Effect of ethinyl estradiol on the differentiation of mouse fetal testis

In an evaluation of the effect of ethinyl estradiol (EE) on the differentiation of fetal mouse testes, the ratio of the seminiferous tubular region to the testicular tissue region, the ratio of Sertoli cells to gonocytes in tubule cross sections, and the size of Leydig cells were determined by the Texture Analyse System (T.A.S., Leitz) in histological preparations of the testes. The testes were those of fetuses taken from dams given orally 0, 0.02, 0.2 or 2.0 mg/kg of body weight of EE in olive oil from day 11 through day 17 of gestation and killed at term. From experimental and the control testes, five sections were taken at 40-micron intervals. The areas of the seminiferous tubular region and the testicular region were determined and the Sertoli cells and gonocytes in tubule cross section were counted in each of the five sections. The diameters of 100 Leydig cells selected at random were averaged. These data were analyzed by Student's t test. The seminiferous tubular region was significantly increased in the testes treated with 0.02 mg/kg of EE and significantly decreased in those treated with 0.2 mg/kg of EE. The number of gonocytes per tubule cross section was significantly increased in the testes treated with 0.02 or 2.0 mg/kg of EE. The number of Sertoli cells per tubule cross section and the number of Sertoli cells per gonocyte were significantly decreased in the experimental testes. The size of the Leydig cells was significantly decreased in the testes treated with 0.2 mg/kg of EE. These findings suggest that prenatal exposure to EE before testicular differentiation affects tubular formation, the proliferation of fetal Sertoli cells, and Leydig cell differentiation, resulting in disturbances of spermatogenesis.     

Morphologic study of the testes from spontaneous unilateral and bilateral abdominal cryptorchid boars

Macroscopical and histological characteristics were examined in both testes from three healthy boars, three boars with unilateral abdominal cryptorchidism on the right side, and three boars with bilateral abdominal cryptorchidism. Abdominal cryptorchidism, unilateral and bilateral, provoked a significant decrease of the weight and volume of the ectopic testes. The scrotal testis of the unilateral cryptorchid boars showed an increase in its volume and weight. Cryptorchidism also induced abnormalities in the histological structure of seminiferous tubules, lamina propria, and interstitial tissue of the abdominal testes. The number of seminiferous tubules decreased; the seminiferous epithelium was constituted by few spermatogonia with an atypical pattern and by abnormal Sertoli cells. The lamina propria showed a variable degree of thickening and collagenization. The interstitial tissue was very developed but displayed a decrease in the Leydig cell population. These abnormalities were more critical in bilateral cryptorchidism than in unilateral cryptorchidism. The scrotal testis of the unilateral cryptorchid boars showed normal appearance, but a decrease of the number of seminiferous tubules was observed. Moreover, the seminiferous tubules showed impaired spermatid maturation. The alterations observed in the abdominal testes of the unilateral and bilateral cryptorchid boars were attributed to defective proliferation and differentiation of Sertoli cells and Leydig cells. The anomalies in the scrotal testis of the unilateral cryptorchid boars were due to disturbances in the Sertoli cell activity.     

Cytology of the interstitial tissue in scrotal and abdominal testes of post-puberal boars

The interstitial tissue of the testes from healthy boars, and unilateral and bilateral abdominal cryptorchid boars was examined by light and transmission electron microscopy. The left and right testes of healthy boars, and the left (scrotal) testis of unilateral cryptorchid boars had abundant mature Leydig cells, few fibroblasts and mast cells, scarce and small blood vessels, and little lymphatic areas. The right (abdominal) testis of unilateral cryptorchid boars contained abundant Leydig cells, fibroblasts and erythrocytes, scarce mast cells, and frequent blood vessels; Leydig cells exhibited either a mature but degenerative appearance or an immature appearance, and fibroblasts displayed immaturity signs. The interstitial tissue of the left (abdominal) testes of bilateral cryptorchid boars had small blood vessels surrounded by erythrocytes, lymphocytes, and few plasma cells, and abundant mature and immature Leydig cells, immature fibroblasts, and mast cells. Mature Leydig cells showed mid or advanced degeneration, and immature Leydig cells displayed either non-degenerative or degenerative patterns. The right (abdominal) testes of bilateral cryptorchid boars contained scarce immature Leydig cells in advanced degeneration, large fibrous and adipose areas, and blood vessels. These results indicated that unilateral abdominal cryptorchidism affect neither the structural nor the cytologic features of the interstitial tissue in scrotal testes. Unilateral and bilateral cryptorchidism induced abnormal differentiation of Leydig cells and fibroblasts leading to decreased steroid production and increased collagenization in abdominal testes.     

