help button home button Endocrine Society JCEM JCEM Call for Nominations for EIC
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
Journal of Clinical Endocrinology & Metabolism, doi:10.1210/jc.2006-2161
This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow All Versions of this Article:
92/4/1438    most recent
Author Manuscript (PDF)
Right arrow Submit a related Letter to the Editor
Right arrow Purchase Article
Right arrow View Shopping Cart
Right arrow Alert me when this article is cited
Right arrow Alert me when eLetters are posted
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow Request Copyright Permission
Citing Articles
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Dumesic, D. A.
Right arrow Articles by Abbott, D. H.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Dumesic, D. A.
Right arrow Articles by Abbott, D. H.
Related Collections
Right arrow Female Endocrinology
Right arrow Obesity
The Journal of Clinical Endocrinology & Metabolism Vol. 92, No. 4 1438-1441
Copyright © 2007 by The Endocrine Society

BRIEF REPORT

Increased Adiposity Enhances Intrafollicular Estradiol Levels in Normoandrogenic Ovulatory Women Receiving Gonadotropin-Releasing Hormone Analog/Recombinant Human Follicle-Stimulating Hormone Therapy for in Vitro Fertilization

Daniel A. Dumesic, Timothy G. Lesnick and David H. Abbott

National Primate Research Center (D.H.A., D.A.D.), University of Wisconsin, Madison, Wisconsin 53715; Department of Obstetrics/Gynecology (D.H.A.), University of Wisconsin, Madison, Wisconsin 53792; Reproductive Medicine and Infertility Associates (D.A.D.), Woodbury, Minnesota 55125; and Department of Biostatistics (T.G.L.), Mayo Clinic, Rochester, Minnesota 55905

Address all correspondence and requests for reprints to: Daniel A. Dumesic, M.D., Reproductive Medicine and Infertility Associates, 2101 Woodwinds Drive, Woodbury, Minnesota 55125. E-mail: danieldumesic{at}aol.com.


   Abstract
Top
 Abstract
Introduction
 Subjects and Methods
 Results
 Discussion
 References
 
Context: Body mass index (BMI) reflects the amount of insulin in the human follicle and may enhance insulin action as a cogonadotropin.

Objective: This study examined whether increased adiposity enhances intrafollicular steroidogenesis in normoandrogenic ovulatory women receiving GnRH analog/recombinant human FSH therapy for in vitro fertilization.

Design, Setting, and Participants: Study participants were from an institutional practice and comprised 30 normoandrogenic ovulatory women who were lean (n = 17; BMI < 25 kg/m2) or overweight (n = 13; BMI ≥ 25 kg/m2). Women received GnRH analog after basal serum hormone determinations and oral glucose tolerance testing, followed by recombinant human FSH therapy and human chorionic gonadotropin administration when two or more follicles 18 mm or larger in diameter were present.

Intervention: Follicle fluid was aspirated at oocyte retrieval from the first follicle of each ovary.

Main Outcome Measures: Follicle fluid was assayed for estradiol (E2), progesterone, 17-hydroxyprogesterone, androstenedione, testosterone, dihydrotestosterone, insulin, glucose, and lactate.

Results: Overweight women had hyperinsulinemia (P = 0.03) with decreased serum SHBG (P = 0.001) and increased serum free testosterone levels (P = 0.02). Elevated intrafollicular insulin levels in overweight women (P = 0.004) were accompanied by normal glucose and lactate levels. Intrafollicular E2 levels were greater in overweight vs. lean women (P = 0.03), whereas the remaining intrafollicular steroid levels were similar in both female groups.

Conclusion: In normoandrogenic ovulatory women undergoing in vitro fertilization, increased adiposity elevates insulin and E2 levels in terminally differentiated follicles without altering intrafollicular androgen levels or luteinization. Additional studies are required to determine whether these abnormalities impair oocyte development.


