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.     

Expression patterns of male germ cell markers in cryptorchid pig testes

Male germ cell apoptosis has been described in heat-damaged testes by cryptorchidism. In the present study, wild type pig testes were compared with cryptorchid testes via histological and immunohistological analyses. Spermatozoa were not detected in two cryptorchid testes and the diameters of seminiferous tubules were significantly reduced in cryptorchid pig testes compared with wild type pig testes. Cells expressing marker genes for undifferentiated spermatogonia, such as protein gene product 9.5 was significantly decreased in cryptochid pig testes. In addition, the numbers of cells expressing DEAD-box polypeptide 4 (VASA), synaptonemal complex protein 3, protamine, and acrosin (a biomarker of spermatocyte, spermatid, and spermatozoa) were significantly reduced in cryptochid pig testes. However, the number of vimentin-expressing Sertoli cells was not changed or was significantly increased in cryptorchid pig testes. This result indicates that male germ cells are specifically damaged by heat in cryptorchid pig testes and not Sertoli cells. These findings will facilitate the further study of spermatogenesis and the specific mechanisms by which cryptorchidism causes male infertility.     

Changes in the arrangement of actin filaments in myoid cells and sertoli cells of rat testes during postnatal development and after experimental cryptorchidism

Background: Abundant actin filaments are present in myoid cells and Sertoli cells in the testis. In the adult rat, the filaments form a lattice arrangement within the myoid cell, and show a hexagonal pattern in the basal junctional regions of Sertoli cells. Methods: Isolated seminiferous tubules and frozen sections were prepared from juvenile to adult Wistar rat testes, stained with FITC-conjugated phalloidin, and observed by confocal microscopy. Unilateral cryptorchidism was induced in adult rats, and seven days later, their testes were also examined. Results: In the myoid cell, parallel actin filaments running circularly around the seminiferous tubules were observed at 15 and 20 days of age. Then, at 30 days, actin filaments arranged longitudinally along the tubular long axis appeared in addition to the circular bundles. A lattice arrangement of actin-filament bundles in myoid cells became obvious at 40 days, when elongated spermatids are found in the tubule. Actin filaments in the basal junctional regions of Sertoli cells did not acquire the hexagonal pattern seen in the adult testis until 30 days of age. In the cryptorchid testes, the arrangement of actin filaments in the both cells showed a remarkable change compared to the control testis; the filaments became thinner and disrupted. Conclusions: A lattice arrangement of the actin filaments in the myoid cell appear at around 30 days, before the completion of spermatogenesis. A hexagonal pattern of the filaments in the junctional regions of Sertoli cells has already developed at this age. Cryptorchidism affects the actin filaments of the both cells. © 1995 Wiley-Liss, Inc.     

Intermediate filaments in the Sertoli cells of the ageing human testis

The presence and distribution of intermediate filaments (vimentin, keratin, desmin) was studied in the Sertoli cells of elderly men by means of quantitative immunohistochemical methods. Sertoli cells from young men showed moderate immunogold labelling to vimentin throughout the entire cytoplasm between the cell organelles in tubules showing complete spermatogenesis. Immunogold particles were more numerous in the perinuclear cytoplasm and beneath the plasma membrane in all its faces. The testes from elderly men showed different tubule types; some showed complete spermatogenesis and a normal lamina propria, while others had spermatogenic arrest at different levels (spermatids, spermatocytes, spermatogonia). The immunohistochemical reaction to vimentin in the Sertoli cells of tubules with complete spermatogenesis (type a) was similar to that in the cells of young men. In the Sertoli cells of severely damaged tubules (type b) the immunohistochemical reaction was more intense and immunogold particles extended in similar proportions throughout the whole cytoplasm. When immunolabelling intensity was compared between the three groups of tubules, by counting the number of immunogold particles per square micrometre of cytoplasm, it was found to be significantly higher (P≤0.05) in type b tubules of elderly men than either in tubules of young men or in type a tubules of elderly men. Since the average cell surface of Sertoli cells was similar in all tubule types, these data suggest that an actual vimentin increase occurs in Sertoli cells of germ-cell-depleted tubules. Sertoli cell immunogold labelling to keratin was found neither in young men nor in type a tubules of ageing men, whereas a positive immunohistochemical reaction was observed in the Sertoli cells of type b tubules of elderly men. Immunogold particles were localized mainly in the perinuclear cytoplasm, and beneath the lateral and basal cell surfaces. The observation of vimentin increase and keratin re-expression in ageing Sertoli cells only in germ-cell-depleted tubules suggests that the changes in intermediate filaments are related to the local factors associated with completion of spermatogenesis, causing functional changes in Sertoli cells.     

