Oxidative stress increases in liver of lactating rats after BPF-low-dose exposure: perinatal effects in the offspring.

Bisphenol F (BPF) is replacing Bisphenol A (BPA) in the manufacture of products due to endocrine-disrupting effects. BPF monomers can also be released into the environment and enter the food chain, resulting in human exposure to low doses. Since bisphenols are primarily metabolized by the liver, this organ is more vulnerable to lower doses of bisphenols than others. Exposure during prenatal development may increase the risk of diseases in adulthood. The aim was to evaluate whether BPF administration could generate oxidative stress in liver of lactating rats, and whether these effects may be also observed in female and male postnatal day 6 (PND6) offspring. Long Evans rats received oral treatment: Control, BPF-low-dose (LBPF) 0.0365 mg/kg b.w./day, and BPF-high-dose (HBPF) 3.65 mg/kg b.w./day. The levels of antioxidant enzymes (CAT, SOD, GR, GPx and GST), glutathione system (GSH, GSSG) and lipid damage markers (MDA, LPO) were measured using colorimetric methods in liver of both lactating dams and in PND6 offspring. Mean values were analyzed using Prism-7. LBPF affected liver defense mechanisms (antioxidant enzymes and glutathione system), increasing ROS levels and producing lipid peroxidation in lactating dams. Similar effects were found in female and male PND6 offspring as a consequence of perinatal exposure.

Influence of aging and growth hormone on different members of the NFkB family and IkB expression in the heart from a murine model of senescence-accelerated aging.

Inflammation is related to several pathological processes. The aim of this study was to investigate the protein expression of the different subunits of the nuclear factor Kappa b (NFkBp65, p50, p105, p52, p100) and the protein expressions of IkB beta and alpha in the hearts from a murine model of accelerated aging (SAM model) by Western blot. In addition, the translocation of some isoforms of NFkB from cytosol to nuclei (NFkBp65, p50, p52) and ATP level content was studied. In addition we investigated the effect of the chronic administration of growth hormone (GH) on these age-related parameters. SAMP8 and SAMR1 mice of 2 and 10 months of age were used (n = 30). Animals were divided into five experimental groups: 2 old untreated (SAMP8/SAMR1), 2 young control (SAMP8/SAMR1) and one GH treated-old groups (SAMP8). Age-related changes were found in the studied parameters. We were able to see decreases of ATP level contents and the translocation of the nuclear factor kappa B p50, p52 and p65 from cytosol to nuclei in old SAMP8 mice together with a decrease of IKB proteins. However p100 and p105 did not show differences with aging. No significant changes were recorded in SAMR1 animals. GH treatment showed beneficial effects in old SAMP8 mice inducing an increase in ATP levels and inhibiting the translocation of some NFkB subunits such as p52. Our results supported the relation of NFkB activation with enhanced apoptosis and pro-inflammatory status in old SAMP8 mice and suggested a selective beneficial effect of the GH treatment, which was able to partially reduce the incidence of some deleterious changes in the heart of those mice.

Reduced apurinic/apyrimidinic endonuclease 1 activity and increased DNA damage in mitochondria are related to enhanced apoptosis and inflammation in the brain of senescence- accelerated P8 mice (SAMP8).

The senescence- accelerated mouse prone 8 (SAMP8) is a well- characterized animal model of senescence that shows early age- related neurodegeneration with impairment in learning and memory skills when compared with control senescence- resistant mice (SAMR1). In the current study, we investigated whether such impairment could be partly due to changes in mitochondrial DNA (mtDNA) repair capacity and mitochondrial DNA damage in the brain of SAMP8 mice. Besides we studied whether these potential changes were related to modifications in two major processes likely involved in aging and neurodegeneration: apoptosis and inflammation. We observed that the specific activity of one of the main mtDNA repair enzymes, the mitochondrial APE1, showed an age- related reduction in SAMP8 animals, while in SAMR1 mice mitochondrial APE1 increased with age. The reduction in mtAPE1 activity in SAMP8 animals was associated with increased levels of the DNA oxidative damage marker 8oxodG in mtDNA. Our results also indicate that these changes were related to a premature increase in apoptotic events and inflammation in the brain of SAMP8 mice when compared to SAMR1 counterparts. We suggest that the premature neurodegenerative phenotype observed in SAMP8 animals might be due, at least in part, to changes in the processing of mtDNA oxidative damage, which would lead to enhancement of apoptotic and inflammatory processes.

