Injection Of Follicle-stimulating Hormone Research Articles

FSH preserves the viability of hypoxic granulosa cells viaactivating the HIF-1α-GAS6-Axl-Akt pathway.

The developmental fate of ovarian follicles is primarily determined by the survival status (proliferation or apoptosis) of granulosa cells (GCs). Owing to the avascular environment within follicles, GCs are believed to live in a hypoxic niche. Follicle-stimulating hormone (FSH) has been reported to improve GCs survival by governing hypoxia-inducible factor-1α (HIF-1α)-dependent hypoxia response, but the underlying mechanisms remain poorly understood. Growth arrest-specific gene 6 (GAS6) is a secreted ligand of tyrosine kinase receptors, and has been documented to facilitate tumor growth. Here, we showed that the level of GAS6 was markedly increased in mouse ovarian GCs after the injection of FSH. Specifically, FSH-induced GAS6 expression was accompanied by HIF-1α accumulation under conditions of hypoxia both in vivo and in vitro, whereas inhibition of HIF-1αwith smallinterfering RNAs/antagonist repressed both expression and secretion of GAS6. As such, Luciferase reporter assay and chromatin immunoprecipitation assay showed that HIF-1α directly bound to a hypoxia response element site within the Gas6 promoter and contributed to the regulation of GAS6 expression in response to FSH. Notably, blockage of GAS6 and/or its receptor Axl abrogated the pro-survival effects of FSH under hypoxia. Moreover, phosphorylation of Axl by GAS6 is required for FSH-mediated Akt activation and the resultant pro-survival phenotypes. Finally, the in vitro findings were verified in vivo, which showed that FSH-induced proliferative and antiapoptotic effects in ovarian GCs were diminished after blocking GAS6/Axl using HIF-1α antagonist. These findings highlight a novel function of FSH in preserving GCs viability against hypoxic stress by activating the HIF-1a-GAS6-Axl-Akt pathway.

Testosterone identifies hatchling sex for Mojave desert tortoises (Gopherus agassizii)

The threatened Mojave desert tortoise (Gopherus agassizii) exhibits temperature-dependent sex determination, and individuals appear externally sexually monomorphic until sexual maturity. A non-surgical sex identification method that is suitable for a single in situ encounter with hatchlings is essential for minimizing handling of wild animals. We tested (1) whether plasma testosterone quantified by enzyme-linked immunosorbent assay differentiated males from females in 0–3 month old captive hatchlings, and (2) whether an injection of follicle-stimulating hormone (FSH) differentially elevates testosterone in male hatchlings to aid in identifying sex. We validated sex by ceolioscopic (laparoscopic) surgery. We then fit the testosterone concentrations to lognormal distributions and identified the concentration below which individuals are more likely female, and above which individuals are more likely male. Using a parametric bootstrapping procedure, we estimated a 0.01–0.04% misidentification rate for naïve testosterone samples, and a 1.26–1.39% misidentification rate for challenged (post-FSH injection) testosterone samples. Quantification of plasma testosterone concentration from small volume (0.1 mL) blood samples appears to be a viable, highly accurate method to identify sex of 0–3 month old hatchlings and could be a valuable tool for conservation measures and investigation of trends and variation in sex ratios for in situ wild nests.

Potential role for human menopausal gonadotropin in ovine superovulatory regimens during the breeding season.

Human menopausal gonadotrophin (hMG) has been reported to produce a comparable superovulatory response to that of follicle stimulating hormone (FSH). Furthermore, hMG has a long half-life as compared with FSH. The present study was designed to compare hMG administered once daily and FSH administered twice daily over a 4 - day period on superovulatory response of Suffolk ewes. During the mid-luteal phase, twenty-four Suffolk donor ewes received intravaginal sponges at day 0 for 12 days. The superovulatory regimens in the Control group (n=12) and the Treatment group (n=12) consisted of eight injections of FSH given at twice daily and four injections of hMG given at once daily, respectively. At day 13, the donor ewes were subjected to laparoscopic insemination. Embryos were recovered, classified, and transferred to recipient ewes at day 19. Pregnancy status was determined by ultrasound examination 40 days after transfer. Lambing rate was calculated after all the ewes had delivered. No significant differences were observed between the two groups in terms of the structures recovered, transferable embryos, degenerated embryos, unfertilized oocytes, pregnancy rate and lambing rate. The results showed that once daily injection of hMG can produce a comparable superovulatory response and embryo transfer outcomes to those obtained by twice daily injection of FSH over a 4 - day period. It is feasible that hMG is used to replace FSH and reduce the number of injection treatments in ovine superovulatory regimens.