Androgen aromatization in cryptorchid mouse testis

Estrogens play an important role in germ cell development. Therefore, we have studied expression patterns of aromatase that converts testosterone into estrogens in 2 recombinant inbred mouse strains that differ in efficiency of spermatogenesis. In order to show whether germ cells are a target for estrogens, estrogen receptors (ER)alpha and beta were localized as well. Adult male CBA and KE mice were made unilaterally cryptorchid to determine alterations in testicular steroidogenesis and spermatogenesis. Differences between control and cryptorchid testes have been studied with respect to (1) cellular sites of aromatase, the enzyme responsible for estrogen formation, (2) the presence of ERalpha and ERbeta in various types of testicular cells, and (3) steroidogenic activity in the testes. Additionally, unilaterally control testes of cryptorchid mice were compared with bilaterally descended testes. Histological or hormonal differences were not found between control testes of cryptorchid and untreated mice. In cryptorchid testes from both strains, degeneration of germ cells was observed as well as a decrease in size of the seminiferous tubules, whereas the amount of interstitial tissue increased, especially in testes of CBA mice. Using immunohistochemistry, aromatase was localized in Leydig cells and germ cells in both control and cryptorchid testes. Sertoli cells were immunopositive in control testes only. In cryptorchid testes of KE mice, aromatase was strongly expressed in spermatids, that were still present in a few tubules. Other cell types in tubules were negative for aromatase. In both control and cryptorchid testes of both mouse strains, ERalpha were present in Leydig cells only, whereas ERbeta were found in Leydig cells and in germ cells in early stages of maturation. In homogenates of testes of CBA control mice, testosterone levels were 3-fold higher than in those of control KE mice, whereas the difference in estradiol levels between both strains was small. Cryptorchidism resulted in decreased testosterone levels and increased estradiol levels. The results of the present study show functional alterations due to cryptorchidism in both mouse strains. Strong aromatase expression in germ cells in control and cryptorchid testes indicates an additional source of estrogens in the testis besides the interstitial tissue and the relevance of estrogen in spermatogenesis.     

Immunohistochemical analysis of connexin43 expression in infertile human testes

Connexin43 (Cx43) is abundantly expressed in mammalian testes and implicated in the regulation of cell-to-cell interaction between germ cells and Sertoli cells, which is essential to the normal process of spermatogenesis. In the present study, we investigated the relation between Cx43 expression and the degree of spermatogenesis in infertile human testes. Immunohistochemical analysis of Cx43 was performed on testicular biopsies from 29 patients with azoospermia (n=23) and severe oligospermia (n=6), who gave informed consent to this experiment. The degree of testicular spermatogenesis was evaluated by Johnsen score. In the interstitium, immunostaining for Cx43 was localized to some focal parts of plasma membrane between neighboring Leydig cells. In seminiferous tubules with normal spermatogenesis, Cx43 expression was found between Sertoli cells and germ cells. However, Cx43 expression in maturation arrest was decreased and located mainly in the basal compartment of seminiferous tubules. Finally, there was a significant positive correlation between histological score of spermatogenesis and intensity of Cx43 (p=0.0294). These data suggest that the alteration of Cx43 expression may be involved in spermatogenic impairment, and that the communication between Sertoli cells and germ cells through Cx43 may be important for maturation of spermatogenesis.     