Articulating the value of health research.

Previous contributions to Quarterly Journal of Medicine have drawn attention to the work of FEAM, the Federation of European Academies of Medicine, in collaboration with others, in exploring and explaining the issues that will ensure an appropriate European Union (EU) policy framework for health research and innovation. In this article, we present a proposal for an archive of important research conducted in the EU that will act as a resource for illustrating and guiding the development of the necessary regulatory framework.

Growth hormone and melatonin prevent age-related alteration in apoptosis processes in the dentate gyrus of male rats.

It has been suggested that the age-related decrease in the number of neurons in the hippocampus that leads to alterations in brain function, may be associated with an increase in apoptosis due to the reduced secretion of growth hormone (GH) and/or melatonin in old animals. In order to investigate this possibility, male Wistar rats of 22 months of age were divided into three groups. One group remained untreated and acted as the control group. The second was treated with growth hormone (hGH) for 10 weeks (2 mg/kg/d sc) and the third was subjected to melatonin treatment (1 mg/kg/d) in the drinking water for the same time. A group of 2-months-old male rats was used as young controls. All rats were killed by decapitation at more than 24 month of age and dentate gyri of the hippocampi were collected. Aging in the dentate gyrus was associated with an increase in apoptosis promoting markers (Bax, Bad and AIF) and with the reduction of some anti-apoptotic ones (XIAP, NIAP, Mcl-1). Expressions of sirtuin 1 and 2 (SIRT1 and 2) as well as levels of HSP 70 were decreased in the dentate gyrus of old rats. GH treatment was able to reduce the pro/anti-apoptotic ratio to levels observed in young animals and also to increase SIRT2. Melatonin reduced also expression of pro-apoptotic genes and proteins (Bax, Bad and AIF), and increased levels of myeloid cell leukemia-1 proteins and SIRT1. Both treatments were able to reduce apoptosis and to enhance survival markers in this part of the hippocampus.

Effect of estrogens on base excision repair in brain and liver mitochondria of aged female rats.

Changes in the endocrine system have been suggested to act as signaling factors in the regulation of age-related events. Among the different hormones that have been linked to the aging process, estrogens have been widely investigated. They have been associated with inflammatory and oxidative processes and several investigations have established a relationship between the protective effects of estrogens and the mitochondrial function. Mitochondrial DNA is subjected to continuous oxidative attack by free radicals, and the base excision repair (BER) pathway is the main DNA repair route present in mitochondria. We have investigated the effect of estrogen levels on some of the key enzymes of BER in brain and liver mitochondria. In both tissues, depletion of estrogens led to an increased mitochondrial AP endonuclease (mtAPE1) activity, while restoration of estrogen levels by exogenous supplementation resulted in restitution of control APE1 activity only in liver. Moreover, in hepatic mitochondria, changes in estrogen levels affected the processing of oxidative lesions but not deaminations. Our results suggest that changes in mtAPE1 activity are related to specific translocation of the enzyme from the cytosol into the mitochondria probably due to oxidative stress changes as a consequence of changes in estrogen levels.

Higher levels of endogenous estradiol are associated with frailty in postmenopausal women from the toledo study for healthy aging.

BACKGROUND: Adverse effects of higher endogenous estradiol (E2) levels on various clinical outcomes and on determinants of the frailty syndrome have recently been reported. However, there are no data about the potential relationship between E2 and frailty. We aimed to study the association between E2 levels and frailty among older postmenopausal women not taking hormonal therapy. METHODS: We used data from the Toledo Study for Healthy Aging, a Spanish population-based cohort study. Frailty was defined according to Fried's approach. Multivariate odds ratios (OR) and 95% confidence intervals (CI) associated with E2 levels were estimated using polytomous logistic regression. RESULTS: E2 levels decreased significantly with age and educational level, whereas they increased with body mass index, high-sensitivity C-reactive protein (hs-CRP), and impairment in Katz activities of daily living. Higher E2 levels were associated with the prevalence of frailty among women younger than 79 yr, but not in the oldest group (p interaction = 0.047). After adjustment, OR of frailty associated with a 1 sd increase of E2 was 1.51 (95% CI, 1.04-2.20; P = 0.03). We identified an interaction between E2 and hs-CRP on the prevalence of frailty (P value = 0.042). Women with both higher E2 and hs-CRP (defined as values into the upper tertile) had an age-adjusted OR of 4.2 (95% CI, 1.7-10.5; P = 0.002), compared with women with low levels of both E2 and hs-CRP. CONCLUSION: Higher E2 levels were associated with frailty in postmenopausal women. The synergism between higher E2 and hs-CRP levels suggests the existence of physiopathological mechanisms connecting inflammation and estrogen to frailty.

