Laura K. Arbogast

Follicular fluid vascular endothelial growth factor concentrations are elevated in women of advanced reproductive age undergoing ovulation induction

OBJECTIVE(S) To determine whether follicular fluid (FF) from women of advanced reproductive age had a relative deficiency of the angiogenic cytokine vascular endothelial growth factor/vascular permeability factor. Furthermore, we sought to determine whether luteinized granulosa cells secrete vascular endothelial growth factor/vascular permeability factor in response to hypoxia. DESIGN Retrospective cohort study. SETTING University teaching hospital. PATIENTS Women undergoing follicular aspiration after superovulation in preparation for IVF-ET. Women of advanced reproductive age consisted of 21 women > or = 38 years old (range, 38 to 46 years); 15 subjects < or = 30 years served as the control population. INTERVENTION(S) Granulosa cells and FF were collected by transvaginal aspiration 35 hours after hCG. Granulosa cells from two women were cultured for 24 and 48 hours in M199 + 10% fetal bovine serum in 1% O2-5% CO2-94% N2 (hypoxic) or 95% air-5% CO2 (normoxic) without or with 0.1 mol/L cobalt chloride. MAIN OUTCOME MEASURE(S) Pooled FF vascular endothelial growth factor/vascular permeability factor concentrations and media vascular endothelial growth factor/vascular permeability factor accumulation at 24 and 48 hours were determined. RESULT(S) Follicular fluid vascular endothelial growth factor/vascular permeability factor concentrations were higher in advanced reproductive age women compared with younger women (3,735 +/- 2,155 versus 2,205 +/- 952 pg/mL, mean +/- SD). Accumulation of vascular endothelial growth factor/vascular permeability factor at 24 and 48 hours was 391 +/- 54 and 744 +/- 2 pg/mL in media maintained in 5% CO2 and air. Cobalt chloride induced a marked increase in vascular endothelial growth factor/vascular permeability factor (2,008 +/- 52 pg/mL at 24 hours and 3,630 +/- 519 pg/mL at 48 hours). An intermediate but significant increase in vascular endothelial growth factor/vascular permeability factor (733 +/- 35 pg/mL at 24 hours and 2,675 +/- 864 pg/mL at 48 hours) was observed with 1% O2 compared with normoxic controls. CONCLUSION(S) After hMG and hCG administration the FF from women of advanced reproductive age showed increased vascular endothelial growth factor/vascular permeability factor concentrations compared with younger women. Increased vascular endothelial growth factor/vascular permeability factor concentrations could be consistent with a hypoxic environment within follicles of older women.

Vascular Endothelial Growth Factor Stimulates Preantral Follicle Growth in the Rat Ovary

Abstract The regulation of preantral follicle growth in mammals is poorly understood. The availability of an adequate vascular supply to provide endocrine and paracrine signals may be important during the early states of follicle growth as well as the later states of follicle selection and dominance. The objective of the present study was to investigate whether vascular endothelial growth factor (VEGF) plays a role in preantral follicular development in the rat ovary. Immature (age, 21 days) Sprague-Dawley rats were injected with 500 ng of VEGF in saline or 50 μg of diethylstilbestrol (DES) in oil under the bursa of one ovary. The contralateral ovary was injected with a corresponding volume of vehicle. Rats were killed 48 h later, and the ovaries were removed and analyzed histologically. Intrabursal administration of VEGF significantly increased the number of primary and small secondary, but not of large secondary, preantral follicles in the ovary, similar to the effect of DES (P < 0.05). The VEGF stimulated preantral follicle growth in a time- and dose-dependent manner. Subcutaneous DES administration increased the number of primary and secondary follicles, and both s.c. and intrabursal estrogen administration stimulated VEGF protein expression in the rat ovary. These data indicate that VEGF stimulates preantral follicular development in the rat ovary, is regulated by estrogen, and may be one of the factors that participate in the regulation of early follicle growth in the rat.

