated. The essential oil composition, analysed by GC-FID and GC–MS, highlighted a peculiar chemotype, with linalool and methyl eugenol its main constituents. We also investigated the main chemical variations between dry and fresh batches collected in two different years. Furthermore, from the ethanolic extract, triterpenic acids, quinic acid derivatives, polyphenolic acids and, interestingly, an iridoid precursor, were isolated and structurally characterized by NMR. Study of the plant indumentum showed that the glandular trichomes vary in structure, distribution, secretion mode and in the chemical composition of the secretory products. Finally, the biological assays conducted on the essential oil highlighted moderate antioxidant and antimicrobial activity, noticeable cytotoxicity on human tumor cells and potential immunostimulant effects that are worthy of further investigations.

The aim of this study was to determine the effects of age on chemical composition and tenderness of donkey meat. These characteristics were determined on Longissimus thoracis samples taken from 40 entire donkey males of Martina Franca breed slaughtered at 12 (Group 1, n = 20) and at 18 months (Group 2, n = 20) of age. Tenderness was evalu-ated at two and seven days post slaughter using a Warner-Bratzler shear force device. The chemical composition showed a significant increase in protein content (P < 0.05) in the meat of donkeys slaughtered at 18 months of age (22.3 g/100 g) compared to the meat of younger animals (21.4 g/100 g). Intramuscular fat content was significantly lower (P < 0.05) in Group 1 (2.41 g/100 g) compared to Group 2 (3.71 g/100 g). Shear force values at seven days post slaughter were significantly (P < 0.01) lower (5.15 ± 0.31 kg/cm2) in Group 1 compared with the results obtained at two days post slaughter (6.25 ± 0.53 kg/cm2). In Group 2 shear force values obtained seven days post mortem were higher (5.88 ± 0.23 kg/cm2) compared with Group 1 after the same ageing period but significantly (P < 0.05) lower if compared with those (6.53 ± 0.23 kg/cm2) determined in the same animals two days post slaugher.

A study based on 15 entire donkey males was carried out to evaluate carcass quality and nutritional characteristics of meat obtained by these animals slaughtered at 15 months of age and a mean final body weight of 181 kg. The meat had a low (2.02 g/100 g) fat content, an appreciable (22.8 g/100 g) protein content, and cholesterol content was 68.7 mg/100 g. Glycogen was also determined (0.45 g/100 g) within 12 h of sampling. Potassium was the mineral with the highest content (343 mg/100 g), followed by phosphorus (212 mg/100 g), sodium (52 mg/100 g) and magnesium (24 mg/100 g). Donkey meat obtained from young animals can be considered a good alternative to other red meats and not only for the production salami, or other fermented meat products.

Objectives Growing evidence suggests that IL-17-producing T cells, lacking both CD4 and CD8 molecules and defined as double negative (DN) cells, play a pivotal role in the pathogenesis of a number of systemic autoimmune disorders. We recently demonstrated that this T-cell subset is expanded in the peripheral blood (PB) of patients with primary Sjogren's syndrome (pSS), produces IL-17 and accumulates in minor salivary glands (MSGs). We aimed to investigate glandular and PB DN T cells in early pSS in order to verify a possible correlation with MSGs histological patterns and clinical parameters. Methods Paired samples of PB mononuclear cells and MSGs from pSS patients were evaluated at the diagnosis by flow cytometry and immunofluorescence staining respectively. Histological analysis to identify histological scores, B/T cell segregation and the presence of germinal center (GC)-like structures was also performed. Results In early stages of pSS, circulating DN T cells appear to be not yet expanded and inversely correlated with circulating CD4+Th17 cells. The number of infiltrating DN T cells were associated with extent of glandular involvement, presence of GC-like structures and dryness symptoms and were inversely correlated with circulating DN T cells. Conclusions Our findings suggest that DN T cells are actively involved in the pathogenic mechanisms leading to glandular dysfunction and damage in pSS and may play a role in ectopic lymphoneogenesis development occurring during the disease.

article i nfo Article history: The effects of a pre-formulated commercial plant extract mix, composed of equal parts of oregano essential oil and sweet chestnut wood extract, on performance, oxidative status and pork quality traits were evaluated. In two155-dstudies,60 pigs(meanliveweight: 42.9 kg)were assignedtoeithera controldiet(CTR)or anidentical diet supplemented (0.2%) with the plant extract mix (OC). No differences in the growth rate were observed. Glutathione peroxidase and glutathione reductase activities in the OC muscles (Longissimus lumborum) were higher than in CTR muscles. The lipid oxidation of meat was lower in the OC group. In the cooked meat samples, OC animals had the lowest L* and H° values and the highest a* values. The OC meat received higher scores for colour, taste and overall liking in both the blind and the labelled consumer tests.