Effect of a combined treatment with growth hormone and melatonin in the cardiological aging on male SAMP8 mice.

The effect of a chronic combined treatment with growth hormone (GH) plus melatonin (Mel) on different age-related processes in cytosolic and nuclear fractions of hearts from SAMP8 mice (2 and 10 months) has been investigated. The parameters studied have been messenger RNA expressions of IL-1, IL-10, NFkBp50, NFkBp52, TNFα, eNOS, iNOS, HO-1, HO-2, BAD, BAX, and Bcl2 and protein expressions of iNOS, eNOS, TNFα, IL-1, IL-10, NFkBp50, NFKbp52, and caspase activity (3 and 9). Our results supported the existence of a proapoptotic and oxidative status together with inflammatory processes in the heart of old mice, with increases of inflammatory cytokines, caspase activity, HO-1, BAX, NFkBp50, and NFkBp52 and decreases of eNOS and Bcl2. Also, we were able to observe the translocation of NFkB to nuclei. The combined treatment was able to partially reduce the incidence of these deleterious changes, showing differences with the separated treatments with GH and Mel as were investigated in previous articles from our group.

Effects of oestrus induction with progestagens or prostaglandin analogues on ovarian and pituitary function in sheep.

The aim of the current study was to determine possible differences in ovarian and pituitary features explaining lower fertility rates in sheep with oestrus induced with intravaginal progestagens or prostaglandin analogues (group FGA and PGF, n=8 in both) when compared to a control group (group C, n=8). The growth profiles and the mean individual sizes of preovulatory follicles were similar between groups; however, the number of preovulatory follicles per ewe and, consequently, the number of ovulations were higher in groups FGA and PGF (2.3±0.3 and 2.0±0.1, respectively) than in group C (1.4±0.1, P<0.05). However, plasma oestradiol concentrations were similar between groups suggesting a defective function in some preovulatory follicles of groups FGA and PGF. In group FGA, the basal LH levels during the follicular phase were lower (0.21±0.0 ng/mL, P<0.005) than in groups C (0.41±0.1 ng/mL) and PGF (0.55±0.1 ng/mL); the onset of preovulatory discharge being later (21.0±2.3h vs. 12.8±1.5 in C and 14.5±1.5 in PGF; P<0.05 for both). Finally, luteal activity was also found to be affected in group FGA; the rate of progesterone secretion per total luteal tissue was lower (range: 0.46-0.65 ng/mL/cm(2)) than in ewes treated with cloprostenol (2.1-3.3 ng/mL/cm(2)) and control sheep (2.0-3.4 ng/mL/cm(2)).

Beneficial effects of melatonin on cardiological alterations in a murine model of accelerated aging.

This study investigated the effect of aging-related parameters such as inflammation, oxidative stress and cell death in the heart in an animal model of accelerated senescence and analyzed the effects of chronic administration of melatonin on these markers. Thirty male mice of senescence-accelerated prone (SAMP8) and 30 senescence-accelerated-resistant mice (SAMR1) at 2 and 10 months of age were used. Animals were divided into eight experimental groups, four from each strain: two young control groups, two old untreated control groups, and four melatonin-treated groups. Melatonin was provided at two different dosages (1 and 10 mg/kg/day) in the drinking water. After 30 days of treatment, the expression of inflammatory mediators (tumor necrosis factor-alpha, interleukin 1 and 10, NFkBp50 and NFkBp52), apoptosis markers (BAD, BAX and Bcl2) and parameters related to oxidative stress (heme oxygenases 1 and 2, endothelial and inducible nitric oxide synthases) were determined in the heart by real-time reverse transcription polymerase chain reaction (RT-PCR). Inflammation, as well as, oxidative stress and apoptosis markers was increased in old SAMP8 males, when compared to its young controls. SAMR1 mice showed significantly lower basal levels of the measured parameters and smaller increases with age or no increases at all. After treatment with melatonin, these age-altered parameters were partially reversed, especially in SAMP8 mice. The results suggest that oxidative stress and inflammation increase with aging and that chronic treatment with melatonin, a potent antioxidant, reduces these parameters. The effects were more marked in the SAMP8 animals.

