| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
Other Original Articles |
Department of Obstetrics and Gynecology, Hospital Universitari Sant Joan de Déu (J.C., C.S., S.V., J.M.L.), Cancer Research Institute (A.M.), and Institut Clínic of Gynecology, Obstetrics and Neonatology (J.B.), Hospital Clínic-Institut d’Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Faculty of Medicine–University of Barcelona, 08950 Esplugues, Barcelona, Spain
Address all correspondence and requests for reprints to: Justo Callejo, M.D., Department of Obstetrics and Gynecology, Hospital Universitari Sant Joan de Déu, Po Sant Joan de Déu no 2, 08950 Esplugues, Barcelona, Spain. E-mail: jcallejo{at}medicina.ub.es
Abstract
The transplantation of ovarian tissue has recently been the focus
of intense investigation with the aim of avoiding premature ovarian
failure mainly in patients receiving chemotherapy or radiotherapy for
malignant disease. Here, we present an evaluation of the long-term
function of both fresh (patients 1, 2, and 3) and cryopreserved
(patient 4) ovarian autografts in four premenopausal patients aged
46–49 yr who underwent heterotopic ovarian transplantation and were
followed over a 1-yr period without receiving gonadotropins to
stimulate follicular growth. In patients 1 and 2, 
1 cm3
ovarian cortical autograft was placed sc in the inner aspect of the
arm, whereas and in patients 3 and 4 minced ovarian tissue was placed
into a muscle pocket in the abdominal wall. In patients 1, 2, and 4 the
ovarian hormone secretion (as suggested by sequential estradiol and FSH
serum measurements) was reestablished 3–4 months after
autotransplantation, and graft function was not improved by immediate
rather than delayed heterotopic ovarian autografting. Despite a
reestablished ovarian function, a 2- to 7-fold increase in peripheral
FSH concentration was evidenced. The cases reported here suggest that
hormonal protection can be restored after fresh or cryopreserved
heterotopic ovarian transplantation in women, albeit for only a short
reproductive span.
MODERN MANAGEMENT OF childhood malignancies is so successful that many children are cured. As survival rates for young cancer patients continue to improve, protection against iatrogenic ovarian failure and infertility caused by chemotherapy and/or radiotherapy assumes higher priority (1, 2). Thus, at present, a great interest exists on the potential use of ovarian autotransplantation for preservation of ovarian function (3, 4, 5, 6). This has been accomplished successfully by us (7) and others (8) in rats, sheep, and other animals. Such therapy could provide a source of ovarian tissue that, when autotransplanted, would maintain an adequate estrogenic milieu that protects against postmenopausal complications. A second role for this therapy could be in the form of "oocyte banking" as a strategy to preserve the reproductive potential of younger women or girls before cancer therapy (1, 2). However, before offering such an option clinically, it would be necessary to know when hormonal "protection" is reestablished after autografting and how long such grafts might survive.
Very recently, the first case of successful transplantation of cryopreserved autologous ovarian tissue into a previously oophorectomized woman with nonmalignant disease has been reported (9). However, this patient underwent stimulation of follicular development with human menopausal gonadotropins shortly after ovarian tissue was sutured beneath the pelvic peritoneum with the use of a laparoscope. Here, we present an evaluation of the long-term function of both fresh and cryopreserved ovarian autografts in four premenopausal patients who underwent heterotopic ovarian transplantation and were followed over a 1-yr period without receiving gonadotropins to stimulate follicular growth.
Patients and Methods
Patients
A total of 14 women under 50 yr (range, 46–49) of age were
included in the present study, which was approved by the Institutional
Review Board of the Hospital Universitari Sant Joan de Déu. All
of them were regularly menstruating premenopausal women having a body
mass index below 30 kg/m2 and no ovarian failure
on the basis of early follicular phase (cycle d 2–4) FSH
concentrations of less than 15 IU/liter (standard International
Reference Preparation 78/549) measured within 3 months of surgery in
the four experimental subjects. The latter were four premenopausal
women aged 46–49 yr who underwent abdominal hysterectomy because of
uterine leiomyomas, with bilateral salpingo-oophorectomy performed as a
prophylactic measure for ovarian cancer according to the patients’
choice after appropriate informed consent was obtained. These four
women volunteered for ovarian autotransplantation with fresh or
cryopreserved ovarian tissue (Table 1
).
