Guido Invernizzi

In vitro modulatory effect of ω-3 polyunsaturated fatty acid (EPA and DHA) on phagocytosis and ROS production of goat neutrophils

An in vitro study was carried out to examine the influence of two fish-oil-derived long chain omega-3 polyunsaturated fatty acids (PUFAs), eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), on goat polymorphonuclear leukocytes (PMN). Twelve Saanen healthy goats were used as blood donors. Neutrophils were isolated from blood and incubated with increasing concentration of EPA and DHA (25, 50, 100, 200muM). Control samples were incubated in the absence of omega-3 PUFAs. Phagocytosis was evaluated by fluorescein-labeled Escherichia coli incorporation, while extracellular Reactive Oxygen Species (ROS) production was determined by cytochrome c reduction assay, which was selected among the others due to its specificity for extracellular superoxide anion release. Phagocytic activity was significantly increased by EPA (P<0.05) and DHA (P<0.01). Treating PMN with EPA does not affect extracellular ROS production which is, on the contrary, down-regulated by DHA. This effect was increased in experimental conditions which mimic pro-inflammatory challenges (stimulation with PMA). This study demonstrates that EPA and DHA may have beneficial effect on neutrophil function by increasing their phagocytosis activity and, in the meanwhile, decreasing the tissue damages due to extracellular release of ROS.

Effects of inclusion of selenium-enriched yeast in the diet of laying hens on performance, eggshell quality, and selenium tissue deposition

The aim of this study was to evaluate the bioavailability of ingested selenium (Se) yeast in laying hens and its effects on performance, eggshell quality, and tissue Se distribution. Forty-eight ISA brown laying hens were divided into 3 treatment groups: Group C, fed a basal diet containing 0.11 mg Se/kg of feed; Group SS, fed a basal diet plus 0.4 mg/kg of feed of Se from sodium selenite; and Group SY, fed a basal diet plus 0.4 mg/kg of feed of Se from selenium yeast. Feed intake, egg mass ratio, and production performance were not affected by Se supplementation, regardless of the Se source. Egg weight (+3.61% and +2.95%), eggshell weight (+4.26% and +5.38%), and eggshell surface (+2.43% and +1.96%) were higher (P<0.05) in SS and SY than C, whereas breaking strength was increased in SY (P<0.01). Breast muscle, liver and skin Se levels were higher in SY than in C, while kidney Se content was higher in SS hens. Eggs from SY had higher Se levels than SS. Blood metabolites were not affected in SS or SY groups than C. A higher Se level was detected in eggs and breast muscle of SY hens (P<0.05). Seleniumenriched eggs and edible tissues from organic Se sources in poultry diet could improve antioxidant status in humans and reduce possible Se deficiency-related diseases.

Effects of EPA and DHA on lipid droplet accumulation and mRNA abundance of PAT proteins in caprine monocytes.

The present study investigated the in vitro effects on caprine monocytes of two ω-3 PUFAs, namely eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) on lipid droplet formation, an emerging process of fundamental importance in innate immunity regulation. The mRNA abundance of PAT protein family (PLIN1, PLIN2 and PLIN3), involved in the formation and trafficking of the droplets, was also assessed. The effects of EPA and DHA on monocyte apoptosis were studied as well. The number of lipid droplets per cell was found to be dependent on both type and concentration of fatty acid. ω-3 PUFAs upregulated PLIN3 and PLIN2 gene expression, as well as apoptosis rate. The present findings suggest that PUFA might modify innate immune functions of goat monocytes by interfering with the formation of lipid droplets and by upregulating proteins belonging to PAT protein family.

In vitro modulation of caprine monocyte immune functions by ω-3 polyunsaturated fatty acids.

The in vitro effects of the ω-3 polyunsaturated fatty acids (PUFAs) eicosapentanoic acid (EPA) and docosahexanoic acid (DHA) on phagocytosis and the extracellular respiratory burst in caprine monocytes were assessed. Blood monocytes incubated with increasing concentrations of EPA or DHA (25-200 μM) demonstrated increased phagocytosis compared to unexposed monocytes. Generation of reactive oxygen species (ROS) was not markedly affected in the presence of EPA and DHA, except at 200 μM, at which concentrations monocyte viability was also reduced.

