Electroacupuncture facilitates implantation by enhancing endometrial angiogenesis in a rat model of ovarian hyperstimulation.

Controlled ovarian hyperstimulation (COH) impairs the synchronized development of endometrium and embryo, resulting in the failure of embryo implantation. Here, we investigated what effects electroacupuncture had on embryo implantation in COH rats. Female rats were randomly assigned to four groups: normal (N), model (M), electroacupuncture (EA), and electroacupuncture pretreatment (PEA). Rats in groups M, EA, PEA were injected with pregnant mare serum gonadotropin (PMSG) and human chorionic gonadotropin to establish the COH model. Rats in group EA received electroacupuncture treatment from the PMSG injection day to the 3rd day of pregnancy (D3), while those in group PEA received electroacupuncture treatment for 3 days before the PMSG day and continuing to D3. Furthermore, another 30 female rats who received the same treatment as the rats in group PEA were injected with siVEGFR2 into uterine lumen. The endometrial microvascular density (MVD) and the expression levels of vascular endothelial growth factor-A, angiopoietin-1, and fibroblast growth factor-2 were significantly lower in groups M than in groups N and PEA. The percentage of dolichos biflorus agglutinin positive uterine natural killer cells in groups N, EA and PEA was higher than that in group M. After the siVEGFR2 injection, the protein expression levels of vascular endothelial growth factor receptor 2 (VEGFR2), PI3K, p-AKT and p-ERK, the embryo number and the MVD were significantly reduced. In conclusion, electroacupuncture can facilitate embryo implantation in COH rats by activating the VEGFR2/PI3K/AKT and VEGFR2/ERK signaling pathways which have a positive relationship with endometrial angiogenesis.

Ovarian reaction and estrus manifestation in delayed puberty gilts after treatment with equine chorionic gonadotropin.

BACKGROUND: Prolonged pre-insemination anestrus (i.e. delayed puberty) is a major contributing factor for culling up to 30% of the replacement gilts at large breeding farm units in Vojvodina. It is imperative to determine if these gilts are acyclic (prepubertal) or cyclic, but just fail to exhibit behavioural estrus. Recent investigations demonstrate that treatment with equine chorionic gonadotropin (eCG) can increase the diestrous phase duration in sexually mature gilts. Based on these finding, the aim of the present studies was to determine the reproductive status of delayed puberty gilts following injection with eCG. METHODS: Two experiments were conducted on a swine breeding farm in Vojvodina. In Exp. 1, 20 prepubertal (acyclic) gilts, and 120 sexually mature (cyclic) gilts were injected with a single injection of 400 IU eCG + 200 IU human chorionic gonadotropin (hCG) or with 1000 IU eCG (cyclic gilts), at d5, d11 or d17 after spontaneous estrus detection, to determine their ovarian reaction and induced estrus manifestation. In Exp. 2, sixty delayed puberty gilts (estrus not detected until 8 month of age, av. 258 days) were culled from breeding herd and slaughtered to determine their reproductive status based on ovarian anatomical features. The second group of gilts (n = 60) was treated with a single 1000 IU eCG injection to determine their reproductive status, based on the interval between eCG injection to estrus detection and duration. The data were analyzed by descriptive statistics, t-test, analysis of variance and Duncan's test in the software package Statistics 10th. RESULTS: Ovulations were induced in 90% of acyclic (sexually immature) and, on average, 93.3% of cyclic (sexually mature) gilts after the eCG injection. On average, 4 days after the eCG injection, estrus was detected in 85% of the treated acyclic (sexually immature) gilts and in 95% (19/20) of the cyclic (sexually mature) gilts, treated with eCG on day 17 after spontaneous estrus detection. The interval from eCG to induced estrus detection was prolonged (av. 25 days) in 95% (19/20) of the sexually mature gilts treated with eCG on day 5 and in 90% (18/20) of gilts treated on day 11 after spontaneous estrus detection (Exp. 1). Forty anestrous gilts reached cyclic pubertal ovarian activity. Estrus manifestation was detected in 56 gilts (93.3% of the total 60 treated prolonged anestrous gilts, av. 259 days of age), after a single 1000 IU eCG injection. Thirty-four gilts (60.7% of the total gilts in estrus) with prolonged eCG to estrus interval (av. 24.7 days) were considered spontaneously cyclic (sexually mature), but behaviourally anestrous before treatment. The remaining 22 (39.3% of the total gilts in estrus) were considered truly sexually immature (acyclic) before the treatment or were eCG injected in the late luteal or proestrous phase of spontaneous estrous cycle (Exp. 2). CONCLUSIONS: In 66.7% of the delayed puberty gilts, pre-ovulatory follicles (PoF), corpora hemorrhagica (CH), corpora lutea (CL), or corpora albicantia (CA) were found on the ovaries upon post mortem examination. These gilts were considered as sexually mature before slaughtering. In 60.7% of the delayed puberty gilts, behavioural estrus was detected an average of 24.7 days following eCG injections. These gilts were considered as eCG treated during the luteal phase (diestrus) of the spontaneous estrus cycle. Both findings suggest that delayed puberty gilts actually reached cyclic pubertal ovarian activity (sexual maturity) before culling from the breeding herd.

