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.2005-0639
This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow All Versions of this Article:
90/10/5647    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 HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Matthiesson, K. L.
Right arrow Articles by McLachlan, R. I.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Matthiesson, K. L.
Right arrow Articles by McLachlan, R. I.
Related Collections
Right arrow Male Endocrinology
Right arrow Neuroendocrinology and Pituitary
The Journal of Clinical Endocrinology & Metabolism Vol. 90, No. 10 5647-5655
Copyright © 2005 by The Endocrine Society

Effects of Testosterone and Levonorgestrel Combined with a 5{alpha}-Reductase Inhibitor or Gonadotropin-Releasing Hormone Antagonist on Spermatogenesis and Intratesticular Steroid Levels in Normal Men

Kati L. Matthiesson, Peter G. Stanton, Liza O’Donnell, Sarah J. Meachem, John K. Amory, Richard Berger, William J. Bremner and Robert I. McLachlan

Prince Henry’s Institute of Medical Research and Department of Obstetrics and Gynecology, Monash University, Monash Medical Center (K.L.M., P.G.S., L.O., S.J.M., R.I.M.), Clayton, Victoria 3168, Australia; and Center for Research in Reproduction and Contraception, University of Washington (J.K.A., R.B., W.J.B.), Seattle, Washington 98195

Address all correspondence and requests for reprints to: Dr. Kati Matthiesson, Prince Henry’s Institute of Medical Research, P.O. Box 5152, Clayton, Victoria 3168, Australia. E-mail: kati.matthiesson{at}phimr.monash.edu.au.


   Abstract
Top
 Abstract
Introduction
 Subjects and Methods
 Results
 Discussion
 References
 
Context: Combination of a GnRH antagonist (acyline), types I and II, 5{alpha}-reductase inhibitor (dutasteride) or levonorgestrel (LNG) with testosterone (T) treatment may augment the suppression of spermatogenesis and intratesticular (iT) steroids.

Objective: The objective of this study was to assess the effects of combined hormonal contraceptive regimens on germ cell populations and iT steroids.

Design, Setting, and Participants: Twenty-nine normal health men enrolled in this prospective, randomized, 14-wk study at the University of Washington.

Intervention(s): Twenty-two men (n = 5–6/group) received 8 wk of T enanthate (TE; 100 mg, im, weekly) combined with 1) 125 µg LNG daily, orally; 2) 125 µg LNG plus 0.5 mg dutasteride daily, orally; 3) 300 µg/kg acyline twice weekly, sc; or 4) 125 µg LNG daily, orally, plus 300 µg/kg acyline twice weekly, sc. Subjects then underwent a vasectomy and testicular biopsy. Control men (n = 7) proceeded directly to surgery.

Main Outcome Measure(s): The main outcome measures were germ cells and iT steroids [T, dihydrotestosterone, 3{alpha}- and ß-androstanediol (Adiol), and estradiol (E2)].

Results: High iT levels of all androgens (6- to 123-fold serum levels) and E2 (407-fold serum levels) were found in control men. iTT (1.9–2.6% control; P < 0.001) and iT3ßAdiol (16–34% control; P < 0.05) levels decreased with all treatments. iT dihydrotestosterone (13–29% control; P < 0.05) and iT3{alpha}Adiol (44–47% control; P < 0.05) levels decreased with all but the TE plus LNG treatment. iTE2 levels decreased only in the TE plus acyline group (28% control; P = 0.01). Germ cells from type B spermatogonia onward were suppressed, with no differences between groups found. Variable sites of impairment of germ cell progression were evident between men (spermagonial maturation, meiosis 1 entry, and spermiation). Other than a negative correlation between iT3{alpha}Adiol and haploid germ cell number (P < 0.006), no correlations between germ cell number and gonadotropins, sperm concentration, or iT steroids were found.

Conclusions: A similar high testicular:serum gradient exists for E2 and T in normal men, and 8 wk of gonadotropin suppression markedly reduces iTT, with 5{alpha}-reduced androgens and E2 levels decreasing to a much lesser degree. The heterogeneity of the germ cell response, regardless of treatment, gonadotropins or iT steroids, points to the individual sensitivity of sites in germ cell development, which is worthy of additional exploration.


   Introduction
Top
 Abstract
 Introduction
Subjects and Methods
 Results
 Discussion
 References
 
MALE HORMONAL CONTRACEPTION (MHC) regimens are based on the use of exogenously administered testosterone (T) or synthetic androgens (1) acting to suppress the pituitary gonadotropins (FSH and LH) and thereby spermatogenesis while maintaining adequate androgen action at extragonadal sites. Additional agents have been included to augment gonadotropin suppression, such as progestins [depot medroxyprogesterone acetate (DMPA) (2, 3), levonorgestrel (LNG) (4, 5), desogesterel (6, 7, 8), etonorgestrel (9), norethisterone (10, 11), and cyproterone acetate (12, 13)], GnRH antagonists (acyline, cetrorelix, and teverolix) (14, 15, 16), and 5{alpha}-reductase inhibitors (finasteride and dutasteride) (8, 16, 17). Such combined regimens potentially offer faster and more consistent contraceptive efficacy.

The reduction of intratesticular T (iTT) levels is essential to the interruption of spermatogenesis. Levels have been previously described to fall by about 98% during MHC treatment (2, 18, 19). However there are few data comparing the effects of combined regimens on iTT or regarding the relationships between germ cell development and the levels of iTT or other sex steroid metabolites, such as dihydrotestosterone (DHT), 3{alpha}- and ß-androstanediol (Adiol), and estradiol (E2). Some men administered MHC are able to maintain a low level of sperm production despite marked suppression of LH and iTT, leading to speculation that they are able to convert remaining iTT, through 5{alpha}-reductase action, to the more potent androgen DHT, which then supports spermatogenesis (20). The role of E2 in the spermatogenic process in man is uncertain (21). Estrogen receptor-ß (ERß) has been localized to human germ and Sertoli cells (22, 23, 24), with ER{alpha} localization being less certain (22, 23, 25). Data from rodent ER{alpha}, ER{alpha}ß, and aromatase knockout models displaying disruption of spermatogenesis suggest a regulatory or supportive role of E2 (26, 27, 28, 29).

Recently, two new potential contraceptive agents have become available. Acyline, a member of the azaline B family (30), is a potent GnRH antagonist administered sc every 14 d, which results in rapid and profound gonadotropin suppression when coadministered with T enanthate (TE) and/or LNG (16). Dutasteride, a new 5{alpha}-reductase inhibitor used in the treatment of benign prostatic hypertrophy (31), inhibits both type 1 and 2 isoenzymes of 5{alpha}-reductase and therefore permits the testing of the hypothesis that the withdrawal of DHT support from the seminiferous epithelium will further impair spermatogenesis.

