UROL_V3_No1_001_Editorial.qxd


Urology Journal

UNRC/IUA

38

Introduction

Infertility occurs in approximately 14% of

couples, and abnormalities in the male partner

are estimated to be present in up to 50% of

cases.(1,2) Efforts to evaluate the causes of

azoospermia have shown that regardless of

traditionally recognizable causes (abnormal

karyotype, obstruction, varicocele, hormonal

defect, etc), most cases (50% to 75%) are

unexplained and are considered idiopathic.(3) The

existence of an essential spermatogenesis factor

called azoospermia factor (AZF) was suspected as

early as 1976, from de novo Yq deletions in

azoospermic patients.(4) Polymerase chain

reaction (PCR) studies of sequence-tagged sites

(STSs) distributed every 30 kb enabled detection

of small deletions in the AZF region that had

been undetectable with classical cytogenetic

techniques.(5,6) This led to the identification of 3

loci in Yq11 carrying genes involved in the

control of spermatogenesis, corresponding to 3

deleted regions: AZFa, AZFb, and AZFc.(7,8) Y

Y Chromosome Microdeletions in Idiopathic Infertile Men

from West Azarbaijan

Mir Davood Omrani,1* Saied Samadzadae,2 Mortaza Bagheri,1 Kiarash Attar2

1Department of Genetics, Urmia University of Medical Sciences, Urmia, Iran

2Department of Urology, Imam Khomeini Hospital, Urmia University of Medical Sciences,

Urmia, Iran

ABSTRACT

Introduction: Although assisted reproduction techniques are used extensively in

Iran, screening for Y chromosome microdeletions before intracytoplasmic sperm

injection is often undervalued. Our aim was to investigate Y chromosome

microdeletions in men with idiopathic azoospermia or severe oligospermia.

Materials and Methods: In 99 selected patients with azoospermia or severe

oligospermia and elevated levels of follicle-stimulating hormone and luteinizing

hormone in combination with low serum testosterone levels, 20 pairs of sequence-

tagged site-based primer sets specific for the Y microdeletion loci were analyzed.

Primers were chosen to cover azoospermia factor (AZF) regions as well as deleted in

azoospermia (DAZ) and the sex-determining region on Y chromosome (SRY) genes.

Also, 100 healthy men served as a control group.

Results: Twenty-four patients (24.2%) had microdeletions in AZF genes, but no

microdeletions were found in men in the control group. In 15 patients (62.5%), 1

deletion was found. Six patients (25%) had 2, and 3 (12.5%) had 3 deletions. The

deletions mainly comprised the AZFc region (in 21 of 24 patients; 87.5%), which

corresponds to the DAZ gene. Deletions in AZFb were found in 7 patients (29.2%), and

4 (16.7%) had deletions in the proximal part of AZF regions near SRY gene. No

microdeletions were seen in the AZFa or SRY gene. 

Conclusion: Our results emphasize that Y chromosome microdeletion analysis

should be carried out in all patients with idiopathic azoospermia or severe

oligospermia who are candidates for intracytoplasmic sperm injection. 

KEYWORDS: oligospermia, microdeletion, infertility, Y chromosome

Vol. 3, No. 1, 38-43 Winter 2006

Printed in IRAN

Received December 2004

Accepted December 2005

*Corresponding author: Department of Genetics,

Mottahary Hospital, Kashani St, Urmia, Iran.

Tel: +98 441 224 0166, Fax: +98 441 223 4125

E-mail: davood_omrani@umsu.ac.ir



Omrani et al 39

chromosome microdeletions are the most

frequently encountered genetic abnormality in

male infertility. Up to 30% of men with idiopathic

azoospermia have microdeletions of the Y

chromosome,(3,9-13) and the incidence of Y

chromosome microdeletions in infertile men

varies between studies, from 1% to 55%.(14) The

molecular diagnostics of the Y chromosome have

been restricted mostly to selected patients

presenting with either azoospermia or severe

oligospermia (sperm concentration of less than

5 × 106/mL), so that the majority of Y

chromosome microdeletions have appeared in

this group of infertile men.

Screening for Y chromosome microdeletions is

recommended in patients with severely impaired

spermatogenesis, in particular, before

intracytoplasmic sperm injection (ICSI).(15) In the

West Azarbaijan province of Iran, at least 5

infertility centers are involved in treating infertile

couples, but none of them checks their male

candidates for Y chromosome microdeletions.

Our study aimed to determine the incidence of Y

chromosome microdeletions in our patients in

Urmia, the center of West Azarbaijan. The

findings of this study may provide enough

evidence for clinicians to decide whether or not

to screen all idiopathic infertile men for Y

chromosome microdeletions before attempting

ICSI procedures.

Materials and Methods

In a case-control study, 99 consecutive men

examined in our infertility clinics from November

2001 to December 2003 were screened for Y

chromosome microdeletions. The inclusion

criteria were azoospermia or severe oligospermia

(sperm concentration of less than 5 × 106/mL),

small testis volume, elevated serum levels of

follicle-stimulating hormone (FSH) and

luteinizing hormone (LH), low serum

testosterone level, and the 46,XY karyotype.