Effect of live yeast (Saccharomyces cerevisiae) administration on apparent digestibility of horses

Abstract Six mares were divided into two groups of three subjects each in a crossover design on the basis of body weight and age: Control (C) fed a basal diet, Treatment (T) fed a basal diet and 2 g/head/d of live yeast (S. cerevisiae 4.6x1010 CFU/day The experimental design was divided into two periods named period 1 and period 2 respectively of 35d each, and consisting of 3 different phases. During each period all animals were subject to an adaptation phase of 14d (phase1); during phase 2 (18d) and phase 3 live yeast was administered (T) or not (C). Phase 3 consisted in a three days individual fecal collection period all the groups, in order to determine dry matter, organic matter, crude protein, crude fat, NDF and ADF apparent digestion rates using acid insoluble ash (AIA) as internal marker (Bergero et al., 2005). Results obtained evidenced as the administration of S. cerevisiae to mature horses resulted in increased digestibility of dry matter, organic matter, NDF, and ADF. ).

Selenium and Poultry Products: Nutritional and Safety Implications

Selenium (Se) is both an essential nutrient for humans and animals and a toxicant at excess levels in foods. Its content in animal products reflects that of the feeds consumed.

Nuclear factor E2-related factor 2’s activation in transgenic mice fed with dosage of saturated or unsaturated fatty acids using in vivo bioluminescent imaging

To counteract oxidative stress cells developed several mechanisms, including the transcription factor Nuclear Factor E2-related factor 2 (Nrf2). The aim of the study was to evaluate the activation of Nrf2 in transgenic mice fed saturated or polyunsaturated fatty acids and the anti-inflammatory effect of estrogens on organism. Forty-eight ARE CRE OMO reporter mice were divided into 3 groups, consisting of 16 animals, based on presence/absence of estrogens (ovariectomized or sham female, OVX - SH; male, MA). Each group was further split in 4 subgroups of 4 animals each and fed different diets (7.5% lard, 7.5% tuna oil, 20.0 % lard and 20.0% tuna oil). Two times a week animals were anaesthetized and injected i.p. with 100µL luciferin 15 min before the imaging session. Using the Living Image Software, photon emission was mapped for selected body areas. On day 70, animals were sacrificed after a challenge with Sodium Arsenite. Specific organs were dissected and immediately subjected to ex vivo imaging session. MIXED and GLM procedures of SAS software were used for statistical analysis. Dietary treatments did not affect body weight and feed intake as well as Nrf2 expression in both pre- and post-challenge phases, with the exception of the abdominal region (P=0.031 pre-challenge); in this area, during the pre-challenge phase, OVX showed lower Nrf2 activation (P<0.001). Ex vivo results outlined a significant effect of the challenge on all the considered organs (P<0.001), while OVX subjects had higher Nrf2 expression on urinary bladder and kidney (P<0.05) and high fat diet increased Nrf2 in urinary bladder (P<0.05). The present trial shows how saturated or polyunsaturated fatty acids supplementation in the diet do not exert significant effects on oxidative stress in mice, but confirms the protective role of estrogens under physiological condition.

Can lipid supplementation modulate inflammatory state and immune response in periparturient goats? A case study on hepatic and adipose miRNA expression