The inclusion of progestins is known to be T dose sparing (32) and to increase the rate, but not depth, of gonadotropin suppression (33). It is also postulated that they may have some direct testicular effects independent of gonadotropin suppression (34). Recently, the presence of a membrane-bound progesterone receptor (PR) protein and intracellular PR-A and PR-B mRNA has been reported in human testis (35), but there remains no clear evidence for direct progestin effects on spermatogenesis.

Sperm concentration and serum reproductive hormone levels are usually the end points for most MHC trials, with relatively few contraceptive efficacy studies performed to date (36, 37, 38, 39). However, events within the seminiferous epithelium may not be reflected in semen quality, but be revealed by germ cell enumeration using stereological approaches. There has only been one study using T plus progestin that has reported germ cell numbers as an end point (2). Such data allow the appreciation of the events leading up to sperm release and provide insight into the variable degree of suppression of sperm concentration and the ability of the seminiferous epithelium to recover when expected (after cessation of MHC) or when unwelcome (escape or rebound during treatments). In addition, the procurement of testicular tissue (2, 19) allows morphological, chromatographic, and genetic analyses that may provide novel insights into the development of more universally effective MHC strategies.

We recently reported a prospective, randomized, four-arm, 8-wk study of novel MHC combinations aimed at improving spermatogenic suppression (16). We found similar suppression of serum gonadotropin levels and sperm concentrations, but the question of whether differences exist in spermatogenesis or iT steroid levels remained unanswered. We now report the analysis of testicular biopsy end points in these men and seven normal untreated men serving as controls. We aimed to determine whether the addition of the new GnRH antagonist, acyline, or that of the dual type I and II, 5{alpha}-reductase inhibitor, dutasteride, produced greater suppression of particular germ cell subtypes, spermatid release (spermiation), or iT sex hormone metabolism when added to a conventional T plus progestin MHC regimen. To explore whether the inclusion of a progestin produced gonadotropin-independent testicular effects in the presence of profound gonadotropin suppression, we included TE plus acyline as a comparator to TE, LNG, plus acyline. In conjunction, we report the iT profiles of androgens and E2 in normal human testes and after gonadotropin withdrawal.


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

Twenty-nine men (planning to undergo elective vasectomy) were recruited through media advertisement to participate in this study conducted at the University of Washington. Details regarding consent, inclusion, and exclusion criteria have been previously described (16).

Study design

This research study was divided into three phases: a 2-wk screening phase, an 8-wk treatment phase, and a 4-wk recovery period. After screening, men were randomly assigned to one of the following four treatments groups (n = 5–6/group) for 8 wk or to a control group (n = 7): 1) 100 mg TE (Delstestryl; Bristol-Myers Squibb, Princeton, NJ) weekly, im, plus 125 µg LNG (Wyeth, Madison, NJ) daily, orally (n = 5); 2) 100 mg TE weekly, im; 125 µg LNG daily, orally; plus 0.5 mg dutasteride (GlaxoSmithKline, Research Triangle Park, NC) daily, orally (n = 6); 3) 100 mg TE weekly, im, plus 300 µg/kg acyline (Multiple Peptide Systems, San Diego, CA) every 2 wk, sc (n = 6); 4) 100 mg TE weekly, im; 125 µg LNG daily, orally; plus 300 µg/kg acyline every 2 wk, sc (n = 5); and 5) controls (n = 7). At the completion of the treatment phase, men underwent testicular biopsy in conjunction with a previously planned vasectomy. Men in the control group proceeded directly to surgery.

Testicular biopsies

The testicular biopsies were performed by a board-certified urologist (R.B.) using local anesthesia. Tissue was obtained from one testis after incision of the tunica albuginea. Once the testis was visualized, individual biopsies were taken through a single tunical incision with curved scissors to ensure minimal disruption to testicular architecture. Approximately 250 mg of tissue was taken for stereological analysis, and 200 mg was taken for determination of testicular steroid concentrations.

Stereology

Tissue processing and estimation of germ and Sertoli cell number. Tissue for stereological analysis was directly immersion-fixed in Bouin’s fluid for approximately 3 h and then, using a dissecting microscope, divided into two parts along a plane lying at right angles to the dominant tubule orientation. One piece was then further processed and embedded in hydroxyethyl-methacrylate resin (Technovit 7100; Kulzer and Co., Friedrichsdorf, Germany) according to the manufacturer’s instructions, and 3 x 25-µm sections were cut, with a space of at least 100 µm in between, using a Supercut microtome (model 2050; Reichert Jung, Nossloch, Germany). The sections were then stained with periodic acid-Schiff using Schiff’s reagent (Amber Scientific, Belmont, Australia) and counterstained with hematoxylin.

Germ and Sertoli cells were identified based on their nuclear morphology and were counted using the optical disector technique as previously described in human biopsies (20). Germ cell data were expressed as number per Sertoli cell. Due to the helical arrangement of human spermatogenesis, germ cells could not be counted within each stage; thus, numbers were not corrected for stage frequency. Germ cells were grouped as follows: type A dark spermatogonia (Ad), type A pale spermatogonia (Ap), type B spermatogonia, preleptotene spermatocytes, leptotene spermatocytes, zygotene spermatocytes, pachytene spermatocytes, step 1–2 round spermatids (rST), step 3–6 elongating spermatids (3–6 ST), and step 7–8 elongated spermatids (7–8 ST).

Testicular and serum steroids. iTT, DHT, and 3{alpha}-Adiol were measured in testis homogenates (~110 mg tissue/sample) using HPLC and RIA, as previously described (2). In addition, iT3ß-Adiol and iTE2 were quantitated. Steroid recoveries per sample were monitored using radiolabeled [1,2,6,7,16,17-3H]T ([3H]T; 121 Ci/mmol), [1,2,3,4,5,6-3H]DHT ([3H]DHT; 110 Ci/mmol; NEN Life Sciences, Boston, MA), and [1,2,3,4,5,6-3H]3{alpha}- or 3ß-Adiol ([3H]Adiol) as previously described (2). Percent recoveries were (mean ± SD for serum and testis, respectively; n = 31 for all groups): T, 53.2 ± 10.5 and 77.4 ± 5.2; DHT, 43.3 ± 9.9 and 30.2 ± 6.2; 3{alpha}-Adiol, 38.8 ± 10.8 and 40.1 ± 4.3; and 3ß-Adiol, 39.8 ± 12.7 and 43.2 ± 9.0. E2 recoveries could not be assessed using a tritiated tracer, because both T and E2 coeluted from the HPLC column. Independent experiments in this laboratory showed that E2 recoveries paralleled those for T, which were used for all calculations. T, DHT, 3{alpha}-Adiol, and 3ß-Adiol were quantitated using [125I]T as tracer and unlabeled T, DHT, 3{alpha}-Adiol, or 3ß-Adiol as standards, respectively (2). E2 concentrations were determined using a double-antibody RIA format similar to that used for the androgens, with an in-house iodinated histamine-E2 tracer and an anti-E2 primary antibody (no. 4410; Diagnostic Systems Laboratories, Webster, TX). The cross-reactivity of this antibody for T was less than 0.01% as stated by the supplier and 0.00009% as determined by this RIA. The cross-reactivities for estrone and estriol were 2.40% and 0.64%, respectively (Diagnostic Systems Laboratories), and the sensitivity of the assay was 0.8 pg/ml. All samples were assayed in two or three assays. The intraassay variations (n = 3) from parallel fragments from the same testis sample were 8.7% for T, 6.8% for DHT, 2.1% for 3{alpha}-Adiol, 8.8% for 3ß-Adiol, and 6.0% for E2. Interassay variations were 14.1% for T, 13.9% for DHT, 12.7% for 3{alpha}-Adiol, 8.5% for 3ß-Adiol, and 23% for E2.

