Early effects of equine FSH (eFSH) treatment on hormonal and reproductive parameters in mares intended to carry their own pregnancy

Superovulatory treatment may potentially increase the embryo recovery rate and the per-cycle pregnancy rate in normal or subfertile mares that are managed properly. However, some studies suggest a possible negative effect of superovulatory treatment on ovarian follicular maturation and embryo viability. Objectives of the present study were to investigate the early effects of eFSH treatment in reproductively normal mares in terms of: folliculogenesis, pregnancy rate, early embryonic development, reproductive tract parameters (tone and edema), and serum estradiol-17β and progesterone concentrations. Reproductively sound mares (n = 26) were evaluated daily by transrectal palpation and ultrasonography. Five days after spontaneous ovulation, mares were randomly assigned to one of two treatment groups. In the eFSH group, mares (n = 16 estrous cycles) were administered eFSH twice daily; beginning when a follicle ≥20 mm was detected, and continuing until at least one follicle reached a diameter of ≥35 mm. PGF2α was administered 2 days following initiation of eFSH therapy, and hCG was administered approximately 36 h after cessation of eFSH therapy. In the control group, mares (n = 26 estrous cycles) were administered PGF2α 7 days after spontaneous ovulation, and hCG when a follicle ≥35 mm was detected. All mares were bred with fresh semen, monitored for ovulation (Day 0), and evaluated for pregnancy on Days 11–16. Serum estradiol-17β and progesterone concentrations were analyzed using radioimmunoassay on the Day of hCG administration, and Days 8, 11 and 16. Mares treated with eFSH had more follicles ≥30 mm at the time of hCG administration (2.6 ± 0.4 compared with 1.1 ± 0.1; P < 0.01), and more ovulations (2.3 ± 0.5 compared with 1.1 ± 0.3; P < 0.01). However, pregnancy rates were not significantly different between groups (50%; 8/16 compared with 62%; 16/26). Mean overall daily growth rate of embryonic vesicles from Day 11 to 16 was not statistically different between the two groups (3.3 ± 0.3 compared with 3.7 ± 0.1 mm/day) (P = 0.2); however, was more variable (P < 0.01) in the eFSH group (95%CI: 2.6–3.8 mm/day) than in the control group (95%CI: 3.5–3.9 mm/day). Administration of eFSH modified the reproductive tract variables and serum concentrations of progesterone and estradiol-17β on the days that oocyte maturation, fertilization, and early embryonic development are expected to occur. These alterations may be related to the greater incidence of non-ovulatory follicles (25% compared with 0%), fewer embryos per ovulation rate (0.3 ± 0.1 compared with 0.6 ± 0.1), and the lesser than expected pregnancy rates in the eFSH-treated mares.     

Circulating anti-heat-shock-protein antibodies in normal pregnancy and preeclampsia