Fas expression correlates with human germ cell degeneration in meiotic and post-meiotic arrest of spermatogenesis

Degeneration of human male germ cells was analysed by means of light (LM) and transmission electron (TEM) microscopy. The frequency of degenerating cells was correlated with that of Fas-expressing germ cells in human testes with normal spermatogenesis (n = 10), complete early maturation arrest (EMA) (n = 10) or incomplete late maturation arrest (LMA; n = 10) of spermatogenesis. LM analysis of testis sections with normal spermatogenesis indicated that degenerating germ cells were localized in the adluminal compartment of the seminiferous epithelium. TEM showed that apoptotic cells were mostly primary spermatocytes and, to a lesser extent, round or early elongating spermatids. Apoptotic germ cells appeared to be eliminated either in the seminiferous lumen or by Sertoli cell phagocytosis. An increased number of degenerating cells was observed in testes with LMA as compared with normal testes and testes with EMA of spermatogenesis (P < 0.001, Wilcoxon's rank sum test). Comparison of these results with those obtained from immunohistochemistry experiments demonstrated a tight correlation between the number of apoptotic cells and the number of Fas-expressing germ cells (P = 0.001, Spearman's rank = 0.69). These findings suggest that altered meiotic and post-meiotic germ cell maturation might be associated with an up-regulation of Fas gene expression capable of triggering apoptotic elimination of defective germ cells.     

Crystalloids of actin-like filaments in the Sertoli cell of the swine testis.

Normal swine testes, congenital cryptorchid swine testes, and normal human tests were exposed to HMM (heavy meromyosin) after either glycerination or saponin treatment in order to determine whether the fine filaments composing the crystalloids in the Sertoli cells of the cryptorchid swine testes bind HMM to form arrowhead complexes. Short bundles of microfilaments observed in the basal part of the Sertoli cells in both normal and cryptorchid testes also bind HMM. Similar bundles of HMM-bound filaments are observed in the vicinity of spermatocytes. The periodicity of the arrowhead complexes is about 35 nm, and all arrowheads on a given filament point in the same direction. In addition, the polarity of the HMM-bound filaments in a given crystalloid or bundle is uni-directional. A mechanism for the formation of the swine crystalloids has been strongly support this hypothesis. Fine filaments of Charcot-Boettcher's crystalloid in human Sertoli cells did not bind HMM. Therefore the fine filaments of the human crystalloid are not actin-like in nature.     

Atypical development of Sertoli cells and impairment of spermatogenesis in the hypogonadal (hpg) mouse

Testes of hypogonadal (hpg) mice show arrested postnatal development due to congenital deficiencies of gonadotrophin-releasing hormone (GnRH) and gonadotrophin synthesis and secretion. Follicle-stimulating hormone (FSH), androgen or oestrogen treatment restore qualitatively normal spermatogenesis in hpg testes. Understanding the cellular and molecular changes accompanying hormone-induced spermatogenesis in hpg mice requires detailed morphological analyses of the germ cells and Sertoli cells in the untreated hpg testis. We compared seminiferous epithelial cytology in adult hpg, immature and adult wild-type mice using unbiased optical disector-based stereology, immunolocalization of Sertoli cell microtubules (MT), espin (a component of the blood-testis barrier), markers of Sertoli cell maturity (p27(kip1) and WT-1), and electron microscopy. Hpg testes had marked reductions in weight, seminiferous cord volume and length, and severe spermatogenic impairment with germ cells per testis < 1% of adult wild-type testes. Sertoli cell nuclei expressed WT-1 in hpg testes, but often were centrally located, similar to 9-14-day-old wild-type testes, and they expressed p27(kip1), indicating that hpg Sertoli cells were post-mitotic. Hpg testes had significantly (P < 0.05) reduced Sertoli cells per testis (0.56 million) compared with 10-day wild-type (1.15 million) and adult wild-type testes (2.06 million). Immunofluorescence labelling of normal adult Sertoli cells showed supranuclear MT columns and basally located espin, but these features were absent in 10-day-old and hpg Sertoli cells. Hpg Sertoli cells showed pleomorphic nuclear ultrastructure with mature-type nucleoli, similar to normal adult-type Sertoli cells, but hpg Sertoli cells exhibited incomplete tight junctions that lacked ectoplasmic specializations. We conclude that in hpg mice, chronic gonadotrophin insufficiency restrains Sertoli cell proliferation and maturation, forming pseudo-adult-type Sertoli cells that are incapable of supporting germ cell proliferation and maturation.     