Elevated level of follicular fluid vascular endothelial growth factor is a marker of diminished pregnancy potential

OBJECTIVE To evaluate whether differences in follicular fluid vascular endothelial growth factor (FF VEGF) concentrations are observed between women achieving a clinical pregnancy and those failing to conceive. DESIGN Retrospective chart review and analysis of FF VEGF concentrations. SETTING University teaching center. PATIENT(S) Fifty-seven women < or =42 years of age undergoing follicular aspiration in preparation for IVF or GIFT. INTERVENTION(S) Analysis of FF VEGF concentrations and chart review of a single IVF or GIFT cycle. MAIN OUTCOME MEASURE(S) Follicular fluid VEGF concentrations, clinical pregnancy rate, age, ampules of gonadotropins used, oocytes retrieved, peak estradiol serum concentrations, day 3 FSH levels, and fertilization rate. RESULT(S) Women who did not conceive had higher FF VEGF concentrations than women achieving a clinical pregnancy (4.409 + 2,387 versus 2.793 +/- 1,180 pg/mL: P < .001). A negative correlation was observed between FF VEGF concentrations and peak estradiol levels and number of oocytes retrieved. A positive correlation was found for FF VEGF and patients age and ampules of gonadotropins used. CONCLUSION(S) Elevated FF VEGF concentrations are associated with poor conception rates after IVF or GIFT.

Neutralization of endogenous vascular endothelial growth factor depletes primordial follicles in the mouse ovary

Abstract The regulation of early follicular growth and development involves a complex interaction of autocrine, paracrine, and endocrine signals. The ability of these factors to regulate follicle growth may depend in part on the extent of vascular delivery to and perfusion of the ovary. Vascular endothelial growth factor A (VEGFA) is a major regulator of vascular physiology in the ovary. VEGFA is produced in numerous ovarian compartments and likely plays a role in the regulation of all phases of follicular growth, from preantral through preovulatory. The aim of the present study was to further evaluate the role of VEGF in early follicle growth by neutralization of endogenous VEGF or VEGF receptors. Adult mice were injected systemically and prepubertal mice were injected directly under the ovarian bursa with antibodies designed to neutralize VEGF or block interaction with its receptors in the ovary. Both systemic and intrabursal injections of VEGF antibody significantly reduced the number of primordial follicles within 1–3 days after administration without affecting primary or secondary follicle numbers. Primordial follicle numbers were not different from control levels by 30 days after VEGFA antibody administration. Administration of antibodies to the kinase domain receptor (KDR), but not the FMS-like tyrosine receptor (FLT1), for VEGF also resulted in a significant decrease in primordial follicles. These data suggest that VEGF plays a vital role in the maintenance and growth of the primordial follicle pool.

Identification of gonadotrophin surge-inhibiting factor (GnSIF)/attenuin in human follicular fluid

Evidence from several laboratories suggests that the ovaries of many species produce a non-steroidal factor called gonadotrophin surge-inhibiting or attenuating factor (GnSIF) which may regulate the response of the pituitary to gonadotrophin-releasing hormone (GnRH) and as such, may modulate the timing and/or amplitude of the luteinizing hormone (LH) surge. We have recently isolated a candidate GnSIF from porcine follicular fluid (PFF). Porcine GnSiF is a 69 kDa protein which has undetectable inhibin and follistatin immunological and biological activity. The present study was designed to purify and identify GnSIF from human follicular fluid. GnSIF activity was measured as suppression of GnRH-stimulated LH secretion from rat pituitary cells in primary culture. Human follicular fluid (approximately 500 ml) was recovered from patients undergoing in-vitro fertilization (IVF). GnSIF was purified by heparin-sepharose, Q-sepharose, S-sepharose, and hydrophobic interaction chromatography followed by isoelectric focusing. Gel electrophoresis and Western blot were used to identify human GnSIF and compare it with porcine GnSIF. Using these steps, we obtained a highly-purified preparation of GnSIF that manifests in-vitro bioactivity and chromatography characteristics similar to those observed for porcine GnSIF and that hybridizes with a porcine GnSIF antibody. Following treatment with human chorionic gonadotrophin/human menopausal gonadotrophin (HMG/HCG), human follicular fluid contained roughly 25% of the GnSIF (per mg protein) present in porcine follicular fluid. We conclude that GnSIF is present in human follicular fluid and may participate in the regulation of gonadotrophin secretion in this species.