eFSH in clinical equine practice.

Equine follicle stimulating hormone (eFSH) has been used to induce follicular development in transitional mares and problem acyclic mares, as well as superovulate cycling mares. The most efficacious protocol is to administer 12.5 mg eFSH, intramuscularly, twice daily beginning 5 to 7 days after ovulation when the diameter of the largest follicle is 20 to 25 mm. Prostaglandins are to be administered on the second day of eFSH therapy. Treatment with eFSH is continued for 3 to 5 days until follicle(s) are >or=35 mm in diameter. The mare is subsequently allowed to 'coast' for 36 h, after which human chorionic gonadotropin is administered to induce ovulation.

Luteal P4 synthesis in early pregnant gilts after induction of estrus with PMSG/hCG.

The present study was designed to examine whether an estrus induction with gonadotropins could affect luteal P4 synthesis in early pregnant gilts. Sixteen prepubertal gilts received 750IU of PMSG and 500IU of hCG 72h later. Prepubertal gilts in the control group (n=17) were observed daily for estrus behavior. All gilts were inseminated in their first estrus. Corpora lutea (CLs) were collected on days 10, 12 and 15 of pregnancy and analyzed for (1) the mRNA and protein expression of steroidogenic acute regulatory protein (StAR), cytochrome P450 family 11 subfamily A polypeptide 1 (CYP11A1), and 3ß-hydroxysteroid dehydrogenase (3ßHSD); (2) the tissue concentration of P4; and (3) the mRNA expression of luteinizing hormone receptor (LHR) and estrogen receptors (ESR1 and ESR2). Additionally, P4 concentration was analyzed in blood serum of all animals. PMSG/hCG injections to induce estrus decreased mRNA expression of StAR, CYP11A1 and 3ßHSD on day 10 and CYP11A1 on day 12 of pregnancy compared with the control group, while CYP11A1 and 3ßHSD proteins were down-regulated on day 10 in the hormonally-treated gilts. Concentrations of P4 in luteal tissue and blood serum were also lower in animals after gonadotropin-induced estrus. In contrast, LHR and ESR1 mRNA expression was greater in PMSG/hCG-treated than control gilts on day 15 of gestation. In conclusion, induction of estrus with a PMSG/hCG protocol in prepubertal gilts impaired expression of the luteal P4 synthesis system. Low P4 content may, in turn, induce local mechanisms involving LHR and ESR1 expression to support CL function.

Gonadotropin stimulation for in vitro fertilization and embryo transfer in insulin-dependent diabetics: follicular response, oocyte quality, embryo development, and follicular environment.

