1 Prince Henrys Institute of Medical Research, Monash Medical Centre and 2 Department of Obstetrics and Gynaecology, Monash University, Clayton, Victoria 3168, Australia
3 To whom correspondence should be addressed at: Prince Henrys Institute of Medical Research, PO Box 5152, Clayton, Victoria 3168, Australia. e-mail: rob.mclachlan{at}med.monash.edu.au
![]() |
Abstract |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Key words: FSH/inhibin/LH/Sertoli cell/testosterone
![]() |
Introduction |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
In-situ hybridization and immunocytochemical studies have revealed different patterns of - and
B-subunit expression in the tissue depending on the developmental age and species studied (Bergh and Cajander, 1990
; Forti et al., 1992
; Vannelli et al., 1992
; Majdic et al., 1997
; Andersson et al., 1998
). Using immunocytochemical methods, the
-subunit is localized to human spermatocytes, Sertoli and Leydig cells, while the
B-subunit is localized to Sertoli and Leydig cells and more controversially to spermatocytes and early spermatids (Bergh and Cajander, 1990
; Forti et al., 1992
; Vannelli et al., 1992
; Majdic et al., 1997
; Andersson et al., 1998
).
The secretion of inhibin is controlled by many factors including gonadotrophins and intrinsic factors involving Sertoli, Leydig and germ cells. In normal men, a single large dose (3000 IU) of recombinant human FSH (rhFSH) led to a doubling of serum inhibin B levels, reaching a peak within 72 h (Anawalt et al., 1996) with more recent studies showing that a dose of 225 IU was ineffective (Kinniburgh and Anderson, 2001
). A doseresponse study showed that administration of rhFSH (1000, 2000 and 3000 IU) led to significant increases in inhibin B, with the later two doses showing a significantly greater area under the curve than placebo or 1000 IU rhFSH (Kamischke et al., 2001
). Both FSH and hCG have been shown to significantly increase pro-
C levels (Kamischke et al., 2001
; Kinniburgh and Anderson, 2001
).
It is also recognized that the relationship between serum FSH and inhibin B is modulated by other factors. Several studies have shown that inhibin B levels fall markedly following severe testicular insults (chemotherapy, irradiation) correlating with the disappearance of germ cells (Wallace et al., 1997; Foppiani et al., 1999
; Petersen et al., 1999
). Similar findings have been observed in rats following methoxyacetic acid treatment (Allenby et al., 1991
). On the other hand, gonadotrophic suppression following exogenous sex steroid administration led to little or only a partial suppression of serum inhibin levels (Anderson et al., 1997
; Zhengwei et al., 1998
; Buchter et al., 1999
; Martin et al., 2000
; McLachlan et al., 2002
) with no relationship with either the duration of treatment or the extent of suppression of sperm count (Anderson et al., 1997
). The basis for this differential effect may be that the extent of inhibin B suppression is related to the degree of damage to the spermatogenic process with toxic agents leading to a more profound loss of germ cells, compared with that achieved with gonadotrophin suppression.
To examine the potential influence of germ cells on inhibin release from the testis (of both Sertoli and Leydig cell origin) we examined the acute effects of FSH and/or LH (using hCG as an LH substitute) administration on serum inhibin B and pro-C levels in normal men, and compared this response with that seen following spermatogenic suppression induced by a regimen developed for the purpose of hormonal contraception (Handelsman et al., 1996
). In addition we wished to examine the relationships between spermatogenic recovery, induced by FSH and/or hCG treatment, and serum inhibin B.