Growth variation, final height and secular trend. Proceedings of the 17th Aschauer Soiree, 7th November 2009.

Growth and body height have always been topics interesting to the public. In particular, the stupendous increase of some 15-19cm in final adult height during the last 150 years in most European countries (the "secular trend"), the concomitant changes in body and head proportions, the tendency towards early onset of sexual maturation, the changes in the age when final height is being reached, and the very recent trend in body mass index, have generated much scientific literature. The marked plasticity of growth in height and weight over time causes problems. Child growth references differ between nations, they tend to quickly become out of date, and raise a number of questions regarding fitting methods, effects caused by selective drop-out, etc. New findings contradict common beliefs about the primary importance of nutritional and health related factors for secular changes in growth. There appears to be a broad age span from mid-childhood to early adolescence that is characterised by a peculiar insusceptibility. Environmental factors that are known to influence growth during this age span appear to have only little or no impact on final height. Major re-arrangements in height occur at an age when puberty has almost been completed and final height has almost been reached, implying that factors, which drive the secular trend in height, are limited to early childhood and late adolescence.

Cardiological aging in SAM model: effect of chronic treatment with growth hormone.

The purpose of this study was to investigate the effect of aging on different parameters related to inflammation, oxidative stress and apoptosis in hearts from two types of male mice models: senescence-accelerated mice (SAM-P8) and senescence-accelerated-resistant (SAM-R1), and the influence of chronic administration of Growth Hormone (GH) on old SAM-P8 mice. Forty male mice were used. Animals were divided into five experimental groups: two 10 month old untreated groups (SAM-P8/SAM-R1), two 2 month old young groups (SAM-P8/SAM-R1) and one 10 month old group (SAM-P8) treated with GH for 30 days. The expression of tumor necrosis factor-alpha, interleukin 1, interleukin 10, heme oxygenases 1 and 2, endothelial and inducible nitric oxide synthases, NFkB, Bad, Bax and Bcl-2 were determined by real-time reverse transcription polymerase chain reaction (RT-PCR). Results were submitted to a two way ANOVA statistical evaluation using the Statgraphics program. Inflammation, as well as, oxidative stress and apoptosis markers were increased in the heart of old SAM-P8 males, as compared to young controls and this situation was not observed in the old SAM-R1 mice. Exogenous GH administration reverted the effect of aging in the described parameters of old SAM-P8 mice. Our results suggest that inflammation, apoptosis and oxidative stress could play an important role in the observed cardiovascular alterations related to aging of SAM-P8 mice and that GH may play a potential protective effect on the cardiovascular system of these animals.

Hormonal regulation of pro-inflammatory and lipid peroxidation processes in liver of old ovariectomized female rats.

