Assessment of the utility of testicular FNAC in infertile males with special reference to differential counts

Madhu S Agarwal; Atul Gupta; Kiran Chaturvedi; Prashant Lavania

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RESEARCH ARTICLE

Year : 2004 \| Volume : 20 \| Issue : 2 \| Page : 148-153

Assessment of the utility of testicular FNAC in infertile males with special reference to differential counts

Madhu S Agarwal, Atul Gupta, Kiran Chaturvedi, Prashant Lavania
Department of Surgery & Pathology, S.N. Medical College, Agra, India

Correspondence Address:
Madhu S Agarwal
4/18c, Bagh Farzana, Civil Lines, Agra - 282 002
India

Source of Support: None, Conflict of Interest: None

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Abstract

Objectives: This study was conducted to make morphological assessment of spermatogenesis on testicular FNAC in infertile males, along with quantitative analysis of cytological smears using various cell indices, and to correlate the cytological findings with histology.
Methods: Testicular FNAC was performed in 155 azoospermic and oligozoospermic patients, and smears were examined qualitatively and quantitatively. Various cell indices including `Spermatic index', 'Sertoli cell index' and 'sperm-Sertoli cell index' were studied in these smears, and correlation made with histology.
Results: On the basis of morphological assessment and differential cell counts, the cases were classified as 'normal spermatogenesis' in 54 cases, `hypospermatogenesis' in 37 cases, `maturation arrest' in 18 cases, 'Sertoli cell only' syndrome in 33 cases, `scanty aspirate /absence of spermatogenesis' in 10 cases, and discordant results in bilateral testicular aspirates in 3 patients. 'Sertoli cell index' was found to be the most important index in distinguishing hypospermatogenesis from normal spermatogenesis. The differences between the indices in normal spermatogenesis and other cytological categories were statistically significant. In most cases, complete agreement was found between cytological and histological findings.
Conclusions : Fine needle aspiration cytology of the testis is a safe, reliable and minimally invasive modality for evaluation of spermatogenic activity in infertile males, with excellent correlation with histology. Use of quantitative studies accurately and reproducibly segregate the cases into clinically and therapeutically useful groups.

Keywords: Male infertility, FNA cytology, testis, spermatogenesis.

How to cite this article:
Agarwal MS, Gupta A, Chaturvedi K, Lavania P. Assessment of the utility of testicular FNAC in infertile males with special reference to differential counts. Indian J Urol 2004;20:148-53

How to cite this URL:
Agarwal MS, Gupta A, Chaturvedi K, Lavania P. Assessment of the utility of testicular FNAC in infertile males with special reference to differential counts. Indian J Urol [serial online] 2004 [cited 2021 Sep 27];20:148-53. Available from: https://www.indianjurol.com/text.asp?2004/20/2/148/21532

Introduction

Male factor is responsible, either wholly or partially, in nearly half the cases of infertility in couples. The diagnostic work-up in all patients of male infertility includes detailed clinical evaluation and a standard battery of laboratory investigations including semen analysis, serum hormone assays, immunological studies, etc. Finally, in cases where the semen analysis is abnormal, the status of spermatogenesis in the testis needs to be evaluated by microscopic examination of testicular tissue, conventionally performed by open testicular biopsy.

In recent years, several workers have advocated the use of fine needle aspiration cytology as an alternative to biopsy, and have found a good correlation between cytology and histology of the infertile testis.^{[1],[2],[3],[4],[5],[6],[7],[8],[9],[10],[11],[12]} However, only a few have tried quantitative assessment with calculation of various indices on fine needle aspiration smears.^{[12],[13],[14]} The current study of testicular cyto-morphology by fine needle aspiration in azoospermic and oligozoospermic males was therefore undertaken to make a morphological assessment of spermatogenesis on routine microscopic examination and to carry out quantitative analysis for cytological smears.

Patients and Methods

The study was conducted in 155 infertile males, aged 24-45 years, attending the out-patient department of S.N. Medical College, Agra. There were 81 cases of azoospermia and 74 of oligozoospermia. Following detailed clinical work-up, all patients were subjected to FNAC of the testes. The procedure was performed under local anaesthesia. 2 to 3 punctures were made with 22 G needle in each testis and smears were stained with May-GruendwaldGiemsa (MGG) stain.

Two hundred consecutive spermatogenic and Sertoli cells ^{More Details} were counted from a well spread area with good cellularity in a random manner using light microscopy.^[12],[16] Cell identification was done with the help of criteria given by Papic et al,^[3] Foresta et al^[14] and Schenck and Schill.^[15] The following cells were identified in the counts and expressed as a percentage: Sertoli cells, spermatogonia, primary spermatocytes, spermatids and spermatozoa.

The percentage of spermatozoa per 100 spermatogenic cells was expressed as `Spermatic index'. The 'Sertoli cell index' was expressed as number of Sertoli cells per 100 spermatogenic cells. The 'Sperm-Sertoli cell index' was the ratio of spermatozoa to Sertoli cells. Statistical significance was verified with Student's t test. Probability values <0.05 were regarded as statistically significant.