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.     

Correlation between testicular biopsies (prepubertal and postpubertal) and spermiogram in cryptorchid men

Twenty-one young men who underwent testicular biopsy and orchidopexy in infancy consulted owing to infertility and had biopsies again. The first and second biopsy specimens from these patients were compared by means of a semiquantitative study of the seminiferous tubules to evaluate the evolution of germ cells and to correlate these data with spermatozoon numbers. The infant testes showing lesions were classified into 3 types according to the mean tubular diameter and tubular fertility index: (1) slight lesions, (2) marked germinal hypoplasia, and (3) severe germinal hypoplasia. In the adult testes, spermatogenesis was evaluated by calculating the average numbers of spermatogonia, primary spermatocytes, young spermatids, and mature spermatids. These testes were classified as (1) normal; (2) having lesions in the adluminal compartment; (3) having lesions in the basal compartment; and (4) mixed atrophy. The number of differentiated spermatids was correlated with the expected number of spermatozoa in the ejaculate by a power regression curve. The observation of certain histologic lesions in the seminiferous tubules was assumed to indicate excretory duct obstruction: ectasia, indented outline of the seminiferous epithelium, intratesticular spermatocele, apical cytoplasmic vacuolation of Sertoli cells, and mosaic distribution of testicular lesions. There was a correlation between the prepubertal lesions and the degree of spermatogenesis in postpubertal biopsy specimens. The evolution of the 40 testes without regard to their location in infancy (cryptorchid or scrotal) was as follows. The 14 infant testes with a normal histologic pattern (5 testes) or minor lesions (9 testes) evolved to testes with lesions of the adluminal compartment (8 testes), mixed atrophy (4 testes), or lesions of the basal and adluminal compartments (2 testes). The 6 testes with marked germinal hypoplasia evolved to testes with mixed atrophy. The 20 testes with severe germinal hypoplasia evolved to testes with mixed atrophy (17 testes), Sertoli-cell-only tubules (2 testes), or lesions in the basal compartment (1 testis). In the 9 patients with a histologic pattern of obstruction bilaterally (6 men) or unilaterally (3 men), the expected number of spermatozoa according to the correlation curve was much higher than the actual number in the spermiogram. This means that the testes of many azoospermic men produce spermatozoa, and this finding corroborates the importance of testicular biopsy in infertility studies.     

Morphological and histochemical characteristics of the lamina propria in scrotal and abdominal testes from postpubertal boars: correlation with the appearance of the seminiferous epithelium

This study was undertaken to investigate the morphological characteristics and lectin affinity of the testicular lamina propria in healthy boars and in unilateral and bilateral abdominal cryptorchid boars. The lamina propria of scrotal testes from healthy boars and unilateral cryptorchid boars was constituted by an innermost noncellular layer, the basal lamina, and by 2 layers of peritubular cells, each separated by a fibrous layer. The noncellular layers contained collagen fibres and glycoconjugates with abundant N‐acetylgalactosamine, galactose, fucose, N‐acetylglucosamine and neuraminic acid residues. The inner peritubular cell layer was composed of myoid cells, the outer layer of fibroblasts. In the abdominal testes of unilateral and bilateral cryptorchid boars, the lamina propria of nondegenerating and degenerating seminiferous tubules appeared thickened due to an increased content of collagen fibres and glycoconjugates. Glycoconjugates showed decreased amounts of fucose, neuraminic acid and galactose, and increased amounts of N‐acetylglucosamine residues. The basal lamina formed infoldings toward the seminiferous epithelium and contained small cells. Both inner and outer peritubular cells were fibroblasts of immature appearance. In degenerated seminiferous tubules of bilateral cryptorchid boars, the lamina propria was composed of a thickened and collagenised basal lamina, without peritubular cells and with a low content of glycoconjugates. In scrotal testes, therefore, the lamina propria was implicated in tubular contractility and in mediating the communication and the substrate diffusion between seminiferous tubules and interstitial tissue. Cryptorchidism induced morphological and histochemical alterations in the lamina propria of abdominal testes, which may be linked to evidence from other studies of lack of tubular contractility and defective cell–cell communication and substrate diffusion. The severity of these anomalies correlated with the severity of Sertoli cell alterations.     