There is now a large body of evidence suggesting that the decline in ovarian function with menopause is associated with spontaneous increases in pro-inflammatory cytokines. On the other hand, oxidative stress has been implicated in the pathogenesis of several alterations due to menopause, and can arise through the increased production of lipid peroxides (LPO) and/or a deficiency of antioxidant defense. The aim of the present study was to investigate the effect of aging and ovariectomy on various physiological parameters related to inflammation and oxidative stress in livers obtained from old female rats and the influence of chronic exogenous administration of estrogens, phytoestrogens and growth hormone on these. Thirty-six female Wistar rats of 22 months of age were used in the present study. Twelve of them remained intact, and the other 24 had been ovariectomized at 12 months of age. Intact animals were divided into two groups and treated for 10 weeks with GH or saline, and ovariectomized animals were divided into four groups and treated for the same time with GH, estrogens, phytoestrogens or saline. A group of 2 month old intact female rats was used as young control. Protein expression of iNOS, HO-1, IL-6, TNFalpha, and IL-1beta were determined by Western blot analysis. The levels of NO( x ), LPO, TNFalpha, IL-1beta, IL-6 and IL-10 were determined in different fractions of the liver. Levels of LPO in the liver homogenates as well as iNOS protein expression and NO( x ) levels were increased in old rats as compared to young animals; this effect was more evident in ovariectomized animals. Pro-inflammatory cytokines TNF-alpha, IL-1beta and IL-6 were significantly increased and anti-inflammatory IL-10 decreased during ageing and after ovariectomy. Aging also significantly increased expression of HO-1 protein and ovariectomized rats showed an additional increase. Hormonal administration to the ovariectomized groups decreased NO( x ), LPO levels and pro-inflammatory cytokines as compared with untreated rats. Significant rise in IL-10 and reductions in the iNOS, IL-6, TNFalpha and IL-1beta proteins expression were also found. Oxidative stress and inflammation induced during aging in the liver are more marked in castrated than in intact old females. Administration of the different hormonal replacement therapies was able to inhibit the induction of pro-inflammatory cytokines and iNOS, decreased the levels of oxidative stress markers and had therapeutic potential in the prevention of liver injury.

Ovarian follicular dynamics and plasma steroid concentrations are not significantly different in ewes given intravaginal sponges containing either 20 or 40 mg of fluorogestone acetate.

Although various progestagens are often used to induce and synchronize estrus and ovulation in ruminants, concerns regarding residues are the impetus to develop alternative approaches, including reduced doses of progestagens. Therefore, the objective was to determine whether ovarian function was affected by halving the dose of fluorogestone acetate in intravaginal sponges for synchronizing ovulation in sheep during the physiologic breeding season. Twenty Manchega ewes, 4-6-year-old, were randomly allocated to receive an intravaginal sponge containing either 20mg (P20, n=10) or 40 mg of fluorogestone acetate (P40, n=10). Cloprostenol (125 microg) was given at sponge insertion, and all sponges were removed after 6d. Ovarian follicular dynamics (monitored by daily ultrasonography) and other aspects of ovarian function did not differ significantly between the two groups. Ovulatory follicles (OF) grew at a similar growth rate (r=0.62; P<0.001), with comparable initial and maximum diameters (4.2+/-0.4 to 6.0+/-0.3mm in P20 vs. 4.6+/-0.6 to 5.7+/-0.2 mm in P40, mean+/-S.E.M.). Plasma estradiol concentrations (determined once daily) increased linearly during the 72 h interval after sponge removal (1.3+/-0.1 to 3.3+/-0.1 pg/mL for P20, P<0.005 and 1.4+/-0.1 to 3.1+/-0.2 pg/mL for P40, P<0.005). Ten days after sponge removal, ovulation rates (1.2+/-0.2 for P20 and 1.4+/-0.3 for P40), and plasma progesterone concentrations (3.8+/-0.35 ng/mL for P20 and 3.9+/-0.38 ng/mL for P40) were similar. In conclusion, reducing the dose of fluorogestone acetate from 40 to 20mg did not affect significantly ovarian follicular dynamics or other aspects of ovarian function.

The role of taste and appetite regulation in the understanding of overweight and obesity. Proceedings of the 16th Aschauer Soiree, 26th April 2008.

Overweight and obesity have developed into major illnesses in most Western societies and significantly contribute to the financial burden of modern public health systems. Almost daily, new therapeutic proposals are published in the lay press, and also the scientific literature has increased dramatically in recent years. E.g., when searching MEDLINE (1966 - May 2008 (1)), the key word "obesity" meanwhile appears in more than 108,000 articles. Primary focus however, is put upon aspects of treatment, neglecting the role of taste and appetite regulation. Combining keywords like "obesity + treatment" results in over 50.000 citations, "obesity + diet" in over 23.000, "obesity + energy + expenditure" in over 13.000 citations (even "obesity + gastric + bypass" still evoke 2.600 citations), whereas "obesity + appetite + regulation" result in some 3.000, "obesity + NPY" - neuropeptid Y being one of the major chemical stimulators of appetite - evoke some 500 and "obesity + Arc + nucleus" - the arcuate nucleus being the anatomical centre of appetite regulation - no more than 370 scientific publications. The apparent scarcity of literature about taste and appetite regulation and the amazing lack of interest in neuronal information processing in overweight and obesity, has prompted the authors to brainstorm new aspects of the world-wide derailment of weight control.