   Introduction
Top
 Abstract
 Introduction
Subjects and Methods
 Results
 Discussion
 References
 
INSULIN ACTS PRIMARILY on its own receptors located on theca cells, surrounding stroma and granulosa cells (1, 2) to stimulate ovarian steroidogenesis. In vitro studies suggest that insulin acts alone or with IGFs to amplify the actions of gonadotropins within the maturing follicle (3, 4). We previously have shown that fasting serum insulin levels and body mass index (BMI) positively correlate with intrafollicular insulin levels in women undergoing GnRH analog/recombinant human (rh) FSH therapy for in vitro fertilization (IVF) (2). This study extended these findings to determine whether increased adiposity alters intrafollicular steroidogenesis in normoandrogenic ovulatory women receiving similar ovarian stimulation for IVF. Its purpose was to understand the steroid environment of terminally differentiated follicles in overweight women and how it might relate to the outcome of the oocyte it contains.


   Subjects and Methods
Top
 Abstract
 Introduction
 Subjects and Methods
Results
 Discussion
 References
 
Experimental subjects

After approval by the Mayo Institutional Review Board, 30 normoandrogenic ovulatory women undergoing gonadotropin therapy for IVF were recruited. Normoandrogenic ovulatory women were receiving assisted reproduction for nonovarian indications [male factor (n = 18), endometriosis (n = 4), tubal factor (n = 6), and combined tubal and male factor infertility (n = 2)]. All women were Caucasian and signed informed consent before study participation.

Characteristics of our normoandrogenic ovulatory women previously have been described (5). Briefly, general inclusion criteria included age less than 38 yr, normal serum prolactin levels, and normal thyroid function studies. No woman had galactorrhea, endometriomas, or ovarian cysts greater than 18 mm in diameter. All normoandrogenic ovulatory women had regular menstrual cycles occurring every 21–35 d, luteal serum progesterone (P4) values [>3 ng/ml (SI conversion, 3.18 nmol/liter)], absence of hirsutism (modified Ferriman-Gallwey score < 8), and normal midfollicular serum androgen levels. None had polycystic ovaries by transvaginal ultrasonography. Seventeen women were lean (BMI < 25 kg/m2, 21.8 ± 2.1 kg/m2) and 13 women were overweight (BMI ≥ 25 kg/m2, 31.3 ± 6.4 kg/m2), of whom six were obese (BMI ≥ 30 kg/m2) (6).

Baseline blood sampling

Blood sampling for FSH, LH, estradiol (E2), 17-hydroxyprogesterone (17-OHP4), dehydroepiandrosterone sulfate (DHEAS), androstenedione (A4), total and free testosterone (T), dihydrotestosterone (DHT), and SHBG was performed between cycle d 5–10 of the menstrual cycle preceding IVF. On the same day, blood sampling for glucose and insulin was performed under fasting conditions and was repeated at 30-min intervals during a 75-g, 2-h oral glucose tolerance test.

Gonadotropin stimulation for IVF and oocyte retrieval

Leuprolide acetate (Lupron; TAP Pharmaceuticals, Lake Forest, IL) therapy was started on menstrual cycle d 21 to induce pituitary down-regulation and was initiated at a dose of 1.0 mg sc daily until pituitary down-regulation was established. The leuprolide acetate dose was then reduced to 0.5 mg daily until the day of human chorionic gonadotropin (hCG) administration.

Treatment with rhFSH (Gonal-F; Serono Laboratories, Rockland, MA) then was initiated sc with a starting dose of 225 IU daily for the first 3 d of stimulation, after which daily dosing was increased or decreased as clinically indicated. Serial E2 levels and two-dimensional follicle measurements by transvaginal ultrasonography were performed until at least two dominant follicles reached 18 mm or greater in diameter and serum E2 levels reached approximately 300 pg/ml/per dominant follicle. Human chorionic gonadotropin (10,000 IU, im) was then administered followed by transvaginal oocyte retrieval 36 h later.

At oocyte retrieval, follicular fluid was aspirated from the first follicle of each ovary, chosen by size (at least 15 mm in diameter) and accessibility. The collection tube was changed and the oocyte retrieval was continued based on routine clinical practice. The same procedure was repeated on the contralateral ovary and fluid uncontaminated by blood from each of the two follicles was individually assayed for hormone determinations (total follicle number, n = 55).

