Neuroendocrine characteristics of human Leydig cell tumours

Summary. The neuroendocrine nature of a subset of Leydig cells has already been established. The present investigation deals with neuroendocrine characteristics of Leydig tumour cells. A number of neuroendocrine and neuronal markers were demonstrated in Leydig cell tumours of 7 men aged 25–41 years. The following substances were immunocytochemically tested in Leydig tumour cells: the monoamine-synthesizing enzymes tyrosine hydroxylase and aromatic L-amino acid decarboxylase, the indoleamine serotonin, the calcium-binding protein parvalbumin, the microtubule associated protein-2, neurofilament protein 200, synaptophysin, neuron specific enolase, substance P and neuronal nitric oxide synthase (NOS). Compared to the normal interstitial cells beyond the tumours, all neoplastic cells showed a significantly weaker immunoreactivity for nerve cell markers as well as for testosterone and cyclic guanosine monophosphate (cGMP), which is usually accumulated by nitric oxide (NO). This provides evidence for a certain dedifferentiation of Leydig tumour cells.
However, these results suggest that tumourous development of Leydig cells does not include loss of neuronal phenotype. Moreover, on the assumption that 'neuronal' Leydig cells exist beside 'non-neuronal' ones in normal testicular tissue, we propose the hypothesis that 'neuronal' Leydig cells can transform to tumour cells.     

Non-tumoural parenchyma in Leydig cell tumours: pathogenetic considerations

Little is known about the pathogenesis of Leydig cell tumours (LCTs) of the testis. The observation of several associated dysgenetic features in the non-tumoural parenchyma and in the contralateral testes of men with testicular germ cell neoplasms has served as the basis to propose that there may be a common mechanism for different male reproductive disorders. However, the possible relationship between LCTs and other testicular lesions has not been explored. Here we describe the presence of primary lesions in the non-tumoural parenchyma of testes with LCT, from which we try to establish possible pathogenetic associations. We studied the non-tumoural parenchyma adjacent to 16 LCT specimens. Parameters as Leydig cell hyperplasia (LCHY), qualitative evaluation of the germinal epithelium and spermatogenesis, the presence of Sertoli cell-only tubules (SCOT), and the Sertoli cell nuclear morphology were consistently assessed in all cases. SCOT associated with Sertoli cell dysgenetic morphology was the most frequent finding, present in 50% of the cases. Another interesting finding was the presence of LCHY in four cases (25%). Abnormal spermatogenesis was found in 81.25% of the cases, and it consisted of lesions of the adluminal or basal compartments of seminiferous tubules. The occurrence of either dysgenetic Sertoli cells or LCHY adjacent to LCTs could represent primary anomalies, resulting from a common insult also involved in tumourigenesis. The abnormalities in spermatogenesis observed here are likely to represent consequences of either tumour compression or abnormal hormonal production. The significance of these associations merits further investigation regarding a common pathogenesis.     

Is testosterone involved in the initiation of spermatogenesis in humans? A clinicopathological presentation and physiological considerations in four patients with Leydig cell tumours of the testis or secondary Leydig cell hyperplasia

The purpose of this study was to evaluate the role of testosterone on the puberal development of spermatogenesis and to present additional clinicopathological data which bring about new information to this controversial subject. Four pre-pubertal patients are presented, 2 of them bearing Leydig cell tumours of the testis in the form of nodular masses. In both cases seminiferous tubules in the immediate vecinity to the tumours showed complete development of spermatogenesis, while those located away from the tumours were infantile in nature. Gonadotrophic levels were within the normal pre-pubertal range in these 2 cases. In one of the patients, testosterone concentration in the testis showed higher values than normal, and a concentration gradient was detected between the tumoral nodule and non-tumoral parenchyma. The 3rd patient had a pineal choriocarcinoma producing high amounts of hCG and consequently a diffuse hyperplasia of Leydig cells with high levels of plasma testosterone. Seminiferous tubules showed development up to pachytene spermatocytes. The last case was a precocious puberty in a boy with a tumour of the 3rd ventricle area. He had elevated levels of testosterone in the testis and plasma. In the testicular biopsy, stimulation of Leydig cells was detected. The seminiferous tubules showed mature Sertoli cells and pachytene spermatocytes. FSH levels were abnormally low. These 4 cases present in common different situations in which abnormally high amounts of testos-happens in the immature rat, the interaction between testosterone and gonadotrophins is essential for the normal initiation of spermatogenesis in normal puberty. Considerations are discussed on the possible synergistic role of gonadotrophins or other factors in relation with stimulation of seminiferous tubules by testosterone.     

