The Sertoli cell junctional specializations and their relationship to the germinal epithelium as observed after efferent ductule ligation.

Seminiferous tubules from testes of normal and efferent ductule ligated mice were examined with the electron microscope. The tubules in the ligated animals were markedly distended and at most stages of the seminiferous cycle the epithelium exhibited a series of circumferentially-oriented ridges. Cross-sectional profiles of these ridges were studied with particular emphasis on the Sertoli cell junctional specializations and their relationship to the germinal cells. In the ligated specimen the basal cytoplasm of the Sertoli cells is highly attenuated, often appearing as a thin process resting on the basement lamina. Where the cytoplasm of one Sertoli cell ends, it meets in apposition with the cytoplasm of an adjoining Sertoli cell, and at these sites, junctional specializations are present. The ridges are comprised of a stalk of apical Sertoli cell cytoplasm, often appearing like an inverted cone, with young spermatids aligned along the lateral surfaces and the more mature spermatid population embedded within the apical cytoplasm. Junctional specializations were observed along these lateral Sertoli cell surfaces. In some instances, they formed a free surface, but usually early spermatids were in contact with the junctional specializations. With respect to the more mature spermatids, the acrosomal component was typically found in relation to a junctional specialization. Germ cells at the spermatocyte stage were also noted in relation to the Sertoli cell junctional specializations. The findings suggest that spermatocytes cross the Sertoli cell barrier and gain access to the adluminal compartment of the seminiferous tubule through the disengagement of the inter-Sertoli cell junctional complex. It is proposed that when the inter-Sertoli cell junctional specializations separate, the spermatocytes come in apposition with the newly freed junctional surfaces and remain in relation with them through the ensuing divisions. It appears that at some point, firm adhesion between germ cells and the junctional specializations occurs; the spermatid progeny may thus maintain contact with the original inter-Sertoli cell junctional specializations until their release into the tubule lumen.     

Spermatogenesis in the vasectomized monkey: Quantitative analysis

The seminiferous epithelium in mature vasectomized Macaca fascicularis was examined quantitatively to assess spermatogenesis. Monkeys were bilaterally vasectomized and controls were bilaterally sham operated. At postoperative periods of 10 and 18 months, groups of monkeys were castrated and their testes prepared for morphologic analysis. Diameters were measured in 100 cross sections of seminiferous tubules from each animal. Numbers of spermatogonia (Ad and Ap), preleptotene spermatocytes, pachytene spermatocytes, and step 7 spermatids, relative to Sertoli cell nucleoli, were counted in stage VII tubules. Tubule diameter and germ cell numbers per Sertoli cell nucleoli were not altered by vasectomy. Our study demonstrates quantitatively that spermatogenesis in the monkey is not inhibited up to 18 months following vasectomy.     

Macro-orchidism: light and electron microscopic study of four cases.

A hormonal and quantitative light microscopy study of one man with macro-orchidism associated with mental retardation and fragile X chromosome (case no. 1) and three men with idiopathic macro-orchidism (cases no. 2 to 4) is reported. Hormonal study revealed slightly increased follicle-stimulating hormone serum levels in cases no. 1 to 3. The testes from cases no. 1 (orchidoepididymoectomy specimen) and 2 (testicular biopsy) presented interstitial edema and three different tubular patterns that were arranged in a mosaic-like manner. Type I tubules had an increased diameter (less than 220 microns), dilated lumen, and thin seminiferous epithelium usually consisting of Sertoli cells, spermatogonia, primary spermatocytes, and sometimes a few spermatids. Type II tubules had a normal diameter (180 to 220 microns) and germ cell development varied between complete spermatogenesis and Sertoli-cell-only tubules. Type III tubules had decreased diameter (less than 180 microns), atrophic seminiferous epithelium, and thickened tunica propria. The appearance of the nuclei of the Sertoli cells in the three types of tubules could be either mature or immature. Some of the mature Sertoli cells presented a granular cytoplasm. A few of these granular cells grouped together, forming nests that protruded into the tubular lumen. The testicular biopsies from cases no. 3 and 4 only presented type II tubules that contained both mature and immature Sertoli cells. Quantitative study revealed that the large testicular size was principally due to an increased tubular length in all four cases. Although the seminiferous tubule lesions and interstitial edema suggest an obstructive process, the testicular excretory ducts (studied in case no. 1) appeared normal or only slightly dilated. It is possible that the seminiferous tubule lesions (dilated lumen and germ cell depletion) might be secondary to the Sertoli cell lesions (granular cytoplasm and nuclear immature-like pattern.     