Testicular biopsy was performed in all cases following FNAC, under local anaesthesia. The specimen was placed immediately in Bouin's solution and delivered to the histopathology laboratory for processing, and routine Hematoxylin and Eosin sections were studied, and correlated with cytological findings.

Results

Cases were classified, after cyto-morphological assessment, into 6 categories [Table - 1]:

`Normal spermatogenesis' [Figure - 1]A was the commonest category observed in this study accounting for 54 cases. Spermatogenic cells in all stages were well represented in all cases. Sertoli cells were frequent but less than Spermatogenic cells, the mean 'Sertoli cell index' being 42.4 ± 11.4. Spermatozoa were the predominant cell type, and the `spermatic index' was 51.6 ± 12.4.

'Hypospermatogenesis' [Figure - 1]B was found in 37 cases. In these cases number of spermatogenic cells was diminished relative to the number of Sertoli cells ('Sertoli cell index' >100). However spermatogenic cells in all stages including mature spermatozoa could always be discerned. `Spermatic index' was found to be significantly lower in cases of hypospermatogenesis.

`Maturation arrest' [Figure - 2]A was found in 18 cases in both testes in azoospermic males. In 17 cases, arrest was at the level of primary spermatocytes, while in one case it was at the level of spermatids. As a rule there was a relative excess of Sertoli cells ('Sertoli cell index' > 100 in all cases), suggesting some degree of hypospermatogenesis as well.

'Sertoli cell only' syndrome (syn. `germinal cell aplasia') [Figure - 2]B was present in 33 cases in both the testes. In these cases Sertoli cells were seen in abundance, while no spermatogenic cells at any of the stages of maturation were encountered, the `spermatic index' being 0.

Ten cases showed very `scanty cellularity' with the presence of only few Sertoli cells in both the testicular aspirates. This category was named as `scanty cellularity / absence of spermatogenesis'. It was distinguished from `sertoli cell only' syndrome by markedly reduced cellularity (less than 200 cells in a well spread smear), crumpled sertoli cells and fragments of atrophic tubules. On an average 2-3 passes were made in each attempt, and three attempts made at FNAC, before calling it "scanty cellularity".

Three cases had discordant findings in bilateral testicular aspirates, one case having hypo-spermatogenesis in one and normal spermatogenesis in the other testis, and two having maturation arrest in one testis and 'Sertoli cell only' syndrome in the other testis.

Significant differences were found between various cell indices in different categories [Table - 2]. The 'Sertoli cell index' in cases of hypospermatogenesis and maturation arrest was significantly higher than normal spermatogenesis (P <.01). However, the difference was not significant between hypospermatogenesis and maturation arrest (P>.05). 'Sperm-Sertoli cell index' showed significant differences between cases of hypospermatogenesis, normal spermatogenesis and maturation arrest (P < .01 & < .001 respectively). The `spermatic index' in cases of hypospermatogenesis and maturation arrest was also significantly different from that of normal spermatogenesis (P < .01).

All cases underwent testicular biopsy under local anaesthesia in the same sitting, and the and the cyto-diagnosis was verified against histology in each instance. Diagnosis of normal spermatogenesis on histology was made when testis contained at least 60% tubules with a score of 10, and less than 10% of tubules with a score lower than 8 (Johnson's criteria)^[19] . Hypospermatogenesis was identified by the presence of hypoplastic germinal epithelium with normal maturation of spermatogenic cells. On account of thinning of germinal epithelium, the tubular lumen appeared dilated and empty.

Almost complete correlation was found between histology and cytopathological findings, specially from the point of view of identifying presence or absence of spermatogenesis. In the 10 cases with cyto-diagnosis of `scanty aspirate / absence of spermatogenesis', histology showed changes of testicular atrophy (i.e. hyalinisation and sclerosis of the tubules and interstitial fibrosis) in 8 cases, and 'Sertoli cell only' syndrome in the remaining 2 [Table - 3].

There were no significant complications of the FNAC in our experience, except for minor pain and tenderness lasting 1-3 days. No patients required hospitalisation or any surgical intervention on this account.

Discussion

Open testicular biopsy has remained the cornerstone in the diagnosis of male infertility for decades. Interest in testicular FNAC has picked up in recent years following the pioneering works of researchers like Obrant and Persson,^[1] Papic et al,^[3] Schenck and Schill,^[15] and Foresta et al.^[13],[14] who characterised different cell types in cytological smears, and demonstrated good correlation of cytological diagnosis with histological categories.

The procedure of FNAC of testis differs from other parts of the body in only some minor detail. Testicular FNAC is best performed under local anesthesia^{[12],[13],[14],[15],[16],[17],[18]} in the form of `cord block', which reduces discomfort and improves patient compliance. Other factors, which in our experience improve adequacy of aspirate from testis, include multiple (two or three) punctures, and use of a 22G needle, slightly larger bore than the commonly used 23 or 24G size,^[11],[18] as it gave us the best smear as compared to thinner needles, without any increased morbidity. On-the-spot screening of the unstained smear under microscope by the cytopathologist, performed routinely in our practice, allowed immediate re-aspiration in cases where smear appeared cytologically inadequate.