Finasteride-related Leydig cell tumour: report of a case and literature review

Leydig cell tumours (LCTs) of the testis are rare. Their origin is still unknown. This case report describes a potential relationship between LCT and prolonged exposure to Finasteride.     

Local differences in Leydig cell morphology in the adult rat testis: evidence for a local control of Leydig cells by adjacent seminiferous tubules

In order to test the hypothesis that Leydig cell function in the adult rat testis is influenced by the surrounding tubules, Leydig cell morphology was compared in different types of interstitial areas. Triangular interstitial areas surrounded by 3 cross-sectioned tubules in nearly the same stage of spermatogenesis were chosen for quantitative light microscopy. It was found that the volume density of Leydig cells in such areas was about 30%, except when the surrounding tubules were in stages IX-X or XI-XII, when it was only about 20%. This variation in total Leydig cell mass seemed to be due to a variation in Leydig cell size and not in Leydig cell number. The largest Leydig cell profile area, 118 pL 6 μm² (mean pL SE n = 6 rats), was observed when the surrounding tubules were in stages VII-VIII, i.e. just prior to sperm release. The smallest Leydig cells were seen when the surrounding tubules were in stages IX-X and XI-XII (68 pL 3 and 66 pL 4 μm²). The present results indicate that there may be a Leydig cell cycle in the adult rat testis, which is regulated by the adjacent tubules.     

Early signs of Sertoli and Leydig cell dysfunction in the abdominal testes of immature unilaterally cryptorchid rats

Testicular descent was prevented unilaterally by cutting the gubernaculum testis of newborn rats. When 20 days old unilaterally cryptorchid rats were injected intraperitoneally with 2 μg bFSH per gram body weight and killed 6 h later when testicular testosterone (T) and oestradiol (E₂) concentrations were determined. The increase in E₂ was subnormal in abdominal testes. In 18-day-old unilaterally cryptorchid rats the efferent ducts were ligated bilaterally, and the rats were killed 48 h later. The weight increase, due to accumulation of seminiferous tubule fluid, was significantly greater in the abdominal testes. In contrast, the ABP content of the abdominal epididymis was subnormal in 20-day-old unilaterally cryptorchid rats. Unilateral orchidectomy was performed in 16-day-old unilaterally cryptorchid rats and at 20 days of age intratesticular T and E₂, and plasma FSH and LH concentrations were determined and compared to that in 20-day-old control unilaterally cryptorchid rats. Removal of an abdominal testis resulted in increased plasma FSH and intratesticular E₂, whereas plasma levels of LH and intratesticular levels of T were unaffected. Removal of a scrotal testis resulted in increased plasma FSH and LH coupled with increased intratesticular T and E₂. Rats with a single abdominal testis had higher plasma FSH and LH and intratesticular T, but similar intratesticular E₂, than rats with a single scrotal testis. It is concluded that Sertoli and Leydig cell function are influenced by cryptorchidism at a stage when the temperature difference, and the morphological differences between the testes are very discrete.     

大鼠睾丸间质细胞移植的实验研究

目的:观察大鼠同种异体睾丸间质细胞移植后的血清睾酮变化水平。方法:采用Percoll方法分离提取SD大鼠两侧睾丸间质细胞,异体移植后每月测定受体血清睾酮1次,连续3次。结果:睾丸间质细胞移植后,受体动物血清睾酮含量逐渐升高。移植术后3个月,血清睾酮水平明显上升,与出生后2个月内的动物比较,差异具有显著性。结论:异体睾丸间质细胞移植治疗男性原发性性腺功能低下症可能具有良好的应用前景。     