Preovulatory follicle development in goats following oestrous synchronization with progestagens or prostaglandins.

The study reports on differences in the dynamics of growth and functionality of preovulatory follicles in response to oestrous synchronization, either by the administration of two doses of prostaglandin or by an intravaginal progestagen sponge, in goats. The progestagen-treated group (n = 8) showed more follicles of preovulatory size (> or =5.5 mm) than the cloprostenol group (n = 8) during the follicular phase (4.5 +/- 0.6 vs 1.9 +/- 0.2, p < 0.01). The diameters of the largest follicles (LF1, LF2 and LF3) were also larger in the progestagen group (LF1, 7.8 +/- 0.3 vs 7.0 +/- 0.2 mm, p < 0.05; LF2, 6.7 +/- 0.2 vs 5.6 +/- 0.2 mm, p < 0.01; LF3, 5.5 +/- 0.3 vs 4.2 +/- 0.2 mm, p < 0.01). The study of the preovulatory follicles showed that 27.2% (3/11) of the follicles were in the static phase in the cloprostenol group, whilst 71.4% (10/14) were static in progestagen group (p < 0.05). Higher plasma oestradiol levels were recorded in the progestagen-treated goats during the 48 h prior to cloprostenol injection or progestagen withdrawal (4.2 +/- 0.4 vs 3.0 +/- 0.2 pg/ml, p < 0.05). In conclusion, goats with oestrus synchronized by progestagen showed a higher number of preovulatory-sized follicles, but a decreased oestradiol secretion when compared with does with oestrus synchronized by using prostaglandin analogues. These would support the development of alternative protocols for assisted reproduction.

Effect of chronic treatments with GH, melatonin, estrogens, and phytoestrogens on oxidative stress parameters in liver from aged female rats.

The aging theory postulates that this process may be due to the accumulation of oxidative damage in cells and molecules. The present study has investigated the effect of castration in old female rats on various parameters related to the antioxidant properties of several cellular fractions obtained from the liver, and the influence of several chronic treatments on it, both in intact and castrated animals. Sixty-one 22-month-old Wistar female rats, were used. About 21 intact animals were divided into three groups and treated for 10 weeks with GH, melatonin or saline, and 40 ovariectomized (at 12 months of age) animals were divided into five groups and treated for the same time with GH, melatonin, estrogens (Eos), phytoestrogens (Phyt) or saline. All animals were sacrificed at 24 months of age by decapitation. The activity of glutathione peroxidase (GPx) in cytosolic fraction, glutathione-S-transferase (GST) in cytosol and microsomal fractions, and the levels of nitric oxide (NO) and cytochrome C in mitochondrial and cytosol fractions of liver were determined. A decrease in GST activity was detected in cytosol and in the microsomal fraction in ovariectomized animals as compared to intact rats. The activity of GPx was also decreased in ovariectomized as compared with the intact group. NO level was increased and cytochrome C decreased in the mitochondrial fraction of the liver in ovariectomized females as compared with the intact group, respectively. No significant changes after melatonin or GH treatments were found in GPx, GST activity and NO level in mitochondrial fraction in the intact group. Administration of GH, melatonin, Eos and Phyt in the ovariectomized groups significantly increased the GPx, and GST activity in the cytosol and microsomal fraction and decreased the level of NO in the mitochondrial fraction as compared with the untreated rats. A significant increase in the level of cytochrome C in the mitochondrial fraction and a decrease in the cytosol fraction were also found with all treatments. The administration of GH, melatonin, Eos and Phyt to castrated females seem to reduce oxidative changes in the liver from old ovariectomized rats.

Effect of exogenous administration of melatonin and growth hormone on pro-antioxidant functions of the liver in aging male rats.