![]() |
Methods and materials |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Study design
Phase 1: Responses of serum inhibin B and pro-C to single dose gonadotrophin administration. Development of stimulation test protocol. Stimulation test 1
The overall study design is displayed in Figure 1. Twenty-five men were randomized into five groups (n = 5/group) to receive a single injection of one of the following: rhFSH (either Gonal-F, Serono, Sydney, Australia; or Puregon, Organon, Sydney, Australia) 600, 1200, 2400 IU s.c.; hCG (Profasi, Serono; or Pregnyl, Organon) 5000 IU s.c.; or FSH 1200 IU plus hCG 5000 IU. Blood was drawn (08001100 h) daily for 8 days beginning on the day prior to the injections in order to cover the likely period of inhibin rise (Anawalt et al., 1996).
|
For assessing the stimulatory response of serum inhibin, single doses of 1200 IU FSH alone or 5000 IU hCG or a combination of the two were administered by the investigators on day 0 in weeks 1 and 3 of the recovery phase. These doses were chosen based on the data from phase 1 with blood being obtained for each stimulation test on days 0, 4, 5 and 6. The control response (stimulation test 1) was used for comparison with those obtained after spermatogenic suppression (stimulation test 2) and week 3 in recovery (stimulation test 3). In the remaining weeks of the recovery phase (i.e. 2 and 412), men receiving FSH had their FSH doses administered on day 0 by the investigators and self-administered their second FSH dose on day 4 of each week. Men receiving hCG had their dose administered on day 0 of each week by the investigators. Semen analyses and blood samples were obtained weekly throughout the recovery phase.
Baseline characteristics for participants are shown in Table I. Following 12 weeks of gonadotrophin stimulation, men were followed at regular intervals to monitor recovery of FSH, LH and sperm counts.
|
Serum hCG was measured in an automated immunoassay carried out on a Dade Behring Dimension RxL Clinical Chemistry system with reagents and calibrators supplied by Dade Behring Diagnostics (Sydney, Australia). The assay sensitivity was 1.0 IU/l with an inter-assay variation of 3.96.1%. Testosterone was measured by an automated chemiluminescent immunoassay (Chiron Diagnostics, East Walpole, MA, USA) with a sensitivity of 0.3 nmol/l and inter-assay variations of 5.59.9%.
The pro-C enzyme-linked immunosorbent assay (ELISA) (Groome et al., 1995
) was employed using reagents [INPRO monoclonal antibody (Mab) as capture and R1 Mab as label] and a standard reference preparation provided by Oxford Bio-Innovation Ltd (Upper Heyford, Oxon, UK). The sensitivity of the assay was 2 ng/l. The within-assay variation and the between-assay variation were 89% (eight assays) and 16.5% (eight assays) respectively. The inhibin B ELISA method (Groome et al., 1996
) was employed using kit reagents and inhibin B standard reference provided by Oxford Bio-Innovation. The sensitivity of the assay was 4 ng/l. The within-assay and between-assay variations were 58% (eight assays) and 15% (eight assays) respectively.
Statistical analyses
Data are shown as mean ± SEM or mean percentage baseline. Statistical comparisons were made using Sigma Stat (SSPS Inc., Chicago, IL, USA). All data were log-transformed prior to analysis. Non-parametric statistics were used when equal variance testing failed. Serum hormone and semen data were analysed by either repeated measures analysis of variance (ANOVA) or Friedman repeated measures ANOVA followed by Tukey test to examine differences across time between treatment groups. Differences in percentage baseline inhibin B (day 0 compared with days 46) and pro-C (day 0 compared with days 45) in stimulation tests 1, 2 and 3 in each treatment group was evaluated using one-way ANOVA.
![]() |
Results |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
|
|
|
|
|
|
Spermatogenic suppression and recovery
Testosterone plus DMPA treatment led to marked suppression of spermatogenesis (0.080.21x106/ml) in all 15 men (Figure 6C). All recovery treatments led to a partial restoration in spermatogenesis with sperm counts in the hCG group ranging from 0.21 to 8.4x106/ml, in the FSH group between 0.07 and 10x106/ml and in the combined group between 0.21 and 69x106/ml. However, despite all groups showing partial restoration of spermatogenesis, serum inhibin B levels either fell with hCG, or returned toward control levels with FSH alone or combined with hCG. No clear relationship was apparent between serum inhibin B levels and sperm counts over the 12 week-induced recovery phase (Figure 6A, C).
