Assessment of spermatogenesis by fine needle aspiration of testes in patients with azoospermia

Background and objectives: Recent advances of Invitro Fertilization (IVF) and Intracytoplasmic Sperm Injection (ICSI) combined with the progress in sperm retrieval from testes have provided a hope for azoospermic patients to become fathers. The aim of this study is to describe the cytological features of the cells aspirated from testes of patients with azoospermia and to select those who have mature spermatozoa in their aspirate for assisted fertilization. Design and setting: A descriptive study (case series) conducted in private clinic from Jan. 2007 to Sept. 2012. Patients and methods: Under local anasthesia Fine Needle Aspiration Cytology (FNAC) was performed on 250 patients whose sperm count was zero in at least three consecutive semen samples. The size of the testes was measured and recorded for each case. The cytological features of the aspirated cells were described and the patients who have mature spermatozoa were identified for assisted fertilization. Results: Normal spermatogenesis was found in 54 patients (21.6%), complete maturation arrest at primary spermatocytes in 95 patients (38%), complete maturation arrest at secondary spermatocytes in 32 patients (12.8%), germ cell aplasia in 62 patients (24.8%), and atrophic testis in 7 patients (2.8%). The mean age of the patients was 29.8 years. Regarding the greatest dimension of the testes, there was a significant difference in the greatest dimension of atrophic testes and in testes with germ cell aplasia, from the other groups. The procedure was well tolerated by the patients and no complications were encountered. Conclusions: The technique can differentiate obstructive from intrinsic causes of azoospermia. It is simple, quick, cost effective and free from complications.


INTRODUCTION
nfertility is a common problem worldwide. 1 Azoospermia is the most severe form of male factor infertility. It is present in 5% of infertile couple in Netherland, 2 in 9% of infertile couple in France 3 and in 38.3 % of infertile men in India. 4 Azoospermia was a hopeless cause of infertility, but after the introduction of invitro fertilization (IVF), sperm retrieval from the testis and surgical management of obstruction, gave a hope for fatherhood in those couples. 5,6 Fine needle aspiration of the testis as an alternative to open biopsy has been used to characterize the state of infertility for the last 40 years 7

PATIENTS AND METHODS
From Jul. 2007 to Jul. 2012, two hundred and fifty patients with azoospermia, (sperm count is zero in at least three semen analysis) were received in the private clinic, referred from gynecologist and andrologist colleaques from various units. The procedure was explained for the patient and consent of the patient was obtained.
The age of the patients and the greatest dimension of the testis were recorded.
FNAC from one testis is performed under local anesthesia using disposable syringe and fine needle (gauge 23 G). A smear of the aspirated material is prepared, fixed in 95% ethanol and stained by H & E stain.
Statistical analysis was performed using one way ANOVA test. F=306.46 to see if there is any significant correlation between size of testes and state of spermatogenesis.

RESULTS
The technique is simple, quick and well tolerated by most of the patients. No complications were encountered.
The age of the patients ranged from 20 to 45 years with a mean of 29.8 years ( Table 1).
The greatest dimension of the testis varied according to the underlying lesion from 1 to 5 cm ( Table 2). The testes were significantly smaller in atrophic testis and those with germ cell aplasia than the other groups of lesions ( Table 2).   Table 1). Complete maturation arrest at primary spermatocyte, was characterized by absence of mature spermatozoa and lack of maturation beyond primary spermatocytes. Primary spermatocytes are large rounded cells with relatively large nuclei present singly and in small groups. Ninety five patients (38%) were diagnosed as such ( Table 1).
Complete maturation arrest at secondary spermatocytes revealed secondary spermatocytes, with absence of mature spermatozoa. Secondary spermatocytes are rounded cells smaller than primary spermatocytes with dense nuclei. Thirty two patients were encountered in this group (12.8%). (Table1).
Germ cell aplasia (Sertoli cell only syndrome) showed absence of germ cells. The smear contained Sertoli cells and interstitial cells of Leydig only. It was found in 62 patients (24.8%) (Table1). Sertoli cells are elongated cells (columnar cells) with basal nuclei. Interstitial cells are large polyhedral cells with rounded nuclei and esinophilic cytoplasm.
Atrophic testis revealed absence of germ cells, Sertoli cells and interstitial cells of Leydig. It was found in 7 patients (2.8%) ( Table 1).

DISCUSSION
Although the technique was reported by many others, 6,[8][9][10][11] however it is the first study of this kind in our locality. It is an outpatient procedure and tolerable by most of the patients. Assessment of the accuracy of the procedure was not the aim of this study as it has been assessed repeatedly in other studies with accuracy rate ranging from 87% to 100% . 5,6,8,9 By FNAC it was possible to differentiate post testicular (obstructive) cases from intrinsic testicular cases in 54 patients (21.6% of the cases) A figure which is lower than that reported by Dajani YE and Kilani Z, 12 this difference can be attributed to the type of sample i.e number of aspirates, and the regional variation of underlying cause of azoospermia. 1,4,13 The finding of mature spermatozoa in the aspirate is an important step in the management of infertility as it provides a hope for biologic fatherhood either by invitro fertilization (IVF), 6 or by surgical correction of vaso-epididymal junction, 5 eliminating unnecessary and useless medical treatment in such cases. The age of infertile men seeking advice for infertility in this study is comparable to those reported by Kurien et al. 8 (Table1).
Regarding the greatest dimension of the testes, in germ cell aplasia, atrophic testis and Sertoli cell only syndrome, the testes were significantly smaller than the testes in the other groups (p value is 0.0001) ( Table 2). A finding which is helpful in predicting the underlying pathology.
Bleeding and epididymitis were reported by Piato et al 14 after the use of spring-loaded biopsy. Such complications were neither encountered in this study nor reported by the use of Fine needle Aspiration. 10,14