Effects of Extra-Long-Acting Recombinant Bovine FSH (bscrFSH) on Cattle Superovulation

Over the last few years, several commercial FSH products have been developed for cattle superovulation (SOV) purposes in Multiple Ovulation and Embryo Transfer (MOET) programs. The SOV response is highly variable among individuals and remains one of the main limiting factors in obtaining a profitable number of transferable embryos. In this study, follicle stimulating hormone (FSH) from different origins was included in two SOV protocols, (a) FSH from purified pig pituitary extract (NIH-FSH-p; two doses/day, 12 h apart, four consecutive days); and (b) extra-long-acting bovine recombinant FSH (bscrFSH; a single dose/day, four consecutive days), to test the effects of bscrFSH on the ovarian response, hormone profile levels, in vivo embryo production and the pluripotency gene expression of the obtained embryos. A total of 68 healthy primiparous red Angus cows (Bos taurus) were randomly distributed into two experimental groups (n = 34 each). Blood sample collection for progesterone (P4) and cortisol (C) level determination was performed together with ultrasonographic assessment for ovarian size, follicles (FL) and corpora lutea (CL) quantification in each SOV protocol (Day 0, 4, 8, and 15). Moreover, FSH profiles were monitorised throughout both protocols (Day 0, 4, 5, 6, 7, 8, 9, 10, and 15). In vivo embryo quantity and quality (total structures, morulae, blastocysts, viable, degenerated and blocked embryos) were recorded in each SOV protocol. Finally, embryo quality in both protocols was assessed by the analysis of the expression level of crucial genes for early embryo development (OCT4, IFNt, CDX2, BCL2, and BAX). P4 and cortisol concentration peaks in both SOV protocols were obtained on Day 15 and Day 8, respectively, which were statistically different compared to the other time-points (p < 0.05). Ovarian dimensions increased from Day 0 to Day 15 irrespective of the SOV protocol considered (p < 0.05). Significant changes in CL number were observed over time till Day 15 irrespective of the SOV protocol applied (p < 0.05), being non- significantly different between SOV protocols within each time-point (p > 0.05). The number of CL was higher on Day 15 in the bscrFSH group compared to the NIH-FSH-p group (p < 0.05). The number of embryonic structures recovered was higher in the bscrFSH group (p = 0.025), probably as a result of a tendency towards a greater number of follicles developed compared to the NIH-FSH-p group. IFNt and BAX were overexpressed in embryos from the bscrFSH group (p < 0.05), with a fold change of 16 and 1.3, respectively. However, no statistical differences were detected regarding the OCT4, CDX2, BCL2, and BCL2/BAX expression ratio (p > 0.05). In conclusion, including bscrFSH in SOV protocols could be an important alternative by reducing the number of applications and offering an improved ovarian response together with better embryo quality and superior performance in embryo production compared to NIH-FSH-p SOV protocols.

Thyroid blood profile in high-yielding dairy cows with different ovarian reactions to superovulatory treatment

The reaction of cow ovaries to superovulatory treatment significantly depends on the physiological state of the animal, primarily on its endocrine status. The available information indicates that thyroid hormones can affect the processes of ovarian folliculogenesis in cattle.Objective: To study the thyroid profile in the blood of high-yielding dairy cows in relation to the response of the ovaries to superovulatory treatment.Materials and methods. Thirty four cycles of superovulation stimulation in 16 Holstein cows were studied. After synchronization of the estrous cycle and superovulatory treatment, on the first day of insemination (Day 0), the number of follicles with a diameter of more than 5 mm was recorded in each ovary using an ultrasound scanner. Furthermore, on Day 0 and on the 7th day after insemination (Day 7), the blood was taken from the cows to analyze the content of sex steroids and thyroid hormones by ELISA. The ovarian reaction was assessed by the total number of detected follicles and was divided into 3 types: I – more than 12 follicles (strong), II – 8-12 follicles (moderate), III – less than 8 follicles (weak).Results. The content of sex steroid hormones in the blood of cows on Day 0 did not depend on the type of the ovarian response. By Day 7, the concentration of progesterone increased in most animals, but was 2.7-fold higher (P <0.01) in the case of the strong reaction (type I) than in the case of the weak reaction (type III). Conversely, the serum concentration of estradiol-17β decreased 1.1 times (P <0.05) in cows with the moderate ovarian response (type II). In animals with the type I reaction, the content of thyroxine (T4) in the blood declined 1.2 times (P <0.05) one week after insemination. During the study period, there was also a reduction in the concentration of total triiodothyronine (T3) and reverse T3 (rT3) in cows with ovarian responses of types I and II (1.1-1.3 times, P <0.01-0.05). Concurrently, on Day 0, the T3 content in the blood of individuals with the type II reaction was 1.2-fold higher (P <0.05) than in individuals with the type III reaction. Totally, a positive relationship was found between the concentration of estradiol-17β and the concentration of thyroid hormones: for T4 – in the case of the type II ovarian response (r = 0.44, P <0.05), for T3 – in the case of the type I and II responses ( r = 0.57-0.64, P <0.001-0.05), for rT3 – in the case of the type III response (r = 0.52, P <0.01).Conclusions. Thus, the content of triiodothyronine in the blood of high-yielding dairy cows on the day of insemination is associated with the ovarian response to superovulatory treatment. In addition, during one week after insemination, the state of the thyroid system and its relationship with the level of estradiol-17β in the blood differ with different reactions of the ovaries to the introduction of exogenous gonadotropic hormones.