Focal orchitis in undescended testes: discussion of pathogenetic mechanisms of tubular atrophy.

OBJECTIVE: To evaluate seminiferous epithelium lesions in adult cryptorchid testes showing lymphoid infiltrates in seminiferous tubules and interstitium (i.e., focal orchitis). Also, to consider the possible role of this lesion in the etiology of tubular atrophy. METHODS: We performed a histopathologic study of the cryptorchid testes and adjacent epididymides removed from 50 adult men who had not been previously treated for cryptorchidism. The study included morphologic and semiquantitative evaluation of seminiferous tubule pathology (according to germ cell numbers), Sertoli cell morphology, tubular lumen dilation, rete testis pattern (normal, hypoplastic, or cystic), and epididymal pattern (normal or epididymal duct hypoplasia). The study also included immunohistochemical evaluation of immune cell markers. The results were compared with clinical and laboratory findings. RESULTS: Focal lymphoid infiltrates (mainly lymphocytes) in seminiferous tubules and interstitium were found in 22 patients (44%), all of whom had unilateral cryptorchidism. The course of orchitis was asymptomatic, and laboratory data were normal. According to the seminiferous tubule pathology, a variety of histopathologic diagnoses, were made: (1) mixed atrophy consisting of Sertoli cell-only tubules intermingled with tubules showing maturation arrest of spermatogonia (11 testes, 4 of which also showed hyalinized tubules); (2) Sertoli cell-only tubules plus hyalinized tubules (4 testes); (3) Sertoli cell-only tubules (3 testes); (4) intratubular germ cell neoplasia (2 testes, 1 of which also showed hyalinized tubules); (5) complete tubular hyalinization (1 testis); and (6) tubular hyalinization plus some groups of tubules with hypospermatogenesis (all germ cell types were present although in lower numbers, 1 testis). Dysgenetic Sertoli cells, that is, Sertoli cells that had undergone anomalous, incomplete maturation, were observed in all nonhyalinized seminiferous tubules with inflammatory infiltrates. Tubular ectasia was observed in 13 cases. The rete testis was hypoplastic and showed cystic transformation in 18 testes, and the epididymis was hypoplastic in 15 testes. CONCLUSIONS: The causes of these focal inflammatory infiltrates are unknown. It is possible that tubular ectasia and Sertoli cell dysgenesis are involved and that these alterations cause a disruption of the blood-testis barrier and allow antigens to enter the testicular interstitium, giving rise to an autoimmune process.     

A scanning electron microscopic study of the Sertoli cell and spermiation in the syrian hamster

The scanning electron microscope was used to examine the Sertoli cells in normal and germ-cell-depleted testes. The Sertoli cells appear to attain characteristic configurations in the various stages of the cycle of the seminiferous epithelium. In tubules containing maturation-phase spermatids, Stages I-IV, the Sertoli cells exhibit column-like bases which give rise to lamellae measuring 25–30 μm in width which ensheath the spermatids and residual cytoplasmic bodies. In the Stages VIII-IX long flat sheets of Sertoli cytoplasm rest atop the step-8 and -9 spermatids. These sheets are oriented parallel to the basement membrane of the tubule with their long axis parallel to the long axis of the tubule. In Stages V-VI the head and proximal portion of the tail of the maturation-phase spermatids are ensheathed in sleeve-like Sertoli cell processes. Cuff-like terminations demarcate the terminus of these sleeves that surround maturing spermatids up to spermiation. In tubules undergoing spermiation, the sleeves retract so that only the tip of the spermatid head remains in the sleeve. Appendicular processes extend from the dorsum of the Sertoli cell sleeves in the tubules undergoing spermiation. After spermiation the appendix elongates while the sleeve evaginates until the everted sleeve is a finger-like process that extends into the tubular lumen. In tubules in which the seminiferous epithelium has been depleted with epinephrine injections the Sertoli cells attain two configurations. The first is characterized by having long attenuated lamellar processes that orient perpendicular to the basement membrane, with numerous ramifying processes arising from the lamellae. In the second configuration the Sertoli cells' lamellae orient parallel to the basement membrane of the tubule and lack the elaborate ramifications seen in the first configuration.     