Follicular fluid sampling on the day of oocyte retrieval

Follicular fluid was centrifuged at 1800 g for 5 min to pellet follicular debris and was stored in 2.0 ml cryovials (Sarstedt, Inc., Newton, NC) at –70 C. Follicular fluid samples were transported on dry ice to the National Primate Research Center (University of Wisconsin, Madison, WI) for steroid, insulin, glucose, and lactate determinations, with correction for total protein concentration. There was no detectable insulin (<0.1 µIU/ml) present in any media used for cell preparation (2).

Hormone assays

Baseline serum FSH, LH, high-sensitivity T, SHBG, and DHEAS were measured at the Immunochemical Core Laboratory of the Mayo General Clinical Research Center (Rochester, MN). Serum free T was calculated using the ratio of total T to SHBG (7). Follicular fluid FSH, bioactive (bio) LH, E2, P4, 17OHP4, A4, T, DHT, glucose, insulin, and lactate were measured at the National Primate Research Center Hormone Assay Services Laboratory (2, 5, 8). bioLH was measured in a single assay by mouse Leydig cell bioassay (8). Follicular fluid hormone values were adjusted for protein content (per milligram BSA) to quantitatively reflect the volume of follicular fluid present (2, 5).

Statistical analysis

Basal serum hormone determinations and patient/IVF cycle characteristics were compared between lean and overweight women using Student t test, with logarithmic transformations performed when necessary to meet assumptions in regression modeling. Regression models with estimation by generalized estimating equations (9) were used to compare intrafollicular hormone levels between lean and overweight women and adjusting for intrasubject correlations owing to more than one follicle per patient (2, 5). Values are expressed as mean ± SD; P < 0.05 was considered significant.


   Results
Top
 Abstract
 Introduction
 Subjects and Methods
 Results
Discussion
 References
 
Patient and IVF cycle characteristics

There were no significant differences between female groups in age or basal serum LH, 17-OHP4, A4, T, DHEAS, DHT, and fasting serum glucose levels (Table 1Go). Overweight women had significantly elevated basal serum levels of fasting insulin and 2-h postprandial glucose as well as insulin, accompanied by decreased serum SHBG and increased serum free T levels. Despite blood sampling being performed on similar menstrual cycle days [lean women, d 7.3 ± 1.9; overweight women, d 7.2 ± 1.5 (SD), P = 0.9], overweight women had significantly decreased basal serum E2 levels and increased basal serum FSH levels, compared with lean women.


View this table:
[in this window]
[in a new window]

  		TABLE 1. Characteristics of normoandrogenic ovulatory women undergoing GnRH analog/rhFSH therapy for IVF

 
The amount of rhFSH administered (lean women, 2040 ± 542; overweight women, 2276 ± 768 IU, P = 0.3) and the duration of rhFSH treatment (lean women, 9.5 ± 1.5 d; overweight women, 10.2 ± 1.4 d, P = 0.2) were similar in both female groups, as were maximum serum E2 levels on the day of hCG administration (lean women, 2026 ± 790 pg/ml; overweight women, 1893 ± 684 pg/ml, P = 0.6), average follicle diameter (lean women, 19.5 ± 2.6 mm; overweight women, 20.6 ± 3.1 mm, P = 0.3) and total number of oocytes retrieved (lean women, 14.5 ± 7.1 oocytes; overweight women, 11.7 ± 4.1 oocytes, P = 0.2).

Intrafollicular hormone concentrations

Follicular fluid insulin levels were significantly greater in overweight than lean women, whereas follicular fluid glucose and lactate levels were similar in both female types (Table 2Go). Despite comparable intrafollicular FSH levels and sizes of follicles aspirated, E2 levels in follicles of overweight women were significantly greater than those of lean women. Conversely, intrafollicular bioLH levels were significantly lower in overweight vs. lean normoandrogenic ovulatory women, although follicular fluid P4, 17-OHP4, and androgen levels were similar in both types of patients.