The expression of aromatase, estrogen receptor alpha and estrogen receptor beta in mouse Leydig cells in vitro that derived from cryptorchid males

A broad expression of aromatase and estrogen receptors (ERs) in the testis suggests an important role for estrogens in regulating testicular cell function and reproductive events. The aim of the present study was to show whether Leydig cells in vitro isolated from cryptorchid testes of two inbred strains of mice, KE and CBA, are a site of estrogen synthesis. Using immunocytochemistry, aromatase, estrogen receptor alpha(ERalpha), and estrogen receptor beta(ERbeta) were localized in cultured Leydig cells. Immunoreactive aromatase was found in the cytoplasm of control Leydig cells and those isolated from cryptorchid males, however the intensity of immunostaining was different, being stronger in Leydig cells deriving from cryptorchid mice. The strongest aromatase immunostaining was found in cryptorchid-KE Leydig cells. Strong immunoexpression of ERalpha was detected in the nuclei of both KE-and CBA-Leydig cells. The intensity of ERalpha immunostaining was stronger in cultured cells deriving from cryptorchid testes. ERbeta immunoexpression was detected predominantly in KE-Leydig cells. Control CBA-Leydig cells were negative for ERbeta or the result was inconclusive, whereas in cryptorchid CBA-Leydig cells a weak immunostaining was present in their nuclei. Western blot analysis confirmed the results obtained by immunocytochemistry. In KE- and CBA-Leydig cells aromatase as a band of 55 kDa protein was present, whereas ERalpha molecular weight was 67 kDa on Western blots. No band was detected for ERbeta. Radioimmunological analysis revealed that androgen and estrogen levels secreted by Leydig cells in vitro were strain-dependent. Additionally, in KE-Leydig cells that derived from cryptorchid mice estrogen level was distinctly higher in comparison with that of the respective control.     

双峰驼睾丸和隐睾细胞外基质相关蛋白的组织化学特征及超微结构比较

为探索细胞外基质相关蛋白在隐睾双峰驼的分布情况及其组织化学特征,应用电镜技术和多种组织化学方法比较了隐睾和正常睾丸的超微结构,组织化学特点及层粘连蛋白（LN）、Ⅳ型胶原（Col Ⅳ）和硫酸乙酰肝素糖蛋白（HSPG）的分布特征。结果显示：(1)与正常睾丸间质结构相比,光镜下隐睾生精小管发育不全,间质内胶原纤维稀疏,网状纤维分布明显,间质血管及生精小管固有膜PAS及AB-PAS阳性反应较弱。电镜下,隐睾生精上皮基膜明显增生,外围I型胶原纤维较少,管周肌样细胞不典型;间质毛细血管及Leydig细胞周围纤维细胞多见,而正常睾丸在间质毛细血管及Leydig细胞周围多分布有成纤维细胞。(2) 免疫组织化学染色显示,正常睾丸组织的Col Ⅳ、LN及HSPG在Leydig细胞内均为强阳性表达,Col Ⅳ和LN在毛细血管内皮细胞强阳性表达,后者在Sertoli细胞的表达尤为明显,HSPG在精原细胞无表达;隐睾时Col Ⅳ、LN及HSPG在Leydig细胞内阳性表达均明显减弱,Col Ⅳ、LN在管周肌样细胞及毛细血管内皮细胞阳性表达也减弱明显,HSPG在精原细胞较强阳性表达,且在精子细胞呈强阳性表达。免疫组织化学图像分析结果显示,双峰驼正常睾丸组织中Col Ⅳ和LN的分布显著高于隐睾组织（P<0.05）,HSPG检测结果在正常睾丸与隐睾之间无统计学差异（P>0.01）。该研究表明,双峰驼隐睾生精小管发育异常,间质组织中合成胶原纤维的能力下降,睾丸细胞外基质的重要成分Col Ⅳ,LN与正常组差异显著与生精小管及Leydig细胞异常发育有关,而HSPG在隐睾生精上皮的强阳性表达与精原细胞发育不成熟密切相关。     

Ovarian follicular cell hyperplasia in fetal mice treated transplacentally with ethinyl estradiol