Aging is accompanied by changes in the morphology and physiology of organs and tissues, such as the liver. This process might be due to the accumulation of oxidative damage induced by reactive oxygen (ROS) and reactive nitrogen species (RNS). Hepatocytes are very rich in mitochondria and have a high respiratory rate, so they are exposed to large amounts of ROS and permanent oxidative stress. Twenty-four male Wistar rats of 22 months of age were divided into three groups. One group remained untreated and acted as the control group. The second was treated with growth hormone (GH) (2 mg/kg/d sc) and the third was submitted to treatment wit 1 mg/kg/d of melatonin in the drinking water. A group of 2-months-old male rats was used as young controls. After 10 wk of treatment the rats were killed by decapitation, and the liver was dissected and homogenized. Mitochondrial, cytosolic and microsomal fractions were obtained and cytochrome C, glutathione peroxidase, s-transferase and nitric oxide (NO) were measured. Aging induced a significant increase in mitochondrial nitric oxide. An increase in cytochrome C in the cytosolic fraction and a reduction in the mitochondrial fraction with age was also observed. Both GH and melatonin treatments significantly reduced the enhanced measures and increased the reduced values. A reduction in glutathione peroxidase and glutathione S-transferase was found in old control rats when compared with the group of young animals. Treatment for 2.5 months of old rats with GH and melatonin were able to increase the enzymes reaching values similar to those found in young animals. In conclusion, GH and melatonin treatment seems to have beneficial effects against age-induced damage in the liver.

Effects of breed on follicular dynamics and oestradiol secretion during the follicular phase in sheep.

Current study evaluates effects from breed and management background on follicular dynamics and endocrine output during the follicular phase of sheep oestrous cycle. Follicular phases were synchronized with three cloprostenol doses, 10 days apart, in three groups of 10 females of different non-prolific Spanish breeds (Manchega, Rubia del Molar and Negra de Colmenar). Development of all follicles reaching antral diameters >or=2 mm was assessed by daily transrectal ultrasonographies, whereas follicular function was evaluated by measurement of plasma oestradiol concentrations. All the ovulatory follicles were present at induced luteolysis in Manchega sheep, while a 93.7% were detected in Rubia del Molar and Negra de Colmenar ewes. The mean size of these ovulatory follicles was similar between breeds at 0 h, but their growth rates were higher in Manchega ewes, reaching a larger size at oestrous detection than in Negra de Colmenar and Rubia del Molar sheep (p < 0.05). Conversely, the oestradiol levels increased with time in Rubia del Molar and Negra de Colmenar (p < 0.05); whilst remained stable in Manchega females. However, the patterns of follicular turnover were similar between breeds. These results indicate that, though differences in follicular size and size distribution, patterns of follicular turnover in sheep are affected neither by the breed nor by the background of management and selection.

Effects of progestagens on follicular growth and oocyte developmental competence in FSH-treated ewes.

Previous research has reported evidence for negative effects of progestagens on follicular growth and oocyte competence. In the present study, negative effects of progestagens on follicular growth and oocyte developmental competence were assessed. During the breeding season, 20 Sarda ewes were treated with two doses of cloprostenol, 10 days apart, to assure the presence of a corpus luteum (CL). On day 5 after the second cloprostenol dose, 10 ewes were treated with a progestagen sponge while 10 females remained untreated. Starting on day 7 after the second cloprostenol dose, all the ewes were treated with 6 equal doses of 24 I.U. of FSH (Ovagen, ICP, NZ), every 12h. The number of follicles > or =2mm in diameter increased (P<0.0005) in all the ewes from 24 h before to 60 h after the first FSH dose (from 12.8+/-1.1 to 23.4+/-1.3 in treated and from 12+/-0.6 to 22+/-1.2 in untreated ewes, n.s.). There were no significant differences in follicle dynamics between groups, but concentrations of estradiol in control ewes were higher than in the progestagen group (P<0.05). Twelve hours after the last FSH dose, oocytes were collected by ovum pick-up. Recovery rates were lower for progestagen-treated ewes (71.1 versus 83%; P<0.001). After IVP procedure, cleavage rate was also lower in the progestagen group (39.1 versus 82.6%; P<0.001). Furthermore, blastocysts output revealed that oocyte developmental competence was lower in progestagen group (17.3 versus 30.4%; P=0.245), although differences were not significant. These results suggest deleterious effects from progestagen on oocyte developmental competence and set the basis for new protocols for in vitro embryo production.