Crystalloids of actin-like filaments in the sertoli cell of the swine testis

Normal swine testes, congenital cryptorchid swine testes, and normal human testes were exposed to HMM (heavy meromyosin) after either glycerination or saponin treatment in order to determine whether the fine filaments composing the crystalloids in the Sertoli cells are actin-like. The microfilaments of the crystalloids in the Sertoli cells of the cryptorchid swine testes bind HMM to form arrowhead complexes. Short bundles of microfilaments observed in the basal part of the Sertoli cells in both normal and cryptorchid testes also bind HMM. Similar bundles of HMM-bound filaments are observed in the vicinity of spermatocytes. The periodicity of the arrowhead complexes is about 35 nm, and all arrowheads on a given filament point in the same direction. In addition, the polarity of the HMM-bound filaments in a given crystalloid or bundle is uni-directional. A mechanism for the formation of the swine crystalloids has been suggested by Toyama (′75), and the results of this study strongly support this hypothesis. Fine filaments of Charcot-Boettcher's crystalloid in human Sertoli cells did not bind HMM. Therefore the fine filaments of the human crystalloid are not actin-like in nature.     

Immunohistochemical analysis of the vimentin filaments in Sertoli cells is a powerful tool for the prediction of spermatogenic dysfunction

The close interaction between male germ cells and Sertoli cells, a type of somatic cell found in the seminiferous tubules of mammalian testis, is essential for the normal progression of spermatogenesis in mammals. Vimentin is an intermediate filament protein that primarily provides mechanical support, preserves cell shape, and maintains the nuclear position, and it is often used as a marker to identify Sertoli cells. Vimentin is known to be involved in many diseases and aging processes; however, how vimentin is related to spermatogenic dysfunction and the associated functional changes is still unclear. In a previous study, we reported that vitamin E deficiency affected the testes, epididymis, and spermatozoa of mice, accelerating the progression of senescence. In this study, we focused on the Sertoli cell marker vimentin and explored the relationship between the cytoskeletal system of Sertoli cells and spermatogenic dysfunction using testis tissue sections that caused male reproductive dysfunction with vitamin E deficiency. The immunohistochemical analysis showed that the proportion of the vimentin-positive area in seminiferous tubule cross-sections was significantly increased in testis tissue sections of the vitamin E-deficient group compared with the proportion in the control group. The histological analysis of testis tissue sections from the vitamin E-deficient group showed that vimentin-positive Sertoli cells were greatly extended from the basement membrane, along with an increased abundance of vimentin. These findings suggest that vimentin may be a potential indicator for detecting spermatogenic dysfunction.     

Ectoplasmic specialization, a testis-specific cell-cell actin-based adherens junction type: is this a potential target for male contraceptive development?