As controls for hormone measurements we included a group of 10 women
fulfilling the above mentioned inclusion criteria and who underwent
abdominal hysterectomy with prophylactic bilateral removal of the
adnexa because of uterine fibroids, as did experimental subjects.
|
The ovaries were transferred to Flushing Medium (1076;
Medi-Cult, Leti, Barcelona, Spain) at room temperature for preparing
cortical autografts. Cortex that was free of obvious follicles or
luteal tissue was cut with a scalpel to a thickness of 1–2 mm,
avoiding the hilar area. In patients 1 and 2, after histological
confirmation of the presence of primordial ovarian follicles and the
absence of malignant pathology in the remaining ovarian tissue by using
several frozen sections, 1 cm3 ovarian
cortical autograft was placed sc in the inner aspect of the arm at the
time of surgery. This was performed over the superficial venous bed
(basilic median and/or basilic) through an incision of 2 cm. Before
fixing the implant on its vascular bed, two sections dividing the
ovarian tissue into four parts were performed as a potential measure to
enhace revascularization (3, 10). The sc cellular tissue
was closed with two stitches of vicryl, thus serving as a cushion
protecting the autograft.
In patients 3 and 4, 1 cm3 pathologically
confirmed cortical ovarian tissue was minced into 40–45 fragments 2–3
mm3 and then placed into a muscle pocket in the
rectus muscle of abdomen according to the technique principles used for
the heterotopic parathyroid autotransplantation in the
sternocleidomastoid muscle (11). The muscle pocket and
fascia were closed with stiches of vicryl. In patient 3 the procedure
was performed using fresh ovarian tissue despite that histopathologic
examination indicated that primordial ovarian follicles were very
scanty in the tissue sample analyzed. In patient 4 that procedure was
performed under local anesthesia 3 months after hysterectomy and
bilateral salpingo-oophorectomy using frozen-thawed ovarian tissue
prepared according to protocols reported below.
Freezing and thawing protocol
Pieces of ovarian tissue of 2–3 mm3 were frozen using dimethylsulphoxide (D2650; Sigma, St. Louis, MO) as cryoprotectant. First, the tissue was equilibrated at 4 C for 10 min in a 0.7-M dimethylsulfoxide solution containing 20% human serum (5009; Irvine Scientific, Santa Ana, CA) in RPMI 1640 medium (01-104-1A; Biological Industries, Kibbutz beth Haemek, Israel). They were then transferred to the freezing solution with similar composition but containing 1.5 M dimethylsulfoxide and were left to equilibrate for 10 min at 4 C. The tissue was then transferred to freezing vials (377267, Nunc freezing vials; Nuclon, Roskilde, Denmark) filled until 1.8 ml with freezing solution and placed into a programmable freezer (CM25; Carburos Metálicos, Barcelona, Spain). After 5 min for equilibration at 4 C cooling was performed at 0.5 C/min to -7 C, at which temperature seeding was automatically induced. Ovarian tissue was further cooled at a rate of 0.5 C/min to -50 C and subsequently at 5 C/min to -80 C, and finally at 8 C/min to -120 C. The tissue was then transferred to liquid nitrogen (-196 C) and stored until autografting.
Thawing was performed by immersion of the vials in a water bath at 37 C until the ice was melted. The freezing medium was stepwise diluted during 7 min at 4 C to a final concentration of 0.14 M dimethylsulfoxide (final volume, 18 ml) by adding each minute a double volume of RPMI 1640 medium supplemented with 4% of human serum albumin (860668; Instituto Grifols, Barcelona, Spain). Ovarian tissue was then equilibrated during 3 min at 4 C before replacing the diluted medium by Flushing Medium. Thereafter, the ovarian tissue and Flushing Medium were equilibrated for 25 min at 37 C in a 5% of CO2 to allow a more complete cell rehydration before grafting.