The blood–testis barrier in the toad (Bufo arenarum hensel): A freeze-fracture and lanthanum tracer study

Intercellular junctions between Sertoli cells in the toad testis were studied by freeze-fracture and electron-opaque intercellular markers. These junctional specializations are characterized in thin sections by a series of focal fusions on the outer leaflets of both adjacent cell plasmalemmas, associated with bundles of fine filaments in the subjacent Sertoli cell cytoplasms. However, the wide subsurface cisterna of the endoplasmic reticulum, a component constantly associated with Sertoli cell junctions in mammals, is absent in the toad. The intravascularly injected lanthanum hydroxide, used as a tracer compound, gains access to the seminiferous tubules and surrounds spermatogonia and leptotene spermatocytes, but is persistently excluded from germ cells in later stages of development. This indicates that, as is the case in the mammalian testis, a permeability barrier to lanthanum is established which isolates all germ cells beyond leptotene spermatocytes. Freeze-fracture reveals the characteristic occluding junctions between Sertoli cells, but a variation in their geometric patterns was clearly observed in different regions of the toad seminiferous epithelium. The membrane-fractured faces of Sertoli cells embracing differentiating spermatids exhibit a deep junctional complex: up to 50 rows of particles between adjacent Sertoli cells separate these late germ cells from the periphery of the seminiferous tubules. Sertoli cells surrounding early germ cells generally exhibit, instead, a discontinuous, poorly developed network of interconnected rows of particles with few widely spaced strands. This seems to permit the percolation of the intercellular marker in areas of the seminiferous epithelium containing spermatogonia and leptotene spermatocytes.     

The blood-testis barrier in the toad (Bufo arenarum Hensel): a freeze-fracture and lanthanum tracer study

Intercellular junctions between Sertoli cells in the toad testis were studied by freeze-fracture and electron-opaque intercellular markers. These junctional specializations are characterized in thin sections by a series of focal fusions on the outer leaflets of both adjacent cell plasmalemmas, associated with bundles of fine filaments in the subjacent Sertoli cell cytoplasms. However, the wide subsurface cisterna of the endoplasmic reticulum, a component constantly associated with Sertoli cell junctions in mammals, is absent in the toad. The intravascularly injected lanthanum hydroxide, used as a tracer compound, gains access to the seminiferous tubules and surrounds spermatogonia and leptotene spermatocytes, but is persistently excluded from germ cells in later stages of development. This indicates that, as is the case in the mammalian testis, a permeability barrier to lanthanum is established which isolates all germ cells beyond leptotene spermatocytes. Freeze-fracture reveals the characteristic occluding junctions between Sertoli cells, but a variation in their geometric patterns was clearly observed in different regions of the toad seminiferous epithelium. The membrane-fractured faces of Sertoli cells embracing differentiating spermatids exhibit a deep junctional complex: up to 50 rows of particles between adjacent Sertoli cells separate these late germ cells from the periphery of the seminiferous tubules. Sertoli cells surrounding early germ cells generally exhibit, instead, a discontinuous, poorly developed network of interconnected rows of particles with few widely spaced strands. This seems to permit the percolation of the intercellular marker in areas of the seminiferous epithelium containing spermatogonia and leptotene spermatocytes.     

Degeneration of germ cells in normal,hypophysectomized and hormone treated hypophysectomized rats