Statistical analyses

Data are shown as the mean ± SEM (n = 5–7/group) for all parameters. Statistical comparisons were performed using SigmaStat (Systat Software, Inc., Point Richmond, CA). All data were log transformed before analysis, and the variances were analyzed for homogeneity. Differences between control and treatment groups were analyzed by one-way ANOVA, followed by the Student-Newman-Keuls method or, when normality testing failed, by Kruskal-Wallis one-way ANOVA on ranks, followed by Dunn’s method for post hoc comparison. Relationships among germ cell subtypes, serum and iT steroid levels, and sperm concentration were analyzed by linear regression.


   Results
Top
 Abstract
 Introduction
 Subjects and Methods
 Results
Discussion
 References
 
Germ cell numbers

Germ cell data for control and MHC-treated men are shown expressed as number per Sertoli cell (Table 1Go) and as a percentage of the control value (Fig. 1Go). No significant differences in germ cell number between treatment groups were seen. Relative to controls, all germ cell subtypes from type B spermatogonia onward were significantly suppressed by the MHC treatment combinations. There were several settings in which treatment response differed relative to controls: 1) only the acyline-treated groups showed suppression of type B spermatogonia and preleptotene spermatocytes, and 2) only the TE, LNG, plus dutasteride and TE, LNG, plus acyline groups significantly reduced 3–6 ST.


View this table:
[in this window]
[in a new window]
  		TABLE 1. Germ cell numbers expressed as number per Sertoli cell (N/SC) in testicular biopsies from men receiving either no treatment (control) or 8 wk of MHC treatments [TE plus LNG; TE plus LNG plus dutasteride (D); TE plus acyline (Acy); TE plus LNG plus Acy]

 


View larger version (20K):
[in this window]
[in a new window]
  		FIG. 1. Changes in germ cell number over the 8-wk treatment phase in men (n = 5–6/group) randomized to receive TE (100 mg weekly, im) combined with 1) 125 µg LNG daily, orally; 2) 125 µg LNG plus 0.5 mg dutasteride (D) daily, orally; 3) 300 µg/kg acyline (ACY) every 2 wk, sc; or 4) 125 µg LNG plus 300 µg/kg ACY every 2 wk. Data are expressed as a percentage of the value in control subjects (n = 7). Germ cell subtypes are described along the x-axis as follows; all type A spermatogonia (All A), Ad, Ap, type B spermatogonia (B), preleptotene spermatocytes (Pl), leptotene to zygotene spermatocytes (L-Z), pachytene spermatocytes (PS), rST, 3–6 ST, 7–8 ST, and sperm concentration in the ejaculate on the day of testicular biopsy (sperm). Data are shown as the mean ± SEM. *, P < 0.05, two or more treatments significantly reduced germ cell numbers from control values. No differences between treatment groups were detected.

 
Individual men displayed widely variable patterns of spermatogenic suppression within each of the four treatment groups (Fig. 2Go). A general reduction across all germ cell subtypes was seen in some men, whereas in others, discernible patterns of impaired developmental progression were apparent: failed spermatogonial maturation (Fig. 3AGo), failed meiotic entry (Fig. 3BGo), and either complete or partial spermiation failure (Fig. 3CGo). No significant correlation between any germ cell subtype and sperm concentration on the day of biopsy could be detected.



View larger version (31K):
[in this window]
[in a new window]
  		FIG. 2. Changes in germ cell number over the 8-wk treatment phase in men (n = 5–6/group; numbers represent individual subjects) randomized to receive TE (100 mg weekly, im) combined with either 125 µg LNG daily, orally (A); 125 µg LNG plus 0.5 mg dutasteride (D) daily, orally (B); 300 µg/kg acyline (ACY) every 2 wk, sc (C); or 125 µg LNG plus 300 µg/kg ACY every 2 wk (D). Data are expressed as a percentage of the control values (n = 7). Germ cell subtypes are described along the x-axis as follows: all type A spermatogonia (All A), Ad, Ap, type B spermatogonia (B), preleptotene spermatocytes (Pl), leptotene to zygotene spermatocytes (L-Z), pachytene spermatocytes (PS), rST, 3–6 ST, 7–8 ST, and sperm concentration in the ejaculate on the day of testicular biopsy (sperm).

 


View larger version (19K):
[in this window]
[in a new window]
  		FIG. 3. Changes in germ cell number over the 8-wk treatment phase in individual men displaying failed spermatogonial maturation [A; subjects 116 (TE plus acyline) and 136 (TE, LNG, plus acyline)], failed meiotic entry [B; subjects 107 and 108 (TE, LNG, plus dutasteride)], and spermiation failure [C; complete and partial; subjects 114 (TE plus acyline), 123 (TE plus LNG), 132 (TE, LNG, plus dutasteride), and 137 (TE plus acyline)]. Data are expressed as a percentage of the control values (n = 7). Germ cell subtypes are described along the x-axis as follows: all type A spermatogonia (All A), Ad, Ap, type B spermatogonia (B), preleptotene spermatocytes (Pl), leptotene to zygotene spermatocytes (L-Z), pachytene spermatocytes (PS), rST, 3–6 ST, and 7–8 ST. The logarithmic second y-axis depicts the sperm concentration in the ejaculate on the day of testicular biopsy (sperm), with sperm concentrations of 0 million/ml assigned a value of 0.001.

 
Serum and iT steroid levels

Serum and iT steroid levels are shown in Tables 2Go and 3Go. In controls, the iT steroids levels compared with serum levels were as follows: T was 123-fold higher (2,460 vs. 20 nmol/liter), DHT was 6-fold higher (26 vs. 4.7 nmol/liter), 3{alpha}-Adiol was 10-fold higher (81 vs. 8.1 nmol/liter), 3ß-Adiol was 33-fold higher (158 vs. 4.8 nmol/liter), and E2 was 407-fold higher (11,000 vs. 27 pmol/liter). A 17-fold difference in iTT levels was noted within the control group (range, 227–3,841 nmol/liter) despite a lack of differences in baseline characteristics, serum hormones, or sperm concentrations. Other iT steroids also displayed variability, but to a much lesser extent than iTT (iTDHT, 10–48 nmol/liter; iT3{alpha}Adiol, 50–126 nmol/liter; iT3ßAdiol, 54–290 nmol/liter; iTE2, 5–14 nmol/liter).