Hormone monitoring and analyses
All women included in the study underwent a blood sampling in the early to mid-follicular phase (cycle d 5–7) for baseline E2 and FSH serum measurement. Both experimental subjects and control patients were operated in the next week following this blood sampling. A second blood sampling for serum E2 and FSH determination was obtained in control women 4–5 wk after surgery. In the four experimental subjects blood samples for sequential evaluation of serum concentrations of E2 and FSH were collected at 8–10 d after ovarian grafting and afterward monthly during a 1-yr period. Ultrasonographic scans of ovarian tissue were performed on the same days that hormone sampling once ovarian function was reestablished as suggested by E2 serum levels. In patient 4 ultrasonographic scans and serum inhibins A and B and progesterone measurements were also carried out every 8–10 d over a 1-month period once E2 and FSH serum levels suggested that hormonal ovarian function had been reestablished.
Serum E2 and FSH were measured by a solid-phase chemiluminiscent assay using an Immulite Automated Analyzer (Diagnostic Products, Los Angeles, CA). The intra- and interassay coefficients of variation were 5.4% and 8.1% for FSH and 8.6% and 9.3% for E2, respectively. The sensitivity of the assays was 90 pmol/liter for E2 and 0.2 IU/liter for FSH. Dimeric inhibin A and inhibin B measurements were performed by a solid-phase sandwich ELISA, which uses two monoclonal antibodies (Serotec Ltd., Oxford, UK). The assay sensitivity for inhibin A was 2 pg/ml, and the intra-assay coefficient of variation was less than 7%. The cross-reactivity with activin A was less than 0.1%. The sensitivity of the assay for inhibin B was 15 pg/ml, and the intra-assay coefficient of variation was less than 11%. Ultrasonic scans were performed using a multifrequency 7–5-MHz transducer attached to an Acuson Aspen System (Acuson, Mountain View, CA).
Results
Sequential serum concentrations of E2 and FSH in patients 1, 2,
and 4 plotted to hormone levels obtained in the control group of women
before and 4–5 wk after ovarian removal are shown in Figs. 1 and 2. As
expected, after hysterectomy and bilateral adnexectomy in the four
experimental subjects, there was a significant fall of serum E2 from
normal premenopausal values to levels equivalent to those observed in
the castrated group of women (data not shown for patient 3). The four
women developed menopausal symptoms within 3 wk after surgery but none
of them was given hormone replacement therapy during the study
period.
|
|
and 2). In this patient a growing ovarian
follicle 16 mm in diameter was detected by ultrasonography (Fig. 3) in association with an E2 serum level
of 383 pmol/liter, but with inhibin A and inhibin B serum
concentrations below the detection limit of the assay. Ten days later,
the follicle still had a diameter of 20 mm and serum E2 concentration
was 303 pmol/liter (serum inhibin A and inhibin B again could not be
detected), at which time 5000 IU human CG was administered im (after an
additional informed consent was obtained from the patient) to
investigate the luteinization potential of the follicle. One week after
the human CG injection E2 the serum level was 289 pmol/liter and serum
progesterone concentration was 7.1 nmol/liter (normal values in the
luteal phase, 6–64 nmol/liter), but circulating inhibin A and inhibin
B could not be detected. After one additional week serum levels of E2
and progesterone were 230 pmol/liter and 2.9 nmol/liter, respectively,
both inhibin A and inhibin B remaining undetectable.
|
Discussion
This study confirms that simple ovarian graft, either fresh or
after cryopreservation, without vascular anatomoses can form functional
tissue in a heterotopic site in women. Interestingly, graft function
was not improved by immediate rather than delayed heterotopic ovarian
autotransplantation, a fact that has also been reported recently with
parathyroid autografting (12). An important advantage of
cryopreservation is that it permits long-term storage of ovarian tissue
(cryobanking) for a patient (1, 13, 14, 15). Remarkably,
50% of the follicular population survives implantation and the
majority of follicles are lost acutely after grafting rather than at
the freeze-thaw stage (5, 8).