The seminiferous tubule of the mammalian testis is largely composed of Sertoli and germ cells, which coordinate with Leydig cells in the interstitium and perform two major physiological functions, namely spermatogenesis and steroidogenesis respectively. Each tubule is morphologically divided into (i) the seminiferous epithelium composing Sertoli and germ cells, and (ii) the basement membrane (a modified form of extracellular matrix); underneath this lies the collagen fibril network, the myoid cell layer, and the lymphatic vessel, which collectively constitute the tunica propia. In the seminiferous epithelium, of rodent testes each type A1 spermatogonium (diploid, 2n) differentiates into 256 elongated spermatids (haploid, 1n) during spermatogenesis. Additionally, developing germ cells must migrate progressively from the basal to the luminal edge of the adluminal compartment so that fully developed spermatids can be released into the lumen at spermiation. Without this timely event of cell movement, spermatogenesis cannot reach completion and infertility will result. Yet developing round elongating/elongated spermatids must remain attached to the epithelium via a specialized Sertoli-germ cell actin-based adherens junction (AJ) type known as ectoplasmic specialization (ES), which is crucial not only for cell attachment but also for spermatid movement and orientation in the epithelium. However, the biochemical composition and molecular architecture of the protein complexes that constitute the ES have only recently been studied. Furthermore, the signalling pathways that regulate ES dynamics are virtually unknown. This review highlights recent advances in these two areas of research. It is expected that, if adequately expanded, these studies should yield new insights into the development of novel contraceptives targeted to perturb ES function in the testis. The potential to specifically target the ES may also mean that contraceptive action could be achieved without perturbing the hypothalamic-pituitary-testicular axis.     

Granular transformation of Sertoli cells in testicular disorders.

In order to study the granular transformation of Sertoli cells the following testicular specimens were reviewed: 58 postmortem biopsies from 21 children and 37 young adult males with normal histologic pattern; 165 biopsies from prepubertal cryptorchid testes; 38 biopsies and 18 surgical specimens from postpubertal-cryptorchid testes; bilateral biopsies from eight men with Del Castillo's syndrome, 14 men with retractile testes, and five men with obstructive azospermia; 17 bilateral and seven unilateral biopsies from 24 men with varicocele; seven unilateral biopsies plus five surgical specimens from 12 men with male pseudohermaphroditism; one biopsy and one surgical specimen from two men with macroorchidism; and the autopsy specimens from 28 adult men with acquired immunodeficiency syndrome (AIDS). Sertoli cells with eosinophilic granular cytoplasm were found in the testes of one prepubertal and four postpubertal cryptorchid males, two males with Del Castillo's syndrome, two males with retractile testes, four males with varicocele, two male pseudohermaphrodites, two males with macroorchidism, and one male with AIDS and interstitial orchitis. Histochemical and ultrastructural examination of granular Sertoli cells revealed that these cells accumulate secondary lysosomes and show scant cytoplasmic organelles. In the males with varicocele or retractile testes, these lysosomes were probably heterolysosomes that had degraded the germ cells and testicular fluid accumulated in the lumen of the ectatic seminiferous tubules of these testes. A similar mechanism is also probable in the male with interstitial orchitis that had caused germ cell destruction. In the other cases, in which the tubules showed reduced lumen and severe germ cell depletion, the abundant lysosomes are probably cytolysosomes. The development of these cytolysosomes might be related to the Sertoli cell dysgenesis present in these testes.     

Cytoskeleton in sertoli cells of the mosquito fish (Gambusia affinis holbrooki)