Two IDD patients were stimulated with gonadotropins for IVF and ET. Both patients had high E2 response and greater than or equal to 5 preovulatory oocytes retrieved, normal fertilization and cleavage rates, and transfer; neither conceived. Follicular fluid showed levels of E2, P, A, hCG, and PRL similar to non-IDD. Epidermal growth factor could not be detected in FF. Insulin-dependent diabetes patients can undergo gonadotropin stimulation for IVF with customary responses. Their follicular milieu resembles that of non-IDD patients except for a lack of EGF.

Increased production of sheep embryos following superovulation of Merino ewes with a combination of pregnant mare serum gonadotrophin and follicle stimulating hormone.

In a factorially designed experiment (N = 321), 0, 800 or 1600 I.U. pregnant mare serum gonadotrophin (PMSG) were administered in combination with 0, 12 or 18 mg follicle stimulating hormone (FSH-P) to superovulate Merino ewes in autumn and spring. A moderate dose of PMSG (800 I.U.) in conjunction with 12 or 18 mg FSH-P increased the ovulation rate above that observed when FSH-P was used alone. This was accomplished by (i) increasing the proportion of ewes that exhibited a superovulatory response (greater than 3 corpora lutea (CL) or persistent large follicles (LF): 69/70 (99%) v. 55/74 (74%), P less than 0.001), and (ii) in those ewes that exhibited a superovulatory response, by an additive effect of exogenous gonadotrophin (14.8 +/- 0.9 CL (69) v. 11.3 +/- 0.9 CL (55), P less than 0.01) without increasing the incidence of LF. The use of 1600 I.U. PMSG in conjunction with 12 or 18 mg FSH-P was characterized by an increase in the number of LF and, in comparison with 800 I.U. PMSG, a reduction in ovulation rate. Season had no effect on the numbers of CL, but total ovarian response (CL + LF) was higher in autumn than in spring (P less than 0.01), because of a greater incidence of LF (P less than 0.001). The proportion of ewes with regressed CL was higher in autumn than in spring (53/143 (37%) v. 32/156 (21%), P less than 0.01), and increased with increased dose of gonadotrophin. Furthermore, a nutritional component to the incidence of ewes with regressed CL was suggested by the observation that the mean concentration of plasma glucose was higher for ewes with normal CL than for ewes with regressed CL (P less than 0.05). Rates of ova or embryo recovery, fertilization and embryo development generally declined with an increase in the incidence of LF as a result of increases in the dose of gonadotrophin and season of administration.

Early development and location of embryos in the reproductive tract of Nili Ravi buffalo (Bubalus bubalis): a retrospective analysis.

One year data on embryo recovery were analyzed to study the development and descent of preimplantation embryos in Nili Ravi water buffalo. Forty-five superovulatory attempts were performed on 23 buffalo. A total of 45 embryos were recovered either nonsurgically or after slaughtering the animals at various time intervals (85 to 176 h) post estrus. Embryos were located in the oviducts at 85 h after estrus. At 108 h post estrus, most of them (78%) were recovered from the uteri. The embryos had 8 to 16 cells at 85 h post estrus, grew to morulae at 108 h and to compact morulae at 125 h post estrus. Early blastocysts were observed at 141 h post estrus. Blastocysts were predominant (69%) at 156 to 176 h after estrus; no hatched blastocysts were recovered during this time interval. Based on our findings, embryo recovery at around 150 h post estrus (i.e., Day 6 of the cycle) is recommended for compact morulae or blastocysts in the water buffalo.

Influence of flushing on LH secretion, follicular growth and the response to estrus synchronization treatment in suckled beef cows.