Recovery and adverse events
Recovery to pre-study baseline was monitored in 13 of the 15 participants, one of the subjects in the FSH plus hCG group discontinued at 6 weeks for personal reasons. Thirteen men recovered normal sperm counts by 12 months. One man in the hCG group discontinued follow-up with a last recorded sperm count of 13x106/ml at 14 months post treatment. FSH, LH and testosterone levels recovered in all 14 men between 1 and 12 months. Three men required additional testosterone replacement (12 injections of testosterone esters 100250 mg) in the recovery phase. Two participants in the suppression phase developed mild gynaecomastia, one transiently, the other having persistent 1 cm non-tender unilateral enlargement. No other serious adverse events occurred.
![]() |
Discussion |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
There is considerable evidence that germ cells modify inhibin responses in both animal (Pineau et al., 1989, 1990; Maddocks et al., 1992
) and human models (Carreau, 1995
; Andersson et al., 1998
). The mechanism of this effect is unclear but germ cells may regulate the synthesis and incorporation of the
B subunit into dimeric inhibin by the Sertoli cell, the rate-limiting step in inhibin B synthesis (Clifton et al., 2002
).
In normal men, a positive relationship is recognized between serum inhibin and sperm output (Jensen et al., 1997; Klingmuller and Haidl, 1997
). Therefore in our model of spermatogenic suppression in normal men, one could postulate several possible impacts of germ cells on the Sertoli cell inhibin B secretory response. The most likely relationship would be that germ cells potentiate the ability of the Sertoli cell to secrete inhibin B, resulting in decreased secretion in the setting of germ cell depletion. Alternatively, if germ cells inhibit Sertoli cell inhibin B secretion, then in the setting of germ cell depletion, there may be an increase in its secretion, either basally or in response to FSH. Finally if there are non-germ cell factors modulating the ability of the Sertoli cell to secrete inhibin B, then there may be no significant change in Sertoli cell inhibin B response.
We have shown that the combined reduction in germ cell number and suppression of gonadotrophins resulted in significant falls in inhibin B and pro-C. However, germ cell depletion in normal young men did not modify the ability of the Sertoli cell inhibin B response to gonadotrophins. Comparison of the inhibin B response with FSH showed no significant differences between control (stimulation test 1) and following germ cell suppression (stimulation test 2).
The hypothesis that specific germ cell types might regulate inhibin in normal men (Pineau et al., 1990; Allenby et al., 1991
; Guitton et al., 2000
; Clifton et al., 2002
) is difficult, if not impossible, to assess as specific and complete loss of germ cell subtypes cannot be achieved. Germ cell types are suppressed to a variable degree by chronic gonadotrophin suppression in man (McLachlan et al., 2002
). Type A and B spermatogonia were suppressed to
70 and
20% of control levels respectively, with spermatocytes, round and elongated spermatids remaining at 20%, indicating that the type A pale to B spermatogonial maturation is the key site of effect. The reduction of sperm counts to levels <1% of control is in large part attributed to the failure of sperm release (McLachlan et al., 2002
). In contrast, iatrogenic spermatogenic failure, e.g. by cancer treatments, generally induces total spermatogenic failure, although these treatments may also alter Sertoli/Leydig cell function and interaction.
Alteration in Sertoli cell inhibin secretion in our model could also have resulted from either a chronic FSH deficiency modifying the ability of the Sertoli cells to respond to acute FSH stimulation and/or to the effect of changes in the germ cell complement. We attempted to address this issue by repeating the stimulation test in week 3 of FSH-induced recovery, by which time we speculated that any FSH-related defect in Sertoli cell function would have been corrected and that there would still be a low germ cell complement. However, there was no suggestion of a priming effect of FSH treatment leading to an increased inhibin B response in the third week.