The impact of FSH stimulation and age on the ovarian and uterine traits and histomorphometry of prepubertal gilts

This study investigated the effect of age and follicle stimulating hormone (FSH) treatment on the estradiol (E2) plasma concentration, ovarian follicle development, endometrial histomorphometry, and ultrasonographic parameters of the ovaries and uterus in prepubertal gilts. Thirty-five prepubertal gilts were grouped according to age (140 or 160 d), and within each age, gilts were allotted to receive 100 mg of FSH (treated; G140 + FSH [n = 10] and G160 + FSH [n = 7]) or saline solution (control; G140 + control [n = 10] and G160 + control [n = 8]). The total dose of FSH was divided into 6 similar doses administered every 8 h (days 0–2). Before and after FSH treatment, blood sample was collected, and transabdominal scanning of the ovaries and uterus was performed. Twenty-four hours after the last FSH injection, the gilts were slaughtered and their ovaries and uterus were processed for histological and histomorphometric analysis. The histomorphometric parameters of the uterus differed (P < 0.05) between prepubertal gilts at 160 d and 140 d of age. Moreover, changes (P < 0.05) in uterine and ovarian ultrasound images occurred between 140 and 160 d of age. Age and FSH treatment did not affect (P > 0.05) E2 plasma concentrations. Follicle stimulating hormone treatment did not affect (P > 0.05) the early stage of folliculogenesis in the prepubertal gilts; however, the number of early atretic follicles decreased (P < 0.05) after the FSH treatment. Follicle stimulating hormone administration increased (P < 0.05) the number of medium follicles and decreased (P < 0.05) the number of small follicles in 140 and 160 d old gilts. In the endometrium, luminal/glandular epithelium height and glandular diameter increased (P < 0.05) after FSH treatment. Thus, injections of 100 mg of FSH stimulate the endometrium epithelium and induce follicular growth to a medium follicle size without affecting the preantral stages in prepubertal gilts; also, the uterine macroscopic morphometry does not change from 140 to 160 d of age.

The Success of Ovulation Induction with Letrozole and Gonadotropins in Obese and Nonobese Women: A Study from a Tertiary Center.

Background Letrozole, an aromatase inhibitor, is suggested as a first-line drug for ovulation induction in women with polycystic ovary syndrome (PCOS) especially in obese women. Letrozole has also been used in women with unexplained infertility with similar rates of success to clomiphene. However, literature on letrozole and gonadotropins in obese and nonobese women is sparse. Hence, this study was done to assess the success of ovulation induction (OI) with letrozole plus follicle stimulating hormone (FSH) in obese (BMI ≥ 30 kg/m2) and nonobese women (BMI < 30 kg/m2). Methods A retrospective descriptive cohort study was conducted involving 135 women who underwent OI with letrozole plus follicle stimulating hormone therapy and either timed intercourse or intrauterine insemination. The data was collected from the hospital information system, including the age, body mass index, the type of infertility, number of induction cycles with letrozole, number of gonadotropin injections, and the pregnancy occurrence following treatment. SPSS was used to analyze the data. Results There were 135 women who used FSH injections along with letrozole. Of this, 28.5% obese women got pregnant compared to 29.2% nonobese women, but this did not attain statistical significance (P = 0.75). About 70% of obese women and 57% on nonobese women had polycystic ovarian syndrome. The median number of FSH injections was six, and the interquartile range was 3 to 11. Conclusion Of the 135 women undergoing letrozole and FSH, there was almost an equal probability of pregnancy in the obese group (BMI ≥ 30 kg/m2) and nonobese women.

FSH mediates estradiol synthesis in hypoxic granulosa cells by activating glycolytic metabolism through the HIF-1α–AMPK–GLUT1 signaling pathway