The quality of meat obtained by Fabrianese heavy lambs (33 kg) reared in the Italian Sibillini mountains national park under extensive conditions had been investigated. Twenty entire male lambs had been used in this experiment; the Longissimus Thoracis et Lumborum muscle, taken from all the 20 animals, was analyzed for chemical composition, fatty acid, cholesterol and mineral contents. Protein content was 22.1%, fat content 4.5%, cholesterol 63.5 mg/100 g. The most represented fatty acid was oleic acid (37.62% of total fatty acids), followed by palmitic acid (26.31% of total fatty acids). Phosphorus and potassium were the two most represented minerals in muscles examined, respectively 285.6 and 329.5 mg/100 g. The results of this study demonstrated the good quality of the meat obtained by a local ovine breed slaughtered at higher final body weight when compared with the most common suckling lambs normally commercialized in Italy.

Hypoallergenic properties of ass’s milk protein fractions have been recently con- firmed, allowing ass’s milk to be considered as a valid substitute of the available hypoallergenic infant formulas. The objective of this study was to give a further contribution to the knowledge of ass’s milk safety and quality characteristics. A new procedure has been developed with a cytospin centrifuge in differential counts of milk somatic cells. Somatic cells count (SCC), differential somatic cells count (DCC) and cultural examinations have been carried out in 62 milk samples collected from 11 asses at three different stages of lactation. Four major cells populations had been identified in ass’s milk too: lymphocytes (Ly), monocytes/macrophages (MA), polymorphonuclear neutrophils (PMNL), and epithelial cells (CE). The patterns of these cells have been discussed in comparison with cells found in dairy cows and ewes milk. In conclusion, a reproducible standard procedure has been developed to determine cell count of ass’s milk.

Tenderness and mineral levels were determined in the Longissimus thoracis taken from 20 llama and 30 alpaca males reared in Peru and slaughtered at 25 months of age. Mineral contents were determined using an inductively coupled plasma emission spectrometer. Tenderness evaluation was determined two and seven days post slaughter using a Warner-Bratzler shear force device. Potassium is the mineral with the highest content, with a significant difference (P < 0.05) between the two species of camelids. The other mineral contents were, in decreasing order, phosphorus, sodium, magnesium and calcium, in addition to smaller percentages of zinc and iron. Shear force values determined seven days post slaughter were significantly (P < 0.01) lower in both the species compared with the results obtained two days post slaughter.

This research examined the effects of dietary alpha-tocopheryl acetate (50 or 200 mg/kg diet) and selenium (Se, 0 or 0.5 ppm) supplementation on motion characteristics, oxidative stability and fertilizing ability of rabbit spermatozoa, fresh and stored for 24 h at 5 degrees C. The higher amount of dietary alpha-tocopheryl acetate increased the level of Vitamin E in the fresh semen (1.75 mmol/l versus 0.95 mmol/l) and its oxidative stability (thiobarbituric acid reactive substances-TBARS 12.44 nmol malondialdehyde/10(8) sperm versus 21.4 nmol malondialdehyde/10(8) sperm). Dietary Se increased gluthatione peroxidase activity (GPx) in erythrocytes (285 U/g Hb versus 207 U/g Hb), seminal plasma (270 U/l versus 190 U/l) and spermatozoa (1338 mU/10(9) sperm versus 1103 mU/10(9) sperm), whereas it did not show any effect on alpha-tocopherol level and TBARS. No synergy between Vitamin E and Se was shown. Storage for 24 h at 5 degrees C increased the TBARS level in all the experimental groups. Neither live and acrosome reacted spermatozoa, nor kinetic parameters, nor fertility rate were modified by dietary supplementation.

