Effect of the presence of corpus luteum on the ovary and the new oocyte recovery method on the oocyte recovery rate and meiotic competence of ovine oocytes

This study was designed to identify the effects of the presence of corpus luteum (CL) on the meiotic competence of the ewe oocytes. In addition, due to the pivotal roles of oocyte recovery methods on efficiency of in vitro maturation and in vitro fertilization, this study considered the new oocyte recovery method “Oocyte Recovery with Centrifuge” (ORC); Ovine ovaries were obtained from local abattoir at Karaj, Iran. The ovaries were placed in 0.9% saline which was supplemented with penicillin/streptomycin in thermo flask at 37°C and transported to the laboratory within 1 to 2 h of slaughter. The ovaries were assigned to two groups; group-1 with functional CL and group-2 without CL. The oocytes were recovered by means of aspiration pump or ORC. After oocyte recovery, they were cultured in TCM-199 for 24 h. The mean number of oocyte recovery per ovary in group 1 ovaries (1.8 ± 0.01 via aspiration and 3.84 ± 0.05 via ORC) was lower (P<0.05) than group 2 ovaries (2.2 ± 0.00 via aspiration and 5.43 ± 0.01 via ORC). There were no significant differences (P>0.05) between percentage of nuclear maturation in oocytes which were recovered from group 2 ovaries via aspiration and ORC (75.20 ± 0.00 vs. 74.95 ± 0.00, metaphase II; M-II) method. The nuclear maturation in oocytes which were obtained via ORC from group 2 ovaries was higher (P<0.05) than group 1 ovaries (74.95 ± 0.00 vs. 60.07 ± 0.00b). Nuclear maturation for oocytes obtained via ORC from group 1 ovaries was (P<0.05) lower than oocytes obtained via aspiration (60.07 ± 0.00 vs. 74.70 ± 0.00). Result of the present study showed that the presence of CL on ovaries lead to decrease in the quality and the quantity of oocytes. ORC method increased the quantity and quality of recovered oocytes.Key words: Aspiration, corpus luteum, in vitro maturation (IVM), oocyte recovery

Retinoic acid effects on nuclear maturation of bovine oocytes in vitro

In the present study, the effect of all-trans retinoic acid (t-RA) administration during in vitro maturation (IVM) on bovine oocytes maturation was determined. Concentrations of t-RA (RA; 0, 0.25, 0.5 and 1 μM) and 0.1% ethanol (vehicle) were included in the maturation medium. Ovaries collected from the local abattoir were transported to the laboratory in in 0.9% NaCl with 100 IU/ml penicillin and 100 in vitro maturation (IVM) on bovine oocytes maturation was determined. Concentrations of t-RA (RA; 0, 0.25, 0.5 and 1 μM) and 0.1% ethanol (vehicle) were included in the maturation medium. Ovaries collected from the local abattoir were transported to the laboratory in in 0.9% NaCl with 100 IU/ml penicillin and 100  in vitro maturation (IVM) on bovine oocytes maturation was determined. Concentrations of t-RA (RA; 0, 0.25, 0.5 and 1 μM) and 0.1% ethanol (vehicle) were included in the maturation medium. Ovaries collected from the local abattoir were transported to the laboratory in in 0.9% NaCl with 100 IU/ml penicillin and 100 g/ml streptomycin at 30 - 35°C within 1-2 h after collection. The oocytes of antral follicles, 2 to 8 mm in diameter, were recovered by aspiration. After preliminary evaluation, the oocytes were selected and washed four times in HEPES-TCM 199 supplemented with 2% FBS, 0.2 mM sodium pyruvate, 100 IU/ml penicillin and 100 g/ml streptomycin. Then 10 cumulus-oocyte complexes (COCs) were subjected to each droplet of maturation medium and incubated at 38.5°C, 5% CO2 and 95% humidity for 24 h. Maturation medium was bicarbonate-buffered TCM199 supplemented with 10% FBS, 0.2 mM sodium pyruvate, 5 μg/ml bovine FSH, 0.01 IU/ml bovine LH, 100 IU/ml penicillin and 100 g/ml streptomycin. Results show different concentrations of t-RA have no effect on cumulus expansion. The rate of oocytes developing to the MII stage compared to control, vehicle, and 0.25 μM groups was significantly increased with 1 μM t-RA treatment (