Owing to the avascular environment within ovarian follicles, granulosa cells (GCs) are believed to live in a hypoxic niche. Follicle-stimulating hormone (FSH)-mediated steroidogenesis is crucial for normal growth and maturation of ovarian follicles, but it remains unclear how FSH stimulates estradiol (E2) synthesis under hypoxic conditions. Here, we aimed to explore whether FSH affects the ATP production required for estrogen synthesis from the perspective of glucose metabolism. It was observed that the levels of both E2 and HIF-1α were markedly increased in a dose-dependent manner in mouse ovarian GCs after the injection of FSH in vivo, indicating that hypoxia/HIF-1α may be relevant to FSH-induced E2 synthesis. By treating hypoxic GCs with FSH in vitro, we further revealed that the activation of the AMP-activated protein kinase (AMPK)–GLUT1 pathway, which in turn stimulates ATP generation, may be essential for FSH-mediated E2 production during hypoxia. In contrast, inhibition of AMPK or GLUT1 with siRNAs/antagonist both repressed glycolysis, ATP production, and E2 synthesis despite FSH treatment. Moreover, blocking HIF-1α activity using siRNAs/PX-478 suppressed AMPK activation, GLUT1 expression, and E2 levels in FSH-treated GCs. Finally, the in vitro findings were verified in vivo, which showed markedly increased AMPK activity, GLUT1 expression, glycolytic flux, ATP levels, and E2 concentrations in ovarian GCs following FSH injection. Taken together, these findings uncovered a novel mechanism for FSH-regulating E2 synthesis in hypoxic GCs by activating glycolytic metabolism through the HIF-1α–AMPK–GLUT1 pathway.

Plasma profile of follicle-stimulating hormone and sex steroid hormones after a single epidural administration of follicle-stimulating hormone via caudal vertebrae in Holstein dry cows.

The conventional follicle-stimulating hormone (FSH) treatment for bovine superstimulation involves multiple intramuscular injections, which is stressful for animals and onerous. We herein investigated whether a single epidural injection of porcine FSH (pFSH) can induce superovulation and peripheral concentrations of pFSH and steroid hormones after the treatment in Holstein dry cows. We intramuscularly administered pFSH twice daily to three cows for 3 days (control) or a single epidural pFSH administration (epidural). Numbers of follicles (≥10mm in diameter) at estrus and corpora lutea at luteal phase were counted by ultrasonography. Blood was sampled from 0 to 104 h after the first pFSH administration and plasma pFSH, progesterone, androstenedione, testosterone, and estradiol-17β concentrations were measured. Numbers of follicles (control: 18.3 ± 7.5, epidural: 15.7 ± 4.0; mean ± SD) and corpora lutea (control: 7.3 ± 4.2, epidural: 8.0 ± 2.6) were similar between both treatments. Plasma pFSH concentrations were higher in epidural than in control (p < 0.01). Although no significant differences were observed in progesterone, androstenedione, or estradiol-17β concentrations between the groups, testosterone concentrations were slightly lower with the epidural treatment than with the control treatment (p = 0.08). In conclusion, superovulation was induced by a single epidural injection of pFSH, which achieved higher pFSH level than the multiple injections in Holstein dry cows.

The Effect of Epidural Application of FSH on Superovulatory Response in Eastern Anatolian Red Cow

This study aimed to evaluate the superovulation response in Eastern Anatolian Red (EAR) cows treated with a single epidural injection or traditional administration of follicle stimulating hormone (FSH). Eight cows were randomly divided into two groups, as control (CG) and experiment (EG). Four replications were applied for each animal in the groups. A total of 16 studies were carried out in each group. In this study, 300 mg intramuscular (i.m) FSH injection was applied with decreasing doses twice daily for four days to the cows in the CG. (70:70, 50:50, 20:20, 10:10 mg). On the other hand, 300 mg FSH was administered as 150 mg epidural and 150 mg intramuscular injection to the cows in EG. The number of untransferable embryos and the total CLs were found statistically significant (p&amp;lt;0.05). However, the number of transferable embryos and unfertilized oocyte (UFO) were found statistically insignificant among the groups (p&amp;gt;0.05). According to the obtained data, it was observed that the epidural administration of FSH for superovulation in EAR cows did not provide sufficient follicle and oocyte development. In conclusion, different studies with different superovulation protocols and FSH doses in local breed EAR cows will be needed for further studies.