View this table:
[in this window]
[in a new window]
  		TABLE 2. Serum steroid levels in men undergoing no treatment (control) vs. 8 wk of MHC treatments [TE plus LNG; TE plus LNG plus dutasteride (D); TE plus acyline (Acy); TE plus LNG plus Acy]

 

View this table:
[in this window]
[in a new window]
  		TABLE 3. Intratesticular steroid levels in men undergoing no treatment (control) vs. 8 wk of MHC treatments [TE plus LNG; TE plus LNG plus dutasteride (D); TE plus acyline (Acy); TE plus LNG plus Acy]

 
All treatments resulted in similar and marked falls in iTT (1.9–2.6% of control; P < 0.001). All treatments except TE plus LNG resulted in a fall in iTDHT levels (13–29% of control; P < 0.05), with dutasteride providing additional suppression of iTDHT (13% of control; P < 0.05) beyond that of all other treatment groups. Similarly, all treatment groups except TE plus LNG saw a reduction in iT3{alpha}Adiol (44–47% of control; P < 0.05). All treatments produced similar reductions in iT3ßAdiol (16–34% of control; P < 0.001). The mean iTE2 level in controls was 11,000 ± 3,320 pmol/liter (range, 4,804–29,621 pmol/liter), with a significant reduction in iTE2 levels in the TE plus acyline group (28% of control; P = 0.011) and all other treatment groups that approached significance (P = 0.053).

The iTT to iTDHT ratio, relative to the control value (123:1), was lowered by all treatments (4–31:1; P < 0.001–0.016). The addition of dutasteride to TE plus LNG (31:1) also maintained a higher ratio compared with the TE plus LNG group (4:1; P = 0.043). Ratios of iTT to total 5{alpha}-reduced steroids (DHT plus both Adiols) and of iTT to E2 did not differ between treatment groups (data not shown).

The only significant correlation found between iT steroids and germ cell numbers was a negative correlation between iT3{alpha}Adiol and the haploid germ cells (rST: r = 0.592; P = 0.004; 3–6 ST: r = 0.569; P = 0.006; 7–8 ST: r = 0.579; P = 0.005). No other significant correlations were found between the other concurrently measured iT steroids or serum gonadotropins and germ cell number.

Recovery and adverse events

No serious adverse events occurred, and all treatments were well tolerated (16). One study participant experienced a minor wound infection, which resolved rapidly with a 1-wk course of antibiotics.


   Discussion
Top
 Abstract
 Introduction
 Subjects and Methods
 Results
 Discussion
References
 
We report the effects of combination MHC treatment on germ cell subtypes and iT androgen and E2 levels. Treatments resulted in significant suppression of germ cell subtypes from type B spermatogonia onward; however, neither the addition of the dual type I and II 5{alpha}-reductase inhibitor, dutasteride, nor that of the new potent GnRH antagonist, acyline, provided substantial advantage over TE plus LNG treatment, as previously described for sperm concentration and serum gonadotropin levels (16). We also report that a high testicular:serum gradient exists for E2 and 5{alpha}-reduced androgens in addition to the recognized gradient for T (2, 19). Eight weeks of MHC-induced gonadotropin suppression markedly reduced iTT levels, with 5{alpha}-reduced androgens and E2 falling to a much lesser degree. All MHC treatments profoundly suppressed gonadotropins and iTT, but these parameters did not correlate with germ cell numbers. A striking heterogeneity in the germ cell suppression pattern was seen between men across all treatments, suggesting intrinsic differences in the susceptibility of spermatogenesis to gonadotropin withdrawal.

Germ cell numbers

Germ cell numbers in control men were similar to previous reports (20, 40), as was the pattern of germ cell suppression seen after 8 wk of treatment, which was consistent with a wave of spermatogenic failure after gonadotropin withdrawal. The extent and time frame of germ cell changes were in keeping with primary lesions in 1) type A to B spermatogonial development and 2) inhibition of spermiation resulting in the rapid onset of oligo- or azoospermia despite the continued presence of elongated spermatids (20, 40). Germ cell numbers were similarly suppressed by all treatments, but differences between individual spermatogenic suppression patterns were evident.

With regard to type A spermatogonia, we previously observed a rise in the mitotically inactive Ad spermatogonia at 2 and 6 wk (20), but this was not seen with any treatment in the current study, suggesting that either the survival of type Ad spermatogonia or dedifferentiation of type Ap to Ad reaches a critical point between 6 and 8 wk of gonadotropin withdrawal. The maturation of type A to B spermatogonia is dependent on FSH (20, 40, 41, 42, 43, 44), with iTT having a proposed inhibitory role in rodents, but there are no supportive data in primates or humans as yet (45, 46, 47). Given that there were no significant differences seen between treatment groups in FSH and iTT levels, it is not surprising that mean type B spermatogonial numbers were similar.

A novel observation was the profound disruption of meiotic entry demonstrated in some subjects who showed relatively preserved type B spermatogonial numbers, but profoundly suppressed preleptotene spermatocyte numbers. Interestingly, once reaching the preleptotene spermatocyte stage, germ cells appeared to complete the first meiotic prophase in all subjects, as supported by our observation that there was little decline between preleptotene and pachytene spermatocyte numbers.

Spermiogenesis was markedly interrupted by 8 wk of all MHC treatments, as reflected by the reduction in haploid germ cell number (rST, 16–22% of control; 3–6 ST, 26–44% of control; 7–8 ST, 12–25% of control). This is in contrast with our previous report at a 6 wk point after administration of TE with or without DMPA (rST, 71–93% of control; 3–6 ST, 48–54% of control; 7–8 ST, 77–123% control) (20), suggesting the haploid germ cell survival is being critically effected by gonadotropin withdrawal within this 14-d window. The fall in rST is likely to be accounted for by the loss of the spermatogenic wave, but the decline in 7–8 ST without a fall in 3–6 ST, suggests the former may be more vulnerable to gonadotropin and/or iTT withdrawal. The fact that 12 wk of TE with or without DMPA (20) or 19–24 wk of TE treatment profoundly lowers all round and elongating spermatid numbers (e.g. 3–6 STs 7–10% and 6% of control, respectively) (40) indicates that eventually the earlier lesions flow through to all spermatogenic cells.