The occurrence of follicular cell hyperplasia was studied by light and electron microscopy in fetal mouse ovaries exposed to ethinyl estradiol (EE) from day 11 through day 17 of pregnancy. Pregnant mice were given EE in olive oil (0.02, or 0.2 mg/kg of body weight) and were sacrificed on day 18. The female fetuses were examined for ovarian histogenesis. Follicular cell hyperplasia was detected in both of the experimental groups, but the incidence was statistically significant only in fetuses exposed to 0.2 mg/kg of EE. Light and electron microscopic observations of the ovaries showed that the hyperplasia was located in the medullary region, and the follicular cells showed pleomorphism. Accumulation of abundant lipid droplets, enlarged rough endoplasmic reticulum with granular material, dense bodies, and vague masses of fibrous structures were seen in the cytoplasm. These morphological observations indicate that hyperplasia of follicular cells in fetal mouse ovaries at term can be induced by prenatal treatment with EE.     

Expression of anti-Müllerian hormone (AMH) in the equine testis

Anti-Müllerian hormone (AMH) induces regression of Müllerian ducts during male fetal development; in the human male, it is expressed in Sertoli cells during fetal development (and through puberty). The objective was to characterize expression of AMH in the fetal, neonatal, prepubertal, and adult equine testis, as well as in equine cryptorchid testes, in select testicular neoplasms, and in intersex gonads, based upon immunohistochemistry (IHC). Testes were removed from equine fetuses at 5.5, 10, and 11 months of gestation, at 12 months of age, and from adult stallions. In addition, cryptorchid testes, testis tumors (teratomas, seminomas, Sertoli cell tumors), and male intersex gonads were examined by IHC for expression of AMH using a goat polyclonal primary antibody (alpha-AMH) directed against a C-terminal peptide antigen from human AMH. Immunolabeling with alpha-AMH was localized to Sertoli cells within the developing seminiferous tubules of fetal, neonatal and prepubertal equine testes, with no expression detected in Sertoli cells from normal adult equine testes. Furthermore, expression was detected in cryptorchid testes (in animals up to 3-4 years of age) and in Sertoli cell tumors and male intersex gonads. In conclusion, AMH was strongly expressed by Sertoli cells in fetal, neonatal and prepubertal equine testes, but not in normal adult testes. That AMH was expressed in cryptorchid testes may provide a useful biomarker for detection of cryptorchid testes, as well as for immunohistochemical characterization of testicular tumors and intersex gonads in the horse.     

The effects of cryptorchidism on the regulation of steroidogenesis and gap junctional communication in equine testes

INTRODUCTION: Evidence collected over the years has demonstrated that cryptorchidism is associated with a defect in spermatogenesis and, as a consequence, with either reduced fertility or infertility. However, the effect of cryptorchidism on Leydig cell function is less clear. The aim of our study therefore was to investigate the regulation of steroid hormone biosynthesis and, additionally, intercellular communication in the cryptorchid equine testes. MATERIAL AND METHODS: Testes of mature bilaterally cryptorchid horse and healthy stallions were used for this study. The expression of luteinising hormone receptor (LHR), 3beta-hydroxysteroid dehydrogenase (3beta-HSD), aromatase and connexin43 (Cx43) was detected by means of immunohistochemistry. Testosterone and oestradiol levels were measured in testicular homogenates using appropriate radioimmunoassays. RESULTS: In the testes of both normal and cryptorchid stallions, immunostaining for LHR, 3beta-HSD and aromatase was confined to the Leydig cells. In the cryptorchid horse, the intensity of the staining for LHR and 3beta-HSD was weaker, whereas the staining for aromatase was clearly stronger than that of the normal stallion. Radioimmunological analysis revealed disturbance of the androgen-oestrogen balance in the cryptorchid testes. Additionally, in both the seminiferous tubules and interstitial tissue of the cryptorchid a clear reduction of the Cx43 signal was observed. CONCLUSIONS: Decreased expression of LHR and 3beta-HSD and increased expression of aromatase in the cryptorchid testes suggest that hormonal imbalance was caused both by reduced testosterone synthesis and by increased androgen aromatisation. Impaired expression of Cx43 in the seminiferous tubules as well as in the interstitial tissue of the cryptorchid horse indicates that cryptorchidism affects intercellular communication in the testes.     

