Ovarian reserve tests and their utility in predicting response to controlled ovarian stimulation in rhesus monkeys

Abstract

Controlled ovarian stimulation (COS) is an alternative to natural breeding in nonhuman primates; however, these protocols are costly with no guarantee of success. Toward the objective of predicting COS outcome in rhesus monkeys, this study evaluated three clinically used ovarian reserve tests (ORTs): day 3 (d3) follicle-stimulating hormone (FSH) with d3 inhibin B (INHB), the clomiphene citrate challenge test (CCCT), and the exogenous FSH Ovarian Reserve Test. A COS was also performed and response was classified as either successful (COS+) or unsuccessful (COS-) and retrospectively compared with ORT predictions. FSH and INHB were assessed for best hormonal index in conjunction with the aforementioned tests. INHB was consistently more accurate than FSH in all the ORTs used. Overall, a modified version of the CCCT using INHB values yielded the best percentage of correct predictions. This is the first report of ORT evaluation in rhesus monkeys and may provide a useful diagnostic test before costly follicle stimulations, as well as predicting the onset of menopause.

Figure 1. Representative hormone profiles of a controlled ovarian stimulation (COS)

A) COS protocol: onset of menses designated day 1; Antide administered on d1–9; r-hFSH administered on d1–6; r-hFSH and r-hLH administered on d6–9; hCG injected on d10 in induce ovulatory maturation of follicles. B) Representative estradiol (E2) profile of a successful COS (COS+) with accompanying ultrasound. C) Representative E2 profile of an unsuccessful COS (COS−) with accompanying ultrasound (numbers on the ultrasound denote follicular measurements).

Figure 2. Day 3 Hormone Values

A) Protocol for day 3 hormone values: blood was collected from the saphenous vein on menses day 3 and analyzed for FSH and INHB. B) Day FSH: number of correct and incorrect COS outcome predictions with overall percent accuracy indicated (59%). Day 3 FSH predictions did not significantly predict (X2 > 3.84; df=1; P > 0.05) actual COS outcomes. C) Day 3 INHB: number of correct and incorrect COS outcome predictions with overall percent accuracy indicated (77%). Day 3 INHB predictions did not significantly predict (P = 0.06) actual COS outcomes.

Figure 3. Clomiphene Citrate Challenge Test (CCCT)

A) Protocol for CCCT: blood was collected from the saphenous vein on menses day 3 and day 10 and analyzed for FSH and INHB.. 50 mg clomiphene citrate was administered daily on menses days 5–9. B) CCCT using FSH values: number of correct and incorrect COS outcome predictions with overall percent accuracy indicated (45%). CCCT (FSH) predictions did not significantly predict (X2 > 3.84; df=1; P > 0.05) actual COS outcomes. C) CCCT using INHB values: number of correct and incorrect COS outcome predictions with overall percent accuracy indicated (59%). CCCT (INHB) predictions did not significantly predict (p > 0.05) actual COS outcomes.

Figure 4. Modified Clomiphene Citrate Challenge Test (M-CCCT)

A) Flowchart of protocol for M-CCCT: Day 3 FSH or INHB served as an initial screen; FSH values ≤ 1.0 ng/mL or INHB values ≥ 50 pg/mL were excluded from further testing and predicted for a successful COS (COS+). Animals failing to meet the day 3 hormone criteria for a COS+ were further tested by the CCCT using fold change in E2 to determine COS+ or COS−. B) M-CCCT using FSH: number of correct and incorrect COS outcome predictions with overall percent accuracy indicated (77%). M-CCCT (FSH) predictions significantly predicted (X2 = 3.84 or 5.99; P < 0.05; df=1 or 2 respectively) actual COS outcomes. C) M-CCCT using INHB: number of correct and incorrect COS outcome predictions with overall percent accuracy indicated (82%). M-CCCT (INHB) predictions significantly predicted (X2 = 3.84 or 5.99; P < 0.05; df=1 or 2 respectively) actual COS outcomes.

Figure 5. Exogenous Follicle-Stimulating Hormone Ovarian Reserve Test (EFORT)

A) Protocol for EFORT: blood was collected from the saphenous vein on menses day 1 and day 2 and analyzed for FSH and INHB. An intra-muscular injection of recominant human FSH (r-hFSH) was administered following the blood sample on day 1. B) EFORT using FSH values: number of correct and incorrect COS outcome predictions with overall percent accuracy indicated (55%). EFORT (FSH) predictions did not significantly predict (X2 > 3.84; df=1; P > 0.05) actual COS outcomes. C) EFORT using INHB values: number of correct and incorrect COS outcome predictions with overall percent accuracy indicated (80%). CCCT (INHB) predictions did not significantly predict (X2; df=1; P = 0.06) actual COS outcomes.