The effects of energy supplementation (flushing) on LH and estradiol secretion, follicular growth and the response to estrus synchronization treatment (Norgestomet + PMSG initiated 41.9 +/- 3.4 d after calving) were investigated in 16 suckled beef cows fed either 70% (Group C, n = 8) of energy requirements from calving to 3 wk after AI or fed the same restricted diet until 11 d before synchronization and then were supplemented with 2 kg concentrate until 3 wk after AI (Group S, n = 8). Concentrations of LH and estradiol 17 beta were measured from 3 sampling periods: 25 and 39 d after calving and between 29 and 49 h after implant removal. Ovaries were examined by ultrasonography 11 d before treatment to implant withdrawal (IR). The effects of energy level, day (or hour) of observation and corresponding interactions were tested on repeated measurements by split-plot ANOVA. No positive effect of flushing was observed on characteristics of LH secretion on Day 39. However, the size of the largest follicle and the number of large follicles were higher in Group S than in Group C cows, respectively, 7 and 9 d after the beginning of flushing to 2 d after the start of treatment. After IR, the estradiol secretion tended to be higher in Group S than in Group C cows (9.8 +/- 0.4 pg/mL vs 7.2 +/- 0.2 pg/mL; P = 0.06), but no effect on LH secretion was observed. After implant removal 12 cows ovulated (Group S: 7/8 vs Group C: 5/8; P > 0.05), 7 were pregnant at 21 d after AI (Group S: 6/8 vs Group C: 1/8; P < 0.05) and 4 at 45 d after AI (Group S: 4/8 vs Group C 0/8; P > 0.05). To conclude, flushing had a positive effect on follicular growth, which does not seem to be mediated by LH. In cows fed a restricted diet, flushing enhanced follicular growth, increased the fertilization rate and/or reduced early embryonic death.

Changes in plasma testosterone and testicular transferrin concentration, testicular histology and semen quality after treatment of testosterone-depot plus PMSG to 3 dogs with asthenozoospermia.

Three dogs diagnosed as having asthenozoospermia were given three intramuscular injections of 50 mg testosterone(T)-depot plus 250 IU pregnant mare serum gonadotropin (PMSG) at 2-week intervals, and their plasma T and testicular transferrin (Tf) concentrations, testicular histology, and semen quality were examined during the period of hormone therapy. Plasma T concentrations temporarily increased, and there was a slight improvement in spermatogenesis. Increased Tf concentrations suggested that Sertoli cell function in all three dogs was promoted by hormone treatment. The results showed that semen quality, especially the percentages of motile sperm and abnormal sperm, were improved between 1 and 5 weeks after the start of T-depot plus PMSG treatment.

Effects of tokishakuyakusan, keishibukuryogan, shakuyakukanzoto and unkeito on ovarian endothelin, renin and angiotensin II in pregnant mare's serum gonadotropin-treated immature rats.

We have previously proposed the ovarian ERAANPS (endothelin-rein- angiotensin-atrial natriuretic peptide system). The present study was undertaken to examine in vivo the effects of herbal medicines [Tokishakuyakusan (TS), Keishibukuryogan (KB), Shakuyakukanzoto (SK) and Unkeito (UT)] on endothelin-1 (ET), renin and angiotensin II (A II) in the ovaries, of immature rats treated with 10 IU PMS for 48 h. ET and all components of renin-angiotensin system (RAS) were found at high levels in the ovary. Concomitant treatment with PMS plus TS, KB, SK or UT, especially TS and UT, tended to decrease the ET levels in ovary, while components of RAS tended to increase. However, ET, renin and A II levels in plasma were not at all affected after treatment with TS, KB, SK or UT. These results suggest that TS, KB, SK or UT may regulate the ovarian ERAANPS.

Evaluation of a pregnant mares' serum and human chorionic gonadotrophin mixture for alleviating summer infertility in sows.

A controlled on-farm trial was conducted to evaluate the effectiveness of a commercial gonadotrophin hormone mixture at overcoming a seasonal decrease in fertility in pigs in a sub-tropical environment (summer infertility). The average weaning to oestrus interval of the untreated sows was more than double that of the treated sows and this difference was highly significant. However, fewer of the treated sows farrowed and they produced smaller litters. The improvement in fertility in the treated group achieved by reducing the weaning to oestrus interval was more than cancelled out by an overall reduction in the number of weaners produced.

Evaluation of a regimen using a progesterone releasing intravaginal device (CIDR) and PMSG as a treatment for post partum anoestrus in dairy cattle.