Lecithin nanoparticles enhance the cryosurvival of caprine sperm

P. 38-44This study was designed to compare the effects on goat spermatozoa cryosurvival of nano-lecithin-based (NL), lecithin-based (L) and egg yolk-based (EY) extenders. Ejaculates were collected from four fertile goats using artificial vagina and diluted with nine extenders. NL and L were tested at concentrations 1%, 2%, 3% and 4% (w/v), versus 15% (v/v) egg yolk-based extender. Overall, sperm quality (higher motility, viability and HOST, and lower apoptosis) was higher for NL than for L treatments (P < 0.05 for most cases, except for 1%). NL at 1% and especially at 4% showed lower motility and viability than 2% or 3% NL. NL at 2% achieved a better performance (P < 0.05) than EY for VCL (131.5 ± 1.3 vs. 120.3 ± 1.9 μm/s), VSL (43.9 ± 1.5 vs. 35.8 ± 1.4 μm/s), LIN (35.7 ± 0.6 vs. 29.3 ± 0.8%), WOB (47.0 ± 0.5 vs. 43.9 ± 0.9%) and viability (66.4 ± 1.7 vs. 52.7 ± 1.9%). Late apoptotic spermatozoa were also lower in 2% NL compared to EY (16.0 ± 0.5 vs. 26.3 ± 1.1%, P < 0.001). EY and 2% NL were compared in an IVF trial, with no significant differences in cleavage (68.8 vs. 70.8%) or blastocyst ratios (21.3 vs. 20.8%). In conclusion, using 2% nanolecithin in semen dilution could improve sperm cryosurvival of goat.S

Endocrine disruptors and abnormalities of pubertal development

Onset and development of puberty is regulated by the neuroendocrine system. Population-based studies worldwide have observed secular trends towards earlier puberty development. These changes are apparently caused by environmental factors such as improved socio-economic status, improved health care and nutrition. However, they may also partly result from endocrine-disrupting chemicals in the environment. Epidemiological studies have investigated the relationship between pubertal development and exposure to endocrine-disrupting chemicals (polychlorinated biphenyls, polybrominated biphenyls, 1,1,1-trichloro-2,2-bis(p-chlorophenyl)ethane, phthalate esters, furans and the pesticide endosulfan). Associations with both perinatal and postnatal exposure have been reported. Studies in experimental animals support some of these findings and point to differential endocrine regulatory mechanisms linked to pubertal development acting in the perinatal and the pre-pubertal period. Pubertal development is naturally associated with growth and body composition. There is increasing evidence for a link between prenatal development and pubertal onset. In girls born small for gestational age (SGA), pubertal onset and age at menarche often are advanced, especially if there has been an extensive catch-up growth during the first months of life. In utero growth retardation may have multiple causes including exposure to xenobiotic substances as was suggested for some endocrine-disrupting chemicals. An abnormal perinatal environment of children born SGA may alter the endocrine status and the sensitivity of the receptors for endocrine and metabolic signalling that may have effects on maturation of brain and gonads. However, the causal pathways and the molecular mechanisms that may link the pubertal growth pattern of children born SGA, pubertal development and endocrine-disrupting chemicals need further study

Hormonal regulation of puberty onset in female rats: is leptin a missing link?

Numerous factors in the hypothalamus-pituitary-gonadal axis (HPG) are involved in the timing of puberty onset.Leptin signals the nutritional status and the presence of an adequate amount of loaded adipose tissue, as a long-term resource of energy, to the brain which in response switches on the reproduction process (i.e. puberty onset).The growth hormone-insulin-like growth factor I (GH/IGF-I) axis is also thought to be involved in the timing of puberty onset. The aim of this thesis was to test the hypothesis that leptin may directly or indirectly via theGH/IGFI-I axistrigger the onset of puberty onset in female rats. First, we carried out a series of descriptive and basic studies.Time of puberty onset was monitored by scoring the moment of vaginal opening (VO).The data showed a strong positive correlation between body fat and leptin levels, and showed leptin is increasing over the prepubertal period. Then, we aimed to find out the effects of food-restriction (FR) on puberty onset, body fat, plasma leptin levels and body temperature. FR postponed time of VO. Body fat, plasma leptin levels and body temperature in the FR rats were significantly lower than the controls throughout the experiment. To find outif leptin is the signal initiating the onset of puberty, we used FR rats as a model for delayed puberty onset, and centrally (icv) or peripherally (sc) administered leptin, or we centrally (icv) immunoneutralized leptin. Central leptin infusion not only restored the delay in puberty onset caused by food restriction but also advanced sexual maturation in normally fed animals.Like central infusion, also peripheral leptin infusion restored puberty onset in FR animals. So, we showed an advancing effect of both centrally and peripherally infused leptin on puberty onset in prepubertal female rats.The central immunoneutralization of leptin on the other hand, postponed puberty onset. We therefore conclude that leptin is one of the crucial factors triggering puberty onset in female rats.In the FR model system we then centrally (icv) infused GH.The infusion of GH postponed puberty onset in normally fed rats, but advanced puberty onset in FR animals. Also,the plasma leptin levels in the GH-infused animals were significantly higher than their controls and increased as GH infusion proceeded.Central infusion of GH antiserum (AS) advanced puberty onset in the pair-fed animals but not in the ad lib-fed animals. Also the central infusion of somatostatin worked likewise in both groups. These findings match with the results of the central infusion of GH which also advanced puberty onset in FR animals only.Finally, we centrally (icv) infused IGF-I in FR rats. Central infusion of IGF-I significantly postponed puberty onset in ad libitum fed animals.Immunoneutralization of endogenous IGF-I enhanced prevailing plasma leptin levels, but there was no effect on the timing of puberty onset. Centrally-present endogenous IGF-I does not appear to be involved in puberty onset although it seems to inhibit leptin secretion.In conclusion, our findings provide further evidence that leptin, a hormone produced mainly by adipose tissue, is an important and crucial signal between the bodily nutritional status and the brain (i.e. the hypothalamus) to trigger puberty onset. Furthermore, we suggest that there is a functional interaction between growth hormone (GH) and leptin to initiate puberty in female rats