In the present study, eight subjects (spread across all treatment groups) displayed a failure of spermiation, as evidenced by the presence of 7–8 ST on biopsy (13–40% of control) with few or no sperm (0–0.05 million/ml) in the ejaculate. This supports our previous finding that the process of sperm disengagement is a major target of MHC treatment and is the likely basis for sperm concentrations falling more rapidly than would be expected from the interruption of spermatogonial maturation (2). In contrast was subject 117 (in the TE, LNG, plus acyline group), who showed profound suppression of haploid germ cells (6% of control, 7–8 ST), but maintained a biopsy day sperm concentration of 15 million/ml despite no apparent differences in baseline serum FSH or iTT from the study group (16) and supporting the idea that intrinsic differences in his spermatogenic process permitted continued sperm release. These data also point to the possibility that some men on long-term MHC with an ongoing low level of 7–8 ST spermatid development may be prone to appreciable rebounds in sperm concentration when escape from gonadotropin suppression allows a rapid restoration of the spermiation process (9).

A marked heterogeneity of germ cell suppression was observed within all treatment groups, with patterns from complete failure to relatively preserved spermatogenesis noted. In line with previous analyses (37, 48), we saw no differences in the subjects’ phenotypic characteristics, including physical examination, baseline gonadotropins, and sperm concentrations, and yet their responses to MHC treatment, as assessed by germ cell number, was highly variable. No germ cell subtype was related to sperm concentration, indicating that sperm concentration is a poor marker of the state of germ cell development. Germ cell subtypes could not be correlated with serum gonadotropins, iTT, or iTDHT, but a weak correlation between haploid germ cells and iT3{alpha}-Adiol was noted. To date, only a rise in 5{alpha}-reductase activity during treatment (5, 49) and an increased prevalence of extended androgen receptor CAG repeats have been shown to be associated with nonsuppression of sperm concentration in normal fertile men administered MHC treatment (50). The current stereological and iT steroid data do not appear to explain the variability in the suppression of sperm concentration in response to MHC.

Testicular steroids

Similar to previous reports using testicular biopsy or needle aspiration, (2, 19), a marked reduction in iTT levels was seen. Interestingly, a 17-fold difference in iTT levels was observed in control testes despite similar baseline gonadotropin levels and sperm concentrations, indicating widely variable set points for iTT in the maintenance of normal spermatogenesis. Upon reevaluation of past data, we noted that this has also been apparent in three previous studies (two percutaneous aspiration and one testicular biopsy), with iTT estimates covering a 2- to 34-fold range (2, 19, 51).

Levels of iTDHT were relatively preserved in the TE plus LNG group as seen previously with TE and TE plus DMPA treatments (2); however, dutasteride and acyline treatment resulted in significantly lower iTDHT levels compared with control values, with dutasteride providing additional suppression over all other treatment groups. Based on rat and human data, it has been hypothesized that lowering of iTT levels may up-regulate 5{alpha}-reductase activity to relatively maintain iTDHT levels, which are then able to support spermatogenesis (5, 49, 52, 53). However, in the present study the lowering of iTDHT levels in the dutasteride group did not result in significant additional suppression of any germ cell subtype, nor did iTDHT levels show a correlation with germ cell number. Failure to reveal an effect of lowering iTDHT may indicate that it is irrelevant to spermatogenesis or, rather, is a result of small group numbers and limited study duration. The addition of either dutasteride or acyline to TE plus LNG produced the lowest iTDHT levels, and these groups were the only ones to show significant suppression of 3–6 ST, suggesting possible involvement of DHT in midspermiogenesis. Alternatively, the maximal extent of iTDHT suppression reached in the TE, LNG, plus dutasteride group (13% of control) may still have exceeded the threshold for the maintenance of germ cell development.

High levels of iTE2 were found in control testes (mean, 11,000 ± 3,300 pmol/liter), approximately 2.3 times higher than those previously reported in human spermatic vein sampling in normal men (54), but somewhat lower than the results of a biopsy study of previously well men in stage IV coma in an intensive care unit setting during the process of multiple organ sampling (44,100 ± 7,340 pmol/liter) (55). Control men displayed a 3-fold larger testes:serum E2 concentration gradient (x407) than that seen for T (x120). iTE2 levels were relatively preserved (14–37% of control) after 8 wk of MHC treatment despite the marked fall in T substrate levels (to 2% of control). Testicular conversion of T to E2 is mediated through aromatase, controlled by the CYP19 gene with a tissue-specific promoter (PII) (56). FSH, LH, T, and DHT are known to induce aromatase expression in Leydig, Sertoli, and germ cells in rodent models (57); however, the regulation of aromatase and E2 production in the human testis remains to be clarified. We suggest that the relative preservation of iTE2 levels is less likely to be due to up-regulation of aromatase, but, rather, relates to the fact that even the marked reduction of T substrate has relatively little impact on E2 production due to the Km of aromatase for T being approximately 40 nmol (58).

In conclusion, using stereological approaches to estimate germ cell number, we have not found substantial differences as a result of adding dutasteride and acyline to a conventional T plus progestin regimen. Although dutasteride provided a significant reduction in iTDHT over other treatments, this did not translate into greater germ cell suppression. We have confirmed that spermatogonial maturation and spermiation are the principal sites in spermatogenesis affected by MHC, but have also described the range of responses seen between men regardless of their treatment and despite similarly marked reductions in gonadotropin and iT steroid levels. These include a potential lesion at meiosis I entry between type B spermatogonia and preleptotene spermatocytes. The heterogeneity of response, regardless of the MHC regimen, points to intrinsic differences between individuals and the need to explore genetic/proteomic approaches to understand the variable sensitivity of sites in germ cell development, rather than therapeutic paradigms relying solely on gonadotropin suppression for the development of universal and reliable azoospermia.


   Acknowledgments
 
The excellent assistance of Fiona McLean for serum and iT steroid assays, Georgia Balourdos for germ cell counting, clinical research coordinators Ms. Jennifer Bullock and Ms. Amanda Wiseman, and clinical research nurses Ms. Marilyn Busher and Ms. Pam Lovey is acknowledged.


   Footnotes
 
This work was supported by the National Institute of Child Health and Human Development, Male Contraception Research Center Grant 654-HD-42454, and by the Australian National Health and Medical Research Council, Program Grant 241000 and Postgraduate Scholarship Grant ID-241031 (to K.L.M.).

First Published Online July 19, 2005

Abbreviations: Ad, Type A dark spermatogonia; Adiol, androstanediol; Ap, type A pale spermatogonia; DHT, dihydrotestosterone; DMPA, depot medroxyprogesterone acetate; E2, estradiol; ER, estrogen receptor; iT, intratesticular; LNG, levonorgestrel; MHC, male hormonal contraception; PR, progesterone receptor; rST, step 1–2 round spermatid; 3–6 ST, step 3–6 elongating spermatid; 7–8 ST, step 7–8 elongated spermatid; T, testosterone; TE, T enanthate.

Received March 23, 2005.

Accepted July 12, 2005.