View this table:
[in this window]
[in a new window]

  		TABLE 2. Intrafollicular hormone levels of normoandrogenic ovulatory women undergoing GnRH analog/rhFSH therapy for IVF

 

   Discussion
Top
 Abstract
 Introduction
 Subjects and Methods
 Results
 Discussion
References
 
The present study shows that increased adiposity enhances E2 production in follicles of normoandrogenic ovulatory women receiving GnRH analog/rhFSH therapy for IVF. Elevated intrafollicular E2 production in size-matched follicles of overweight vs. lean women were not associated with differences in rhFSH therapy, maximum serum E2 levels, and numbers of oocytes retrieved between female groups. These findings confirm that the microenvironment of the human follicle is not necessarily predicted by circulating steroid levels because follicle fluid E2 represents production by a single follicle, whereas circulating E2 reflects the cohort size of all dominant follicles. Furthermore, the latter is influenced by amounts of steroids entering the blood from ovarian or nonovarian sources and leaving the blood through clearance (10).

Apart from enhanced intrafollicular E2 production in overweight women, similar FSH levels were associated with comparable amounts of P4, 17-OHP4, aromatizable androgens, and DHT in follicles of overweight, compared with lean women. Whereas subtle effects of adiposity on intrafollicular steroidogenesis may have been undetected due to the small number of follicles examined, the increased adiposity of our study participants did not appear to impair the transition from 5{alpha}-reductase to aromatase activity (11) or the shift from androgen to P4 production that normally occur in terminally differentiated follicles (12). Reduced bioLH activity in follicles from overweight women, however, confirm that bioavailability of administered hCG (contributing to most of each bioLH value) negatively correlates with BMI (13, 14). Nevertheless, sufficient hCG was available in follicles of overweight women to induce conversion of glucose to lactate as energy substrate for appropriate luteinization, as evidenced by normal intrafollicular levels of glucose, lactate, and P4 (15). Additional studies are required to determine whether the degree of adiposity affects the intrafollicular microenvironment and whether adiposity-related intrafollicular abnormalities impair oocyte development.


   Footnotes
 
This work was supported by National Institutes of Health Grants U01 HD044650 and R01 RR 013635, Mayo Clinical Research Grant 2123-01, Mayo Grant M01-RR-00585, Grant P51 RR 000167 (to the National Primate Research Center, University of Wisconsin, Madison, a facility constructed with support from Research Facilities Improvement Program Grants RR15459-01 and RR020141-01), and Serono Pharmaceuticals. This work was partially supported by the National Institutes of Health as part of the National Institute of Child Health and Human Development National Cooperative Program on Female Health and Egg Quality under cooperative agreement U01 HD044650.

Author Disclosure Summary: D.H.A. and T.G.L. have nothing to declare. D.A.D. received grant support from Serono, Inc., consisting of medications only from 2003 to 2007.

First Published Online January 23, 2007

Abbreviations: A4, Androstenedione; bio, bioactive; BMI, body mass index; DHEAS, dehydroepiandrosterone sulfate; DHT, dihydrotestosterone; E2, estradiol; hCG, human chorionic gonadotropin; IVF, in vitro fertilization; 17-OHP4, 17-hydroxyprogesterone; P4, progesterone; rh, recombinant human; T, testosterone.

Received October 3, 2006.

Accepted January 17, 2007.


   References
Top
 Abstract
 Introduction
 Subjects and Methods
 Results
 Discussion
 References
 