In normal adult rats some germ cells degenerate at several vulnerable steps of spermatogenesis. These are the type A spermatogonia, midpachytene spermatocytes, primary and secondary spermatocytes which degenerate during their respective maturation divisions and step 7 and 19 spermatids. In the present study, these degenerating cells were examined under the electron microscope, and their frequency was determined in toluidine blue stained semithin sections of testes from normal, hypophysectomized (at 5.5 days after operation) and hypophysectomized rats injected with FSH and LH separately or in combination. With the exception of the step 19 spermatids, the degenerating germ cells underwent necrosis in vacuolated spaces delimited by Sertoli cells. In the case of the affected step 19 spermatids, an apical cytoplasmic process of the Sertoli cell initially ensheathed a long segment of their flagellum, and then each degenerating cell was drawn deep in the seminiferous epithelium where it was phagocytozed by the Sertoli cell. Soon after hypophysectomy the incidence of degenerating mid-pachytene spermatocytes, step 7 and 19 spermatids which are present in stages VII or VIII of the cycle of the seminiferous epithelium, increased significantly. In contrast the number of degenerating primary or secondary spermatocytes during the meiotic divisions seen in stage XIV of the cycle or of any other germinal cell was not significantly modified. While the injection of FSH alone had no influence on the number of degenerating cells in hypophysectomized rats, injections of LH at the two doses administered (0.7 μg or 20 μg) reduced significantly the number of degenerating cells seen in stages VII-VIII of the cycle; combined injections of FSH and LH (20 μg) reduced the number of these degenerating cells to the normal low values. Thus it appeared that the mid-pachytene spermatocytes and the step 7 and 19 spermatids, all present in the adluminal compartment of the seminiferous epithelium in stages VII or VIII of the cycle, were more sensitive to the presence of absence of gonadotropic hormones than the other germ cells present in the seminiferous epithelium.     

Effects of testosterone enanthate on the structure of the male reproductive tract of the rat

The histology and fine structure of the testis, epididymis and sex accessory glands were studied in young adult male rats administered testosterone enanthate, 120 μg/100 g body weight, three times weekly for 4, 8, or 12 weeks. The weights of the testis and epididymis decreased, and animals treated for 11 weeks were infertile. Alterations were found in the seminiferous tubules of all rats treated for 8 or 12 weeks, including the presence of many degenerating germ cells and a-large decrease or absence of late spermatids. Study of different stages of the cycle of the seminiferous epithelium showed that the greatest number of degenerating germ cells, step 7 spermatids and pachytene primary spermatocytes, occurred at stages VII-VIII of the cycle. Some normal appearing spermatogonia, primary spermatocytes and early spermatids remained in most seminiferous tubules. Sertoli cells contained many lipid droplets and lysosome-like bodies, and degenerating cells were surrounded by Ser-toli cell cytoplasm. The Leydig cells of treated animals were greatly reduced in size. Sperm progressively disappeared from the lumen of the middle segment and proximal part of the terminal segment of the epididymis after treatment for 8 or 12 weeks. Changes in the middle segment also included the appearance of intraepithelial cavities containing debris, and the presence within the epithelium of phagocytic cells that resembled leukocytes. The lumen of the proximal part of the terminal segment was often collapsed, while in the distal part of the terminal segment, the lumen was filled with cellular debris and degenerating sperm. Organelles of the principal cells of the epididymal epithelium appeared to be qualitatively unaltered. The weight of the sex accessory glands remained close to normal, and the presence of normal ultrastructural features suggested that production of secretions continued.     

The degenerative fate of germ cells not conforming to stage in the pubertal golden hamster testis

In the golden hamster ( Mesocricetus auratus ), pubertal establishment of spermatogenesis includes a defined period (d 26–30 of life) during which elongation of spermatids is selectively arrested. The resulting appearance of germ cell associations not conforming to stage and the phenomenon of desynchronisation-related germ cell degeneration are analysed both quantitatively and qualitatively by means of light and 'retrospective' electron microscopy. From d 26 onwards, the portion of tubules containing non-stage conforming germ cell associations gradually increases up to 37.5% of sectioned tubules on d 32. Concomitantly, the degree of desynchronisation rises to a maturational gap between spermatids and associated younger germ cells of 7 stages of the seminiferous epithelium cycle, i.e. of fully half a cycle. Beyond d 32, the frequency of desynchronised tubule segments decreases again. Some of the arrested round spermatids and, eventually, all belatedly elongating spermatids degenerate and are lost from the epithelium. Thus a regular maturation of advanced spermatids does not succeed under non-stage conforming conditions. Possibly it is not the desynchronisation between the associated germ cell generations and the spermatids by itself that impedes normal further development of the latter cells. Instead this may be due to the maturational delay of the stage-aberrant cells by several stages compared to the seminiferous epithelium as a whole and, especially, in relation to the stage-conditioned functional state of the neighbouring Sertoli cells.     