It has been previously reported that circulating anti-heat-shock-protein (Hsp) antibody levels are elevated in cardiovascular disorders. The aim of the present study was to determine circulating antihuman Hsp60, antimycobacterial Hsp65, and antihuman Hsp70 antibody levels in healthy pregnant women and preeclamptic patients and to investigate their relationship to the clinical characteristics of the study subjects, as well as to the markers of inflammation (C-reactive protein (CRP)), endothelial activation (von Willebrand factor antigen), or endothelial injury (fibronectin), oxidative stress (malondialdehyde) and to serum Hsp70 levels. Ninety-three preeclamptic patients and 127 normotensive healthy pregnant women were involved in this case control study. Serum anti-Hsp60, anti-Hsp65, anti-Hsp70, and Hsp70 levels were measured with enzyme-linked immunosorbent assay (ELISA). Serum CRP levels were determined by an autoanalyzer using the manufacturer’s kit. Plasma von Willebrand factor antigen levels were quantified by ELISA, while plasma fibronectin concentration by nephelometry. Plasma malondialdehyde levels were measured by the thiobarbituric-acid-based colorimetric assay. For statistical analyses, nonparametric methods were applied. Anti-Hsp60, anti-Hsp65, and anti-Hsp70 antibodies were detected in all of our serum samples. There were no significant differences in serum anti-Hsp60, anti-Hsp65, and anti-Hsp70 antibody levels between the control and preeclamptic groups. Serum levels of Hsp70 and CRP, as well as plasma levels of VWF antigen, fibronectin, and malondialdehyde, were significantly higher in preeclamptic patients than in normotensive healthy pregnant women. Serum anti-Hsp60 antibody levels showed significant correlations with serum anti-Hsp65 antibody levels both in the control and the preeclamptic groups (Spearman R = 0.55 and 0.59; p < 0.001, respectively). However, no other relationship was found between clinical features (maternal age, smoking status, parity, body mass index, gestational age at blood draw, systolic and diastolic blood pressure, gestational age at delivery, and fetal birth weight) and measured laboratory parameters of the study subjects and serum anti-Hsp antibody levels in either study group. In conclusion, anti-Hsp60 and anti-Hsp70 antibodies as naturally occurring autoantibodies are present in the peripheral circulation of healthy pregnant women. Nevertheless, humoral immunity against heat shock proteins was not associated with preeclampsia. Further studies are warranted to explore the role of heat shock proteins and immune reactivity to them in the immunobiology of normal pregnancy and preeclampsia.     

Applying variations of the Ovsynch protocol at the middle of the estrus cycle on reproductive performance of lactating dairy cows during summer and winter

Two modifications of the Ovsynch protocol, GnRH + TAI after PGF_2α 48 h (CO-48) or 72 h (CO-72), were compared with the original protocol (OVS: GnRH–7 d–PGF_2α–2 d–GnRH–16 h–TAI) to study their effects on reproductive performance in 785 lactating dairy cows (Holstein Friesian, Bos Taurus). Results showed that more cows (P < 0.001) returned to estrus within a week after TAI with CO-48 treatment compared with that in OVS and CO-72 treatments. Pregnancy rates were greater (P < 0.001) for the CO-72 cows than those for both OVS and CO-48 treatments and for primiparous cows compared with multiparous cows. Moreover, pregnancy rates were lower in summer compared with those in winter. Pregnancy losses for cows in both CO-48 and CO-72 were greater (P < 0.05) than that for cows in OVS treatment. Pregnancy losses were greater in summer (P < 0.001) than in winter and for multiparous cows (P < 0.001) than for primiparous cows. In conclusion, primiparous and winter-bred cows had greater pregnancy rates and fewer pregnancy losses than those of multiparous cows and summer-bred cows, respectively. Because of the presence of significant treatment, parity, and season interactions, TAI with ovulation synchronization protocols should be tailored according to the season and parity. CO-72 is recommended for primiparous cows but not for multiparous cows, and CO-48 is not recommended for synchronization. Furthermore, cows that exhibited estrus at any time were inseminated to improve pregnancy rates in ovulation synchronization protocols.     

Detection and identification of plasma progesterone metabolites in the female Florida manatee (Trichechus manatus latirostris) using GC/MS/MS