Background The discovery of IL-17-producing CD4 + T helper (Th17) cells challenged the long-standing paradigm of Th1/Th2 cell immune response and shed some light on the pathogenic mechanisms of systemic autoimmune disorders. IL-17 in a pro-inflammatory cytokine which mediates target organs damage during the disease. To note, it has been recently observed that asmall CD3 + T-cell population, that lacks of both CD4 and CD8 molecules, defined as double negative (DN), is expanded in the peripheral blood of patients with systemic lupus erythematosus, produces IL-17 and accumulates in the kidney during lupus nephritis. Objectives Since IL-17 production is enhanced in salivary gland infiltrates of patients with primary Sjogren’s syndrome (SS), we aimed to investigate whether DN T cells may be involved in the pathogenesis of salivary gland damage Methods Thirty patients with SS and 16 normal controls (NC) were enrolled. CFSE-stained PBMCs were cultured in anti-CD3-coated plates in presence or absence of dexamethasone (Dex) at different concentrations. Phenotypic characterization was performed by flow cytometry in freshly isolated cells and after culture using anti-CD3, CD4, αβ-TCR, γδ-TCR, RORγt, IL-17 antibodies and respective isotypes. SS minor salivary glands (MSG) were processed for immunofluorescence staining. Results Total circulating DN T cells were increased in SS compared to NC. This was due to an increase of both αβ-TCR + and γδ-TCR + cells. NC and SS freshly isolated DN T cells express RORγt and produce consistent amounts of IL-17. Notably, DN T cells were found in the SS-MSG infiltrate. αβ-TCR + , but not γδ-TCR + , DN T-cell expansion in SS was dependent on disease activity according to the EULAR Sjogren’s syndrome disease activity index (ESSDAI). Dex was able to down-regulate IL-17 in vitro production in NC and SS CD4 + cells and in NC DN T cells, but not in DN T cells from SS samples. Conclusions DN T cells are expanded in SS PB, produce IL-17 and infiltrate SS MSG. The expansion is dependent on disease activity. In SS, conventional Th17 cells are inhibited by Dex, but DN T cells appear to be resistant to this effect. The recognition of a pathogenic T-cell subset, that appears to be in vivo activated and not sensitive to CS, in an autoimmune disorder such as SS may represent an intriguing finding with potential therapeutic implications of great interest. References Crispin C et al. J Immunol 2008;181:8761 Nguyen CQ et al. Arthritis Rheum 2008;58:734 Disclosure of Interest None Declared

Summary On 700 milk samples from single half udders of Comisana ewes, somatic cell count (SCC) and differential cell count (DCC) were determined, using a Fossomatic 90 cell counter (Foss Electric, Denmark) (SCCF) and milk smears stained with May Grunwald-Giemsa (DCCS). SCC and DCC were also determined with modified KOVA® SYSTEM (Hicor Biomedical Inc. Irvine, CA, USA) (SCCK and DCCK, respectively). Out of 665 milk samples from half udders without clinical signs of mastitis, 640 (Class I) were sterile, while 25 (Class II) were bacteriologically positive. Out of 35 milk samples (Class III) from half udders with clinical signs of mastitis, 25 were bacteriologically positive. Mean results (after logarithmic transformation of cells/ml/103) of SCCF and SCCK for all the 700 milk samples were 1.89 ± 0.58 and 1.86 ± 0.60 with linear correlation coefficient (r) of 0.960, while least squares means for Class I, II and III were 1.78, 2.23 and 3.73 respectively and 1.75, 2.19 and 3.74 with r of 0.894, 0.979 and 0.987. Mean results of DCCS and DCCK were 38.1 ± 23.3, 34.9, 52.1, and 82.2 PMNL% and 41.8 ± 21.7, 38.6, 60.2, and 87.3 PMNL% with r of 0.855, 0.812, 0.697 and 0.805. The results showed high correlation coefficients and a good reliability between SCCK and SCCF and high correlation coefficients for DCC methods. In conclusion, it could be suggested that the possibility of routine use of the KOVAr SYSTEM method is particularly useful in detecting if an abnormal SCC is due to a polymorphonuclear neutrophil leukocytes increase.

Milk is one of the most common causes of food allergies among children under one year of age. No specific therapy exists for this allergy, and thus the only feasible response is to avoid assumption of milk and derived products. Studies conducted on the serum of children with hypersensi- tivity to milk have shown that caseins are the proteins with the greater allergenic potential. However, in some cases, children have also shown hypersensitivity to the β-lactoglobulines and to the α-lactal- bumins. When food intolerance is diagnosed in an infant, it is often necessary to impose a period of total parenteral feeding, followed by breast feeding, considered the most correct method of re-feeding. When human milk can not be given, alternative food sources must be sought. Clinical studies have demonstrated that donkey milk could substitute breast feeding in infants affected by severe Ig-E me- diated milk allergies. In these subjects, donkey milk is not only useful, but also safer than other types of milk. In fact donkey milk composition in lipids (high levels of linoleic and linolenic acid) and pro- teins (low caseins content) is very close to human milk. Lysozyme content in donkey milk resulted to be very high (mean value 1.0 mg/ml) if compared to bovine (traces), caprine (traces) and human milk. The high lysozyme content of donkey milk may be responsible of the low bacterial count reported in literature and also makes this milk suitable to prevent intestine infections to infants. Among seropro- teins, β-lactoglobulin and α-lactalbumin content in donkey milk was respectively 3.75 and 1.80 mg/ml and remained substancially the same during the different stages of lactation.