Proliferation and apoptosis of spermatogonia in postpuberal boar (Sus domesticus) testes with spontaneous unilateral and bilateral abdominal cryptorchidism

Cryptorchidism is a frequent male sexual disorder in mammals, which affects the histology of the tunica propria, interstitial tissue, blood vessels, seminiferous epithelium and testis functioning. In this paper, proliferation and apoptosis were examined in the seminiferous epithelium of both testes from unaffected boars and from boars suffering unilateral and bilateral cryptorchidism. In germ cells, proliferation was studied using the immunohistochemical PCNA technique, and apoptosis was analysed by in situ TUNEL labelling. An index was obtained for the proliferation and apoptosis observed in seminiferous tubules. In abdominal testes the epithelium contained few spermatogonia and Sertoli cells. In the testes of unaffected boars, numerous spermatogonia proliferated, whereas in cryptorchid testes such proliferation was lower and the proliferation/apoptosis ratio diminished. In the unaffected group, the TUNEL-positive germ cells were spermatogonia and spermatocytes in different phases of meiosis. In abdominal testes, the TUNEL-positive germ cells were spermatogonia alone. The apoptosis index of both abdominal and scrotal testes was similar. In conclusion, spontaneous cryptorchid testes showed a lower rate of spermatogonia proliferation in the seminiferous epithelium.     

In-vitro effects of FSH and testosterone withdrawal on caspase activation and DNA fragmentation in different cell types of human seminiferous epithelium

BACKGROUND: Caspases are downstream elements of apoptosis-mediating pathways initiated by the Fas ligand/Fas receptor system which is supposed to play a central role in the regulation of apoptosis in the human seminiferous epithelium. However, caspase activity in different cell types of this epithelium has never been addressed. METHODS AND RESULTS: We evaluated caspase activity and DNA integrity in Sertoli and germ cells within in-vitro cultured segments of human seminiferous tubules after induction of apoptosis by FSH or testosterone withdrawal. FSH withdrawal increased the incidence of DNA fragmentation in meiotic (primary spermatocytes) and post-meiotic (spermatids) germ cells without producing any detectable effect on caspase activity in these cells and without affecting DNA integrity or caspase activity in Sertoli cells. Testosterone withdrawal stimulated caspase activity and produced DNA fragmentation in Sertoli cells, but showed only a weak effect on DNA fragmentation in germ cells and did not alter germ cell caspase activity. CONCLUSIONS: These findings confirm the central role of caspases in apoptosis of Sertoli cells. However, they also suggest that acute apoptosis of germ cells in the adult human testis occurs in a caspase-independent way and is controlled by Sertoli cells via an as yet undetermined mechanism.     

Morphometric studies on rat seminiferous tubules

The goal of this morphometric study was to obtain quantitative information on the seminiferous tubules of Sprague-Dawley rats, including changes seen at various stages of the cycle of the seminiferous epithelium. Tissue from perfusion-fixed testes was embedded in Epon-Araldite; and sections were subjected to morphometric measurements at the light microscopic level, using point counting for volume densities and the Floderus equation for numerical densities. Changes occur in the diameter of the seminiferous tubule, as well as in the volume of the seminiferous epithelium and tubule lumen, from stage to stage during the cycle. A significant constriction of the seminiferous tubule accompanies spermiation. The volume of the seminiferous epithelium per unit length of the tubule begins to increase after stage XIV, and peaks at stage V of the next cycle. The tubule lumen increases dramatically from stages V to VII, at the expense of the epithelium. The number of Sertoli cells is constant per unit length of the seminiferous tubule at all stages of the cycle. This is also true for primary spermatocytes of various developmental phases and for round spermatids from step 1 through step 10 of spermiogenesis. The average number of younger (preleptotene, leptotene, zytgotene) primary spermatocytes per Sertoli cell is 2.34 ± 0.082 (SEM), the number of older (pachytene, diplotene) primary spermatocytes per Sertoli cell is 2.37 ± 0.064, and the ratio of step 1–10 spermatids to Sertoli cells is 7.89 ± 0.27. By studying tangential views of serially sectioned seminiferous tubules at stage V, it is shown that the number of step-17 spermatids associated with each Sertoli cell averages 8.35 ± 0.128, although the counts ranged from 6 to 11. The only appreciable occurrence of cell death after the last spermatogonial mitosis appears to be a 15% loss during the first meiotic division. From our morphometric results, corrected for volume changes during preparation for microscopy, there are 15.7 million (± 0.99 million) Sertoli cells per gram of fresh rat testis. The length of seminiferous tubule per gram of testis is estimated to be 12.4 ± 0.56 meters, and the tubule surface area per gram testis is 119.7 ± 2.57 cm². The daily production of mature spermatids is 9.61 million (± 0.615 million) per gram of testis.     

