Grape pomace improves performance, antioxidant status, fecal microbiota and meat quality of piglets.

In the present study, grape pomace (GP) was used as feed additive in the diet of weaned piglets in order to develop innovative feedstuffs and to investigate their potential beneficial effects on welfare, productivity and meat quality. For examining the antioxidant capacity of the experimental feeds, 24 piglets of 20 days old were assigned to two experimental groups receiving standard or experimental diet for 30 days. Blood and tissues collections were performed at four different time-points, 2, 20, 35 and 50 days post birth. The collected tissues were brain, heart, kidney, liver, lung, quadriceps muscle, pancreas, spleen and stomach. The following oxidative stress markers were assessed: reduced glutathione (GSH), catalase activity, total antioxidant capacity (TAC), thiobarbituric acid reactive substances (TBARS), protein carbonyls (CARB) and H2O2 decomposition activity. The effect on bacterial growth was assessed by examining microbial populations in piglets' fecal microbiota. Furthermore, the average daily gain (ADG) was calculated and the fatty acid profile of quadriceps muscle was assessed. The results showed that piglets fed with the diet supplemented with GP, had significantly increased antioxidants mechanisms in almost all the tissues as shown by increases in GSH, H2O2 decomposition activity and TAC compared with control group. Piglets fed with the experimental diet exhibited decreased oxidative stress-induced damage to lipids and proteins as shown by decreases in TBARS and CARB in GP group compared with control. In addition, the experimental diet increased significantly ADG (by 23.65%) (P<0.05) and enhanced the growth of facultative probiotic bacteria (by up to 1.2 log colony forming units (CFU)/g) (P<0.05) and lactic acid bacteria (by up to 2.0 log CFU/g) (P<0.05) in GP group compared with the control group. GP supplementation inhibited the growth of pathogen populations such as Enterobacteriacae (by up to 1.8 log CFU/g) (P<0.05) and Campylobacter jejuni (by up to 1.0 log CFU/g) (P<0.05). Regarding fatty acid composition of meat, GP inclusion in piglets' diet increased significantly n-3 fatty acids (EPA; C20 : 5n-3, DHA; C22 : 6n-3, α-linolenic acid; C18 : 3n-3) and decreased significantly n-6/n-3 ratio compared with control (P<0.05). The results suggested that dietary GP supplementation may have a beneficial impact on piglets' welfare and may improve productivity as well as meat quality.

Grape pomace improves antioxidant capacity and faecal microflora of lambs.

Grape pomace is a by-product of winemaking process and rich in bioactive compounds such as plant polyphenols having antioxidant and antimicrobial activities. As known, oxidative stress may cause a number of pathological conditions in farm animals and thus affecting animal welfare and production. Moreover, pathogenic bacteria affect animals' health status. Thus, the objective of this study was to investigate whether lambs' feed supplemented with grape pomace enhances the antioxidant mechanisms and reduces the growth of pathogenic bacteria. For this purpose, redox status was assessed in blood and tissues using oxidative stress markers in lambs fed with diet supplemented with grape pomace. Moreover, the effect on bacterial growth was assessed by examining microbial populations in lambs' faecal microflora. The following oxidative stress markers were assessed: reduced glutathione (GSH), catalase activity (CAT), total antioxidant capacity (TAC), thiobarbituric acid-reactive substances (TBARS) and protein carbonyls (CARB). Twenty-four lambs were assigned to two experimental groups receiving standard or experimental diet for 55 days. Blood and tissues (liver, quadriceps muscle, spleen, heart and brain) were collected at two different time-points, 42 and 70 days post-birth, after ration administration. The results showed that lambs fed with experimental diet had significantly increased antioxidants mechanisms in blood and tissues as shown by increases in CAT and GSH compared to control. Moreover, lambs fed with the experimental diet exhibited decreased oxidative stress-induced damage to lipids and proteins as shown by decreases in TBARS and CARB respectively. In addition, the experimental diet enhanced the growth of facultative probiotic bacteria and inhibited the growth of pathogen populations such as Enterobacteriacae and E. coli. This is the first study investigating the effect of diet supplemented with grape pomace on the redox status and microbiota of lambs.

