Low levels of serum inhibin B do not excludesuccessful sperm recovery in men with nonmosaicKlinefelter syndrome

Klinefelter syndrome (KS) is the most common chromosomal aberration in infertile men and occurs in 1of 600 newborn males. Traditionally, men with nonmosaic KS were considered sterile, but the introductionof ICSI with surgically recovered testicular spermatozoa has given these men new hope of achieving geneticfatherhood. In 1996, the first successful sperm recovery with testicular sperm extraction (TESE) in azoosper-mic men with nonmosaic KS was reported (1).

In men with nonobstructive azoospermia, such as men with nonmosaic KS, no clinical variable has so farproven to be predictive for successful sperm recovery (2, 3). A reliable method of predicting the presence orabsence of testicular spermatozoa is highly desirable in this group of patients, who currently must undergodiagnostic TESE. Recently, two studies have indicated that serum inhibin B is a useful predictor ofspermatogenesis in men with oligozoospermia (4) and azoospermia (5). The purpose of this prospective studywas to assess the predictive value of inhibin B before diagnostic TESE in men with nonmosaic KS.

Between January 1998 and June 2000, 18 men (mean age, 33.4 � 4.9 years) with nonmosaic KS had adiagnostic TESE performed as part of their infertility workup. At least two semen analyses had previouslyshown them to have azoospermia, and clinical examination had revealed small testes of 2–5 mL, verified byscrotal ultrasonography. For comparison, 18 healthy men of similar age (mean age, 33.2 � 4.7 years), allfathers of babies recently delivered at our hospital, were recruited to the study. None of these men had ahistory of infertility, and each pregnancy was naturally conceived. Informed consent was obtained from eachparticipant, and the study had the approval of the human ethics committee of Goteborg University, Sweden.

Testicular sperm extraction was performed as described elsewhere (6). Briefly, the operations wereperformed under general anesthesia. Depending on testicular volume, one or two biopsies were taken fromeach side, placed in a sterile tube containing culture medium (IVF-50, Scandinavian IVF Science, Goteborg,Sweden), and sent for examination in our laboratory. In addition, a single biopsy from each testicle was sentfor histopathological examination.

Blood samples of serum inhibin B were analyzed after storage at �70°C. Inhibin B levels were measuredby a solid-phase sandwich ELISA (Serotec Limited, Oxford, UK). The lowest detectable inhibin B concen-tration found was 5 pg/mL. The intra-assay and interassay variations of the kit were �9% and 15%,respectively.

All statistical analyses were based on individuals. Data are presented as mean � SD and range. Men withinhibin B concentrations below the detection level were included in the statistical analysis and assigned a valueof 5 pg/mL. For comparison between two groups, the Mann-Whitney U test was used. All significance testswere two-tailed and conducted at the 5% significance level.

Diagnostic TESE successfully identified the presence of spermatozoa in 5 (27.8%) of the 18 men with KS.These men were consequently accepted for ICSI treatment. All TESE procedures in subsequent ICSI cycleswere successful, resulting in two pregnancies. Serum concentrations of inhibin B of all men included in thestudy are shown in Table 1. Serum inhibin B levels were below the detection limit in 15 of the men with KS.In 4 of these men, sperm recovery was successful. The fifth male with successful sperm retrieval had a seruminhibin B concentration of 5.5 pg/mL. In the Klinefelter group with unsuccessful sperm recovery, 11 of thesemen had serum inhibin B levels below the level of detection. The remaining two patients had serum inhibinB concentrations of 6.0 pg/mL and 6.5 pg/mL, respectively. In the fertile control group, serum inhibin B wassignificantly higher (P�.0001) compared with the case of the total Klinefelter group. There was no overlapin serum inhibin B levels between the Klinefelter and the control group. Testicular histopathology revealedtubular sclerosis and atrophy in all men with KS, but in the men with successful sperm recovery, heteroge-neous focal spermatogenesis within the atrophic tissue was found.

The role of inhibin B as a predictor for finding sperm by TESE in men with nonobstructive azoospermiais still not clarified. Two studies have indicated that inhibin B had a clear predictive value for finding spermby surgical retrieval procedures (4, 5). One of these studies claimed that a serum inhibin B concentration of�40 pg/mL indicated no presence of testicular spermatozoa after surgical procedure (5). However, the number

Received October 15,2002; revised andaccepted February 11,2003.Supported by grants fromthe Goteborg MedicalSociety, the MerchantHjalmar SvenssonFoundation, and theFaculty of Medicine,Goteborg University,Sweden.Reprint requests: GoranWestlander, M.D., Ph.D.,Center for ReproductiveMedicine, SahlgrenskaUniversity Hospital, SE-41345 Goteborg, Sweden(FAX: 46-31-829248;E-mail: goran.westlander@medfak.gu.se).