Superovulation response of Peranakan Ongole (PO) and Simmental cows after FSH stimulation in multiple ovulation and embryo transfer program

Multiple ovulation and embryo transfer (MOET) is a reproductive technology to increase the livestock population in a short period. The success of ovary stimulation programs is influenced by an individual’s response to follicle-stimulating hormone (FSH) stimulation. In this study embryo production was carried out on local cows represented by Peranakan Ongole (PO) cows and exotic cows represented by Simmental cows. FSH stimulation was performed on 10 PO cows and 10 Simmental cows. On Day-1 Cue Mate® (progesterone) was inserted intravaginally. FSH injection (400 mg) was carried out intramuscularly from D-10 at reduced dose with 12 hours intervals (Day-10: 100 mg; Day-11: 60 mg; Day-12: 40 mg per injection). On Day-12, prostaglandin (PGF2α) was injected and Cue Mate® was removed. Artificial insemination (AI) was performed 48 hours after PGF2α injection for 3 times at 12 hours intervals. The embryo was collected 6 days after the last AI (Day-21). Superovulation response was detected on 70% of PO cows and 90% of Simmental cows. The average number of transferable embryos in Simmental cows (9.11±7.27) was higher than PO cows (7.86±7.78). This research shows that Simmental cows are more responsive to FSH stimulation, and can produce more transferable embryos than PO cows.

Süperovulasyon Öncesi GnRH ve Kısa Süreli Progesteron Uygulamalarının Sığır Embriyolarının Sayısı ve Kalitesi Üzerine Etkisi

Bu çalışmada, insan koryonik gonadotropini (hCG) ve prostaglandin F2-alfa (PGF2α) ile östruslarının indüklenmesinden sonra gonadotropin salgılatıcı hormon (GnRH) ve kısa süreli progesteron uygulamasını takiben süperovulasyon uygulanan Holştayn donörler (modifiye protokol) ile klasik süperovulasyon uygulanan Holştayn donörlerden elde edilen embriyoların kalitesi ve sayısı karşılaştırıldı. Araştırmada, üç yaşlı 20 adet Holştayn ırkı inek iki eşit gruba ayrıldı. Grup I’ deki hayvanlara 11 gün ara ile iki defa kloprostenol enjeksiyonu yapıldı ve östrus sonrası takip eden siklusun 9. gününden itibaren, günde iki kez dört gün süresince azalan dozlarda FSH enjeksiyonları uygulandı. Beşinci ve 6. FSH enjeksiyonları ile birlikte kloprostenol uygulamaları yapıldı.Grup II’ deki hayvanlara ise siklusun herhangi bir döneminde hCG uygulaması yapıldı. Bundan 7 gün sonra kloprostenol enjeksiyonu yapıldı. Östrus sonrası devam eden siklusun 7. günü buserelin enjeksiyonu yapıldı. PRID siklusun 9. günü yerleştirildi ve siklusun 14. günü çıkarıldı (6. FSH enjeksiyonu sırasında). Grup II’de FSH ve kloprostenol uygulamaları siklusun 12-15. günlerinde Grup I’ deki gibi yapıldı. Tüm inekler son FSH enjeksiyonundan sonra 12 saat ara ile üç kez tohumlandı. İkinci suni tohumlama ile eş zamanlı olarak buserelin enjeksiyonu yapıldı. İkinci tohumlama sonrası 7. günde embriyolar toplandı, morfolojilerine bağlı kalite ve gelişim evreleri değerlendirildi. Bu çalışmada donör başına sırasıyla Grup I ve Grup II’ de toplam CL (8.50±1.90 ve 9.40±1.52), toplam oosit ve embriyo (7.5±2.03 ve 7.0±1.61), toplam embriyo (6.00±2.00 ve 6.10±1.47), transfer edilebilir embriyo ı (5.50±1.84 ve 5.50±1.59), dejenere embriyo (0.50±0.22 ve 0.60±0.40) ve unfertilize oosit sayıları (1.60±0.97 ve 0.90±0.31) tespit edildi. Sonuç olarak, iki protokol arasında embriyo sayısı ve kalitesi açısından istatistiksel olarak fark bulunamadı (P &amp;gt;0,05). Böylece çok sayıda donöre östrus takibine gerek duyulmadan sabit zamanlı süperovulasyona olanak sağladığından dolayı klasik süperovulasyon uygulamalarına alternatif bir yöntem olabileceği sonucuna varılmıştır.

Collection and in vitro maturation of Mazama gouazoubira (brown brocket deer) oocytes obtained after ovarian stimulation.