The effect of spermatogenic disruption on the ability of testicular fluid to stimulate androgen production by normal Leydig cells

Reports from this and other laboratories have concluded that unilateral disruption of spermatogenesis induces a predominantly ipsilateral increase in the responsiveness of Leydig cells to stimulation with luteinizing hormone (LH) and have suggested that if such effects were mediated by locally produced hormones then such "factors" should be detectable in testicular interstitial fluid. We sought to demonstrate such factors in testicular fluid from gonads subjected to a variety of treatments that disrupt gametogenesis. Fluid (TF) was drained from testes of adult rats that had been sham treated, irradiated, or treated with busulfan in utero, made unilaterally or bilaterally cryptorchid, or were unilaterally or bilaterally efferent-duct-ligated. Leydig cells obtained from normal rats basally produced 8 +/- 1 ng androgen/10(6) Leydig cells/2 h and, when maximally stimulated with LH, produced 66 +/- 3 ng. The addition of the various TFs to the incubations significantly increased both basal and LH-stimulated androgen production. TF from lesioned testes was more effective in increasing androgen production than TF from control rats. Unilateral lesions caused an increase in the ability of TF from the disrupted testes to increase the androgen production by normal Leydig cells, as compared to TF from contralateral testes. Thus, locally produced "factor(s)" do appear to modify Leydig cell function. Additional studies using TF from control and bilaterally cryptorchid animals suggest that the "factor' in TF is heat-labile; has a molecular size between bovine serum albumin and ovalbumin; exerts a portion of its action independently of cAMP formation; and does not appear to be LH, follicle-stimulating hormone, prolactin, or gonadotropin-releasing hormone.     

Testosterone restores testicular inhibin content in cryptorchid rats

The effects of testosterone administration on testicular inhibin content and histology were studied in bilaterally cryptorchid rats, in which a marked decrease in testicular inhibin content had been observed. Mature male Wistar rats weighing approximately 300 g were made bilaterally cryptorchid by placing the testes in the abdominal cavity. Testosterone in oil, 0.1, 1.0 or 10 mg, was given i.m. each week. Testicular inhibin and testosterone content, histology and plasma LH, FSH and testosterone were studied 2 weeks later. Abnormally decreased testicular inhibin in cryptorchidism was restored toward normal by testosterone in a dose dependent manner in 2 weeks after surgery. Sertoli cell structure also recovered toward normal with increasing amount of testosterone. Decreased testicular testosterone content and Leydig cell atrophy were observed with suppressed plasma LH and FSH after testosterone. These results showed that the increased plasma concentration of testosterone had a stimulatory effect on the Sertoli cell function in cryptorchidism, in which compensated Leydig cell failure was demonstrated.     

Distribution of extracellular matrix related proteins in normal and cryptorchid ziwuling black goat testes

The Ziwuling black goat is an indigenously in China, their offspring are frequently affected by congenital cryptorchidism. The extracellular matrix (ECM) contains cytokines and growth factors that regulate the development of the testis, and component changes often result in pathological changes. Cryptorchidism is closely related to structural changes in ECM. In this study, the histochemical staining, immunohistochemical, immunofluorescence and Western blot combined with semi-quantitative analysis was used to describe the distribution of the important ECM components Collagen type IV (Col IV), laminin (LN)and heparan sulfate proteoglycans (HSPG) in the normal and cryptorchid testes of Ziwuling black goats. Results showed that: The histochemical staining showed that the dysplasia of seminiferous tubules and decreased number of Sertoli cells in cryptorchidism, as well as sparse collagen fiber. Meanwhile, the distribution of reticular fibers is relatively rich. Furthermore, the PAS and AB staining in the interstitial vessels and lamina propria of seminiferous tubules is weak. The immunohistochemical and immunofluorescence revealed that Col IV, LN was strongly expressed in Leydig, Sertoli cells of normal testes and moderately positive in the spermatogonia and spermatids, but HSPG was not expressed in the spermatogonia. However, cryptorchidism, the expression of Col IV, LN and HPSG in Leydig, Sertoli cells significantly decreased, as well as the expression of Col IV and LN in capillary endothelial cells, but HSPG was moderately expressed in spermatogonia. Based on these data, the underdevelopment of spermatogenic epithelium, decreased synthesis function of collagen fibers and Leydig cells develop usually in the cryptorchidism were shown to be closely related to the abnormal metabolism of Col IV and LN. The positive expressed of HSPG in the spermatogonia of cryptorchid testes is related to the compensatory development of spermatogonia.     