   References
Top
 Abstract
 Introduction
 Subjects and Methods
 Results
 Discussion
 References
 

  1. Nieschlag E, Zitzmann M, Kamischke A 2003 Use of progestins in male contraception. Steroids 68:965–972[CrossRef][Medline]
  2. McLachlan RI, O’Donnell L, Stanton PG, Balourdos G, Frydenberg M, de Kretser DM, Robertson DM 2002 Effects of testosterone plus medroxyprogesterone acetate on semen quality, reproductive hormones, and germ cell populations in normal young men. J Clin Endocrinol Metab 87:546–556[Abstract/Free Full Text]
  3. Turner AI, Tilbrook AJ, Clarke IJ, Scott CJ 2001 Progesterone and testosterone in combination act in the hypothalamus of castrated rams to regulate the secretion of LH. J Endocrinol 169:291–298[Abstract]
  4. Anawalt BD, Bebb RA, Bremner WJ, Matsumoto AM 1999 A lower dosage levonorgestrel and testosterone combination effectively suppresses spermatogenesis and circulating gonadotropin levels with fewer metabolic effects than higher dosage combinations. J Androl 20:407–414[Abstract/Free Full Text]
  5. Anderson RA, Wallace AM, Wu FC 1996 Comparison between testosterone enanthate-induced azoospermia and oligozoospermia in a male contraceptive study. III. Higher 5{alpha}-reductase activity in oligozoospermic men administered supraphysiological doses of testosterone. J Clin Endocrinol Metab 81:902–908[Abstract]
  6. Anawalt BD, Herbst KL, Matsumoto AM, Mulders TM, Coelingh-Bennink HJ, Bremner WJ 2000 Desogestrel plus testosterone effectively suppresses spermatogenesis but also causes modest weight gain and high-density lipoprotein suppression. Fertil Steril 74:707–714[CrossRef][Medline]
  7. Wu FC, Balasubramanian R, Mulders TM, Coelingh-Bennink HJ 1999 Oral progestogen combined with testosterone as a potential male contraceptive: additive effects between desogestrel and testosterone enanthate in suppression of spermatogenesis, pituitary-testicular axis, and lipid metabolism. J Clin Endocrinol Metab 84:112–122[Abstract/Free Full Text]
  8. Kinniburgh D, Anderson RA, Baird DT 2001 Suppression of spermatogenesis with desogestrel and testosterone pellets is not enhanced by addition of finasteride. J Androl 22:88–95[Abstract]
  9. Anderson RA, Kinniburgh D, Baird DT 2002 Suppression of spermatogenesis by etonogestrel implants with depot testosterone: potential for long-acting male contraception. J Clin Endocrinol Metab 87:3640–3649[Abstract/Free Full Text]
  10. Kamischke A, Venherm S, Ploger D, von Eckardstein S, Nieschlag E 2001 Intramuscular testosterone undecanoate and norethisterone enanthate in a clinical trial for male contraception. J Clin Endocrinol Metab 86:303–309
  11. Kamischke A, Heuermann T, Kruger K, von Eckardstein S, Schellschmidt I, Rubig A, Nieschlag E 2002 An effective hormonal male contraceptive using testosterone undecanoate with oral or injectable norethisterone preparations. J Clin Endocrinol Metab 87:530–539[Abstract/Free Full Text]
  12. Meriggiola MC, Bremner WJ, Costantino A, Pavani A, Capelli M, Flamigni C 1997 An oral regimen of cyproterone acetate and testosterone undecanoate for spermatogenic suppression in men. Fertil Steril 68:844–850[CrossRef][Medline]
  13. Meriggiola MC, Bremner WJ, Paulsen CA, Valdiserri A, Incorvaia L, Motta R, Pavani A, Capelli M, Flamigni C 1996 A combined regimen of cyproterone acetate and testosterone enanthate as a potentially highly effective male contraceptive. J Clin Endocrinol Metab 81:3018–3023[Abstract]
  14. Erb K, Pechstein B, Schueler A, Engel J, Hermann R 2000 Pituitary and gonadal endocrine effects and pharmacokinetics of the novel luteinizing hormone-releasing hormone antagonist teverelix in healthy men–a first-dose-in-humans study. Clin Pharmacol Ther 67:660–669[Medline]
  15. Behre HM, Klein B, Steinmeyer E, McGregor GP, Voigt K, Nieschlag E 1992 Effective suppression of luteinizing hormone and testosterone by single doses of the new gonadotropin-releasing hormone antagonist cetrorelix (SB-75) in normal men. J Clin Endocrinol Metab 75:393–398[Abstract]
  16. Matthiesson KL, Amory JK, Berger R, Ugoni A, McLachlan RI, Bremner WJ 2005 Novel male hormonal contraceptive combinations: the hormonal and spermatogenic effects of testosterone and levonorgestrel combined with a 5{alpha}-reductase inhibitor or gonadotropin-releasing hormone antagonist. J Clin Endocrinol Metab 90:91–97[Abstract/Free Full Text]
  17. McLachlan RI, McDonald J, Rushford D, Robertson DM, Garrett C, Baker HW 2000 Efficacy and acceptability of testosterone implants, alone or in combination with a 5{alpha}-reductase inhibitor, for male hormonal contraception. Contraception 62:73–78[CrossRef][Medline]
  18. Morse HC, Horike N, Rowley MJ, Heller CG 1973 Testosterone concentrations in testes of normal men: effects of testosterone propionate administration. J Clin Endocrinol Metab 37:882–886[Medline]
  19. Coviello AD, Bremner WJ, Matsumoto AM, Herbst KL, Amory JK, Anawalt BD, Yan X, Brown TR, Wright WW, Zirkin BR, Jarow JP 2004 Intratesticular testosterone concentrations comparable with serum levels are not sufficient to maintain normal sperm production in men receiving a hormonal contraceptive regimen. J Androl 25:931–938[Abstract/Free Full Text]
  20. McLachlan RI, O’Donnell L, Meachem SJ, Stanton PG, de Kretser DM, Pratis K, Robertson DM 2002 Identification of specific sites of hormonal regulation in spermatogenesis in rats, monkeys, and man. Recent Prog Horm Res 57:149–179[Abstract/Free Full Text]
  21. O’Donnell L, Robertson KM, Jones ME, Simpson ER 2001 Estrogen and spermatogenesis. Endocr Rev 22:289–318[Abstract/Free Full Text]
  22. Pentikainen V, Erkkila K, Suomalainen L, Parvinen M, Dunkel L 2000 Estradiol acts as a germ cell survival factor in the human testis in vitro. J Clin Endocrinol Metab 85:2057–2067[Abstract/Free Full Text]
  23. Saunders PT, Sharpe RM, Williams K, Macpherson S, Urquart H, Irvine DS, Millar MR 2001 Differential expression of oestrogen receptor {alpha} and ß proteins in the testes and male reproductive system of human and non-human primates. Mol Hum Reprod 7:227–236[Abstract/Free Full Text]
  24. Taylor AH, Al-Azzawi F 2000 Immunolocalisation of oestrogen receptor ß in human tissues. J Mol Endocrinol 24:145–155[Abstract]
  25. Pelletier G, El-Alfy M 2000 Immunocytochemical localization of estrogen receptors {alpha} and ß in the human reproductive organs. J Clin Endocrinol Metab 85:4835–4840[Abstract/Free Full Text]
  26. Dupont S, Krust A, Gansmuller A, Dierich A, Chambon P, Mark M 2000 Effect of single and compound knockouts of estrogen receptors {alpha} (ER{alpha}) and ß (ERß) on mouse reproductive phenotypes. Development 127:4277–4291[Abstract]
  27. Couse JF, Hewitt SC, Bunch DO, Sar M, Walker VR, Davis BJ, Korach KS 1999 Postnatal sex reversal of the ovaries in mice lacking estrogen receptors {alpha} and ß. Science 286:2328–2331[Abstract/Free Full Text]
  28. Eddy EM, Washburn TF, Bunch DO, Goulding EH, Gladen BC, Lubahn DB, Korach KS 1996 Targeted disruption of the estrogen receptor gene in male mice causes alteration of spermatogenesis and infertility. Endocrinology 137:4796–4805[Abstract]
  29. Robertson KM, O’Donnell L, Jones ME, Meachem SJ, Boon WC, Fisher CR, Graves KH, McLachlan RI, Simpson ER 1999 Impairment of spermatogenesis in mice lacking a functional aromatase (cyp 19) gene. Proc Natl Acad Sci USA 96:7986–7991[Abstract/Free Full Text]