Background: There is little information about the distrib ution of cytoskeletal components in the testes of teleost fish. The aim of this (tublin, actin, vimentin, desmin, and cytokeratins) in the sertoli cells of Gambusia affinis holbrooki and in their efferent duct epithelial cells which are possibly orginated from the Sertoli cells Methods: Light and electron microscopic immunocytochemical studies and Western blotting analysis were performed in G. affinis testis. Results: Actin immuncor eaction was observed in the Sertoli cells at all spermatogenic stages, although the intensity of the reaction varied from one state to another. Sertoli cells that support supermatogonia or supermatocytes showd a weak immunoreaction which was uniformly distributed throughout the cytoplasm and somewhat more concentrated at the level of the inter-Sertoli specialized junctions, Immunoreaction to actin increased during the first states of supermiogenesis and was manly localized beneath the plasma membrane. This immunoreacction was more intense in the basal than in the aical ctoplasm of Sertoli cells. In a more advanced stage of supermiogenesis, actin immunoreaction become stroger in the apical cytoplasm where Sertoli cells displayed cytoplasmic projections around each supermatid. After sperm release, the apical Sertoli cell cytoplasm still showed an intense actin immunoreaction. Intense immuncreation to actin was also observed in the epithelial cells lining the efferent ducts. Immunoreaction to tubulin was diffuse throughout the Sertoli cell cytoplasm. No immunocreation to vimentin or desmn was observed in the Sertoli cells during the spermatogenic process. Immunoreation to both vimentin and desmin was observed in the efferent ducts cells. Desmin immunoreaction was also observed in the seminiferous tubule boundary cells, mainl in the sections showing germ cell cysts at the last stages of spermiogenesis and in the peritubular cells that surrounded the efferent duct epitheium. Immunoreaction to cytokeratins was found in the endothelium of testicular blood vessels but not in the Sertoli cells or in the efferent duct epithelium. Conclusions: Immunoreaction pattern to cytoskeletal proteins in the Sertoli cells of G. afinis: differs from that reported in mammalian Sertoli cells. These differences include the distribution of action filaments and the absence of dectectable vimentin immunoreaction in G. affinis: Sertoli cells. © 1995 Wiley-Liss, Inc.     

The distribution pattern of cytokeratin and vimentin immunoreactivity in testicular biopsies of infertile men

Testicular biopsies of infertile patients are often characterized by a mixed atrophy, in which different types of spermatogenic lesions are found in adjacent tubules. In order to evaluate a possible involvement of the state of differentiation of the Sertoli cells, the distribution pattern of cytokeratin and vimentin intermediate filaments within the seminiferous epithelium of 228 biopsy specimens with normal spermatogenesis (n=10), mixed atrophy (n= 206) or Sertoli Cell Only Syndrome (n=12) were investigated by means of immunohistochemical techniques. Sertoli cells were regularly found to show vimentin expression in tubules with normal spermatogenesis as well as in tubules with any kind of spermatogenic impairment including SCO. Cytokeratin expression as a marker showing lack of differentiation was common in Sertoli cells of tubules with arrest of spermatogenesis at the level of spermatogonia, and was occasionally associated with arrest at the level of primary spermatocytes or with SCO. Ultrastructural examination of tubules with spermatogonial arrest revealed Sertoli cells with features of typical fetal or prepubertal Sertoli cells, such as round to ovoid nuclei without indentations, stacks of rough ER and spot desmosomes. These data suggest that spermatogenic arrest at the level of spermatogonia might be due to functional impairment of the associated Sertoli cells, which have maintained or regained an undifferentiated state and are not able to initiate or trigger the process of spermatogonial differentiation.     

Changes in the distribution of microtubules in rat Sertoli cells during spermatogenesis

I have studied the distribution of microtubules in rat Sertoli cells during spermatogenesis. The distribution of microtubules was determined by indirect immunofluorescence in fragments of seminiferous epithelium which were mechanically dissociated from perfusion-fixed testes. The presence of microtubules in regions indicated by immunofluorescence was confirmed with electron microscopy. Microtubules are not evident in Sertoli cell regions surrounding early spermatogenic cells, but are abundant in cytoplasm adjacent to apical crypts containing elongate spermatids. In regions surrounding crypts, microtubules are uniformly aligned parallel to the long axis of Sertoli cells. Microtubules in these more central, or columnar, regions of the cells extend from supranuclear levels to the apex of the epithelium. In apical processes surrounding late spermatids, microtubules conform to the contours of the hook-shaped spermatid heads. These microtubules appear continuous with those in the columnar regions of the Sertoli cells. Changes in microtubule distribution observed in the rat are generally similar to, but less elaborate than, those reported previously in the squirrel. These changes may be characteristic of mammalian spermatogenesis in general.