Florida manatees (Trichechus manatus latirostris) have relatively low peripheral concentrations of progesterone (P4). The objective of this study was to determine if these relatively low P4 concentrations are associated with a high ratio of progestin metabolites and to document metabolite concentrations from individual blood samples obtained from manatees during diestrus or pregnancy. Metabolites known to exist in elephants—terrestrial manatee relatives—were targeted. These included 5α-reduced progestins (5α-pregnane-3,20-dione [5α-DHP] and 3α-hydroxy-5α-pregnan-20-one [5α-P3-OH]) and 17α-hydroxyprogesterone (17α-OHP), which occurs in Asian elephants. An additional, inactive metabolite, 20α-hydroxyprogesterone (20α-OHP), indicative of P4 overproduction, was also targeted. Progesterone itself was the predominant progestin detected in pregnant and nonpregnant manatee plasma (n = 10) using gas chromatography-mass spectrometry with tandem quadrupole detectors (GC/MS/MS). Progesterone concentrations in pregnant females varied from early (moderate to high) through mid and late (low) pregnancy. Progesterone concentrations ranged from low to high in nonpregnant, nonlactating females. The most commonly detected metabolite was 5α-P3-OH (n = 7), which occurred in pregnant (lower limit of detection [LLOD] to high) and nonpregnant (trace to high) females. The 5α-DHP metabolite was also detected in pregnant (LLOD to moderate) and nonpregnant (low) females. The 17α-OHP metabolite was not detected in any tested female. The 20α-OHP metabolite was detected in one nonpregnant, nonlactating, captive female (LLOD). Metabolites were most prevalent during early pregnancy, concurrent with maximum P4 concentrations. Based on their concentrations in peripheral circulation, we inferred that these metabolites may have, opposite to elephants, a limited physiologic role during luteal, pregnant, and nonpregnant phases in the manatee.     

Progesterone concentrations, exogenous equine chorionic gonadotropin, and timing of prostaglandin F2α treatment affect fertility in postpuberal Nelore heifers

Two experiments were performed to test the hypothesis that elevated progesterone concentrations impair pregnancy rate to timed artificial insemination (TAI) in postpuberal Nelore heifers. In Experiment 1, postpuberal Nelore heifers (n = 398) received 2 mg estradiol benzoate (EB) and either a new progesterone-releasing intravaginal device containing 1.9 g of progesterone (CIDR) (first use) or a CIDR previously used for 9 d (second use) or for 18 d (third use) on Day 0, 12.5 mg prostaglandin F_2α (PGF_2α) on Day 7, 0.5 mg estradiol cypionate (ECP) and CIDR withdrawal on Day 9, and TAI on Day 11. Largest ovarian follicle diameter was determined on Day 11. The third-use CIDR treatment increased largest ovarian follicle diameter and pregnancy rate. Conception to TAI was reduced in heifers with smaller follicles in the first- and second-use CIDR treatments, but not in the third-use CIDR treatment. In Experiment 2, postpuberal Nelore heifers received the synchronization treatment described in Experiment 1 or received 12.5 mg PGF_2α on Day 9 rather than Day 7. In addition, 50% of heifers received 300 IU equine chorionic gonadotropin (eCG) on Day 9. Heifers were either TAI (Experiment 2a; n = 199) or AI after detection of estrus (Experiment 2b; n = 125 of 202). In Experiment 2a, treatment with eCG increased pregnancy rate to TAI in heifers that received PGF_2α on Day 9 but not on Day 7 and in heifers that received a first-use CIDR but not in heifers that received a third-use CIDR. Treatments did not influence reproductive performance in Experiment 2b. In summary, pregnancy rate to TAI in postpuberal Nelore heifers was optimized when lower concentrations of exogenous progesterone were administered, and eCG treatment was beneficial in heifers expected to have greater progesterone concentrations.     

Reproductive toxicologic evaluations of Bulbine natalensis Baker stem extract in albino rats