In vitro production of embryos has gained prominence as a tool for use in wildlife conservation programmes in situ and ex situ. However, the development of this technique depends on steps that include ovarian stimulation, collection and oocyte maturation. The purpose of this study was to assess the feasibility of an ovarian stimulation protocol for follicular aspiration, the efficiency of videolaparoscopy for follicular aspiration and test a medium for in vitro oocyte maturation for the species Mazama gouazoubira. Five females were submitted to repeated ovarian stimulation (hormone protocol using controlled internal drug release), and estradiol benzoate on D0 and eight injections of follicle-stimulating hormone, once every 12 h, from D4 onwards at 30-day intervals. Fourteen surgical procedures were performed in superstimulated females, resulting in the collection of 94 oocytes and an average of 17.1 ± 9.1 follicles observed, 13.5 ± 6.6 follicles aspirated and 7.2 ± 3.7 oocytes collected per surgery. After collection, the oocytes were submitted to in vitro maturation for 24 h and stained with Hoechst 33342 dye to evaluate their nuclear status; 64.5% of the oocytes reached MII and 16.1% were spontaneously activated by parthenogenesis. The nuclear status of oocytes that did not undergo in vitro maturation was evaluated; 80.9% were found to be immature.

Evaluation of the relationship between serum paraoxonase-1 activity and superovulation response/embryo yield in Holstein cows.

In this study, the effect of serum paraoxonase-1 (PON-1) activity on superovulation response and embryo yield was evaluated. The study material comprised 50 Holstein cows aged 3–4 years on postpartum day 90–120 with a body condition score of 3–3.25. A progesterone-based estrus synchronization protocol was initially administered to the selected donors. For this purpose, progesterone source was inserted intravaginally (day 0) and gonadotropin-releasing hormone injection was performed (day 6). Seven days after the insertion of progesterone device, follicle-stimulating hormone injections (total dose of 500 µg in decreasing doses for 4 days) were administered for superovulation. On the morning of the ninth day, prostaglandin (PG) F2α was administered, and the progesterone device was removed from the vagina in the evening on the same day. Two days after PGF2α administration, fixed-time artificial insemination was performed in the morning and in the evening. On the day of artificial insemination, blood samples were taken from the donors to determine the serum PON-1 activity. Uterine flushing was performed seven days after insemination. The results revealed that the serum PON-1 activity (mean ± SD, 562.71 ± 140.23 U/l) of the cows that responded to superovulation (donors with total corpus luteum count of ≥3 in both ovaries) was higher than those (389.91 ± 80.51 U/l) that did not (P<0.05). On the day of insemination, a positive correlation was determined between serum PON-1 activity and the counts of total corpus luteum (r=0.398), total oocyte/embryo (r=0.468), transferable embryo (r=0.453), and Code I embryos (r=0.315, P<0.05). Unlike the Code I embryos, there was no significant correlation between serum PON-1 activity and the number of Code III embryos. Moreover, no significant difference in the number of Code III embryos between the two PON-1 groups was observed. However, embryo yield and quality were found to have increased with increased PON-1 activity. Therefore, it was concluded that serum PON-1 activity may be associated with superovulation response, embryo yield and quality in donor cows.

A novel route of follicle-stimulating hormone administration with a split-single ischiorectal fossa in Thai-Holstein crossbred superovulation programs under heat stress conditions.

The objectives of this study were to compare the efficiency of a split single injection of follicle-stimulating hormone (FSH) given by either intramuscular (split-single IM) or ischiorectal fossa (split-single IRF) injection to the traditional treatment and to determine the concentrations of FSH. The temperature and humidity index (THI) values were interpreted together with the ovarian responses and embryo characteristics. The ovarian responses in the split-single IRF group were similar to those of the control group (p > .05) but higher compared with the split-single IM group (p < .05). Higher peak levels of plasma FSH in the split-single IRF group did not differ compared with the control group (p > .05) but were lower in split-single IM administration (p < .05). The results showed a significant decrease in the numbers of large follicles and corpora lutea (CLs) in the moderate THI compared with low and high THI (p < .05). The high THI affected ovulation rate as well as the numbers of transferable embryos and degenerated embryos (p < .05). In conclusion, the split-single IRF administration had a comparable superovulatory response to the traditional twice-daily protocol. Moreover, the ovulation rate, ovarian follicle responses, and embryo quality were affected by heat stress.