The ideal site for heterotopic ovarian autotransplantation remains to be established. As a potential treatment for the menopause, the site of the graft seems not critical and, as suggested by a recent case report (9) and the present study, with the advent of in vitro fertilization technology and high-resolution ultrasound, heterotopic grafting could also potentially form an alternative approach for the preservation of fertility. Nevertheless, there are differences in surgical technique used in patients reported here that might affect the process of ovarian revascularization essentially related to the functional longevity of the grafts. In patients 1 and 2 the whole ovarian cortical autograft was placed sc in the arm, whereas in patient 4 minced ovarian tissue was transplanted im in the abdominal wall. A major problem is that heterotopic ovarian cortical pieces containing primordial follicles cannot be directly reanastomosed to a blood supply and, therefore, remain hypoxic until new blood vessels grow into them. The impact of the ischemic period that follows grafting on any given follicle is thought to be correlated with its metabolic demands and location within the ovary (8, 16). When the ovary is cut into small pieces, ischemic and degenerative changes are minimized because the process of revascularization depends on the size of the tissue involved (8, 10, 16). On the other hand, the well vascularized abdominal musculature rather than the sc space may provide better graft survival for endocrinologically active glandular tissue as it has been shown with parathyroid autotransplantation (11, 12).
Results in the present study suggest that it is unlikely that
heterotopic graft longevity would be an adequate substitute for hormone
replacement therapy after a normal menopause, and ovarian tissue
banking and transplantation should, therefore, be regarded primarily as
a fertility aid, a view previously emphasized by others
(5). Several facts deserve discussion in this respect.
First, it is necessary to wait 3–4 months until spontaneous recovery
of ovarian function as shown by patients 1, 2, and 4 in the present
study. As discussed above, experimental studies (5, 8, 15)
have shown that the ischemia that occurs in the graft before
revascularization results in the loss of virtually the entire growing
follicle population in addition to the 50% of the primordial
folllicles. The growth of a primordial follicle to the large preantral
stage in humans takes approximately 85 d, whereas the final
maturation of the large preantral follicle to the preovulatory stage
takes approximately 14 d, representing the duration of the
follicular phase (17). The present results are, therefore,
consistent with the hypothesis that the delay from autografting to
resumption of ovarian function represents the period of time required
for the primordial follicles that have survived to develop to a
preovulatory follicle (18). Second, spontaneous follicular
growth seems to be slower in autotransplanted ovarian grafts as
suggested by patient 4, in whom the leading follicle increased from 16
mm to 20 mm in diameter only after 10 d. Interestingly, a third
feature characterizing patients 1, 2, and 4 is that the autograft
resulted in a 2- to 7-fold increase in peripheral FSH concentration in
the face of a reestablished E2 secretion, which is in agreement with
studies using ovarian cortical autografts in sheep (18).
Elevated basal FSH in regularly menstruating women has been reported to
be associated with slower follicular growth (19). In turn,
as suggested by patient 4 in this study and it has been reported in the
sheep (18), the most likely cause of the increase in FSH
associated with restoration of ovarian function after autografting is a
decline in inhibin secretion. Lower inhibin A and inhibin B levels in
older cycling women are considered as early markers of female
reproductive aging (20) and, thus, overall, the endocrine
changes associated with human ovarian autografting are similar to those
observed in aged women. In this respect, our study has an important
potential limitation, which is that all patients included were 46- to
49-yr-old (i.e. they were in the perimenopausal transition)
and this does not fit well with the young patient probable candidate
for cancer treatment and at risk of iatrogenic premature ovarian
failure. This is further stressed by patient 3 where ovarian function
did not recover after fresh ovarian autografting and in whom a very
scanty number of primordial follicles were seen in the ovarian biopsy
in the face of a normal FSH value at baseline but in association with
an elevated E2 on d 2–4, which is a sign of advanced reproductive age.