  30. Rivier JE, Jiang G, Porter J, Hoeger CA, Craig AG, Corrigan A, Vale W, Rivier CL 1995 Gonadotropin-releasing hormone antagonists: novel members of the azaline B family. J Med Chem 38:2649–2662[CrossRef][Medline]
  31. Clark RV, Hermann DJ, Cunningham GR, Wilson TH, Morrill BB, Hobbs S 2004 Marked suppression of dihydrotestosterone in men with benign prostatic hyperplasia by dutasteride, a dual 5{alpha}-reductase inhibitor. J Clin Endocrinol Metab 89:2179–2184[Abstract/Free Full Text]
  32. Handelsman DJ, Conway AJ, Howe CJ, Turner L, Mackey MA 1996 Establishing the minimum effective dose and additive effects of depot progestin in suppression of human spermatogenesis by a testosterone depot. J Clin Endocrinol Metab 81:4113–4121[Abstract/Free Full Text]
  33. Robertson DM, Pruysers E, Stephenson T, Pettersson K, Morton S, McLachlan RI 2001 Sensitive LH and FSH assays for monitoring low serum levels in men undergoing steroidal contraception. Clin Endocrinol (Oxf) 55:331–339[CrossRef][Medline]
  34. McLachlan RI, Robertson DM, Pruysers E, Ugoni A, Matsumoto AM, Anawalt BD, Bremner WJ, Meriggiola C 2004 Relationship between serum gonadotropins and spermatogenic suppression in men undergoing steroidal contraceptive treatment. J Clin Endocrinol Metab 89:142–149[Abstract/Free Full Text]
  35. Shah C, Modi D, Sachdeva G, Gadkar S, Puri C 2005 Coexistence of intracellular and membrane-bound progesterone receptors in human testis. J Clin Endocrinol Metab 90:474–483[Abstract/Free Full Text]
  36. 1990 Contraceptive efficacy of testosterone-induced azoospermia in normal men. World Health Organization Task Force on Methods for the Regulation of Male Fertility. Lancet 336:955–959
  37. 1996 Contraceptive efficacy of testosterone-induced azoospermia and oligozoospermia in normal men. Fertil Steril 65:821–829
  38. Gu YQ, Wang XH, Xu D, Peng L, Cheng LF, Huang MK, Huang ZJ, Zhang GY 2003 A multicenter contraceptive efficacy study of injectable testosterone undecanoate in healthy Chinese men. J Clin Endocrinol Metab 88:562–568[Abstract/Free Full Text]
  39. Turner L, Conway AJ, Jimenez M, Liu PY, Forbes E, McLachlan RI, Handelsman DJ 2003 Contraceptive efficacy of a depot progestin and androgen combination in men. J Clin Endocrinol Metab 88:4659–4667[Abstract/Free Full Text]
  40. Zhengwei Y, Wreford NG, Royce P, de Kretser DM, McLachlan RI 1998 Stereological evaluation of human spermatogenesis after suppression by testosterone treatment: heterogeneous pattern of spermatogenic impairment. J Clin Endocrinol Metab 83:1284–1291[Abstract/Free Full Text]
  41. Narula A, Gu YQ, O’Donnell L, Stanton PG, Robertson DM, McLachlan RI, Bremner WJ 2002 Variability in sperm suppression during testosterone administration to adult monkeys is related to follicle stimulating hormone suppression and not to intratesticular androgens. J Clin Endocrinol Metab 87:3399–3406[Abstract/Free Full Text]
  42. Zhengwei Y, Wreford NG, Schlatt S, Weinbauer GF, Nieschlag E, McLachlan RI 1998 Acute and specific impairment of spermatogonial development by GnRH antagonist-induced gonadotrophin withdrawal in the adult macaque (Macaca fascicularis). J Reprod Fertil 112:139–147
  43. Weinbauer GF, Schlatt S, Walter V, Nieschlag E 2001 Testosterone-induced inhibition of spermatogenesis is more closely related to suppression of FSH than to testicular androgen levels in the cynomolgus monkey model (Macaca fascicularis). J Endocrinol 168:25–38[Abstract]
  44. O’Donnell L, Narula A, Balourdos G, Gu YQ, Wreford NG, Robertson DM, Bremner WJ, McLachlan RI 2001 Impairment of spermatogonial development and spermiation after testosterone-induced gonadotropin suppression in adult monkeys (Macaca fascicularis). J Clin Endocrinol Metab 86:1814–1822[Abstract/Free Full Text]
  45. Meistrich ML, Kangasniemi M 1997 Hormone treatment after irradiation stimulates recovery of rat spermatogenesis from surviving spermatogonia. J Androl 18:80–87[Abstract/Free Full Text]
  46. Meachem SJ, Wreford NG, Robertson DM, McLachlan RI 1997 Androgen action on the restoration of spermatogenesis in adult rats: effects of human chorionic gonadotrophin, testosterone and flutamide administration on germ cell number. Int J Androl 20:70–79[CrossRef][Medline]
  47. Meachem SJ, Wreford NG, Stanton PG, Robertson DM, McLachlan RI 1998 Follicle-stimulating hormone is required for the initial phase of spermatogenic restoration in adult rats following gonadotropin suppression. J Androl 19:725–735[Abstract/Free Full Text]
  48. Handelsman DJ, Farley TM, Peregoudov A, Waites GM 1995 Factors in nonuniform induction of azoospermia by testosterone enanthate in normal men. World Health Organization Task Force on Methods for the Regulation of Male Fertility. Fertil Steril 63:125–133[Medline]
  49. Anderson RA, Kelly RW, Wu FC 1997 Comparison between testosterone enanthate-induced azoospermia and oligozoospermia in a male contraceptive study. V. Localization of higher 5{alpha}-reductase activity to the reproductive tract in oligozoospermic men administered supraphysiological doses of testosterone. J Androl 18:366–371[Abstract/Free Full Text]
  50. von Eckardstein S, Syska A, Gromoll J, Kamischke A, Simoni M, Nieschlag E 2001 Inverse correlation between sperm concentration and number of androgen receptor CAG repeats in normal men. J Clin Endocrinol Metab 86:2585–2590[Abstract/Free Full Text]
  51. Coviello AD, Matsumoto AM, Bremner WJ, Herbst KL, Amory JK, Anawalt BD, Sutton PR, Wright WW, Brown TR, Yan X, Zirkin BR, Jarow JP 2005 Low dose human chorionic gonadotropin maintains intratesticular testosterone in normal men with testosterone induced gonadotropin suppression. J Clin Endocrinol Metab 90:2595–2602[Abstract/Free Full Text]
  52. O’Donnell L, Pratis K, Stanton PG, Robertson DM, McLachlan RI 1999 Testosterone-dependent restoration of spermatogenesis in adult rats is impaired by a 5{alpha}-reductase inhibitor. J Androl 20:109–117[Abstract/Free Full Text]
  53. Pratis K, O’Donnell L, Ooi GT, Stanton PG, McLachlan RI, Robertson DM 2003 Differential regulation of rat testicular 5{alpha}-reductase type 1 and 2 isoforms by testosterone and FSH. J Endocrinol 176:393–403[Abstract]
  54. Weinstein RL, Kelch RP, Jenner MR, Kaplan SL, Grumbach MM 1974 Secretion of unconjugated androgens and estrogens by the normal and abnormal human testis before and after human chorionic gonadotropin. J Clin Invest 53:1–6
  55. Marie E, Galeraud-Denis I, Carreau S 2001 Increased testicular steroid concentrations in patients with idiopathic infertility and normal FSH levels. Arch Androl 47:177–184[Medline]
  56. Lanzino M, Catalano S, Genissel C, Ando S, Carreau S, Hamra K, McPhaul MJ 2001 Aromatase messenger RNA is derived from the proximal promoter of the aromatase gene in Leydig, Sertoli, and germ cells of the rat testis. Biol Reprod 64:1439–1443[Abstract/Free Full Text]
  57. Bourguiba S, Genissel C, Lambard S, Bouraima H, Carreau S 2003 Regulation of aromatase gene expression in Leydig cells and germ cells. J Steroid Biochem Mol Biol 86:335–343[CrossRef][Medline]
  58. Yoshida N, Osawa Y 1991 Purification of human placental aromatase cytochrome P-450 with monoclonal antibody and its characterization. Biochemistry 30:3003–3010[CrossRef][Medline]