Testicular development in cynomolgus monkeys

The testes from 136 male cynomolgus monkeys were examined histopathologically in order to investigate the relationship between the development of spermatogenesis and testis weight, age, and body weight. At Grade 1 (immature), Sertoli cells and spermatogonia were the only cell classes in the testis. At Grade 2 (pre-puberty), no elongated spermatids were observed in the testis, although a few round spermatids and small lumen formation were observed. At Grade 3 (onset of puberty), all classes of germ cells were observed in the testis, although seminiferous tubule diameters and numbers of germ cells were small. Slight debris in the epididymis was observed in almost all animals. At Grade 4 (puberty), almost complete spermatogenesis was observed in the seminiferous tubules and it was possible to ascertain the spermatogenesis stage as described by Clermont, although tubule diameters and numbers of germ cells were small. There was less debris in the epididymis than at Grade 3. At Grade 5 (early adult), complete spermatogenesis was observed in the seminiferous tubules. At Grade 6 (adult), complete spermatogenesis in the seminiferous tubules and a moderate or large number of sperm in the epididymis were observed. Moreover, sperm analysis using ejaculated sperm was possible. Logistic regression analysis showed that testis weight is a good indicator of testicular maturity.     

Relative volume of sertoli cells in monkey seminiferous epithelium: A stereological analysis

Techniques of quantitative stereology have been utilized to determine the relative volume occupied by the Sertoli cells and germ cells in two particular stages (I and VII) of the cycle of the seminiferous epithelium. Sertoli cell volume ranged from 24% in stage I of the cycle to 32% in stage VII. Early germ cells occupied 3.4% in stage I (spermatogonia) and 8.7% in stage VII (spermatogonia and preleptotene spermatocytes). Pachytene spermatocytes occupied 15% (stage I) and 24% (stage VII) of the total volume of the seminiferous epithelium. In stage I the two generations of spermatids comprised 58% of the total epithelium by volume, whereas in stage VII, after spermiation, the acrosome phase spermatids occupied 35% of the total seminiferous epithelial volume.     

Testicular damage caused by inhalation of ethylene oxide in rats: light and electron microscopic studies.

Although testicular damage caused by ethylene oxide vapor (EtO) has been previously reported, the morphological changes occurring in seminiferous tubules remain unclear. We examined the time course of the testicular lesion induced by EtO in order to clarify its morphogenesis. Wistar rats were exposed to 500 ppm EtO for 6 hr per day, 3 times per week for 2, 4, 6, or 13 weeks through inhalation. In the 2-week exposure group, Sertoli cells often showed condensation and retraction of the cytoplasm, and dilatation of the endoplasmic reticulum (ER). In apical Sertoli cells, processes which encapsulated the heads of elongate spermatids, ectoplasmic specializations, and tubulobulbar complexes were often deformed and many elongate spermatids were degenerated. In the 4- and 6-week exposure groups, many degenerated Sertoli cells were present, and deformed germ cells, sometimes with multinucleation, appeared to make direct contact with each other without interlocation of Sertoli cell lateral processes. A few scattered immature Sertoli cells were evident in the 6-week exposure group. In the 13-week exposure group, seminiferous tubules containing almost all types of germ cells reappeared, mixed with atrophic tubules containing Sertoli cells only. In the former tubules, Sertoli cells often possessed regularly regenerated lateral processes, which were interposed between germ cells. These results indicate that the germ cell damage may be associated with damage to Sertoli cells. In spite of the intermittent exposure, focal regeneration of Sertoli cells appeared after 6 weeks of exposure to EtO and preceded patchy recovery of germ cells. Therefore, the data suggest that Sertoli cell regeneration may permit regeneration of germ cells.     

Germ cell transfer into rat, bovine, monkey and human testes.