The effects of oral administration of aqueous extract of Bulbine natalensis Baker stem at daily doses of 25, 50, and 100 mg/kg body weight on the reproductive function of Wistar rats were evaluated. The indices of mating and fertility success as well as quantal frequency increased after 7 days of treatment in all the dose groups except the 100 mg/kg body weight group. The number of litters was not statistically different (P > 0.05) from the control. Whereas the absolute weights of the epididymis, seminal vesicle, and prostate were not affected, that of the testes was significantly increased. The epididymal sperm count, motility, morphology, and viscosity were not different from the control after 7 days of treatment. The male rat serum testosterone, progesterone, luteinizing hormone, and follicle-stimulating hormone significantly increased in the 25 and 50 mg/kg body weight groups, whereas the estradiol concentration decreased significantly at all the doses. The extract dose of 100 mg/kg body weight decreased the serum testosterone and progesterone levels in male rats. The prolactin concentration was not affected by all the doses. All the indices of reproduction, maternal, embryo/fetotoxic, teratogenic, and reproductive hormones in the female rats were not statistically different from that of their control except the resorption index, which increased at the dose of 100 mg/kg body weight of the extract. Histologic examination of the cross section of rat testes that received the extract at all the doses investigated revealed well-preserved seminiferous tubules with normal amount of stroma, normal population of spermatogenic and supporting cells, as well as normal spermatocytes within the lumen. The results revealed that the aqueous extract of Bulbine natalensis stem at doses of 25 and 50 mg/kg body weight enhanced the success rate of mating and fertility due to increased libido as well as the levels of reproductive hormones in male rats. The absence of alterations in the reproductive parameters of female rats at doses of 25 and 50 mg/kg body weight of Bulbine natalensis stem extract suggest that the extract is “safe” for use at these doses by females during the organogenic period of pregnancy, whereas the extract dose of 100 mg/kg body weight portends a negative effect on some reproductive functions of male and female rats.     

Androgen receptors in coeliac ganglion in late pregnant rat

The ovarian function is controlled by endocrine factors and neural influence. In late pregnant rat, androstenedione, from the coeliac ganglion, has a luteotrophic effect in the ex vivo coeliac ganglion-superior ovarian nerve-ovary system. In this work we investigate the presence of androgen receptors in the coeliac ganglion of late pregnant rats by immunohistochemistry. We also explore, from a physiological point of view, the potential participation of these receptors in the androstenedione ganglionic action on progesterone release and metabolism, as well as on nitrites release in the ovary compartment. The coeliac ganglion was isolated after being fixed in situ and immunohistochemistry was performed. In the system, three experimental groups were used with the addition of (a) androstenedione, (b) flutamide, and (c) androstenedione plus flutamide in the ganglion compartment. Progesterone and nitrite concentrations were determined in the ovary compartment at different incubation times. Corpora lutea samples isolated at the end of incubation were used to determine the expressions and activities of the progesterone synthesis (3β-hydroxysteroid-dehydrogenase, 3β-HSD) and degradation (20α-hydroxysteroid-dehydrogenase, 20α-HSD) enzymes. Immunohistochemistry revealed cytoplasmatic androgen receptor immunoreactivity in neural somas in the coeliac ganglion. In the coeliac ganglion-superior ovarian nerve-ovary system, androstenedione addition increased 3β-HSD and decreased 20α-HSD, showed a tendency to decrease 20α-HSD expression, and increased nitrites release in relation to control. Androstenedione plus flutamide decreased progesterone and nitrites release in relation to the androstenedione group. This work demonstrates the presence of androgen receptors in neurons of celiac ganglion and provides evidence for the luteotrophic action of androstenedione via a neural pathway that may be mediated by these receptors.     

GnRH treatment at artificial insemination in beef cattle fails to increase plasma progesterone concentrations or pregnancy rates

Treatment with GnRH at the onset of standing estrus increased pregnancy percentages and circulating concentrations of progesterone in repeat breeder dairy cows. The objective of this study was to determine the effect of treatment with GnRH at AI on concentrations of progesterone and conception rates in beef cattle that exhibited estrus. Two hundred ninety-three heifers at four locations were synchronized with the Select Synch plus CIDR protocol (given GnRH and a CIDR was placed into the vagina, and 7 d later, given PGF_2α and CIDR removed; n = 253) or the 14-19 melengestrol acetate (MGA) protocol (MGA fed at 0.5 mg/head/d for 14 d, with PGF_2α 19 d after MGA withdrawal n = 40) and AI was done after detection of estrus. At Location 1, blood samples were collected on Day 2, 4, 6, 10, 15, and 18 after AI (Day 0 = AI). Two hundred and fifty postpartum cows at two locations were synchronized with the Select Synch plus CIDR protocol, and AI was performed after detection of estrus. At AI, cattle were alternately assigned to one of two treatments: (1) treatment with GnRH (100 μg) at AI (n = 127 heifers and n = 108 cows); or (2) non-treated control (n = 120 heifers and n = 119 cows). Concentrations of progesterone tended to be greater in control heifers compared to GnRH-treated heifers on Days 6 (P = 0.08), 10 (P = 0.07), and 15 (P = 0.11). Overall conception rates were 68% and 66% for GnRH treated and control, respectively, and were not different between treatments (P = 0.72). In summary, treatment with GnRH at time of AI had no influence on conception rates in cattle that had exhibited estrus.     