Anoestrous dairy cows in seasonally calving herds in the Macalister Irrigation District of Gippsland, Victoria were treated at the start of the mating period with a progesterone releasing intravaginal device (CIDR). The CIDR was inserted for 7 days and 400 IU of PMSG was injected intramuscularly at removal. There was no clinically useful difference among cows receiving the CIDR, a placebo and untreated cows in the interval from treatment to either first oestrus or conception, the conception rate to first service or percent pregnant by the end of mating. Analyses of data from 2-year-old cows, older cattle, cows calved at least 45 days or cows calved at least 55 days and cows treated 3 weeks after the start of mating did not show improved reproductive performance following treatment with the CIDR.

Birth of Siberian tiger (Panthera tigris altaica) cubs after transvaginal artificial insemination.

A captive 9-yr-old female Siberian tiger (Panthera tigris altaica) at the Lisbon Zoo was treated with equine chorionic gonadotropin followed 80 hr later by human chorionic gonadotropin to stimulate follicular development and ovulation, respectively. Forty-one hours after the second hormone administration, the tiger was subjected to transvaginal artificial insemination with diluted fresh semen containing 500 x 10(6) motile spermatozoa obtained from an 8-yr-old male Siberian tiger via electroejaculation. After anesthesia, the female tiger was placed in an inclined position with the hind limbs elevated during and after the insemination and was maintained in this position for 15 min to minimize semen reflux. An anesthetic reversal agent was then administered. After a 103-day gestation, the female gave birth to three apparently healthy cubs that survived 24-48 hr. These results demonstrate that transvaginal artificial insemination can be successfully performed in the Siberian tiger to produce full-term offspring.

Acyclicity and dyscyclicity in mares.

Two trotter stud farms were visited on Mondays, Wednesdays, and Fridays during 1972 and 1973. A total of 356 mares were bored on these farms during these breeding seasons. There were 194 nursing, 113 barren, and 49 maiden mares. Of the 356 mares, 78 (21.9%) had functional disorders of the ovaries at one time or another. The functional disorders consisted of: 27 anoestrous mares after early embryonic death, 20 with low ovarian activity, 19 with multifollicular ovaries, and 12 mares with persistent corpora lutea. The following therapies were used: (a) 1000 I.U. PMSG i.v. + 10 mg stilboestrol i.m. Of 13 treated mares, 3 (23.1%) came into heat and all 3 became pregnant in the first oestrus following treatment. (b) Intra-uterine infusion of 0.5 - 1 liter physiological saline solution. Of the 26 treated mares, 19 (73.1%) came into oestrus and 14 (53.9%) became pregnant in the first oestrus following treatment. (c) Chlormadinone acetate, 10 mg. orally for 17 days. Of 38 treated mares, 35 (92.4%) came into oestrus and 22 (57.9%) became pregnant in the first oestrus following treatment. This therapy was also used in early spring (February-March) in 15 mares, of which 10 (66.7%) came into oestrus and 4 (26.7%) became pregant in the first oestrus following treatment. (d) Prostaglandin F2alpha, 25 mg intra-uterine. Of the 9 mares treated, al came into oestrus and 8 (88.9%) became pregnant in the first oestrus following treatment. (e) Curettage of the uterus. In this treatment group, none of the 5 treated mares showed improvement. Altogether, 54.7% of the treated mares became pregnant in the first oestrus after treatment, and 76.9% of the mares became pregnant after one of more breedings.

Gonadotropin stimulation using P.G. 600® on reproductive success of non-lactating anestrous ewes.