However, the acute endocrine changes depicted here after both bilateral
adnexectomy and fresh or frozen-thawed ovarian autografting are
demonstrative enough of the potential features associated with the
procedure.
In summary, the cases reported here suggest that hormonal "protection" and potentially even fertility can be restored after fresh or cryopreserved heterotopic ovarian autografting in women, albeit for only a short reproductive span. However, much more work is necessary on the subject considering: 1) there is minimal information available in the medical literature in regard to hormonal function from transplanted ovarian tissue in humans; 2) the present study includes only a small group of patients undergoing transplants in two different sites and even freezing some and not all of the tissue; and 3) the 1-yr follow-up was an arbitrary interval that may be useful to analyze the functional longevity of the grafts, but we do not know the potential long-term risks of ovarian cancer in these patients. However, considering the small amount of ovarian tissue left in place and that autografts had no vascular pedicle and were nonfunctioning at the end of the experiment, this possibility seems unlikely.
Acknowledgments
We thank Dr. A. Vela for performing ultrasonographic scans and Drs. C. Valls and R. Casamitjana for performing hormonal determinations.
Received February 27, 2001.
Accepted May 26, 2001.
References
This article has been cited by other articles:
 |
|
 |
|
  I. Demeestere, P. Simon, S. Emiliani, A. Delbaere, and Y. Englert Orthotopic and heterotopic ovarian tissue transplantation Hum. Reprod. Update, November 1, 2009; 15(6): 649 - 665. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 V.J. Onions, R. Webb, A.S. McNeilly, and B.K. Campbell Ovarian endocrine profile and long-term vascular patency following heterotopic autotransplantation of cryopreserved whole ovine ovaries Hum. Reprod., November 1, 2009; 24(11): 2845 - 2855. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 X. Deng, H. Zheng, X. Yu, H. Yu, C. Zhang, L. Chao, R. Li, and W. Liu Cryopreserved ovarian tissues can maintain a long-term function after heterotopic autotransplantation in rat Reproduction, September 1, 2009; 138(3): 519 - 525. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
V Isachenko, I Lapidus, E Isachenko, A Krivokharchenko, R Kreienberg, M Woriedh, M Bader, and J M Weiss Human ovarian tissue vitrification versus conventional freezing: morphological, endocrinological, and molecular biological evaluation Reproduction, August 1, 2009; 138(2): 319 - 327. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
V. Fain-Kahn, C. Poirot, C. Uzan, M. Prades, S. Gouy, C. Genestie, P. Duvillard, and P. Morice Feasibility of ovarian cryopreservation in borderline ovarian tumours Hum. Reprod., April 1, 2009; 24(4): 850 - 855. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. A. Bedaiwy, S. A. El-Nashar, A. M. El Saman, J. L.H. Evers, S. Sandadi, N. Desai, and T. Falcone Reproductive outcome after transplantation of ovarian tissue: a systematic review Hum. Reprod., December 1, 2008; 23(12): 2709 - 2717. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Y. Wang, Z. Xiao, L. Li, W. Fan, and S.-W. Li Novel needle immersed vitrification: a practical and convenient method with potential advantages in mouse and human ovarian tissue cryopreservation Hum. Reprod., October 1, 2008; 23(10): 2256 - 2265. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. Meirow, I. Hardan, J. Dor, E. Fridman, S. Elizur, H. Ra'anani, E. Slyusarevsky, N. Amariglio, E. Schiff, G. Rechavi, et al. Searching for evidence of disease and malignant cell contamination in ovarian tissue stored from hematologic cancer patients Hum. Reprod., May 1, 2008; 23(5): 1007 - 1013. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Sanchez, P. Alama, B. Gadea, S.R. Soares, C. Simon, and A. Pellicer Fresh human orthotopic ovarian cortex transplantation: long-term results Hum. Reprod., March 1, 2007; 22(3): 786 - 791. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