The Total Leydig Cell Volume of the Testis in some Common Mammals

Total-Leydig-Zell-Volumen des Hodens bei einigen Säugern
In einer quantitativen Studie wurde mittels der histometrischen Punktzählmethode bei sechs Säugern einschließlich des Menschen das Total-Leydig-Zell-Volumen bestimmt. Obgleich die Größe der Hoden von 1,3 ±0,1 ml bei der Ratte auf 19.8 ± 6.9 ml beim Büffel anstieg, blieb die Zusammensetzung leidlich konstant, wobei die Leydig-Zellen 9–16% des Hodenvolumens ausmachten. In absoluten Werten ergab sich, daß das Total-Leydig-Zell-Volumen progressiv mit dem Körpergewicht anstieg, und zwar von 0.11 ± 0.02 ml/Hoden bei der Ratte auf 2,44 ± 0.64 ml/Hoden beim Büffel; der Wert für den Menschen betrug 2.21 ± 0.40 ml/Hoden. Die Signifikanz dieser Resultate wird unter dem Gesichtspunkt der vergleichenden Physiologie der Reproduktion diskutiert.     

Local regulation of Leydig cells by the seminiferous tubules. Effect of short-term cryptorchidism

Unilateral cryptorchism was induced in adult rats for 24 h, and its effect on testicular morphology and intratesticular testosterone concentration after hCG-stimulation were studied. In seminiferous, tubules from abdominal testes an increased number of degenerating germ cells was noted in stages XIV-III of the spermatogenic cycle and Sertoli cells contained an increased amount of lipid droplets in stages XIV-VIII. However, germ cells and Sertoli cells from tubules at other stages of the cycle appeared unaffected. In scrotal testes the size of peritubular Leydig cells varied in phase with the spermatogenic cycle. The largest cells were found adjacent to stage VII-VIII and the smallest adjacent to stage XI-XII. In abdominal testes no stage-dependent variation in the size of peritubular Leydig cells was seen. Perivascular Leydig cells were of equal size in abdominal and scrotal testes. The testicular testosterone concentration following stimulation with a low dose of hCG was significantly lower in abdominal testes. It is suggested that the seminiferous tubules locally modulate Leydig cell function and that the stage specific stimulatory influence from stage VII-VIII is rapidly lost during experimental cryptorchidism.     

睾丸间质干细胞分化和移植

睾丸间质细胞是男性体内合成雄激素的主要细胞,胚胎发育期中肾胚的间质细胞及生精小管周成纤维样细胞可能是睾丸间质细胞的干细胞。在胚胎期间质干细胞分化为胎儿型间质细胞;出生后间质干细胞经间质祖细胞、未成熟间质细胞分化为成熟间质细胞。老年期间质细胞数量可能不变,但雄激素合成下降。间充质干细胞及脂肪干细胞等干细胞经诱导可分化为分泌雄激素的睾丸间质细胞,因此,间质干细胞移植可望成为治疗男性性腺功能不全和中老年雄激素缺乏的创新方法,本文对睾丸间质干细胞的分化及移植方面研究进行综述。     

Paracrine regulation of Leydig cells by the seminiferous tubules

Testes of adult control and unilateral cryptorchid rats were fixed by vascular perfusion. The cell profile area of peritubular Leydig cells surrounding tubules in different stages of spermatogenesis, and the cell profile area of perivascular Leydig cells were determined. The size of peritubular Leydig cells was dependent on which type of tubulus the cells were surrounding. Some peritubular Leydig cells, especially those surrounding stages VII–VIII (88.1 ± 7.1 μm², mean ± SD, n = 6 rats), were larger than perivascular Leydig cells (69.3 ± 5.9 μm²). The size of Leydig cells surrounding stages IX–XIV was similar to that of perivascular cells. In the abdominal testes no spermatogenic cycle was present and the sizes of peritubular and perivascular Leydig cells were equal (63.0 ± 5.1, vs 66.7 ± 7.3 μm², mean ± SD, n = 5 rats). It is suggested that the tubules and the spermatogenic cycle locally modulate Leydig cell activity and that Leydig cell malfunction in abdominal testes may be due to a decreased stimulatory influence from the damaged tubules.     