145 Comparison of single to multiple injections of follicle-stimulating hormone before ovum pickup in Holstein heifers: Oocyte recovery and embryo production

The demand for invitro-produced embryos from heifers with high genetic merit has increased over time. Synchronization and stimulation of follicular growth before ovum pickup (OPU) has been used to improve oocyte quality and, consequently, embryo production. Multiple injections involve extra labour and stress for both personnel and cattle. The release of FSH can be prolonged by using 0.5% hyaluronan (HA) as a diluent, allowing a decrease in the number of injections. The objective of this study was to compare oocyte recovery and embryo production between single or multiple injections of FSH before OPU of Holstein heifers. During April and May 2020, 20 Holstein heifers (8 to 15 mo old) from Ruann Dairy (Riverdale, CA) were randomly divided and submitted to two different treatments (crossover design). Gonadotrophin-releasing hormone (GnRH; Fertagyl®, Merck, 129µg, IM) was given to synchronize the follicular wave emergence. Treatment 1×FSH consisted of a single intramuscular (IM) injection of 100mg of FSH (Folltropin®, Vetoquinol) 36h after GnRH. The FSH consisted of a 2.5-mL injection of 400mg of FSH diluted in 10mL of 0.5% HA. OPU was performed 48 to 50h after FSH. Treatment 5×FSH consisted of 100mg of FSH divided into 5 equal IM injections (10-14h intervals) 36h after GnRH. The FSH consisted of 5×1-mL injections of 400mg of FSH in 20mL of saline. OPU was performed 18 to 20h after the last FSH injection. All donors received both treatments at a 14-day interval and the recovered oocytes were fertilized with the same sexed female-sorted semen in both rounds. OPU, oocyte classification, IVM, IVF, and culture (IVC) were performed as described by Demetrio et al. (2020 Anim. Reprod. 17, e20200053). All oocytes went into IVM, except for degenerated oocytes. The number of 4-cell (or more) embryos on Day 3 of IVC divided by the number of oocytes in IVC after IVF is defined as the cleavage rate. The number of blastocysts (early to hatched) on Day 7 of IVP divided by the number of oocytes in IVC after IVF is defined as the blastocyst rate. Poisson-normal (count data) and Logistic-normal (proportion data) models were used to analyse the data. Treatment, donor (random effect), and sire were included in the models. The results are summarized in Table 1. There were no differences between the two treatments on the number of oocytes recovered per OPU (total and grade 1 and 2), percentage of grade 1 and 2 oocytes, cleavage rate, blastocyst rate and number of embryos (total and grade 1). Oocyte recovery and embryo production are highly donor dependent. Stimulation of the follicular growth before OPU with one single injection of FSH diluted in 0.5% HA 36h after GnRH can be efficiently used for IVP in Holstein heifers, without decreasing the number of oocytes recovered and/or embryos produced with the advantage of reducing labour and stress of handling cattle. Table 1. Number and quality of oocytes and cleavage and blastocyst rates Treatment OPU Oocytes per donor Grade 1 and 2 oocytes (%) Cleavage rate (%) Blastocyst rate (%) Total embryos per OPU Grade 1 embryos per OPU 1×FSH 20 17.0 45.7 84 39.8 6.2 3.8 5×FSH 20 19.9 46.5 82 35.6 6.3 4.0

146 Single injection of follicle-stimulating hormone before ovum pickup in lactating Holstein donors: Oocyte recovery and embryo production

Lactating donor cows frequently have decreased oocyte quality, lower fertilization rates, and impaired early embryonic development due to their lactational metabolic challenges. Synchronization and stimulation of follicular growth before ovum pickup (OPU) has been used to improve oocyte quality and consequently, embryo production. The objective of this study was to evaluate the effects of a single injection of FSH before OPU on oocyte recovery and embryo production in lactating Holstein donors. During June and July 2020, 22 lactating Holstein donors (open, 40 to 90 DIM, producing &amp;gt;90 lbs of milk) from Ruann Dairy (Riverdale, CA) were randomly assigned to one of two treatments (crossover design). Donors did not receive any injections before OPU when assigned to the No FSH treatment. Treatment 1×FSH consisted of a single intramuscular (IM) injection of 140mg of FSH (Folltropin®, Vetoquinol) 36h after gonadotrophin-releasing hormone (GnRH; Fertagyl, Merck®, 129µg, IM). The FSH consisted of a 3.5-mL IM injection of 400mg of FSH diluted in 10mL of 0.5% hyaluronan (HA). OPU was performed 48 to 52h after FSH. All donors received both treatments on a 14-day interval. The recovered oocytes were fertilized with the same sexed female-sorted semen in both rounds (3 different sires were used). OPU, oocyte classification, IVM, IVF, and culture (IVC) were performed as described by Demetrio et al. (2020 Anim. Reprod. 17, e20200053). All oocytes went into IVM, except for degenerated occytes. The number of 4-cell (or more) embryos on Day 3 of IVC divided by the number of oocytes in IVC after IVF is defined as the cleavage rate. The number of blastocysts (early to hatched) on Day 7 of IVP divided by the number of oocytes in IVC after IVF is defined as the blastocyst rate. Poisson-normal (count data) and Logistic-normal (proportion data) models were used to analyse the data. Treatment, donor (random effect), and sire were included in the models. The results are summarised in Table 1. Oocyte recovery and embryo production were highly donor dependent. There were no differences in the number of recovered oocytes among treatments. Stimulation of the follicular growth before OPU with one single injection of FSH diluted in 0.5% HA 36h after GnRH improved oocyte quality, cleavage rates, blastocyst rates, embryo quality, and the total number of embryos per OPU in lactating Holstein donors. Table 1. Oocyte recovery, cleavage rate, and embryo production results Treatment OPU Oocytes per donor Grade 1 and 2 oocytes Cleavage rate (%) Blastocyst rate (%) Total embryos per OPU Grade 1 embryos per OPU No FSH 22 18.2 30a 69a 22a 3.4a 1.6a 1×FSH 22 17.5 34b 77b 34b 5.3b 3.3b a,bValues with different superscripts in the same column differ (at least P&amp;lt;0.05).