An FSH doseresponse effect on pro-C secretion was seen, with
1200 IU FSH being required for a maximal response. The rise in pro-
C after hCG was more rapid (peak day 12) than with FSH, suggesting that the Leydig cell responds more rapidly to its tropic hormone than does the FSHSertoli cell axis. Additive effects of FSH and hCG on pro-
C were seen in both the single dose acute setting and during recovery, implying that both the Sertoli and Leydig cells contribute during combined treatment. This would also be in keeping with immunohistochemical data which have shown
-subunit staining in both Sertoli and Leydig cells (Bergh and Cajander, 1990
; Forti et al., 1992
; Vannelli et al., 1992
; Majdic et al., 1997
; Anderson et al., 1998
; Andersson et al., 1998
). As previously shown, hCG (as an LH substitute) does not stimulate inhibin B secretion (Kamischke et al., 2001
; Kinniburgh and Anderson, 2001
).
In contrast with the inhibin B response, germ cell loss appeared to enhance pro-C responses to both FSH and hCG. It is possible that germ cells may have an inhibitory effect on the production of the
-subunit by both the Sertoli and Leydig cells which, when removed, results in a greater secretion of this isoform. Equally possible is that the germ cells positively influence the production of the
B-subunit by the Sertoli cell to FSH and that their removal results in less dimerization and thus an apparent increase in the free
-subunit. However, this apparent increase in pro-
C may also relate to the lower baseline value for the test after gonadotrophin suppression (stimulation test 2). It should also be noted that there was a lesser pro-
C response with the presumed return of germ cells in the third week of recovery. In any event, it is clear that both Sertoli (FSH-dependent) and Leydig cell (hCG/LH-dependent) components respond more vigorously in terms of pro-
C secretion than inhibin B with germ cell depletion.
Germ cell/FSH/inhibin relationships may differ in other settings of testicular dysfunction. Despite severe germ cell depletion, high levels of -subunit-containing peptides are found in infertile men, including Klinefelters syndrome (Anawalt et al., 1996
), and chemotherapy-induced azoospermia (Wallace et al., 1997
), in which settings, inhibin B levels are very low or undetectable. This secretion of pro-
C is gonadotrophin sensitive as suppression of FSH/LH reduces the serum levels (Martin et al., 2000
; Kinniburgh et al., 2002
). The threshold for FSH and LH/hCG effects on both inhibin B and pro-
C may differ in these disease settings compared with those in the current study. The rise in pro-
C in male infertility may result from the much more severe loss of germ cell number (e.g. Sertoli cell-only syndrome) than that seen in our model, resulting in increased
-subunit secretion, and/or a loss of inhibin B secretory capacity. It has been previously suggested (Wallace et al., 1997
) that observed differences could be accounted for by the frequently elevated serum LH levels in these subjects leading to an increased secretion of
subunit by the Leydig cell. However, it is also possible that there is a contribution by preferential secretion of
-subunit by the Sertoli cell in the face of severe germ cell loss and high FSH levels.
One interesting incidental finding was the augmentation of hCG-induced testosterone secretion associated with co-administration of FSH, both in the single dose acute setting and over the weeks of recovery. These data support the previous observation (Levalle et al., 1998) that enhancement of Sertoli cell function by FSH stimulation may result in improved Leydig cell function via paracrine effects. Given that there are no FSH receptors on Leydig cells, it has been suggested that the increase in testosterone observed after the administration by FSH is possibly mediated via a Sertoli cell-released non-steroidal factor with a molecular mass of >50 kDa (Levalle et al., 1998
). However, this effect was not seen in subjects with acquired hypogonadotrophic hypogonadism in whom no further rise in serum testosterone was seen following 1 month of treatment with FSH with or without hCG (Young et al., 2000
). One potential explanation for these differences is that in the latter study, testicular size was markedly reduced (410 ml) probably due to spermatogenic failure. In turn this may have altered Sertoli cell function and its ability to augment Leydig cell testosterone secretion which was not restored by 1 month of gonadotrophin treatment.
In conclusion, we found that inhibin B in this setting of experimental germ cell depletion in normal men is a poor marker of spermatogenesis, as germ cell loss led to only minor changes in basal inhibin B levels and no modulation of FSH stimulatory response. Furthermore, during induced gonadotrophin recovery of spermatogenesis, no clear relationship was seen between sperm output and serum inhibin B. However, we caution that this conclusion should not be extrapolated to other settings of genetic or acquired defects of spermatogenesis wherein different FSH/inhibin B/spermatogenic relationships may exist and account for the observed reciprocal relationship between serum inhibin B and FSH in spermatogenic failure. Thus, while we see no benefit in the use of an FSH stimulation test to assess spermatogenic status in normal men, it may be useful in some settings of male infertility.