Gonadal dysgenesis induced by prenatal exposure to ethinyl estradiol in mice

Daily oral administration of ethinyl estradiol (0.02, 0.2, or 2.0 mg/kg of body weight) to pregnant Jc1:ICR mice resulted in ovotestis and intra-abdominal testis with persistent Müllerian duct and Wolffian duct in male fetuses and ovarian hypoplasia in female fetuses when it was given from day 11 through day 17 of gestation (before gonadal differentiation in the fetus). The ovotestis consisted of testicular and ovarian portions. In the testicular portion, a few solid seminiferous tubules containing spermatogonia, some with pachytene nuclei with Sertoli cells and compact interstitial tissue including Leydig cells, were seen. In the ovarian portion, pachytene nuclei were seen. The intra-abdominal testis was smaller and contained more spermatogonia per tubule in cross section than the control testis. These findings suggest that in male fetuses ethinyl estradiol affects Sertoli cell differentiation resulting in suppression of Müllerian inhibiting factor. On the other hand, in the ovarian hypoplasia, the primordial follicles and follicular cells in a primordial follicle were significantly decreased in number, and the number of the degenerated primordial follicles was significantly increased. It seems likely that ethinyl estradiol affects the intimate contact between follicular cells and oocytes to cause degeneration of primordial follicles.     

Structural and functional abnormality of ectopic testes in rats

Some males of a mutant strain of King-Holtzman rats exhibit an anomalous heritable defect manifested as either unilateral or bilateral ectopic testes. In the adult, these testes contain seemingly immature Sertoli and Leydig cells, seminiferous tubules greatly reduced in diameter, and exhibit arrested spermatogenesis. Thus, the affected testis is essentially sterile. An inability to produce normal amounts of testosterone and androstenedione by these gonads is probably a reflection of changes that have been effected in their Leydig cells. Thus, this study suggests that abnormal function of the Leydig and Sertoli cells and seminiferous tubule failure in these mutant animals result from the physiologically cryptorchid condition.     

Intrascrotal temperature, testicular histology and fertility of heart-acclimatized rats

This study investigated intrascrotal and deep body temperatures, reproductive ability, and histological patterns in the testes of heat-acclimatized rats. Albino rats aged 100-110 days were acclimatized to 35 degrees C. and a relative humidity of 25-40% for at least 3 months. There was free access to food and water and 14 hours of light daily. A control group was kept at an ambient temperature of 22 degrees C. and relative humidity of 35-50%. Deep-body temperature was measured with a thermistor probe inserted 4 cm into the rectum. Intrascrotal temperatures were measured with a thermocouple contained in a 27-gauge hypodermic needle inserted into the scrotum after the testes had been displaced. Readings were taken on alternate days for 30 days. Deep-body and intrascrotal temperatures were higher in experminetal than in control animals. Both groups maintained differences between body and intrascrotal temperatures. The intrascrotal temperatures of heat-acclimatized rats resembled the deep-body temperatures of the controls and, therefore, were similar to the environmental temperatures of cryptorchid testes. The mating rate of heat-acclimated males was lower than that of controls and fewer females conceived when mated with experimental males. However, in females which did conceive the type of male had no effect on implantation, pregnancy loss, or number of young. In about 20% of the experimental animals seminiferous tubules showed necrobiosis of germinal epithelium. Slight hyperplasia of Leydig cells was noted. The epididymal epithelium was intact and normal spermatozoa were present in the lumen. It is suggested that the gradient between the internal body termperature and the termperature of the testes is essential for spermatogenesis. Similar temperatures without the gradient, as in cryptorchid testes or heated scrotum, have been shown to be detrimental to spermatogenesis.     