Negative impact of high doses of follicle-stimulating hormone during superovulation on the ovulatory follicle function in small ovarian reserve dairy heifers†.

When women with small ovarian reserves are subjected to assisted reproductive technologies, high doses of gonadotropins are linked to high oocyte and embryo wastage and low live birth rates. We hypothesized that excessive follicle-stimulating hormone (FSH) doses during superovulation are detrimental to ovulatory follicle function in individuals with a small ovarian reserve. To test this hypothesis, heifers with small ovarian reserves were injected twice daily for 4 days, beginning on Day 1 of the estrous cycle with 35, 70, 140, or 210IU doses of Folltropin-V (FSH). Each heifer (n=8) was superovulated using a Williams Latin Square Design. During each superovulation regimen, three prostaglandin F2α injections were given at 12-h interval, starting at the seventh FSH injection to regress the newly formed corpus luteum (CL). Human chorionic gonadotropin was injected 12h after the last (8th) FSH injection to induce ovulation. Daily ultrasonography and blood sampling were used to determine the number and size of follicles and corpora lutea, uterine thickness, and circulating concentrations of estradiol, progesterone, and anti-Müllerian hormone (AMH). The highest doses of FSH did not increase AMH, progesterone, number of ovulatory-size follicles, uterine thickness, or number of CL. However, estradiol production and ovulation rate were lower for heifers given high FSH doses compared to lower doses, indicating detrimental effects on ovulatory follicle function.

Cumulative outcome of pre-implantation genetic diagnosis for sickle cell disease: a 5-year review.

To review the cumulative outcome of pre-implantation genetic diagnosis (PGD) cycles performed for prevention of sickle cell disease (SCD). Couples referred for PGD for SCD between April 2012 and October 2017 were included. Ovarian stimulation was performed using a short gonadotrophin-releasing hormone (GnRH) antagonist protocol and follicle-stimulating hormone injections. The GnRH agonist was used to trigger oocyte maturation. Oocytes were fertilised using intracytoplasmic sperm injection. Trophectoderm biopsy was performed on day 5 or 6 followed by vitrification. Genetic testing was done using pre-implantation genetic haplotyping. A total of 60 couples started 70 fresh PGD cycles (mean 1·2cycles/couple) and underwent a total of 74 frozen-embryo-transfer (FET) cycles (mean 1·3FET/couple). The mean (SD) female age was 33(4·4)years and the mean (SD) anti-müllerian hormone level was 22·9(2·8)pmol/l. The cumulative live-birth rate was 54%/PGD cycle started and 63%/couple embarking on PGD. The rate of multiple births was 8%. The cumulative outcome of PGD treatment for prevention of SCD transmission is high and PGD treatment should be offered to all at-risk couples.

Real-time assessment of the superovulatory effect of FSH and eCG with laparoscopy at different seasons in Akkaraman ewes.