In our study, recognition of various cell types was in general easy. Secondary spermatocyte could not be identified with any degree of confidence because of the variable description available in the literature, and possibly their transient appearance due to short half-life.^{[3],[6],[7],[12]} Sertoli cells were easily distinguished from all other spermatogenic cells by virtue of their fairly abundant light staining vacuolated cytoplasm, granular chromatin and prominent nucleoli. Leydig cells were seen only in 4 cases, as they are known to be encountered very seldom in routine testicular aspirates.^[6],[8]

Normal spermatogenesis was differentiated from hypospermatogenesis by the dominance of spermatogenic cells over Sertoli cells. Among the spermatogenic cells, spermatozoa were the predominant cell type (51.6 ±12.4%). These findings are in agreement with the studies of Batra et al^[12] and Foresta et al.^[14] Demonstration of normal spermatogenesis in patients with azoospermia denotes obstructive pathology, which is potentially correctable surgically.

Cases of hypospermatogenesis were distinguished from normal spermatogenesis by the presence of increased Sertoli cells ('Sertoli cell index' of >100). In cases with maturation arrest the `Spermatic index' was severely diminished with virtually absent spermatids and spermatozoa. In addition, these smears had higher 'Sertoli cell index', indicating associated hypospermatogenesis. Mean 'spermSertoli cell index' in this group was also significantly different from normal spermatogenesis and hypospermatogenesis.

Cases of 'Sertoli cell only' syndrome had good cellularity and were easy to diagnose due to monomorphic appearance of sheets of only Sertoli cells in the smear. No spermatogenic cell could be observed in these specimens. `Spermatic index' and 'sperm-Sertoli cell' indices were zero and 'Sertoli cell index' was °O.

Most of the cases exhibited identical findings in both testes. We found only 3 cases with discordant findings in right and left testicular aspirates. Foresta and Varotto^[13] found different findings in right and left testis in 24% of their 166 patients. None of the other studies have mentioned discordant findings in bilateral testicular aspirates.^{[6],[7],[8],[10],[12],[16]}

We found differential cell counts and `cell indices' useful in correctly classifying the different cytological categories, especially the borderline cases. Significant differences were found between various cell indices in different categories. 'Sertoli cell index' was found to be the most important index in distinguishing hypospermatogenesis from normal spermatogenesis, especially in those cases with only subtle changes of hypospermatogenesis. However, the difference was not significant between hypospermatogenesis and maturation arrest (P >.05), suggesting presence of some degree of hypospermatogenesis in cases of maturation arrest. Progressively decreasing values of `Spermatic index' and 'Sperm-Sertoli cell index' were seen in normal spermatogenesis, hypospermatogenesis, maturation arrest, and 'Sertoli cell only' syndrome.

In nearly all cases the histopathology report matched almost exactly with the cyto-diagnosis, except for minor disparity at both ends of the spectrum, confirming the accuracy and reliability of the procedure in assessment of spermatogenesis [Table - 3]. Cytology reported normal spermatogenesis in one of the 38 cases of hypo-spermatogenesis, possibly due to overlap in sertoli cell index. The findings on cytology in these two categories are otherwise similar to each other except for relative proportions of Sertoli cells to the spermatogenic cells.

This observation corresponds with the findings of several previous studies. ^{[1],[2],[3],[4],[5],[6],[7],[8],[9],[10],[11],[12]} Person et al^[2] opined that the distinction between germinal cell aplasia and testicular atrophy is not always possible on cytological smears. On the other hand, Verma et al^[8] segregated these two groups on the basis of variably scanty cellularity in atrophic testis as opposed to good cellularity in germ cell aplasia. However, since in either instance the prognosis is unaltered, the distinction is prognostically meaningless.

In addition to its usefulness and accuracy in diagnosis of the state of spermatogenesis in the infertile male, FNAC offers many advantages over testicular biopsy. First, it is a painless and minimally invasive outpatient procedure, not requiring any incision or sutures, and thus is more acceptable to the patient. Secondly, it provides a more representative sample of the testis as compared to biopsy, as it allows aspiration from 2-3 sites with the passage of needle through the depth of testis. It is cost-effective, and can provide results in much shorter time as compared to biopsy. In addition, FNAC avoids the problems of post-operative fibrosis and adhesions that are an invariable result of biopsy, making subsequent exploration for microsurgical repair difficult in cases of obstructive azoospermia. Finally, it avoids at least theoretical risk of development of anti-sperm antibodies resulting from the breach of blood-testis barrier during testicular biopsy.

Conclusions

The results of the study provide support for the use of fine needle aspiration of the testis as a useful minimally invasive modality for evaluation of spermatogenic activity in infertile males. It is safe, reliable, cost-effective, and, with the use of quantitative studies in form of various cell indices, it can reproducibly segregate the cases into clinically and therapeutically useful groups. It consistently correlates well with histology, obviating the need for more invasive testicular biopsy.

References

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