Twenty‐nine Leydig cell tumors: Histological features,outcomes and implications for management

Leydig cell tumors are the most common interstitial neoplasm of the testes. Metastatic progression is historically quoted at over 10%, fuelling uncertainty as to the safety of testis sparing surgery. Between June 1998 and March 2009, 29 patients underwent surgery for Leydig cell tumor of the testis in our cancer network. We reviewed their histological features and clinical outcomes. Four patients with sub‐5 millimetre lesions underwent testis sparing surgery and 25 were treated with radical orchidectomy. Histopathological characteristics that have been linked with risk of malignant progression were seen infrequently in our cohort: diameter greater than 50 mm, 0%; nuclear atypia, 14%; >3 mitoses per 10 high‐power fields, 3%; infiltrative borders, 10%; necrosis, 3%; and vascular invasion 0%. No patient developed local or distant recurrent disease over a median follow up of 49 months, including seven and four patients disease‐free at 5 and 10 years, respectively. The rate of metastatic progression is likely to be significantly less than 10%. Our data suggest that, in the absence of high‐risk histopathological features, this tumor can be safely regarded as benign, pending a longer‐term follow‐up evaluation.     

Flow cytometry derived DNA content of the primary lesions of advanced germ cell tumours

We currently lack good prognostic indicators for patients with advanced disseminated germ cell tumours (GCT). An analysis of archival tumour samples of forty-nine patients with advanced GCT for flow cytometry derived DNA content suggests an inverse relationship between tumour proliferative activity and patient survival. The great majority of patients with advanced GCT have aneuploid tumour subpopulations. This analysis suggests that the survival of patients with advanced GCT may depend in part on kinetic variables present in the primary tumour.     

Effect of cryptorchidism on testicular and Leydig cell androgen production in the mouse

Unilateral cryptorchidism was induced surgically in adult mice and the effects on testicular and Leydig cell steroidogenesis were studied after 7 weeks. There was a 60% reduction in weight of the cryptorchid testis and this was associated with a significant reduction in intratesticular androgen content, both under basal conditions and following an injection of hCG. Testicular androgen production in vitro was also significantly lower in the cryptorchid testis compared to the scrotal testis, again under both basal conditions (29 +/- 6% of control) and in the presence of hCG (46 +/- 9% of control). Scrotal testes from the unilaterally cryptorchid animals did not show any significant difference in steroidogenic capacity compared to testes from untreated control animals. The decrease in steroidogenic capacity of the cryptorchid testis was due, at least in part, to a reduction in activity for each Leydig cell. In four experiments, androgen production by Leydig cells isolated from cryptorchid testes was 48 +/- 9% of cells from scrotal testes in the presence of a saturating dose of hCG. Under basal conditions the effect was more variable between experiments with steroid secretion by Leydig cells from cryptorchid testes being 58 +/- 32% of that for cells from scrotal testes. Leydig cell steroidogenesis in the scrotal testes of unilaterally cryptorchid animals did not differ significantly from untreated controls. These results show that induced cryptorchidism in the mouse causes a significant reduction in Leydig cell activity. This is apparently different from the effects of this procedure on the rat and raises the possibility that intratesticular regulation differs between the two species.     

Increased incidence of Leydig cell tumours of the testis in the era of improved imaging techniques

Study Type – Diagnostic (exploratory cohort) Level of Evidence 2b What’s known on the subject? and What does the study add? Leydig cell tumours (LCTs) of the testis are rare tumours, accounting for 1–3% of all testicular neoplasms. Our data indicate that using scrotal ultrasound with high resolution imaging in routine checkups leads to an earlier detection of LCTs. Most patients underwent an organ‐sparing surgery and no androgen deprivation was observed.

OBJECTIVE

? To report an observed high frequency of Leydig cell tumours (LCTs) diagnosed at our centre.

PATIENTS AND METHODS

? Charts of all patients who underwent surgery for a testicular tumour between 1999 and 2008 at our department were searched and data from patients with LCT were collected. ? Before surgery all patients underwent ultrasound and complete staging. In all but two patients with LCT an organ‐sparing surgery was performed. Surgery was performed under ultrasound or palpation guidance. ? All patients underwent postoperative follow‐up. We retrospectively reviewed surgical technique, histology, epidemiology and outcome in all LCT patients.