This article has been cited by other articles:


Home page
 Endocr. Rev.Home page
S. T. Page, J. K. Amory, and W. J. Bremner
Advances in Male Contraception
Endocr. Rev., June 1, 2008; 29(4): 465 - 493.
[Abstract] [Full Text] [PDF]

Home page
 Hum ReprodHome page
S. M. Ruwanpura, R. I. McLachlan, K. L. Matthiesson, and S. J. Meachem
Gonadotrophins regulate germ cell survival, not proliferation, in normal adult men
Hum. Reprod., February 1, 2008; 23(2): 403 - 411.
[Abstract] [Full Text] [PDF]

Home page
 J AndrolHome page
S. T. Page, T. F. Kalhorn, W. J. Bremner, B. D. Anawalt, A. M. Matsumoto, and J. K. Amory
Intratesticular Androgens and Spermatogenesis During Severe Gonadotropin Suppression Induced by Male Hormonal Contraceptive Treatment
J Androl, September 1, 2007; 28(5): 734 - 741.
[Abstract] [Full Text] [PDF]

Home page
 J EndocrinolHome page
S. J Meachem, S. Schlatt, S. M Ruwanpura, and P. G Stanton
The effect of testosterone, dihydrotestosterone and oestradiol on the re-initiation of spermatogenesis in the adult photoinhibited Djungarian hamster
J. Endocrinol., March 1, 2007; 192(3): 553 - 561.
[Abstract] [Full Text] [PDF]

Home page
 J. Clin. Endocrinol. Metab.Home page
K. L. Matthiesson, R. I. McLachlan, L. O'Donnell, M. Frydenberg, D. M. Robertson, P. G. Stanton, and S. J. Meachem
The Relative Roles of Follicle-Stimulating Hormone and Luteinizing Hormone in Maintaining Spermatogonial Maturation and Spermiation in Normal Men
J. Clin. Endocrinol. Metab., October 1, 2006; 91(10): 3962 - 3969.
[Abstract] [Full Text] [PDF]

Home page
 J AndrolHome page
A. O. Hammoud, M. Gibson, C. M. Peterson, B. D. Hamilton, and D. T. Carrell
Obesity and Male Reproductive Potential
J Androl, September 1, 2006; 27(5): 619 - 626.
[Full Text] [PDF]

Home page
 J. Clin. Endocrinol. Metab.Home page
M. J. Walton, R. A. L. Bayne, I. Wallace, D. T. Baird, and R. A. Anderson
Direct Effect of Progestogen on Gene Expression in the Testis during Gonadotropin Withdrawal and Early Suppression of Spermatogenesis
J. Clin. Endocrinol. Metab., July 1, 2006; 91(7): 2526 - 2533.
[Abstract] [Full Text] [PDF]

Home page
 Hum Reprod UpdateHome page
K. L. Matthiesson and R. I. McLachlan
Male hormonal contraception: concept proven, product in sight?
Hum. Reprod. Update, July 1, 2006; 12(4): 463 - 482.
[Abstract] [Full Text] [PDF]

Home page
 J. Clin. Endocrinol. Metab.Home page
K. Bay, K. L. Matthiesson, R. I. McLachlan, and A.-M. Andersson
The Effects of Gonadotropin Suppression and Selective Replacement on Insulin-Like Factor 3 Secretion in Normal Adult Men
J. Clin. Endocrinol. Metab., March 1, 2006; 91(3): 1108 - 1111.
[Abstract] [Full Text] [PDF]

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow All Versions of this Article:
90/10/5647    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