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- Letters to the Editor -

Analysis of the CAG repeat number in exon 1 of the androgen receptor gene in Slovene men with idiopathic azoospermia and oligoasthenoteratozoospermia

Borut Peterlin¹, Branko Zorn², Natasa Teran¹, Tanja Kunej^1,3

¹Division of Medical Genetics, ²Andrology Centre, Department of Obstetrics and Gynecology, University Medical Centre, Ljubljana, Ljubljana 1000, Slovenia
³Biotechnical Faculty, University of Ljubljana, Ljubljana 1000,, Slovenia

Correspondence to: Dr. Borut Peterlin, Division of Medical Genetics, Department of Obstetrics and Gynaecology, University Medical Center, Slajmerjeva 3, 1000 Ljubljana, Slovenia.
Tel/fax: +386-1-5401-137; E-mail: borut.peterlin@guest.arnes.si
Received 2006-07-01 Accepted 2006-08-20

DOI: 10.1111/j.1745-7262.2007.00242.x

Dear Sir,

I am Borut Peterlin, from Division of Medical Genetics, Department of Obstetrics and Gynaecology, University Medical Center, Ljubljana, Slovenia. We write to you to discuss if the number of CAG repeats in the androgen receptor (AR) gene is associated with male infertility in a group of 190 Slovene infertile men compared to 137 men with proven fertility.

Pathophysiology of subfertility involves a complex network of genetic and environmental factors. Stimulation and maintenance of spermatogenesis require androgens and functional AR. The first exon of the AR gene contains a polymorphic CAG repeat coding for a polyglutamine tract. Expansion of the CAG repeats to 40 or more causes spinal and bulbar muscular atrophy (SBMA) [1]. Besides progressive muscle weakness and atrophy, patients also show signs of mild androgen insensitivity: endocrine disturbances (including gynae-comastia), testicular atrophy, oligozoospermia, azoosper-mia, impairment of sperm quality and infertility.

Both in vivo and in vitro studies have demonstrated reduced transactivation potential of the AR as the polyglutamine tract increased in length [2]. Inverse correlation between sperm concentration and the number of androgen receptor CAG repeats was found within the normal fertile population; shorter CAG repeats were associated with higher sperm output [3].

The concept of the potential pathogenic effect of longer CAG repeats in the range of 24_40 (CAG)n is important not only for better understanding of the etiology of male infertility, but also for the treatment of infertile men. Ten studies to date have shown the existence of the correlation between CAG repeat number and male infertility (in Chinese, Japanese, North American, French, Israeli, Taiwan [China] and Spanish populations), but 20 other studies did not confirm the association (in Swedish, Belgian, German, Japanese, Danish, Dutch, Indian, Greek, New Zealander, Finish, Hong Kong [China] and Italian populations) [4]. No data are available for Slavic populations.

In this letter, we would like to provide such data gained from a group of 190 Slovene infertile men compared to 137 men with proven fertility.

Patients with obstructive azoospermia as a result of epididymal stenosis or congenital bilateral absence of deferens and those with Klinefelter and Kallman syndrome, cytogenetic abnormalities and Y chromosome microdeletions were excluded. The study group consisted of 74 men with idiopathic non-obstructive azoospermia (NOA), 116 men with oligoasthenoterato-zoospermia (OAT) and a control group of 137 men with proven fertility (at least one offspring). All patients were Slovene or of Slavic origin. Informed consent was obtained from each patient. The study was approved by the National Ethics' Committee.

The number of CAG repeats of the AR gene was analyzed using the polymerase chain reaction (PCR) method and electrophoresis on Spreadex EL 1200 gels (Elchrom Scientific AG, Cham, Switzerland) [1, 5]. Statistical analysis was performed using the statistical package SPSS (version 11.0, SPSS Inc., Chicago, IL, USA). The difference in the number of CAG repeats between the infertile and the control group was tested by the Mann_Whitney U-test. The CAG repeat number in infertile patients was correlated to clinical characteristics using Spearman's correlation coefficient.