X.-h. Deng, A.-r. Xu, L. Chao, H.-l. Yu, J.-h. Zhen, S. Hashimoto, and Y. Morimoto Effect of different sites for cryopreserved ovarian tissue implantation in rabbit Hum. Reprod., March 1, 2007; 22(3): 662 - 668. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. Donnez, B. Martinez-Madrid, P. Jadoul, A. Van Langendonckt, D. Demylle, and M.-M. Dolmans Ovarian tissue cryopreservation and transplantation: a review Hum. Reprod. Update, September 1, 2006; 12(5): 519 - 535. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
I. Demeestere, P. Simon, F. Buxant, V. Robin, S. A. Fernandez, J. Centner, A. Delbaere, and Y. Englert Ovarian function and spontaneous pregnancy after combined heterotopic and orthotopic cryopreserved ovarian tissue transplantation in a patient previously treated with bone marrow transplantation: Case Report Hum. Reprod., August 1, 2006; 21(8): 2010 - 2014. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. Sonmezer and K. Oktay Fertility preservation in young women undergoing breast cancer therapy. Oncologist, May 1, 2006; 11(5): 422 - 434. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K.L.T. Schmidt, C. Y. Andersen, A. Loft, A.G. Byskov, E. Ernst, and A. N. Andersen Follow-up of ovarian function post-chemotherapy following ovarian cryopreservation and transplantation Hum. Reprod., December 1, 2005; 20(12): 3539 - 3546. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Samuel Kim, H. G. Kang, N. H. Kim, H. C. Lee, and H. H. Lee Assessment of the integrity of human oocytes retrieved from cryopreserved ovarian tissue after xenotransplantation Hum. Reprod., September 1, 2005; 20(9): 2502 - 2508. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
B. Schubert, M. Canis, C. Darcha, C. Artonne, J.-L. Pouly, P. Dechelotte, D. Boucher, and G. Grizard Human ovarian tissue from cortex surrounding benign cysts: a model to study ovarian tissue cryopreservation Hum. Reprod., July 1, 2005; 20(7): 1786 - 1792. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. A. Gook, D.H. Edgar, J. Borg, J. Archer, and J.C. McBain Diagnostic assessment of the developmental potential of human cryopreserved ovarian tissue from multiple patients using xenografting Hum. Reprod., January 1, 2005; 20(1): 72 - 78. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. A. Bedaiwy and T. Falcone Ovarian tissue banking for cancer patients: Reduction of post-transplantation ischaemic injury: intact ovary freezing and transplantation Hum. Reprod., June 1, 2004; 19(6): 1242 - 1244. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. Oktay and M. Sonmezer Ovarian tissue banking for cancer patients: Fertility preservation, not just ovarian cryopreservation Hum. Reprod., March 1, 2004; 19(3): 477 - 480. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Revel and J. Schenker Ovarian tissue banking for cancer patients: is ovarian cortex cryopreservation presently justified? Hum. Reprod., January 1, 2004; 19(1): 14 - 19. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. Hreinsson, P. Zhang, M. L. Swahn, K. Hultenby, and O. Hovatta Cryopreservation of follicles in human ovarian cortical tissue. Comparison of serum and human serum albumin in the cryoprotectant solutions Hum. Reprod., November 1, 2003; 18(11): 2420 - 2428. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. Callejo, C. Salvador, P. Prats, and J. M. Lailla What Treatment Should Be Given to the Cryopreserved Ovarian Tissue after Thawing? J. Clin. Endocrinol. Metab., April 1, 2002; 87(4): 1908 - 1909. [Full Text] [PDF]
|
|
|
|
|
|
K. Oktay, H. C. M. T. Prinsen, W. R. de Vries, H. P. F. Koppeschaar, and M. G. M. de Sain-van der Velden Evidence for Limiting Ovarian Tissue Harvesting for the Purpose of Transplantation to Women Younger than 40 Years of Age J. Clin. Endocrinol. Metab., April 1, 2002; 87(4): 1907 - 1908. [Full Text] [PDF]
|
|
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| 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 |
, progesterone (P) serum levels in patient
4.
