Is diagnostic testicular fine needle aspiration necessary in azoospermic men before sperm aspiration/extraction for intracytoplasmic sperm injection cycles?

PURPOSE: To determine whether diagnostic testicular fine needle aspiration (TEFNA) sampling needs to be performed in azoospermic men prior to obtaining testicular sperm cells for IVF-ICSI procedures. METHODS: Ten azoospermic patients underwent TEFNA in 1993-1996. During 1997, all patients underwent testicular sperm aspiration (TESA) and/or testicular sperm extraction (TESE) to retrieve spermatozoa for IVF-ICSI cycles. The results of the two procedures performed in two separate hospitals were compared. RESULTS: Diagnostic TEFNA revealed spermatozoa in five patients; identical results in four were found during IVF-ICSI cycles. In three patients, only Sertoli cells were found on TEFNA, in two of them TESA/TESE showed identical results, and in one, two spermatozoa were detected by Cyto-SEM. In the remaining two patients, spermatids or spermatocytes were found on both procedures. CONCLUSIONS: There was a very good correlation between the diagnostic and therapeutic procedures. We suggest that in azoospermic patients, diagnostic TEFNA is valuable in order to avoid unnecessary controlled ovarian hyperstimulation in the female partner for IVF. In patients in whom spermatozoa are detected, cryopreservation may be performed for later IVF-ICSI cycles.

[Are testes in oligo/azoospermia homogenous or heterogenous?].

We determined whether a single testicular specimen is sufficient to represent qualitatively the spermatogenic process within the testes of azoospermic or severely oligospermic infertile men. In 191 testes of azoospermic patients and in 26 of those with severe oligospermia, fine needle aspirations at 3 different sites of each testis were performed. Aspirated material from each puncture was stained and in each smear all spermatogenic cells, as well as Sertoli cells, were identified. Testes were classified according to the most mature spermatogenic cell type present, or the presence of only Sertoli cells. The homogeneity of the testicular spermatogenic process was then evaluated. There was an overall intratesticular difference between aspirates in 14.1% of azoospermic testes and in 26.9% of severely oligospermic testes with regard to the most mature spermatogenic cell type. When spermatozoa were the most mature cell type, they were detected in all of the 3 aspirates in 71.4% of the testes. In 18.4% or 10.2% of this group of testes they were retrieved in only 1 or 2 of the aspirates, respectively. In testes in which spermatids or spermatocytes were the most mature spermatogenic stage, these cell types were detected in all 3 aspirates in only 36.4% and 68.0%, respectively. In azoospermic patients with full testicular spermatogenesis, the likelihood of retrieving spermatozoa from the testes was 84.3%, 92.7% and 100% in 1, 2 and 3 specimens, respectively. The following conclusions were drawn: There is a wide range of testicular heterogeneity in azoospermia or very severe oligospermia for diagnosing the testicular spermatogenic pattern. In azoospermia, specimens from several testicular sites are required. It is strongly recommended that no assisted fertilization be offered to azoospermic patients unless prior evaluation of the spermatogenic pattern in the seminiferous tubules is determined.

[Follicle-stimulating hormone in azoospermia in prediction of spermatogenic patterns].

Follicle-stimulating hormone (FSH) is considered to be the most important plasma hormone correlated with spermatogenesis. Elevated FSH plasma levels were shown to be associated with complete damage to testicular seminiferous tubule germinal epithelium. Recently, there have been conflicting reports with regard to the value of FSH plasma levels in predicting seminiferous tubule histology in the azoospermic patient and hence, as a guide for therapy in assisted reproduction using testicular sperm retrieval. The aim of this study was to evaluate whether FSH plasma levels can predict spermatogenic pattern in the testes of the azoospermic infertile patient. 69 infertile men with non-obstructive azoospermia and 18 with very severe oligospermia were studied. In all, plasma levels of testosterone, free testosterone, prolactin, luteinizing hormone and follicle-stimulating hormone were measured by enzyme immunoassay. In the azoospermic patients the seminiferous tubule spermatogenic pattern was determined in testicular aspirates obtained by multiple fine needle aspiration and categorized according to the most mature spermatogenic cell type in the aspirates: Sertoli cells only, spermatogenic maturation arrest or full spermatogenesis. There were no significant differences in plasma levels of any hormone measured except in very severely oligospermic and azoospermic patients. Both normal and elevated levels were detected in all, regardless of seminiferous tubule cytological pattern or plasma FSH in azoospermic patients. It is concluded that plasma levels of FSH can not be used as a predictive parameter, neither for the presence of spermatozoa nor for any other seminiferous tubule cytological pattern in azoospermic infertile men. They cannot serve as guides for selection of azoospermic men for trials of testicular sperm retrieval in assisted reproduction.