Informed consent was obtained from each

patient, according to protocols approved by the

ethics review board of Urmia University of

Medical Sciences. 

Semen analysis was carried out using WHO

criteria(16) with a Nikon phase contrast

microscope (Nippon kogaku, Tokyo, Japan).

Serum hormone levels of FSH, LH, and

testosterone were measured by solid-phase, two-

site chemiluminescent enzyme immunometric

assay (Immulite, Diagnostic Products

Corporation, Los Angeles, Calif, USA). Normal

reference ranges for men were FSH, less than 10

mIU/mL; LH, less than 10 mIU/mL, and

testosterone, 270 ng/dL to 1070 ng/dL. 

Cytogenetic analysis was carried out on

peripheral lymphocytes to rule out cases of

abnormal karyotypes. GTG-banding was

performed according to standard procedures.

One hundred age-matched fertile men with a

normal semen analysis without genital

abnormalities, selected from couples who had

been referred for tubectomy or vasectomy, served

as controls. The PCR and cytogenetic analyses

were carried out in controls, too.

Polymerase chain reaction amplification of

the three AZF loci. Genomic DNA was obtained

from peripheral leukocytes using the Nucleon Kit

II (Scotlab, Wiesloch, Germany). A set of 20

Y-specific STSs spanning the euchromatic region

of Yq from centromere to interval 7, with

particular interest in interval 6 (the AZF region),

was tested in each patient. To check the AZFa

region, PCR amplifications were carried out to

evaluate the sY81, sY83, and sY121 sites. Using

the sY128, sY130, sY133, and sY143 sites, the

AZFb region was checked. The AZFc region was

screened using the sY147, sY149, sY242, sY231,

sY254, sY255, sY182, and sY238 sites. In

addition, sY202, sY158, and sY157 were included,

corresponding to the downstream area of the

DAZ (deleted in azoospermia) gene, as well as

sY14 for the sex-determining region on the Y

chromosome (SRY) gene and sY274 as the site

next to the SRY region. As a negative control,

every PCR reaction included 1 sample of female

genomic DNA. A sample was considered negative

if a product of the expected size was not obtained

after 3 PCR attempts. 

The PCR program was as follows: amplification

of DNA by 35 cycles with 94°C for 50 seconds,

57°C for 30 seconds, and 72°C for 90 seconds;

including an initial denaturation step at 94°C for

2 minutes, and a final extension step at 72°C for

10 minutes. The PCR products were separated on

1.5% agarose gels. 

Statistical analyses. Frequencies of the Y

chromosome microdeletions were compared

between the patients with infertility and controls

using the chi-square test. Values for P less than

.05 were considered statistically significant.

Results

Overall, of 99 infertile men, 39 (39.4%) had



Y Chromosome Deletions in Idiopathic Infertile Men40

oligospermia and 60 (60.6%) had azoospermia.

The paraclinical data of infertile patients are

summarized in Table 1. Of 39 patients with

oligospermia, 6 (15.4%) had deletions, and of 60

patients presenting with azoospermia, 18 (30%)

had deletions in the AZF region of the Y

chromosome. Five of the 6 patients with

oligospermia and Y chromosome microdeletions

had a sperm concentration of less than

0.1 × 106/mL with only a few immotile

spermatozoa observed after centrifugation of the

specimen. The sperm concentration of the sixth

patient in this group was 2.4 × 106/mL. 

In general, 24 infertile men (24.2%) showed

microdeletions of the Y chromosome, while no

microdeletions were detected in controls

(P < .001). Some of the patients' PCR products

that were run on a 1.5% agarose gel are shown in

Figure 1. 

In 15 patients (62.5%), 1 deletion was found.

Six patients (25%) had 2, and 3 (12.5%) had 3

deletions. The deletions mainly comprised the

AZFc region (in 20 out of 24 patients; 83.4%),

which corresponds to the DAZ gene. Deletions in

AZFb were found in 7 patients (29.2%), and 4

(16.7%) had deletions proximal to the AZF

regions near the SRY gene. No microdeletions in

the AZFa or SRY genes were identified. A

schematic diagram of the STS markers used in

this study and the deleted markers are

summarized in Figure 2. 

Chromosome analyses of peripheral

lymphocytes of all selected patients and controls

were normal male karyotype (46,XY). 

Discussion

Interest in Y chromosomal deletion analysis in

infertile men arises largely from the likelihood

that Yq microdeletions will be transmitted by

ICSI and cause the same infertility problem in

male offspring. This transmission of

microdeletions has been described previously,(17)

an observation that underlines the necessity of

proper genetic counseling in infertile men. Some

authors have emphasized the importance of

genetic counseling and testing for Y chromosome

microdeletions in all ICSI candidates, irrespective

of their sperm concentration.