Germ cell transplantation is a potentially valuable technique offering oncological patients gonadal protection by reinitiating spermatogenesis from stem cells which were reinfused into the seminiferous tubules. In order to achieve an intratubular germ cell transfer, intratubular microinjection, efferent duct injections and rete testis injections were applied on dissected testes of four different species: rat, bull, monkey and man. Ultrasound-guided intratesticular rete testis injection was the best and least invasive injection technique with maximal infusion efficiency for larger testes. Deep infiltration of seminiferous tubules was only achieved in immature or partially regressed testes. This technique was applied in vivo on two cynomolgus monkeys. In the first monkey a deep infusion of injected cells and dye into the lumen of the seminiferous tubules was achieved. In the second, transplanted germ cells were present in the seminiferous epithelium 4 weeks after the transfer. These cells were morphologically identified as B-spermatogonia and located at the base of the seminiferous epithelium. In summary, this paper describes a promising approach for germ cell infusion into large testes. The application of this technique is the first successful attempt of a germ cell transfer in a primate.     

Histologic changes in the mouse testis after treatment with gossypol tetra-acetic acid

The effect of oral administrations (20 or 40 mg/kg body weight/day, for 21 days) of gossypol tetraacetic acid on the testis of the Parkes strain mouse was investigated. Gossypol treatment did not affect the body weight or testicular weight, but caused a significant depression in the weight of the seminal vesicle. Histologically, the testes in mice treated with gossypol possessed regressed seminiferous tubules showing the exfoliation of germ cells, the occurrence of giant cells, a disorganization of the germinal epithelium, the degeneration of germinal elements, intraepithelial vacuolation and dislocation of the Sertoli cells into the luminal portion. However, the effect of gossypol was not uniform, and normal features were also observed in the majority of the tubules in the testes of the gossypol-treated mice. When quantitatively analysed, the frequency of regressed seminiferous tubules was significantly higher in the testes in the treated mice than the controls. The results suggest that the gossypol treatment induces non-uniform regressive changes in the seminiferous tubules in the mouse testis.     

Normal and pathological human testes express hormone-sensitive lipase and the lipid receptors CLA-1/SR-BI and CD36

Numerous studies have demonstrated the important role of cholesterol and cholesteryl esters in tumor cell proliferation and progression of cancer. However, few studies have focused on the role of lipid transporters and lipases in cancer development and progression. The present study examined the expression of hormone-sensitive lipase (HSL) and the scavenger receptors CLA-1/SR-BI and CD36 in normal human testis and in nontumor and tumor testicular disorders by immunohistochemistry and Western blotting analysis. In normal young testes, immunoreaction to CLA-1/SR-BI was found in the spermatid acrosomic vesicle and on the surface of Sertoli and Leydig cells. HSL was detected in spermatogonia, the Golgi region of spermatocytes, the nucleus of spermatids, and the cytoplasm of both Sertoli and Leydig cells. Elderly testes and testes with hypospermatogenesis showed a similar staining pattern to that of normal young testes except for CD36, which was expressed in Sertoli cells. Cryptorchid testes demonstrated intense labeling to HSL and weak labeling to SR-BI in Sertoli cells (nucleus and cytoplasm) and Leydig cells (cytoplasm). Seminiferous tubules with intratubular germ cell neoplasia exhibited intense immunolabeling to the 3 lipid receptors in the surface of neoplastic cells and to HSL in the nucleus. In seminoma and spermatocytic seminoma, neoplastic cells labeled to HSL but failed to stain with antilipid receptors; in the seminiferous tubules at the periphery of the tumour, Charcot-B?ttcher crystalloids of Sertoli cells were strongly positive to CLA-1. Testes with mature teratoma showed a weak reaction to CD36 and SR-BI in some cells of enteric-type glands, and immature teratoma were exclusively immunolabeled with HSL. Western blotting analysis revealed that multiple bands were immunolabeled, with differences seen between normal and pathological testes. The results of this study indicate that the presence of lipid receptors (CLA-1/SR-BI) and hormone-sensitive lipase in Leydig cells suggests a role of these proteins in steroidogenesis. Also, these proteins seem to be involved in spermiogenesis, as their labeling in spermatids suggests. In nonmalignant and malignant pathologies, cholesterol metabolism is probably altered, and HSL labeling in neoplastic germ cell nuclei suggests a still-unknown function of this enzyme, probably related to cell cycle regulation.