[Does testicular volume reflect spermatogenic pattern in men with azoospermia?].

The aim of this study was to determine whether testicular volume can serve to predict patterns of spermatogenesis in azoospermia. In 27 tests of azoospermic infertile men, cytological specimens from several sites from each testis were obtained by fine needle aspiration. Testes were classified according to the most mature spermatogenic cell type. Classifications were testes with spermatozoa, with arrested spermatogenic development, and with only Sertoli cells. Prior to fine needle aspiration the 3 dimensions of each testis were determined ultrasonically and its volume calculated. Mean testicular volume (+/- SD) was 7.71 (+/- 5.95) ml for testes with spermatozoa and 7.55 (+/- 2.35) and 7.31 (+/- 4.42) ml for testes with spermatogenic maturation arrest and with only Sertoli cells, respectively (differences not significant). It is concluded that testicular volume can not be used as a predictive parameter, neither for the presence of spermatozoa nor for the cytological pattern of the testes of azoospermic infertile men.

[Seminiferous tubule cytological pattern in infertile, azoospermic men in diagnosis and therapy].

We determined spermatogenic patterns of seminiferous tubules in azoospermic infertile men and evaluated the prevalence of bilateral testicular homogeneity. 185 azoospermic men underwent bilateral testicular fine-needle aspiration (TFNA) in which each testis was punctured at 3 different positions. Aspirated material was stained and classified according to the most mature spermatogenic cell type present or whether only Sertoli cells were present. 35.7% had spermatozoa in their testes, 36.2% had spermatogenic maturation arrest, and 28.1% had only Sertoli cells in their seminiferous tubules. In 15.6% of all patients, the diagnosis in 1 testis differed from that in the other. In only 73.2% of those with testicular spermatozoa was it bilateral. In the remaining 26.9%, only Sertoli cells, spermatocytes or spermatids were found as the most mature cell type in the other testis. The study definitely indicates that fertilization with retrieved testicular spermatozoa should not be offered to azoospermic patients without prior evaluation of the seminiferous tubuespermatogenic pattern in both testes.

[Quantitative analysis of testicular varicocele by scintigraphy].

Testicular varcocele is the most common factor affecting male fertility. The determental effects of the varicocele on semen quality and fertility are codonsidered to be related to its severity. The aim of this study was to establish a quantitative radioisotopic method for the diagnosis and evaluation of testicular varicocele. 31 men attending our infertility clinic underwent scintigraphy with a gamma camera scanner (Apex 415) equipped with a pinhole low energy collimator (zoom factor 2). The evaluation was performed in the supine position and the collimator at 8.5 cm above the testicular area. The patients' red blood cells were labeled in-vivo by injection of SN-pyrophosphate before i.v. administration of To-99. An image of the scrotal area was obtained and the computer processing consisted of a drawing of the region of interest over the area of the varicocele, with background subtraction. The following indices were calculated: testes total count, varicocele area (in pixels) and average count per pixel (ACPP). The patients also underwent high resolution duplex sonography (HRDS) using a Multigon duplex scanner with a 7.5 Mhz transducer. Spermatic vein diameter, and reflux when present, were determined. HRDS was performed to compare and to validate the results obtained by scintigraphy. In men without varicocele mean ACPP was 2.80, in those with mild varicocele 3.76, in those with moderate varicocele 5.40 and in those with severe varicocele 7.48. A significant positive correlation was found between ACPP values and severity of the varicocele, as determined either by the diameter of the spermatic veins or by reflux of blood in the veins. We conclude that the ACPP index obtained by this new technique enables objective diagnosis and quantitative grading of testicular varicocele.

Delivery following intracytoplasmic injection of mature sperm cells recovered by testicular fine needle aspiration in a case of hypergonadotropic azoospermia due to maturation arrest.

This is the first reported delivery following intracytoplasmic sperm injection (ICSI) of mature live testicular sperm cells collected in a case of hypergonadotrophic azoospermia with maturation arrest. The 30 year old couple presented with primary infertility of 11 years duration, the man being submitted in childhood to five orchidopexy operations for the treatment of cryptorchism. He had elevated serum follicle stimulating hormone (FSH; 18.8 IU/I), an atrophic left testis and a normal sized right testis, the biopsy of which diagnosed maturation arrest and focal scarring. The couple refused donor insemination for religious reasons and the only option was an attempt at testicular sperm collection. Multiple testicular and epididymal fine needle aspirations were performed, using an aspiration handle loaded with 20 ml syringe and 21-23 gauge butterfly needles. The mature spermatozoa recovered were used to inseminate the oocytes by ICSI. Prior to this procedure, the patient's wife underwent ovulation induction using a long protocol of mid-luteal gonadotrophin-releasing hormone analogue/human menopausal gonadotrophin (GnRHa/HMG). At oocyte retrieval, ten oocytes were recovered. Eight live sperm cells were recovered from the aspirates of the right testis. Following ICSI into four metaphase II and two metaphase I oocytes, one mature oocyte was fertilized, cleaved and was transferred to the uterus 48 h after oocyte retrieval. The patient conceived and delivered a 3300 g boy at term. In conclusion, our results demonstrate that this novel approach should be considered in cases with hypergonadotrophic azoospermia due to testicular failure. Further experience is needed to establish the exact criteria for its use.