Expression of the NSE,SP,NFH and DβH in normal and cryptorchid testes of Bactrian camel

Neuroendocrine substances play essential roles in regulating the normal physiological functions of testicles. The purpose of this study is to explore the localization and effects of four neuroendocrine markers (NSE, SP, NFH and DβH) in normal and cryptorchid testes of Bactrian camels using western blotting, transmission electron microscopy, immunohistochemistry, and immunofluorescence methods. The results showed that cryptorchidism caused a reduction in layers of spermatogenic epithelium and decreased glycogen positivity in the basement membrane. The ultrastructure revealed that macrophages were always found around the Leydig cells, crowded with swelling mitochondria in cryptorchidism. Expression of NSE in the Leydig cells of cryptorchidism was significantly weakened compared to that in the normal group(p<0.01). We found that SP was always distributed along the nerve fibers in normal testes and was expressed in the Leydig cells of cryptorchidism. However, expression of NFH in the cryptorchidic tissue was strongly positive in the spermatogenic epithelium, with limited expression in Leydig cells and no expression in peritubular myoid cells. Therefore, the expression of DβH in the Sertoli cells was comparatively strong in both the normal and cryptorchidism groups. NFH and DβH expression was significantly increased in the cryptorchidism group compared with the normal group (p<0.01). These findings indicated that the underdeveloped seminiferous epithelium and pathological changes in cryptorchid tissue in Bactrian camels were potentially related to a disorder in glycoprotein metabolism. Our results suggest that NSE and SP could help judge the pathological changes of cryptorchidism. The present study provides the first evidence at the protein level for the existence of NFH and DβH in Sertoli and Leydig cells in Bactrian camel cryptorchidism and provides a more in-depth understanding of neuroendocrine regulation is crucial for animal cryptorchidism.     

Testicular function and Leydig cell ultrastructure in long-term bilaterally cryptorchid rams

This study was conducted to investigate the effects of bilateral cryptorchidism induced in adult rams on testicular function and Leydig cell ultrastructure. The results indicated that long-term bilateral cryptorchidism resulted in decreased testicular size, degeneration of seminiferous tubules, elevated serum LH levels, maintenance of normal testosterone concentrations in peripheral and spermatic vein serum, impairment of the magnitude and duration of androgen response to exogenous luteinizing hormone (LH), a 13-fold reduction in total number of Leydig cells/paired testes, and a 3-fold hypertrophy in the average size of remaining Leydig cells. Based on quantitative morphometry, the hypertrophied Leydig cells exhibited significant increases in the volume of intracellular organelles, including the cell nucleus, mitochondria, smooth and rough endoplasmic reticulum, lysosome-like bodies and lipid vesicles. Quantitatively, the hypertrophy alone was not enough to offset the loss in number of Leydig cells and was insufficient to explain the maintenance of normal levels of testosterone in jugular and spermatic venous blood. The additional mechanisms responsible for production of normal serum testosterone levels in the cryptorchid ram remain to be elucidated.     

Clinical, morphological and endocrinological aspects of cryptorchidism in the horse

The authors analysed clinical, histological and hormonal data obtained from 205 cryptorchid horses. The majority of the unilaterally and bilaterally retained testes were located in the inguinal canal; however, the ratio of inguinal vs abdominal retention appeared to decrease with advancing age. In unilateral cryptorchidism, a pronounced preference was noted for left abdominal retention, whereas for inguinal cryptorchids, the retained testes occurred equally on both sides. Right inguinal retention was found to decrease with advancing age. Histology of cryptorchid testes revealed apparently normal Leydig cells and arrested spermatogenesis. Plasma testosterone concentrations were similar in normal stallions and unilateral cryptorchids, even in those which had the scrotal testis removed. Plasma oestradiol-17beta levels were lower in unilateral cryptorchids than in stallions.     

Effect of LH-RH treatment on hypothalamo-pituitary-gonadal axis and Leydig cell ultrastructure in cryptorchid boys

62 cryptorchid boys aged 2-6 years were selected at random either for surgical or for hormonal LH-RH treatment. As all biopsies from boys operated show typical histological and ultrastructural signs of cryptorchidism, it can be concluded that only true cryptorchid patients are included in our study. LH-RH treatment was successful in 16 (55%) of 31 boys. Median 30-min response values to LH-RH test of LH were initially normal in all boys. Those treated successfully remain normal after treatment whereas unsuccessfully treated patients have significantly lower LH 30-min response values at the end of the 4 weeks of LH-RH treatment. FSH response was not statistically different before or after treatment. This might indicate that a normal LH response is necessary for testicular descent. In the 15 boys operated after 4 weeks of unsuccessful LH-RH treatment, testes biopsies show recruitment of the Leydig cell precursors and development of juvenile Leydig cells with an increase in their content of lipoid droplets and smooth endoplasmic reticulum. No adverse effect on the number of spermatogonia could be observed after 4 weeks of LH-RH treatment.