Poor ovarian response and the possible role of natural and modified natural cycles

About 20% of women undergoing in vitro fertilization struggle with poor ovarian response, indicating a poor prognosis related to low response following ovarian stimulation. Indeed, poor ovarian response, that is associated with both high cancelation rates and low live birth rates, still represents one of the most important therapeutic challenges in in vitro fertilization. In this context, natural cycle/modified natural cycle– in vitro fertilization, as a ‘milder’ approach, could be a reasonable alternative to high-dose/conventional ovarian stimulation in poor ovarian responders, with the aim to retrieve a single oocyte with better characteristics that may result in a single top-quality embryo, transferred to a more receptive endometrium. Moreover, modified natural cycle– in vitro fertilization may be cost-effective because of the reduced gonadotropin consumption. Several studies have been published during the last 20 years reporting conflicting results regarding the use of natural cycle/modified natural cycle– in vitro fertilization in women with poor ovarian response; however, while most of the studies concluded that mild stimulation regimens, including natural cycle/modified natural cycle– in vitro fertilization, have low, but acceptable success rates in this difficult group of patients, others did not replicate these findings. The aim of this narrative review is to appraise the current evidence regarding the use of natural cycle/modified natural cycle– in vitro fertilization in poor ovarian responders.

The two-step process of ovarian follicular growth and maturation in mammals can be compared to a fruit ripening where quality depends on the second step

In human IVF, the main uncertainty factor impacting on success is oocyte quality, which largely depends on the follicular status at the time of collection. Decades of debate ensued to find the perfect stimulation protocol demonstrated the complexity of the ovarian response to exogenous gonadotropins and the dynamic nature of late folliculogenesis. Although several follicular markers, proteins, RNA from granulosa cells or microRNA and follicular fluid metabolites have been associated with outcome, the possibility to influence them during stimulation remains elusive. The heterogeneity of the follicle’s maturity following control ovarian stimulation is also an important factor to explain average poor oocyte quality still observed today. In this review, the analogy between the apple ripening on the tree and follicular development is presented to focus the attention on a biphasic process: growth and differentiation. The molecular analysis of the progressive follicular differentiation indicates 2 competing phenomena: growth and differentiation where a delicate balance must operate from one to the other to ensure proper maturity at ovulation. As long as FSH stimulates growth, follicles remain green, and it is only when FSH is replaced by LH that the ripening process begins, and “apples” become red. Both fruits, follicles and apples, depend on a perfect timing of events to generate offspring.

What Does Unexpected Suboptimal Response During Ovarian Stimulation Suggest, an Overlooked Group?

Unlike poor ovarian response, despite being predicted to be normal responders based on their ovarian reserve markers, many patients respond suboptimally to ovarian stimulation. Although we can improve the number of retrieved oocytes by increasing the recombinant FSH dose and adding LH, the effect of suboptimal ovarian response on cumulative live birth rate (CLBR) and offspring safety is unclear. This study focuses on the unexpected suboptimal response during ovulation induction, and its causes and outcomes are analysed for the first time with a large amount of data used to compare the cumulative pregnancy rate (CPR), CLBR and offspring safety of patients with one complete ART cycle with all embryos used. Our analysis included 5218 patients treated with the GnRH agonist long protocol for their first IVF–embryo transfer (ET) cycles. Patients were divided into two groups according to whether the ovarian response was suboptimal. Propensity score matching (PSM) was utilized for sampling at up to 1:1 nearest-neighbour matching with caliper 0.05 to balance the baseline and improve comparability between the groups. Results showed that age, BMI and basal FSH were independent risk factors for slow response; the initial dosage of Gn, FSH on the first day of Gn, and LH on the first day of Gn were independent protective factors for suboptimal response. Suboptimal responders were also more likely to have irregular menses. Regarding the clinical pregnancy rate of the fresh IVF/ICSI-ET cycles, the adjusted results of the two groups were not significantly different. There was no difference in the CPR, CLBR, or offspring safety-related data, such as gestational age, preterm delivery rate, birthweight, birth-height and Apgar Scores between the two groups after PSM. Age-related changes in the number of oocytes retrieved from women aged 20–40 years old between the two groups were different, indicating that suboptimal response in elderly patients suggests a decline in ovarian reserve. Although we can now improve the outcomes of suboptimal responders, it increases the cost to the patients and the time to live birth, which requires further attention.