  1. Samoto T, Maruo T, Ladines-llave C, Matsuo H, Deguchi J, Barnea E, Mochizuki M 1993 Insulin receptor expression in the follicular and stroma compartments of the human ovary over the course of follicular growth, regression, and atresia. Endocr J 40:715–726[Medline]
  2. Phy JL, Conover CA, Abbott DH, Zschunke MA, Walker DL, Session DR, Tummon IS, Thornhill AR, Lesnick TG, Dumesic DA 2004 Insulin and messenger ribonucleic acid expression of insulin receptor isoforms in ovarian follicles from nonhirsute ovulatory women and polycystic ovary syndrome patients. J Clin Endocrinol Metab 89:3561–3566[Abstract/Free Full Text]
  3. Bergh C, Carlsson B, Olsson JH, Selleskog H, Hillensjo T 1993 Regulation of androgen production in cultured human theca cells by insulin-like growth factor-I and insulin. Fertil Steril 59:323–331[Medline]
  4. Franks S, Gilling-Smith C, Watson H, Willis D 1999 Insulin action in the normal and polycystic ovary. Endocrinol Metab Clin North Am 28:361–378[CrossRef][Medline]
  5. Foong SC, Abbott DH, Zschunke MA, Lesnick TG, Phy JL, Dumesic DA 2006 Follicle luteinization in hyperandrogenic follicles of polycystic ovary syndrome (PCOS) patients undergoing gonadotropin therapy for in vitro fertilization. J Clin Endocrinol Metab 91:2327–2333[Abstract/Free Full Text]
  6. 1998 Executive summary of the clinical guidelines on the identification, evaluation, and treatment of overweight and obesity in adults. Arch Intern Med 158:1855–1867
  7. Harman SM, Metter EJ, Tobin JD, Pearson J, Blackman MR 2001 Longitudinal effects of aging on serum total and free testosterone levels in healthy men. J Clin Endocrinol Metab 86:724–731[Abstract/Free Full Text]
  8. Dumesic DA, Abbott DH, Eisner JR, Goy RW 1997 Prenatal exposure of female rhesus monkeys to testosterone propionate increases serum luteinizing hormone levels in adulthood. Fertil Steril 67:155–163[CrossRef][Medline]
  9. Liang K-Y, Zeger S 1986 Longitudinal data analysis using generalized linear models. Biometrika 73:13–22[Abstract/Free Full Text]
  10. Dumesic DA, Schramm RD, Peterson E, Paprocki AM, Zhou R, Abbott DH 2002 Impaired developmental competence of oocytes in adult prenatally androgenized female rhesus monkeys undergoing gonadotropin stimulation for in vitro fertilization. J Clin Endocrinol Metab 87:1111–1119[Abstract/Free Full Text]
  11. Slater CC, Chang L, Stanczyk FZ, Paulson RJ 2001 Altered balance between the 5{alpha}-reductase and aromatase pathways of androgen metabolism during controlled ovarian hyperstimulation with human menopausal gonadotropins. J Assist Reprod Genet 18:527–533[CrossRef][Medline]
  12. Chaffin CL, Hess DL, Stouffer RL 1999 Dynamics of periovulatory steroidogenesis in the rhesus monkey follicle after ovarian stimulation. Hum Reprod 14:642–649[Abstract/Free Full Text]
  13. Elkind-Hirsch KE, Bello S, Esparcia L, Phillips K, Sheiko A, McNichol M 2001 Serum human chorionic gonadotropin levels are correlated with body mass index rather than route of administration in women undergoing in vitro fertilization-embryo transfer using human menopausal gonadotropin and intracytoplasmic sperm injection. Fertil Steril 75:700–704[CrossRef][Medline]
  14. Chan CCW, Ng EHY, Chan MMY, Tang OS, Lau EYL, Yeung WSB, Ho PC 2003 Bioavailability of hCG after intramuscular or subcutaneous injection in obese and nonobese women. Hum Reprod 18:2294–2297[Abstract/Free Full Text]
  15. Gull I, Geva E, Lerner-Geva L, Lessing JB, Wolman I, Amit A 1999 Anaerobic glycolysis. The metabolism of the preovulatory human follicle. Eur J Obstet Gynecol Reprod Biol 85:225–228[CrossRef][Medline]




This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow All Versions of this Article:
92/4/1438    most recent
Author Manuscript (PDF)
Right arrow Submit a related Letter to the Editor
Right arrow Purchase Article
Right arrow View Shopping Cart
Right arrow Alert me when this article is cited
Right arrow Alert me when eLetters are posted
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow Request Copyright Permission
Citing Articles
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Dumesic, D. A.
Right arrow Articles by Abbott, D. H.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Dumesic, D. A.
Right arrow Articles by Abbott, D. H.
Related Collections
Right arrow Female Endocrinology
Right arrow Obesity

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
Endocrinology Endocrine Reviews J. Clin. End. & Metab.
Molecular Endocrinology Recent Prog. Horm. Res. All Endocrine Journals