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of patients with nonobstructive azoospermia was small in boththese studies. Results of other recently published studies have beenless convincing. In a large study including 185 patients with non-obstructive azoospermia, Vernaeve et al. (7) reported that neitherserum inhibin B nor the combination of serum FSH and inhibin Bwere of predictive value for successful TESE. Histological diagno-sis showed maturation arrest, germ-cell aplasia, or tubular sclerosisand atrophy, and in this study, several men with positive spermretrieval had inhibin B concentrations below the detectable limit.Two other studies reported that the combination of serum inhibin Band FSH improves the sensitivity in predicting recovery of sper-matozoa but is not accurate enough to predict a positive spermretrieval in azoospermic men (8, 9). Further, in a recent case report,a pregnancy after successful surgical sperm recovery in men ofnormal karyotype with nonobstructive azoospermia and undetect-able serum inhibin B levels was described (10). The present studydid not find inhibin B predictive for successful TESE in azoosper-mic men with KS. Despite the limited number of patients, theseresults are of clinical interest because men with KS were excludedin the previously mentioned articles (5, 7–9). The serum levels ofinhibin B measured in our Klinefelter men were low and, in mostcases, below the level of detection. These findings are in accor-dance with those of previous studies (4, 11). Inhibin B levels aremost likely to be dependent on the amount of functioning seminif-erous tubules. In our experience, tubular atrophy in general is moresevere in men with KS compared with in other patient groups withnonobstructive azoospermia. This may explain the high percentageof Klinefelter men who have inhibin B levels below the level ofdetection. Despite the low levels of inhibin B, foci of tubules withgerm cells and small numbers of mature spermatozoa were found in5 of 18 men with KS (27.8%). Four of these men had undetectable

serum inhibin B levels. All 5 men with successful diagnostic spermrecovery had TESE repeated 6–9 months later, in a subsequentICSI cycle. Spermatozoa were found in all cases, and ICSI resultedin two pregnancies.

The normal range of serum inhibin B in fertile men has previ-ously been found to be 94 to 327 pg/mL (8). In our control group,serum levels of inhibin B ranged between 30 and 212 pg/mL. Oneof the fertile men had an inhibin B concentration of only 30 pg/mL.This man also had an elevated concentration of serum FSH (19.3IU/L). Most likely, these results were hormonal signs of testiculardamage. However, his medical history did not disclose the cause ofthese findings. It may indicate that low levels of inhibin B in mendo not always preclude the ability to conceive naturally. On theother hand, the concentration of inhibin B may have been muchhigher at the time of conception.

One suggested advantage of inhibin B measurement has beenthat inhibin B reflects the function of the total testicular tissue,whereas biopsies are not representative of the entire testis. Theabsence of spermatozoa in one testicular biopsy does not precludethe presence of spermatozoa in another part of the testis (12, 13). Toidentify focal spermatogenesis, multiple biopsies are usually ob-tained at TESE. Because of the small testicular volume (2–5 mL) inmen with nonmosaic Klinefelter syndrome, the number of biopsiesand the amount of testicular tissue that can be excised are limited.Apparently, when TESE is successful in men with KS, histologicaldiagnosis of testicular tissue shows heterogeneous focal spermato-genesis within the atrophic tissue. It seems likely that the focalspermatogenic activity is too small to have a substantial stimulatingeffect on serum inhibin B. The present available assay for inhibinB measurement does not detect extremely low inhibin B concen-

T A B L E 1

Clinical and endocrinological variables in men with Klinefelter syndrome and in fertile controls.

Men with Klinefelter syndrome Fertile controls

Patient Age (y)Inhibin B(pg/mL) TESE Patient

Age(y)

Inhibin B(pg/mL)

1 29 ND Failed 19 37 2122 38 ND Failed 20 33 713 30 ND Successful 21 27 724 43 ND Failed 22 41 1395 33 6.5 Failed 23 24 596 31 ND Successful 24 34 1097 34 ND Successful 25 29 1428 38 ND Failed 26 36 729 36 5.5 Failed 27 33 165

10 27 ND Failed 28 28 3011 27 ND Failed 29 36 13412 40 ND Failed 30 30 7313 31 5.5 Successful 31 32 18814 37 ND Failed 32 28 8215 28 ND Successful 33 39 8416 39 ND Failed 34 37 15517 32 ND Failed 35 36 5218 29 ND Failed 36 37 183

ND � not detectable.

Westlander. Inhibin B and successful sperm recovery. Fertil Steril 2003.

FERTILITY & STERILITY� 1681

trations. As in the present study, we do not know whether theinhibin B levels not detectable were 0 pg/mL or were just below thelower level of detection.

In conclusion, this study shows that inhibin B is not predictiveof successful surgical sperm recovery in men with nonmosaicKlinefelter syndrome. Even a concentration of serum inhibin Bbelow the level of detection does not exclude foci of spermatogen-esis. We recommend that all our azoospermic men with Klinefeltersyndrome have a diagnostic TESE before treatment unless donorsperm backup is available and fully accepted by the couple.

Acknowledgments: The authors express their gratitude to midwife CharlotteWerner, B.Sc., University of Adelaide, The Queen Elizabeth Hospital,Woodville SA, Australia, for invaluable help in data collection. The authorsalso thank Rob Norman, M.D., Reproductive Medicine Unit, University ofAdelaide, for his constructive criticism of this manuscript.

Goran Westlander, M.D., Ph.D.Erling Ekerhovd, M.D., Ph.D.Christina Bergh, M.D., Ph.D.Center for Reproductive Medicine, Sahlgrenska University

Hospital, Goteborg, Sweden

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