Effect of the Trends of Estradiol Level on the Outcome of in Vitro Fertilization-embryo Transfer With Antagonist Regimens: a Single Center Retrospective Cohort Study

Background: The outcome of in vitro fertilization-embryo transfer is often determined according to follicles and estradiol levels following gonadotropin stimulation. However, there is no accurate indicator to predict pregnancy outcome, and it has not been determined how to choose subsequent drugs and dosage based on the ovarian response. This study aimed to make timely adjustments to follow-up medication to improve clinical outcomes based on the potential value of estradiol growth rate.Methods: Serum estradiol levels were measured on the day of gonadotrophin treatment (Gn0), four days later (Gn4), seven days later (Gn7), and on the trigger day (HCG). The ratio was used to determine the increase in estradiol levels. According to the ratio of estradiol increase, the patients were divided into four groups: group A1 (Gn4/Gn0≤6.44), group A2 (6.44˂Gn4/Gn0≤10.62), group A3 (10.62˂Gn4/Gn0≤21.33), and group A4 (Gn4/Gn0>21.33); group B1 (Gn7/Gn4≤2.39), group B2 (2.39˂Gn7/Gn4≤3.03), group B3 (3.03˂Gn7/Gn4≤3.84), and group B4 (Gn7/Gn4>3.84). We analyzed and compared the relationship between data in each group and pregnancy outcome.Results: In the statistical analysis, the estradiol levels of Gn4 (P = 0.029, P = 0.042), Gn7 (P< 0.001, P = 0.001), and HCG (P< 0.001, P = 0.002), as well as the ratios of Gn4/Gn0 (P = 0.004, P = 0.006), Gn7/Gn4 (P = 0.001, P = 0.002), and HCG/Gn0 (P< 0.001, P< 0.001) both had clinical guiding significance, and the lower one significantly reduced the pregnancy rate. The outcomes were positively linked to groups A (P = 0.040, P = 0.041) and B (P = 0.015, P = 0.017). The logistical regression analysis revealed that group A1 (OR = 0.440 [0.223–0.865]; P = 0.017, OR = 0.368 [0.169–0.804]; P = 0.012) and B1 (OR = 0.261 [0.126–0.541]; P< 0.001, OR = 0.299 [0.142–0.629]; P = 0.001) had opposite influence on outcomes.Conclusion: Maintaining a serum estradiol increase ratio at least above 2.39 on Gn7/Gn4 may result in a higher pregnancy rate. When estradiol growth is not ideal, gonadotrophin dosage should be adjusted appropriately to ensure the desired outcome.

Growth Hormone Cotreatment for Low-Prognosis Patients According to the POSEIDON Criteria

BackgroundPoor ovarian response (POR) remains one of the most challenging conditions in assisted reproduction technology. Previous studies seemed to indicate that growth hormone (GH) was a potential solution for the dilemma of POR; however, the role GH played on the low-prognosis patients diagnosed and stratified by the POSEIDON criteria remains indistinct.MethodsThis retrospective study was performed among women with POR according to the POSEIDON criteria who failed a previous in vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI) cycle, and the subsequent cycle was under GH cotreatment and conducted within 12 months. These participants were stratified into four groups according to the POSEIDON criteria. The comparison was implemented between the failed cycle and the cycle treated with GH. Generalized estimating equation (GEE) multivariate regression was applied for data analysis.ResultsA total of 428 low-prognosis women were included in this study. GH supplementation improved the live birth rates (47.66%, 28.33%, 45.45%, and 24.07%; in groups 1, 2, 3, and 4, respectively) and the clinical pregnancy rates (OR 19.16, 95% CI 7.87–46.63, p &lt; 0.001; OR 7.44, 95% CI 1.65–33.55, p = 0.009; OR 10.19, 95% CI 2.39–43.52, p = 0.002; OR 27.63, 95% CI 4.46–171.11, p &lt; 0.001; in groups 1, 2, 3, and 4, respectively) in all four POSEIDON groups. The number of oocytes retrieved was significantly elevated in the subgroups with normal ovarian reserve (IRR 1.47, 95% CI 1.36–1.59, p &lt; 0.001; IRR 1.31, 95% CI 1.15–1.49, p &lt; 0.001; in groups 1 and 2, respectively). The number of day-3 good-quality embryos was significantly elevated in the subgroups with either normal ovarian reserve or aged young (IRR 2.13, 95% CI 1.78–2.56, p &lt; 0.001; IRR 1.54, 95% CI 1.26–1.89, p &lt; 0.001; IRR 1.47, 95% CI 1.10–1.98, p = 0.010; in groups 1, 2, and 3, respectively).ConclusionGrowth hormone cotreatment could ameliorate the pregnancy outcome for women with POR under the POSEIDON criteria who failed a previous IVF/ICSI cycle. The application of growth hormone for low-prognosis women who experienced a failed cycle might be considered and further studied.