The effect of stimulation with a gonadotropin preparation with combined follicle stimulating and luteininzing hormone like activity on reproductive success in anestrous ewes was evaluated. In Experiment 1, ewes of mixed breeding were treated with CIDR inserts (0.3g progesterone) for 5 days and were assigned randomly to receive either gonadotropin stimulation (3mL i.m. injection of P.G. 600®, 240IU eCG and 120IU hCG) at CIDR removal or no further treatment. Intact raddled rams were joined at insert removal for 30-35 days, and ewes were observed for indications of estrus after 4 days of ram exposure. Pregnancy diagnosis was conducted via transrectal ultrasonography at the time of ram removal and again 20-25 days. The second experiment was similar to Experiment 1, except treated ewes received the gonadotropin 1 day prior to insert removal. In Experiment 1, incidence of estrus was greater for treated ewes (P=0.01), and prolificacy tended to be greater in treated ewes (P=0.06). In Experiment 2, treated ewes had greater conception rates (P=0.01), pregnancy rates to first service (P=0.0007), and tended to have greater overall pregnancy rates than control ewes (P=0.07). A greater percentage of ewes lambed in the gonadotropin treated ewes than in ewes in the control group (P<0.0001), and overall lambing rates in treated ewes were greater than non-treated controls (P<0.0001). In conclusion, gonadotropin treatment 1 day prior to CIDR removal increased reproductive success in progesterone-treated anestrous ewes.

Resynchronization of estrous cycle with eCG and temporary calf removal in lactating Bos indicus cows.

The aim of this study was to compare four methods of estrus resynchronization performed 23 days after timed artificial insemination (TAI) plus estrus observation in Bos indicus cows. Eight hundred fourteen lactating Nelore cows were submitted to TAI and then randomly assigned to one of the five following treatments: R23 (resynchronization without eCG), R23/200 (resynchronization with 200 IU of eCG), R23/300 (resynchronization with 300 IU of eCG), R23/TCR (resynchronization with temporary calf removal [TCR]), and a control group, with estrus observation followed by AI (with no resynchronization). Treatment consisted of a progesterone device plus administration of estradiol benzoate on Day 0; on Day 8, the device was removed and cloprostenol was applied, together with estradiol cypionate. Also on Day 8, either eCG was administered or TCR was performed in the resynchronized groups, except for R23. The females were inseminated 48 hours after device removal or TCR (33 days after the first TAI). The control group was kept under estrus observation from 18 to 23 days after the first TAI and was inseminated 12 hours after detection of estrus. The first pregnancy evaluation was performed using ultrasound examination 31 days after the first TAI. After 30 days of the resynchronization, a second pregnancy evaluation was performed and the animals in the R23/300 and R23/TCR groups achieved the highest conception rates, 76.6% and 74.0%, respectively (P < 0.05). There were no differences between the conception rates of the animals in the R23/200 (63.3%), R23 (61.3%), and control (54.3%) groups (P > 0.05). These results suggest that estrus resynchronization at 23 days after TAI can effectively improve the conception rate of lactating Bos indicus cows in a short time period. Furthermore, resynchronization with 300 IU of eCG or with TCR provided the best results.

Hormone replacement therapy in children with hypogonadotropic hypogonadism: where do we stand?

OBJECTIVE: To characterize hormone replacement therapy in a cohort of adolescent males and females with hypogonadotropic hypogonadism (HH) with a focus on changes in management during the past 10 years. METHODS: Medical records of patients followed for HH during the past 10 years were reviewed. RESULTS: A total of 45 patients (22 female: 23 male) with HH were identified. The average age at HH diagnosis was 14.48 ± 2.02 years in females and 14.89 ± 1.64 years in males (P = .53). In females, the average age of pubertal induction was 14.53 ± 1.86 years. Conjugated equine estrogen was used in 54.5%, transdermal estradiol in 41%, and oral estradiol in 4.5%. The average duration to cycling was 1.96 ± 0.78 years. A progressive increase in the use of transdermal estradiol was noted over time, with 100% of females being started on this regimen since 2008. In males, the average age of induction was 15.22 ± 1.41 years. All were started on intramuscular testosterone cypionate at various doses. The average duration to full adult replacement was 1.95 ± 0.51 years. CONCLUSION: There is no current standard of care to guide pubertal induction in adolescents with HH. However, a significant increase in the use of transdermal estrogen was noted in females during the past 10 years. While much less variability in pubertal induction was seen in males, wide disparities in doses and escalation schedules were found. Prospective studies aimed at elucidating optimal strategies for sex steroid replacement in this pediatric population are badly needed.