RESULTS

? In the study period, 197 testicular tumours were surgically removed of which 29 were diagnosed as LCT (14.7% of 197; further study group) in 25 patients. Mean age of patients with LCT was 45 years (range 21–68 years). ? Tumour size ranged from 1.2 to 80 mm (mean 10.23 mm). In two patients (8%) the lesion was palpable whereas incidental diagnosis was made in seven patients (28%). ? In the remaining patients diagnosis was made by ultrasound performed for testicular pain (six patients, 24%) or during infertility or erectile dysfunction evaluation (10 patients, 40%). ? Definitive histology reported no malignant histopathological features in all but one patient; this particular patient experienced tumour progression after 2 months and died from advanced disease 1 year later. All other patients are free of disease after a mean follow up of 56 months (range 7–93 months). ? During this period one patient developed a second LCT on the contralateral side; another patient had a recurrence within the same testicle, but on the opposite pole. Both underwent a subsequent organ‐sparing tumour resection.

CONCLUSION

? The percentage of LCT (14.7% of all testicular tumours removed) was significantly higher than expected from the literature. One possible explanation for this phenomenon is the increasing use of better ultrasound technology and the subsequent increased detection of small nodules that have not been found in historical series. Use of ‘observation‐only’ for very small lesions detected at infertility clinics is under debate.     

Influence of rat testicular macrophages on Leydig cell function in vitro

The influence of co-cultures of rat testicular macrophages and Leydig cells (LC) on LC morphology and steroidogenesis was investigated with and without macrophage stimulation by a bacterial lipopolysaccharide (LPS). LC showed an elongated form in the presence of stimulated testicular macrophages. In the presence of non-stimulated testicular macrophages a significant inhibition of testosterone production was observed (decrease of 33%) from 48 h in co-culture while an increase of 16% was obtained at the same culture time, after stimulation of macrophages by LPS. When LC were treated with testicular macrophage-conditioned media (MCM) obtained from LPS-treated macrophages, they became fusiform and there was stimulation (78%) of steroid production. After human FSH stimulation (1-1000 mIU ml-1), MCM from testicular macrophages was no more effective in enhancing testosterone production by LC than was media from untreated LC. Similar experiments with LPS were conducted with macrophages of peritoneal origin. Peritoneal macrophages stimulated or not by LPS in co-cultures with LC or peritoneal MCM did not significantly modify testosterone production. However, these cells were able to modify LC morphology when LPS-MCM was added to LC-culture medium. The present results suggest strongly that testicular macrophage-LC interactions could be important in the control of LC steroidogenesis.     

Cyproterone acetate induced changes in the behaviour of hydroxysteroid dehydrogenases in rat Leydig cells during perinatal development

The influence of cyproterone acetate (CA) upon the behaviour of hydroxysteroid dehydrogenases (HSDH) in the Wistar rat Leydig cells was investigated during the perinatal phase with the help of enzymhistochemical cum morphometrical techniques. The pregnant rats as well as their offsprings were injected with CA (dosage: 35 mg/kg body wt) sc, daily from 14 fetal day upto 31 postnatal day (p.n.d.). The animals were killed on 5, 20, and 32 p.n.d.; the enzymhistochemical reactions for 3-beta-HSDH, 11-beta-HSDH, 17-HSDH and 3-alpha-HSDH were performed in the cryostat sections of the testis, and the morphometric evaluation of HSDH positive Leydig cells was carried out. On 5 p.n.d. the activity of 17-beta-HSDH was slightly impaired in the intertubular Leydig cells of the CA treated animals. On 20 p.n.d., CA prevented nearly completely the HSDH activity in the newsly built peritubular Leydig cells; the activities of 3-beta-HSDH, 17-beta-HSDH, and 3-alpha-HSDH resided mainly in the intertubular Leydig cells. On 32 p.n.d. the HSDH activities in the Leydig cells were observed in the control as well as in treated animals. It seems that the differentiation of peritubular Leydig cells, and thereby the steroid production, is delayed by CA, but not entirely blocked.