Effects of maternal undernutrition during late gestation and/or lactation on colostrum synthesis and immunological parameters in the offspring.

The emerging immune system is vulnerable to insult not only during fetal life, but also through colostrum transfer of maternal factors with immunomodulatory functions. The aim of the present study was to examine the effects of maternal undernutrition during late gestation and/or lactation on colostrum and milk synthesis, as well as on immunological parameters in offspring. Pregnant ewes were fed to 100% of nutrient requirements throughout pregnancy and lactation (Control) or to 50% during lactation (R1) or during the last 20 days of pregnancy and lactation (R2). Colostrum samples were collected 3 and 18h after parturition and thymus glands were obtained from 5-month-old offspring. Lamb birthweight did not differ between groups, whereas growth rate was significantly lower in males in the R1 group and in females in both undernourished groups. There was a significant reduction in lactose percentage in the 18-h colostrum of the R2 group. The IgG concentration, as a percentage of protein, was significantly increased in 3-h colostrum samples of the R2 group. Quantitative polymerase chain reaction analysis revealed a significant increase in the expression of Toll-like receptor (TLR) 2, TLR4 and TLR9 in the thymus gland of female lambs in both undernourished groups. In conclusion, early life nutritional imbalances may impact on immune system function in later life due to programming effects.

The role of early life nutrition in programming of reproductive function.

Accumulating evidence suggest that the concept of programming can also be applied to reproductive development and function, representing an ever expanding research area. Recently issues such as peri- or even preconceptional nutrition, transgenerational effects and underlying mechanisms have received considerable attention. The present chapter presents the existed evidence and reviews the available data from numerous animal and human studies on the effects of early life nutritional environment on adult reproductive function. Specific outcomes depend on the severity, duration and stage of development when nutritional perturbations are imposed, while sex-specific effects are also manifested. Apart from undernutrition, effects of relative overnutrition as well as the complex interactions between pre- and postnatal nutrition is of high importance, especially in the context of our days obesity epidemic. Mechanisms underlying reproductive programming are yet unclear, but may include a role for epigenetic modifications. Epigenetic modulation of critical genes involved in the control of reproductive function and potential intergenerational effects represent an exciting area of interdisciplinary research toward the development of new nutritional approaches during pre- and postnatal periods to ensure reproductive health in later life.

Effects of maternal undernutrition on the hypothalamic-pituitary-gonadal axis function in female sheep offspring.

A study was conducted to evaluate the effects of maternal undernutrition on the hypothalamic-pituitary-gonadal axis in female sheep offspring. Pregnant ewes were fed to 100% throughout pregnancy (Control) or to 50% from 0 to 30 (R1) or from 31 to 100 days of gestation (R2). Female lambs were selected and fed to appetite throughout the study. At 2, 5.5 and 10 months of age a GnRH challenge was conducted. At the age of 10 months lambs were synchronized and blood samples were collected at 3 h intervals for 72 h following sponge removal. At slaughter (10 months) ovaries were removed and examined macroscopically. Maternal undernutrition did not affect the time of the onset of puberty, defined as the first increase in plasma progesterone concentrations >or=1 ng/ml. The magnitude of the pre-ovulatory gonadotrophin surge and the time to surge were unaffected by treatment. The LH and FSH response to GnRH challenge did not differ between groups at 2 and 5.5 months but at 10 months of age a higher (p < 0.05) FSH response was found in R1 group. Although the total number of visible follicles and corpora lutea did not differ between groups, a significant higher (p < 0.05) number of small (2-3 mm diameter) follicles in R1 group and a significant lower number (p < 0.05) of corpora lutea with diameter 8-11 mm and not even one with diameter >12 mm were detected in the ovaries of R2 lambs. In conclusion, maternal undernutrition during the first month of pregnancy resulted in increased pituitary sensitivity to GnRH and increased number of small follicles in the ovary, while during mid to late gestation resulted in a reduction of large corpora lutea in female offspring.