Of 99 patients examined in our study, 24 had

microdeletions in the AZF regions on the Y

chromosome (24.2%), which is in agreement with

previous studies. The Kurd and Azari ethnic

groups are the majority in our region; thus, it can

be inferred that Y chromosome microdeletion

FIG. 1. An example of deletions of the Y chromosome in the AZF region in men with oligospermia: PCR products of the

AZF region from 19 tested patients, which were run on 1.5% agarose gel identified a deleted region in 6 patients.

TABLE 1. Mean serum concentrations of FSH, LH, and testosterone in men with severe oligospermia and

azoospermia

Mean (Range) 

 

Men with oligospermia Men with azoospermia Infertile men 

FSH (mIU/mL) 9.9 (2 to 28) 14.9 (6 to 28) 14.1 (2 to 44) 

LH (mIU/mL) 5.2 (1 to 18) 7.6 (2 to 11) 6.3 (1 to 18) 

Testosterone (ng/dL) 366 (135 to 565) 344 (223 to 482) 355 (135 to 565) 

 



Omrani et al 41

may be relatively prevalent in West Azarbaijan.

The deletions mainly involved the AZFc region

(in 87.5% of the patients), which corresponds with

the DAZ gene, as well as deletion in the AZFb (in

29.2% of the patients). No microdeletions in the

AZFa or the SRY gene have been identified. We

found deletions in the proximal part of the AZF

region near the SRY gene in 16.7% of the

patients. The deletion of this region may affect

SRY gene function, but to check this hypothesis,

we must perform more experiments including

functional studies. Despite of the report of Pryor

and associates,(3) no microdeletions were found in

healthy men of our control group. This variability

in the detection of microdeletions between

studies is probably explained by the different

clinical selection criteria used by different

research groups. Stringent selection of patients

according to histologic, endocrinologic, and

clinical criteria have been found to be associated

with high deletion frequencies.(7,14,18) For

instance, Foresta and colleagues have studied

patients with idiopathic azoospermia and

bilateral Sertoli-cell-only syndrome and found a

very high number of Yq11 microdeletions.(14) By

contrast, less stringent criteria for selection has

been associated with low deletion frequencies in

studies on a large number of men with

oligospermia.(19,20) Our study shows the influence

of the selection criteria on the reported incidence

of microdeletions; we had a high rate of Y

chromosome deletion in our patients since we

used strict patient selection criteria. 

Microdeletion frequency in a sample of infertile

men is not significantly related to the number of

STS loci analyzed.(21) Kent-First and colleagues

have analyzed a large number of STSs in

different Y chromosome regions.(22) They have

shown that each STS is statistically correlated

with male infertility. It seems that patients'

selection criteria have a much more profound

effect on the rate of detection of microdeletions

than do the numbers of STSs analyzed. 

Of the 24 infertile men with microdeletions of

the Y chromosome, 18 had azoospermia and 6

had severe oligospermia. In fact, most

microdeletions have been found in men with

azoospermia and severe oligospermia, because in

FIG. 2. Schematic diagram of a Y chromosome illustrating sequence-tagged sites, AZF regions, and the different deletion

patterns of the patients in this study. A "- -" denotes deletion of a specific sequence-tagged site.



Y Chromosome Deletions in Idiopathic Infertile Men42

the majority of the studies published so far, the

analyses were limited to patients with severe

defects of spermatogenesis. Pryor and colleagues

were the first to examine 200 consecutive

patients who included 102 men with a normal

sperm count. They found deletion frequencies of

23.1%, 9.7%, and 1% in infertile men with

azoospermia, oligospermia, and normal sperm

count, respectively. They concluded that the

microdeletion in men with a normal sperm count

probably indicated a polymorphism, because it

comprised only one STS, which was also found in

fertile men.(3)

Finally, all of the men with Y chromosome

microdeletions in our study were cytogenetically

normal, showing that PCR-based assay is needed

to detect microdeletions in the Y chromosome. 

Conclusion

The correlation between Y chromosome

microdeletions and infertility, and the relative

absence of such deletions in fertile men, suggests

a cause-and-effect relationship between the

deletions and infertility. As compared with other

known causes of infertility, Y chromosome

microdeletions are relatively frequent, and their

frequency increases with the severity of the

spermatogenic defect. However, Y chromosome

microdeletions cannot be predicted on the basis

of clinical findings or even the results of semen

analyses. The role of analyses of Y chromosome

microdeletions in evaluating men with infertility

remains to be determined. With the advent of

ICSI, the potential for passing on these defects to

offspring is serious and should be considered

when infertile couples are counseled about this

procedure.

Acknowledgments

We wish to thank the enrolled families for their

cooperation in the study and also, we gratefully

thank Dr. Agenta Nordenskjold from the

Department of Molecular Medicine and Genetics

of Karolinska University Hospital in Solna,

Sweden  for kindly providing us some of the

material in this project. This study was funded in

part by the research deputy of Urmia's University

of Medical Sciences. 

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