Comparison of accuracy of ultrasonography, progesterone, and pregnancy-associated glycoprotein tests for pregnancy diagnosis in semidomesticated reindeer

The aim of the study was to compare transrectal ultrasound with progesterone (P4) and pregnancy-associated glycoproteins (PAGs) as pregnancy detection methods for semidomesticated reindeer (Rangifer tarandus tarandus) in field conditions. Female reindeer (n = 195) were scanned transrectally by a 7.5-MHz linear array transducer, and blood was sampled either in December 2005 (n = 33), December 2006 (n = 92), or January 2007 (n = 70) during early or mid gestation. Plasma levels of P4 and PAGs were assessed by radioimmunoassay (RIA). Based on calving records, the sensitivity, specificity, predictive values, and the overall accuracy of the three tests were calculated. The overall calving rate calculated from the calving records was 86.2%. The overall accuracy of transrectal ultrasound was 99.5%. The sensitivity and specificity of transrectal ultrasound were 99.4% and 100%, respectively. In the plasma P4 test, the threshold level of 5.0 nmol/L gave the highest overall accuracy (94.9%). The sensitivity of the P4 test decreased from 96.4% to 81.5%, when the threshold level increased from 5.0 nmol/L to 8.0 nmol/L, while the specificity remained at 85.2% over the range of these cutoff values. The overall accuracy of the plasma PAG test decreased from 96.4% to 64.1% when the plasma PAG threshold level increased from 0.5 ng/mL to 3.5 ng/mL, whereas sensitivity decreased from 99.4% to 58.3%. Specificity increased from 77.8% to 100% when the plasma PAG threshold level reached 3.0 ng/mL. Transrectal ultrasound showed higher diagnostic values than those of plasma P4-RIA and PAG-RIA in diagnosing pregnancy of reindeer, with the advantage that diagnoses can be made in real time in field conditions.     

Identification of sow-specific risk factors for late pregnancy loss during the seasonal infertility period in pigs

Reduced farrowing rates due to late pregnancy loss (LPL) is a manifestation of seasonal infertility in pigs. This study was undertaken to determine sow- and gilt-specific risk factors leading to LPL during the seasonal infertility period (January to April) in Australia. Age at first service was considered to be a major gilt-specific risk factor, whereas sow-specific factors considered included parity, prior wean-to-service interval, prior lactation length, and number of piglets weaned in the lactation period immediately preceding the mating/pregnancy event under scrutiny. Logistic regression analysis of these factors was undertaken on 13,213 animals from three farms (Farms A, B, and C). Age at first service for gilts had an effect on LPL (P < 0.05) on Farm C when compared with that for Farms A and B, with those mated at approximately 220 d having the lowest rate of LPL. For older sows, parity was a factor on Farms A and C (P < 0.001), with the proportion of sows with LPL increasing with increasing parity. When the data from each farm were combined and analyzed, there was a significant farm by WSI interaction, with animals from Farm C being most at-risk for LPL. Sows with shorter lactation periods (P < 0.05) and smaller litters (P < 0.05) at the previous lactation had a greater chance of LPL on all farms. Under the conditions of this study, we were able to identify risk factors for LPL that producers can manipulate during the seasonal infertility period to improve breeding herd productivity.