![]() |
Acknowledgements |
---|
![]() |
References |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Anawalt, B.D., Bebb, R.A., Matsumoto, A.M., Groome, N.P., Illingworth, P.J., McNeilly, A.S. and Bremner, W.J. (1996) Serum inhibin B levels reflect Sertoli cell function in normal men and men with testicular dysfunction. J. Clin. Endocrinol. Metab., 81, 33413345.[Abstract]
Anderson, R.A. (2001) Clinical studies: inhibin in the adult male. Mol. Cell. Endocrinol., 180, 109116.[CrossRef][ISI][Medline]
Anderson, R.A., Wallace, E.M., Groome, N.P., Bellis, A.J. and Wu, F.C. (1997) Physiological relationships between inhibin B, follicle stimulating hormone secretion and spermatogenesis in normal men and response to gonadotrophin suppression by exogenous testosterone. Hum. Reprod., 12, 746751.[Abstract]
Anderson, R.A., Irvine, D.S., Balfour, C., Groome, N.P. and Riley, S.C. (1998) Inhibin B in seminal plasma: testicular origin and relationship to spermatogenesis. Hum. Reprod., 13, 920926.[Abstract]
Andersson, A.M., Muller, J. and Skakkebaek, N.E. (1998) Different roles of prepubertal and postpubertal germ cells and Sertoli cells in the regulation of serum inhibin B levels. J. Clin. Endocrinol. Metab., 83, 44514458.
Bergh, A. and Cajander, S. (1990) Immunohistochemical localization of inhibin-alpha in the testes of normal men and in men with testicular disorders. Int. J. Androl., 13, 463469.[ISI][Medline]
Buchter, D., von Eckardstein, S., von Eckardstein, A., Kamischke, A., Simoni, M., Behre, H.M. and Nieschlag, E. (1999) Clinical trial of transdermal testosterone and oral levonorgestrel for male contraception. J. Clin. Endocrinol. Metab., 84, 12441249.
Carreau, S. (1995) Human Sertoli cells produce inhibin in vitro: an additional marker to assess the seminiferous epithelium development. Hum. Reprod., 10, 19471949.[ISI][Medline]
Clifton, R.J., ODonnell, L. and Robertson, D.M. (2002) Pachytene spermatocytes in co-culture inhibit rat Sertoli cell synthesis of inhibin beta B-subunit and inhibin B but not the inhibin alpha-subunit. J. Endocrinol., 172, 565574.
Foppiani, L., Schlatt, S., Simoni, M., Weinbauer, G.F., Hacker-Klom, U. and Nieschlag, E. (1999) Inhibin B is a more sensitive marker of spermatogenetic damage than FSH in the irradiated non-human primate model. J. Endocrinol., 162, 393400.
Forti, G., Vannelli, G.B., Barni, T., Balboni, G.C., Orlando, C. and Serio, M. (1992) Sertoli-germ cells interactions in the human testis. J. Steroid Biochem. Mol. Biol., 43, 419422.[CrossRef][ISI][Medline]
Groome, N.P., Illingworth, P.J., OBrien, M., Priddle, J., Weaver, K. and McNeilly, A.S. (1995) Quantification of inhibin pro-alpha C-containing forms in human serum by a new ultrasensitive two-site enzyme-linked immunosorbent assay. J. Clin. Endocrinol. Metab., 80, 29262932.[Abstract]
Groome, N.P., Illingworth, P.J., OBrien, M., Pai, R., Rodger, F.E., Mather, J.P. and McNeilly, A.S. (1996) Measurement of dimeric inhibin B throughout the human menstrual cycle. J. Clin. Endocrinol. Metab., 81, 14011405.[Abstract]
Guitton, N., Touzalin, A.M., Sharpe, R.M., Cheng, C.Y., Pinon-Lataillade, G., Meritte, H., Chenal, C. and Jegou, B. (2000) Regulatory influence of germ cells on sertoli cell function in the pre-pubertal rat after acute irradiation of the testis. Int. J. Androl., 23, 332339.[CrossRef][ISI][Medline]
Handelsman, D.J., Conway, A.J., Howe, C.J., Turner, L. and Mackey, M.A. (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, 41134121.[Abstract]
Hayes, F.J., Pitteloud, N., DeCruz, S., Crowley, W.F., Jr and Boepple, P.A. (2001) Importance of inhibin B in the regulation of FSH secretion in the human male. J. Clin. Endocrinol. Metab., 86, 55415546.