Saturated or unsaturated fatty acids compounds were administered to second-parity twins-diagnosed alpine dairy goats. Experimental groups were fed either calcium stearate (ST, n.7), fish oil (FO, n.8) or a control diet without any fat supplement (C, n.8) from one wk before (30g/head/d of fatty acids) to three wks after kidding (50g/head/d of fatty acids). ST provided 26% C16:0 and 69.4% C18:0 while FO provided 10.4% EPA and 7.8% DHA. Both ST and FO diets were formulated to be isocaloric and isonitrogenous, with the same calcium content. Previous obtained results on metabolic, productive parameters and mRNA expression of genes related to lipid metabolism and inflammatory response let us to consolidate the hypothesis that in goat lipogenesis is reduced across the transition period, if compared to cow, moreover FO can postpone or reduce lipomobilization. Based on these previous results, miRNA expression was performed on the same hepatic and adipose biopsies (collected on day -7 and 7 and 21 from kidding), as a new perspective in controlling cellular pathways, implicated in adipogenesis and metabolic and endocrine functions. We examined miR-26b and 155 for the infiltration of immune cells, miR-99a, 145 and 221 for the inflammation and lipolysis, miR-143 and 378 for pro-adipogenic function. MIXED and GLM procedures of SAS software were used for statistical analysis. No diet effect, but a time effect for miR-155 and a tendency for miR-221 were found. The increase of their expression over the time after kidding let us to speculate that goats have to face a postponed and more contained inflammation due to the lipolysis, when compared to cows.

Transcriptional regulation of lipid metabolism and inflammation in transition dairy goats by fish oil and stearate

To better understand the interaction between saturated or unsaturated fatty acids and its effect on expression of genes involved in subcutaneous adipose tissue metabolism, 23 second parity alpine dairy goats were enrolled in the experiment and fed either a non fat-supplemented basal diet (C; n=8), the basal diet supplemented with stearic acid (ST; n=7) or the basal diet supplemented with fish oil (FO; n=8). 30g/head/d supplemental fatty acids during the dry period and 50g/head/d during lactation were delivered starting one week before parturition up to 21 days in milk. Subcutaneous adipose tissue samples were harvested at day -7, 7 and 21 relative to kidding and mRNA levels of genes involved in inflammation were measured via qPCR. Data were analyzed using the MIXED procedure of SAS. No significant effects for treatment were observed, however eight genes were significant for time. HP and SAA3 expression peaked at day 7 postpartum, to then return at prepartum level around 21 d relative to kidding, while IL8, IL10, and IL18 expression constantly increased along the transition period. Vice versa, expression of IL1β, IL6R, and RXRA decreased in response to kidding, with a subsequent increase at day 21. The obtained results led us to hypothesize that goats face a postponed lipomobilization after kidding, probably related to their reduced production. The next step will involve the analysis of miRNA related to immune cell infiltration, adipocyte inflammation and lipolysis and positive regulation of adipogenesis to better understand the complex network of lipid metabolism in periparturient goats.

Plasma α-tocopherol content and its relationship with milk somatic cells count in Italian commercial herds.

This work was aimed to investigate relationship between plasma vitamin E concentration and milk somatic cell count in healthy cows in commercial herds. 49 multiparous cows from two commercial dairy herds were monitored from the day of dry off until 90 DIM. BCS was assessed and blood samples were collected at dry off, day 0, 30, 60 and 90 postpartum. Plasma was analyzed for α-tocopherol content. Quantification of NEFA, BOHB, Zn and Se was performed in serum samples. Milk production and composition was obtained from routinely test-day of Italian milk producers association. Somatic Cell Score (SCS) was calculated and included in the dataset. Analysis of data was performed using MIXED repeated and CORR procedures of SAS. We did not observe a correlation between plasmatic vitamin E and somatic cell score, and this can be explained by the low level of somatic cell score (averages 1.64 and 1.26). The lowest value of vitamin E was observed at parturition (1.64 µg/ml and 1.95 µg/ml). A significant (P<0.01) negative (-20%) correlation was observed between NEFA serum content and α-tocopherol plasma concentration. Serum selenium content was positively correlated (+42%, P<0.0001) to zinc concentration. Grouping cows on the basis of their plasma α-tocopherol content higher or lower than 3 μg/mL at dry off, SCS at 30 and 60 DIM tended to be higher in lactating animals with lower content of α-tocopherol (1.12 vs. 1.72, P=0.18 at 30d; 0.92 vs. 1.72, P=0.07 at 60d). However, plasma α-tocopherol content at dry off could be usefully correlated with somatic cell count in fresh cows.