Assessment of testicular spermatozoa morphology by image analysis.

OBJECTIVE: We objectively evaluated the testicular sperm cell morphology. METHODS: The spermatozoa head morphology was evaluated by image analysis as presented on testicular fine-needle aspiration cytology smears. RESULTS: 2,356 spermatozoa heads were classified into six groups, according to different morphology parameters. CONCLUSIONS: This objective method of morphologic assessment of testicular spermatozoa provides an important tool for the evaluation of testicular spermatozoa and can serve as a guide for therapy in male infertility.

The relationship between plasma levels of gonadotropins, androgens, and prolactin in azoospermic men with their testicular spermatogenic pattern.

OBJECTIVE: To assess the relationship between plasma levels of gonadotropins, androgens, and PRL with testicular spermatogenic pattern. DESIGN: Patient series. SETTING: University affiliated medical center. PATIENTS: One hundred twenty azoospermic infertile men. INTERVENTIONS: Testicular fine needle aspirations and determination of plasma levels of FSH, LH, T, free T, and PRL. MAIN OUTCOME MEASURES: Gonadotropins, androgens, and PRL plasma levels as a diagnostic criterion of testicular spermatogenic patterns. RESULTS: No statistically significant differences were detected in plasma levels of LH, androgens, and PRL among patients with Sertoli cell only, spermatogenic arrest, or full spermatogenesis. Elevated plasma levels of FSH threefold above the upper normal limit preclude, with a probability of 95%, the existence of full testicular spermatogenesis, but are not valid for the diagnosis of either Sertoli cell only syndrome or spermatogenic arrest. CONCLUSIONS: Luteinizing hormone, androgens, and PRL plasma levels are of no diagnostic value in predicting any specific spermatogenic pattern, and plasma FSH levels can not be used for diagnosing Sertoli cell only syndrome.

[Relationship between spermatogenic maturation stages and male hormone levels].

The relationship between stages of the spermatogenic maturation process and male hormone levels was evaluated in 41 azoospermic, infertile men. Patients were categorized into groups according to the most mature spermatogenic cell type present in testicular aspirates: spermatocytes, spermatids or spermatozoa. High FSH and LH plasma levels were found in those with spermatocytes. Their hormone levels differed statistically (p < 0.005) from those in patients with maturation arrest at the spermatid stage, or those with spermatozoa in their testes. No statistically significant differences were found between the 3 groups with regard to plasma levels of testosterone, free testosterone and prolactin. However, there were positive correlations, higher for free testosterone than for testosterone, with stage of spermatogenic maturation.

[Severity of testicular varicocele and sperm cell ultramorphologic characteristics].

Ultramorphologic characteristics of sperm cells found in infertile men with varicoceles were compared with the severity of the varicocele. Classification of varicoceles into mild, moderate or marked was determined by spermatic vein diameter and degree of blood reflux. These were evaluated by high resolution duplex sonography. There were no statistical differences between the 3 grades of varicocele severity with regard to abnormalities in the subcellular organelles of the acrosome, karyoplasm, and skeleton of spermatozoal heads, or in the axoneme, outer dense fiber and skeleton of sperm cell tails. The only abnormal ultramorphologic features found to be significant among the 3 groups were differences in the frequencies of karyoplasm agenesis, and of partial agenesis, malformation and degeneration of the acrosome in sperm cell heads. However, these features could not be related to grade of severity and therefore, no ultramorphologic severity pattern could be detected. Thus, no varicocele beneficial score based on grade of severity and sperm cell ultramorphology can be suggested for clinical use.

Is one testicular specimen sufficient for quantitative evaluation of spermatogenesis?