Illingworth, P.J., Groome, N.P., Byrd, W., Rainey, W.E., McNeilly, A.S., Mather, J.P. and Bremner, W.J. (1996) Inhibin-B: a likely candidate for the physiologically important form of inhibin in men. J. Clin. Endocrinol. Metab., 81, 13211325.[Abstract]
Jensen, T.K., Andersson, A.M., Hjollund, N.H., Scheike, T., Kolstad, H., Giwercman, A., Henriksen, T.B., Ernst, E., Bonde, J.P., Olsen, J. et al. (1997) Inhibin B as a serum marker of spermatogenesis: correlation to differences in sperm concentration and follicle-stimulating hormone levels. A study of 349 Danish men. J. Clin. Endocrinol. Metab., 82, 40594063.
Kamischke, A., Simoni, M., Schrameyer, K., Lerchl, A. and Nieschlag, E. (2001) Is inhibin B a pharmacodynamic parameter for FSH in normal men? Eur. J. Endocrinol., 144, 629637.[ISI][Medline]
Kinniburgh, D. and Anderson, R.A. (2001) Differential patterns of inhibin secretion in response to gonadotrophin stimulation in normal men. Int. J. Androl., 24, 95101.[CrossRef]
Kinniburgh, D., Zhu, H., Cheng, L., Kicman, A.T., Baird, D.T. and Anderson, R.A. (2002) Oral desogestrel with testosterone pellets induces consistent suppression of spermatogenesis to azoospermia in both Caucasian and Chinese men. Hum. Reprod., 17, 14901501.
Klingmuller, D. and Haidl, G. (1997) Inhibin B in men with normal and disturbed spermatogenesis. Hum. Reprod., 12, 23762378.[Abstract]
Levalle, O., Zylbersztein, C., Aszpis, S., Aquilano, D., Terradas, C., Colombani, M., Aranda, C. and Scaglia, H. (1998) Recombinant human follicle-stimulating hormone administration increases testosterone production in men, possibly by a Sertoli cell-secreted nonsteroid factor. J. Clin. Endocrinol. Metab., 83, 39733976.
Maddocks, S., Kerr, J.B., Allenby, G. and Sharpe, R.M. (1992) Evaluation of the role of germ cells in regulating the route of secretion of immunoactive inhibin from the rat testis. J. Endocrinol., 132, 439448.[Abstract]
Majdic, G., McNeilly, A.S., Sharpe, R.M., Evans, L.R., Groome, N.P. and Saunders, P.T. (1997) Testicular expression of inhibin and activin subunits and follistatin in the rat and human fetus and neonate and during postnatal development in the rat. Endocrinology, 138, 21362147.
Martin, C.W., Riley, S.C., Everington, D., Groome, N.P., Riemersma, R.A., Baird, D.T. and Anderson, R.A. (2000) Dose-finding study of oral desogestrel with testosterone pellets for suppression of the pituitarytesticular axis in normal men. Hum. Reprod., 15, 15151524.
McLachlan, R.I., ODonnell, L., Stanton, P.G., Balourdos, G., Frydenberg, M., de Kretser, D.M. and Robertson, D.M. (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, 546556.