Conventional methods for determining the reproductive performance of sheep bred either after estrus synchronization during the breeding season or after induction of estrus/ovulation during the non-breeding season take a long time and may give misleading results due to the effect of environmental factors. Laparoscopic observations allow real-time monitoring of ovarian activity around estrus or ovulation. This study was aimed at assessing the superovulatory effects of follicle-stimulating hormone (FSH) and equine chorionic gonadotropin (eCG) treatments by laparoscopy during breeding (September-November, n=12) and non-breeding (April-June, n=12) seasons in Akkaraman sheep. In both seasons, after CIDR withdrawal, the ewes were injected either with 600 IU eCG or 300 μl (20 mg/ml) FSH twice at 12 hour intervals. Plasma P4, E2 and LH concentrations were determined at the time of intra-vaginal CIDR insertion (day 0) and then at its withdrawal (day 12), followed by 3 and 6 days of eCG or FSH injections. After 3 (first observation) and 6 (second observation) days of hormone injections, laparoscopy was performed to record ovarian activity in both seasons. The eCG increased (p⟨0.05) the numbers of large follicles (first observation) and CL (first and second observations) in the breeding season compared to FSH treatment. CL, small-moderate and large follicle numbers of eCG treated ewes were higher (p⟨0.05) than those of FSH at both observations in the non-breeding season. In the breeding season, eCG treated ewes had higher (p⟨0.05) plasma P4 (3 and 6 days after hormones injections) and E2 (3 days after hormones injections) concentrations than those of FSH. In conclusion, the results of the present study indicate that treatment with eCG during the non-breeding season can support ovarian activity, and thus increase ovulation rate and plasma hormone concentrations around induced estrus/ovulation in Akkaraman ewes.

The hypothalamic-pituitary-gonadal axis and thyroid hormone regulation interact to influence seasonal breeding in green anole lizards (Anolis carolinensis)

Reproductive physiology and behavior is mainly regulated by the hypothalamus-pituitary-gonad (HPG) axis, although abnormal thyroid hormone (TH) levels alter HPG axis activity. Seasonally breeding animals, such as green anole lizards (Anolis carolinensis), undergo drastic hormonal and behavioral changes between breeding and non-breeding seasons, with increased sex steroid hormones, larger gonads and increased reproductive behaviors during the breeding compared to non-breeding seasons. Relatively less is known regarding the regulation of gonadal TH in seasonal reproduction. We examined whether the gonadal expression of enzymes involved in TH activation are altered in concert with seasonal reproduction. Type 2 deiodinase (Dio2) mRNA, the TH activating enzyme, was upregulated in breeding compared to non-breeding testes, while type 3 deiodinase (Dio3) mRNA, the TH deactivating enzyme, was upregulated in breeding ovaries. To study the association between the HPG axis and local activation of TH, we manipulated the HPG axis during the non-breeding season by subcutaneously injecting luteinizing hormone (LH) and follicle stimulating hormone (FSH) in male lizards. We found that acute LH and FSH injections induced many aspects of breeding, with increased testes size and testosterone levels. Surprisingly, Dio3 was upregulated in the testes after LH and FSH injections, while Dio2 mRNA levels were unchanged. These results suggest that there might be different roles for local TH activation in developing and maintaining fully mature and functional gonads. Our findings continue to support the role for TH in regulating reproduction.

Follicle Stimulating Hormone and Gonadotropin Releasing Hormone Administration to the Superovulation of Buffalo (Bubalus bubalis)

This study aims to determine the response of superovulation, corpus luteum number, onset and duration of estrus in Swamp Buffalo after administration of Follicle Stimulating Hormone (FSH) and Gonadotropin Releasing Hormone (GnRH). The material used is 16 mother Swamp Buffaloes with an average body weight of 500 kg and age between 3-5 years. The dairy performance index such as the lactation period was 127.5±104.6 days, the milk production accounted for 2.0±0.9 kg/cow/day and the milk yield was 255±209 kg/lactation. The experimental design used was a Completely Randomized Design (CRD) consisting of four replications. Swamp buffaloes are synchronized with 5 mL of PGF2α injection. GnRH hormone injection on the 8th day at a dose of 500 µg with 2 times in the morning and evening. FSH injection begins on the 10th day with 2 times a day i.e., morning and night (12 h interval) with a dose decreased for 3 days intra-muscularly. Artificial insemination was carried out after signs appear estrus. The parameters observed include the superovulation response, the number of corpus luteum, the onset and duration of estrus in the Swamp Buffalo. The results obtained are the superovulation response in 56.25% Swamp Buffalo. The average number of corpus luteum, speed and duration of swamp buffalo estrus at different FSH doses and addition of 500 μg GnRH were 1.75, 20.25 and 145.75 h respectively. It was concluded that superovulation by administering a dose of 16 ml FSH + 500 μg GnRH has shown the best superovulation response and a long duration of estrus. The administration of 22 mL FSH dose and the addition of 500 μg GnRH showed the fastest onset of estrus.