Ascorbic acid effects on in vitro maturation of mouse oocyte with or without cumulus cell

Ascorbic acid has long been associated with fertility. This study was designed to determine the effects of ascorbic acid on in vitro maturation of mouse oocyte with or without cumulus cells. In this study, 508 denuded oocytes (DOs) and 527 cumulus-oocyte complexes (COCs) from mice stimulated with pregnant mare&apos;s serum gonadotrophin (PMSG) were incubated for 24 h in medium containing 0, 80, 250 and 750 µM/ml of ascorbic acid prior to in vitro maturation. Maturation rate was compared. A significant decrease in the maturation rate was observed only when the DOs and COCs were exposed to 750 µM/ml of ascorbic acid (P &lt; 0.05). The maturation rate in COCs was significantly higher than DOs in all groups (P &lt; 0.05). These results indicate that exposure ascorbic acid promotes the development of mouse DOs and COCs from germinal vesicle breakdown (GVBD) to metaphase II (MII) and prevents cumulus cell degeneration at certain levels, especially 250 µM/ml of ascorbic acid (P &lt; 0.05). However, further studies on the potential effects of different concentrations of ascorbic acid on oocyte maturation are needed

Central Application of IGF-1 Postpones Time of Vaginal Opening in Normally Fed, but Not in Food-Restricted Rats

Background/Aims: Central but also peripheral IGF-1 is suggested to play a role in the initiation of puberty as it directly affects GnRH synthesis and release. A possible intermediate in the effects of IGF-1 on puberty might be the adiposity-signaling hormone leptin, whose plasma levels are decreased in food-restricted (FR) rats. Methods: IGF-1 was chronically centrally infused in 23-day-old prepubertal female rats which were either normally fed or 30% FR, and the effects on time of vaginal opening (VO) and plasma leptin levels were monitored. Results: FR treatment postponed time of VO and decreased plasma leptin levels. In normally fed rats centrally infused with IGF-1, time of VO was found to be postponed to the same extent as FR treatment did. The IGF-1 infusion did not affect plasma leptin levels in normally fed animals but increased leptin levels in the FR group compared to controls. Daily food intake was equal between all groups but body weight course was lower in FR rats. IGF-1 treatment did not significantly affect food intake or body weight course. Conclusion: FR treatment delays the moment of vaginal opening to the same extent as observed in normally fed rats that were centrally infused with IGF-1

Chronic leptin infusion advances, and immunoneutralization of leptin postpones puberty onset in normally fed and feed restricted female rats

Does leptin play a vital role in initiating puberty in female rats and can it overrule a nutrionally imposed (i.e. a 30% feed restriction, FR) delay in puberty onset? Prepubertal female rats were chronically infused for 14 days with leptin (icv or sc) or leptin-antiserum (icv) while puberty onset was monitored by means of scoring the moment of vaginal opening (VO). Median VO age was higher (35 days versus 27 days) in FR animals but leptin levels at VO were significantly decreased (1.44 ± 0.17 ng/ml versus 2.79 ± 0.31 ng/ml). Centrally (icv) and peripherally (sc) infused leptin (1 ¿g/day) advanced VO age compared to FR controls (30 days versus 35 days and 31 days versus 41 days, respectively). Congruently, centrally (icv) administered leptin-antiserum (0.6 ¿g/day) delayed puberty onset. In normally fed rats median VO age was only marginally advanced (26 days versus 27 days) but only if leptin was applied centrally. The effects of FR on puberty onset are counteracted or even normalized by the infusion of leptin, whereas immunoneutralization of central leptin postpones puberty onset. We therefore conclude that central leptin is crucial for initiating puberty in female rats