OBJECTIVE: To investigate whether quantitative analysis performed on one testicular specimen is adequate for quantitative evaluation of spermatogenic process. DESIGN: Comparison of quantitative analysis of spermatogenic cell types in testicular cytologic aspirates of various sites of each testis. SETTING: In each aspirate, a total of 500 Sertoli cells and cells at each of the spermatogenic stages were identified, counted, and grouped according to cell type. A quantitative cell type index was calculated for each type of cell in each aspirate. Mean cell type indexes then were calculated for each of the cell types in the three aspirates of each patient, and variations of a given sample from its mean were compared. PATIENTS: Azoospermic or severely oligospermic infertile men. INTERVENTIONS: Fine needle aspiration performed on the upper, middle, and lower poles of each testis. RESULTS: Each of the aspirates showed wide deviations from the mean of the three aspirates for that patient. The deviation ranges of the cell type indexes of each of the spermatogenic stages were as follows: spermatogonia, 0.8% to 200%; spermatocytes, 1.4% to 94.3%; spermatids, 2.9% to 200%; and spermatozoa, 0.7% to 128%. In the majority of the patients, at least one of the three aspirates showed a cell type index score that was statistically different from the others. CONCLUSIONS: These results suggest that more than one testicular specimen is needed to evaluate quantitatively the spermatogenic process.

Assessment of spermatogenic process by deoxyribonucleic acid image analysis.

OBJECTIVE: To evaluate the spermatogenic process through cellular ploidy by image analysis. DESIGN: Twenty-six testicular aspirates from 24 infertile men were examined by fine needle aspiration (FNA) cytologic smears. These results were compared with the ploidy content of the cells using Feulgen stain, determined by image analysis. RESULTS: The results of both methods were divided into three categories: full spermatogenesis, spermatogenic arrest, and only Sertoli cell. There was a good correlation in 25 of 26 smears (96%). Ten patients who had a histogram of diploid showed only Sertoli cell cytologically. From nine patients who had a histogram of diploid and tetraploid, eight cytologically showed spermatogenic arrest and one showed full spermatogenesis. The seven patients who had full spermatogenesis (haploid, diploid, tetraploid) all had normal cytologic smears. CONCLUSIONS: Deoxyribonucleic acid image analysis is an objective qualitative and quantitative method for the evaluation of the spermatogenic process of the infertile male. It has several advantages over the flow cytometry method.

[Ultramorphological changes in sperm cells of infertile men with testicular varicocele].

The ultramorphological characteristics of sperm cells of infertile men with testicular varicocele were evaluated. There was an increased frequency of abnormal changes within their spermatozoa as compared to fertile men. The abnormalities included agenesis, partial agenesis, malformation and degradation of the subcellular organelles of the heads and tails of the sperm cells. These findings suggest the use of these characteristics as a part of a "varicocele score" to determine those who would benefit from varicocelectomy in the treatment of their infertility.

[Vanishing spermatozoa].

The mechanism of spermatozoal degeneration in male genital tract obstruction was evaluated. Epididymal fine-needle aspiration was performed in 17 azoospermic men with distal obstruction of the genital tract, but in whom testicular cytology revealed adequate spermatogenesis. From the contents of each puncture, smears with May-Grunwald-Giemsa and Papanicolaou staining were prepared. All indigenous epididymal cytologic types were identified in the smears. In aspirates which contained spermatozoa, large cells with foamy cytoplasm and round or kidney-shaped nuclei were also found. In some of these foam cells heads of mature spermatozoa were engulfed within the cytoplasm. It is suggested that in male genital tract obstruction, the role of the foam cells within the epididymis may be to phagocytize and disintegrate the trapped spermatozoa. The cellular origin of these foam cells is probably the epithelial lining layer of the epididymis itself.

The use of epididymal fine-needle aspiration for the diagnosis of male genital tract obstruction.

Epididymal fine-needle aspirations (EFNA) were performed on 29 epididymides of 17 infertile, azoospermic men in whom testicular cytology revealed adequate spermatogenesis. Sonograms confirmed the presence of all the excretory male organs. EFNA was performed to diagnose and locate the site of genital tract occlusion. The presence of cuboidal epithelial cells in the aspirate was the sole indication that epididymal content was aspirated. In 10 (34%) of the 29 aspirates there were only cuboidal epithelial cells and no spermatozoa, indicating proximal occlusion in the rete testis, in the canaliculi efferentes or in the most proximal segment of the epididymis itself. Occlusion in the distal segment of the epididymis or in the vas deferens was diagnosed in 13 epididymal aspirates (45%) which contained both cuboidal cells and spermatozoa. From 6 epididymides (21%) no conclusion could be reached because of failed aspirations. There are a variety of treatment modalities available for male genital tract obstruction, such as reconstructive surgery or assisted reproductive technology using sperm aspiration technique. The success rate of treatment is mainly dependent on the site of obstruction. Since vasography has its own limitations and hazards, diagnostic EFNA should be performed before treatment in men in whom genital obstruction is suspected.