Statistical analysis showed no significant difference of the CAG repeat number between the infertile and control groups (P = 0.425; Figure 1). The mean CAG repeat number was 19.65 ± 2.79 (range 11_27; median 20) in infertile males and 19.35 ± 2.79 (range 12_26; median 19) in fertile men.

The number of CAG repeats did not correlate with any clinical characteristics of patients: testicular volume (P = 0.121), sperm concentration (P = 0.726), motility "a" (P = 0.101) and normal morphology (P = 0.886), FSH levels (P = 0.201) and Johnsen's histological score (P = 0.505). The mean CAG repeat number was 19.23 ± 3.0 (range 13_27; median 19.5) in the group of the NOA patients and 19.91 ± 2.64 in the OAT group (range 11_27; median 20). There was no significant difference in CAG repeat number between the groups of NOA and OAT (P = 0.089).

The number of the (CAG)n repeats in the AR gene in infertile men was analyzed in 30 studies with differences in the results in several populations [4]. It was reported that the odds ratio for azoospermia is 7-fold higher in patients with 26 ≥ CAG repeats. Our study revealed that 4.4% of patients had 26 ³ CAG repeats compared to 1.5% of fertile controls. In other studies, the number of men with 26 ≥ CAG repeats varied from 0% in the Israeli population to 49% in the Chinese population, with the average 18.2% [2, 4]. Two studies showed no difference in the mean (CAG)n repeat number, but a statistically significant proportion of men with 26 ≥ CAG repeats [4]. The Italian study showed an association of the (CAG)n repeat number only with joint distribution with another trinucleotide (GGC)n repeat in exon 1 of the AR gene.

The variability of the results by various research groups might be due to different ethnic origins and, hence, different genetic modifiers of the populations studied. The diagnostic criteria used in different studies could also have affected the results. All studies determined that patients with genetic causes of infertility (karyotype mutations, Kallman syndrome, Y chromosome microdeletions and CF mutations) should be excluded from the analysis, however not all known genetic causes for male infertility were determined to be excluded in all studies. Additionally, in our study, after excluding men with obstructive azoospermia, only men with impaired spermatogenesis were taken into account. The size of the study groups is also likely to have contributed to the conflicting findings. We analyzed 190 patients, and in previous studies the size of the study group varied from 33 to 280, with an average of 102 patients being analyzed.

In conclusion, our analysis on the Slovene population indicates that when spermiologic diagnostic categories, NOA and OAT, are taken into consideration and analyzed together they do not correlate with the CAG repeat number in the AR gene. The number of CAG repeats in the AR gene within the normal range is not a clinically relevant genetic risk factor for the development of NOA and OAT in the Slovene population of men entering the ICSI procedure.

References

1 La Spada AR, Wilson EM, Lubahn DB, Harding AE, Fischbeck KH. Androgen gene mutations in X-linked spinal and bulbar muscular atrophy. Nature 1991; 352: 77_9.

2 Tut TG, Ghadessy FJ, Trifiro MA, Pinsky L, Yong EL. Long polyglutamine tracts in the androgen receptor are associated with reduced trans-activation, impaired sperm production, and male infertility. J Clin Endocrinol Metab 1997; 82: 3777_82.

3 Von Eckardstein S, Syska A, Gromoll J, Kamischke A, Simoni M, Nieschlag E. Inverse correlation between sperm concentration and number of androgen receptor CAG repeats in normal men. J Clin Endocrinol Metab 2001; 86: 2585_90.

4 Tufan AC, Satiroglu-Tufan NL, Aydinuraz B, Satiroglu MH, Aydos K, Bagci H. No association of the CAG repeat number in exon 1 of the androgen receptor gene with idiopathic infertility in Turkish men: implications and literature review. Tohoku J Exp Med 2005; 206: 105_15.

5 Kunej T, Teran N, Zorn B, Peterlin B. CTG amplification in the DM1PK gene is not associated with idiopathic male subfertility. Hum Reprod 2004; 19: 2084_7.