Meachem, S.J., Nieschlag, E. and Simoni, M. (2001) Inhibin B in male reproduction: pathophysiology and clinical relevance. Eur. J. Endocrinol., 145, 561571.[ISI][Medline]
Nachtigall, L.B., Boepple, P.A., Seminara, S.B., Khoury, R.H., Sluss, P.M., Lecain, A.E. and Crowley, W.F., Jr (1996) Inhibin B secretion in males with gonadotropin-releasing hormone (GnRH) deficiency before and during long-term GnRH replacement: relationship to spontaneous puberty, testicular volume, and prior treatmenta clinical research center study. J. Clin. Endocrinol. Metab., 81, 35203525.[Abstract]
Petersen, P.M., Andersson, A.M., Rorth, M., Daugaard, G. and Skakkebaek, N.E. (1999) Undetectable inhibin B serum levels in men after testicular irradiation. J. Clin. Endocrinol. Metab., 84, 213215.
Pineau, C., Velez de la Calle, J.F., Pinon-Lataillade, G. and Jegou, B. (1989) Assessment of testicular function after acute and chronic irradiation: further evidence for an influence of late spermatids on Sertoli cell function in the adult rat. Endocrinology, 124, 27202728.[Abstract]
Pineau, C., Sharpe, R.M., Saunders, P.T., Gerard, N. and Jegou, B. (1990) Regulation of Sertoli cell inhibin production and of inhibin alpha-subunit mRNA levels by specific germ cell types. Mol. Cell. Endocrinol., 72, 1322.[CrossRef][ISI][Medline]
Robertson, D.M., Pruysers, E., Stephenson, T., Pettersson, K., Morton, S. and McLachlan, R.I. (2001) Sensitive LH and FSH assays for monitoring low serum levels in men undergoing steroidal contraception. Clin. Endocrinol. (Oxf.), 55, 331339.[CrossRef][ISI][Medline]
Seminara, S.B., Boepple, P.A., Nachtigall, L.B., Pralong, F.P., Khoury, R.H., Sluss, P.M., Lecain, A.E. and Crowley, W.F., Jr (1996) Inhibin B in males with gonadotropin-releasing hormone (GnRH) deficiency: changes in serum concentration after short term physiologic GnRH replacementa clinical research center study. Clin. Endocrinol. Metab., 81, 36923696.[CrossRef]
Vale, W., Rivier, C., Hsueh, A., Campen, C., Meunier, H., Bicsak, T., Vaughan, J., Corrigan, A., Bardin, W., Sawchenko, P. et al. (1988) Chemical and biological characterization of the inhibin family of protein hormones. Recent Prog. Horm. Res., 44, 134.[ISI][Medline]
Vannelli, G.B., Barni, T., Forti, G., Negro-Vilar, A., Vale, W., Serio, M. and Balboni, G.C. (1992) Immunolocalization of inhibin alpha-subunit in the human testis. A light- and electron-microscopy study. Cell Tissue Res., 269, 221227.[ISI][Medline]
Wallace, E.M., Groome, N.P., Riley, S.C., Parker, A.C. and Wu, F.C. (1997) Effects of chemotherapy-induced testicular damage on inhibin, gonadotropin, and testosterone secretion: a prospective longitudinal study. J. Clin. Endocrinol. Metab., 82, 31113115.
World Health Orgnization (1999) Laboratory manual for the examination of human semen and spermcervical mucus interaction. 4th edn, Cambridge University Press, Cambridge.
Young, J., Couzinet, B., Chanson, P., Brailly, S., Loumaye, E. and Schaison, G. (2000) Effects of human recombinant luteinizing hormone and follicle-stimulating hormone in patients with acquired hypogonadotropic hypogonadism: study of Sertoli and Leydig cell secretions and interactions. J. Clin. Endocrinol. Metab., 85, 32393244.
Zhengwei, Y., Wreford, N.G., Royce, P., de Kretser, D.M. and McLachlan, R.I. (1998) Stereological evaluation of human spermatogenesis after suppression by testosterone treatment: heterogeneous pattern of spermatogenic impairment. J. Clin. Endocrinol. Metab., 83, 12841291.
Submitted on September 4, 2002; resubmitted on October 16, 2002; accepted on December 2, 2002.