Reduced Sertoli cell number and altered pituitary responsiveness in male lambs undernourished in utero.

The nutritional status of females during pregnancy can have profound effects on the fetus reproductive system development that could affect the reproductive potential as later as in adulthood (fetal programming). The aim of the present study was to examine the effects of maternal nutrient restriction imposed during different periods of gestation on pituitary responsiveness at different ages postnatal and on Sertoli cell number in male offspring. Pregnant ewes were fed to 100% of Metabolizable Energy and Crude Protein requirements throughout pregnancy (Control) or to 50% from 0 to 30 (R1) or from 31 to 100 days of gestation (R2). Male lambs were selected and fed to appetite throughout the study. At 2, 5.5 and 10 months of age a GnRH challenge was conducted. At slaughter (10 months) testes were removed and examined histologically. Maternal undernutrition did not affect the time of the onset of puberty, defined as the first increase in plasma testosterone concentrations > or =1 ng/ml. The LH and FSH response to GnRH challenge did not differ between groups at 2 and 5.5 months but at 10 months of age a higher (P<0.05) FSH response was found in R2 group. Testes weight did not differ between groups at slaughter. Mean Sertoli cell number was significantly lower in animals of R2 group compared with Control (P<0.01). A smaller seminiferous tubules diameter was detected in R2 group (P<0.05), while the diameter of the lumen was similar in all groups. Collectively, these results provide clear evidence for a direct effect of nutrient restriction during pregnancy on Sertoli cell number in adulthood. The lower number of Sertoli cells is the most candidate factor for the higher pituitary activity through a reduction in the suppressive effect of inhibin.

Impact of maternal undernutrition on the hypothalamic-pituitary-adrenal axis responsiveness in sheep at different ages postnatal.

Epidemiological and experimental data support the hypothesis of 'fetal programming', which proposes that alterations in fetal nutrition and endocrine status lead to permanent adaptations in fetal homeostatic mechanisms, producing long-term changes in physiology and determine susceptibility to later disease. Altered hypothalamic-pituitary-adrenal (HPA) axis function has been proposed to play an important role in programming of disease risk. The aim of the present study was to examine the effects of maternal nutrient restriction imposed during different periods of gestation on the HPA axis function in sheep, at different ages postnatal. Pregnant ewes were fed a 50% nutrient-restricted diet from days 0-30 (group R1, n = 7), or from days 31-100 of gestation (group R2, n = 7) or a control 100% diet throughout pregnancy, (Control, n = 8). Blood samples were collected at 10-day intervals from day 40 of gestation to term. Lambs were born naturally and fed to appetite throughout the study period. At 2, 5.5, and 10 months of age lambs were given an i.v. injection of corticotrophin-releasing hormone (CRH) and blood samples were collected at -15, 0, 15, 30, 60, 120, and 180 min postinjection. Maternal cortisol levels were significantly higher (P < 0.05) in group R1 compared with the other two groups, whereas maternal insulin levels were lower (P < 0.05) in group R2 compared with control. Birth weight of lambs was not affected by the maternal nutritional manipulation. The area under the curve for ACTH and cortisol response to CRH challenge was greater (P < 0.05) in lambs of group R1 at two months of age, whereas no difference was detected at the ages of 5.5 and 10 months. However, significantly higher (P < 0.01) basal cortisol levels were observed in lambs of R1 group at 5.5 months of age. There was no interaction between treatment and sex for both pituitary and adrenal responses to the challenge. A significant sex effect was evident with females responding with higher ACTH and cortisol levels at the age of 5.5 months (P < 0.01, P < 0.001 respectively) and with higher cortisol levels (P < 0.01) at 10 months of age than males. It is concluded that the HPA axis is programmable by altered nutrition in utero. The sensitivity of the axis to exogenous stimulation is enhanced during early postnatal life and attenuated with age, suggesting a role for the postnatal influences in resetting of the HPA axis and emphasizing the importance of identifying the impact of maternal undernutrition at several time points after birth.