Ultrastructural study of the boar seminiferous epithelium: changes in cryptorchidism

The present study compares the ultrastructural features of Sertoli cells and germ cells between scrotal testes of healthy boars and abdominal testes of unilateral and bilateral cryptorchid boars. In healthy boars, spermatogonia are flat cells lying in close association with the basal lamina. As differentiation progresses, spermatogonia acquire an oval profile and lose their contact with the basal lamina. Spermatocytes are round cells moving from the basal compartment of the seminiferous epithelium to the luminal compartment. Spermatids exhibit complex morphological changes leading to the formation of spermatozoa. Sertoli cells extend from the basal lamina to the tubular lumen. The nucleus encloses fine euchromatin and one or two nucleoli; the nuclear envelope has a few deep infoldings. The lateral cell membranes form junctional specializations that constitute the blood-testis barrier. The cytoplasm encloses smooth endoplasmic reticulum, vesicles, aggregates, and scattered mitochondria. The seminiferous epithelium of abdominal testes from unilateral and bilateral cryptorchid boars contains few spermatogonia with an abnormal appearance; the alteration in germ cell number is more severe in the bilateral disease. In unilateral cryptorchid boars, spermatogonia appear as either large pyramidal cells or roundish cells; in bilateral cryptorchid boars, spermatogonia show roundish profiles and degenerative patterns. Abdominal testes of both unilateral and bilateral cryptorchid boars are constituted by immature Sertoli cells that show abnormal cytoplasmic content, defective development of the blood-testis barrier, and atypical nuclear appearance; in bilateral cryptorchid boars, immature Sertoli cells exhibit degenerative signs. At postpubertal age, unilateral and bilateral cryptorchidism induce total arrest of spermatogenesis at spermatogonial stage as a result of an abnormal differentiation of the Sertoli cells. Moreover, the degeneration of abdominal testes initiates earlier in bilateral cryptorchidism than in unilateral cryptorchidism.     

In vitro male germ cell cultures of zebrafish

Transgenic modification of sperm before fertilization has the advantages of a much shorter timeline for the production of transgenic animals. A culture system using primary cultures of zebrafish male germ cells, in which the differentiation of spermatogonia to functional sperm can occur in vitro, allows us to introduce foreign DNA into the cultured sperm and to produce transgenics from the sperm. This chapter describes methods for the co-culture of male germ cells and a Sertoli cell feeder layer and the introduction of foreign DNA with retroviruses. This male germ cell culture system should prove useful not only in producing genetically modified sperm, but also in analyzing the regulatory function of Sertoli cells for spermatogenesis in vertebrates.     

Association study and expression analysis of CD9 as candidate gene for boar sperm quality and fertility traits

Cluster-of-differentiation antigen 9 (CD9) gene expressed in the male germ line stem cells is crucial for sperm-egg fusion, and was therefore selected as candidate gene for boar semen quality. The association of CD9 with boar sperm quality and fertility trait was analyzed using a total of 340 boars both from purebred Pietrain and Pietrain×Hampshire crosses. A single nucleotide polymorphism (g.358A>T) in intron 6 was significantly associated with sperm motility (MOT) (P<0.001), plasma droplet rate (PDR) (P<0.001) and abnormal spermatozoa rate (ASR) (P<0.01). Boars were divided into two groups with group 1 (G-I) boars having a higher SCON and SMOT, lower SVOL (sperm volume) and group 2 (G-II) having a lower SCON and SMOT, higher SVOL. The mRNA and protein expression levels were evaluated in reproductive, non-reproductive tissues and spermatozoa from G-I and G-II animals by using quantitative real-time PCR and western blotting. When both reproductive and non-reproductive tissues were examined, highest mRNA was expressed in prostate gland, then in the body of the epididymis, vas deferens and tail of the epididymis. In case of reproductive tissues, CD9 expression was higher in tissues and spermatozoa collected from G-I boars than those collected from G-II boars. The mRNA expression was significantly different (P<0.05) in body of epididymis from G-I and G-II boars. The CD9 protein expression results from western blot were coincided with the results of qRT-PCR. Moreover, CD9 protein localization in Leydig cells, Sertoli cells, epithelial cells and spermatozoa was remarkable which indicated the important role of CD9 in spermatogenesis process. By using mRNA and protein expression profiles, it could be shown that CD9 plays a crucial role during sperm development, especially within the epididymis where the maturation of the sperm, a key process for the sperm quality and motility takes place. These results will improve the understanding of the functions of the CD9 in spermatogenesis within the reproductive tracts and will shed light on CD9 as a candidate gene in the selection of good sperm quality boars.     

Sertoli cell development and function in an animal model of testicular dysgenesis syndrome

Pregnancy exposure to di(n-butyl) phthalate (DBP) in rats induces a testicular dysgenesislike syndrome (TDS) in male offspring. Earlier studies suggested altered Sertoli cell development/maturation may result, especially in testes that become cryptorchid. This study quantitatively assessed Sertoli cell numerical and functional development in DBP-exposed rats and compared (unilaterally) cryptorchid and scrotal testes. Pregnant rats were gavaged with 500 mg/kg/day DBP or corn oil from embryonic (E) Days 13.5 to 21.5. Male offspring were sampled on E21.5 or Postnatal Day 6, 10, 15, 25, or 90. Sertoli cell number in DBP-exposed males was reduced by approximately 50% at E21.5 but recovered to normal by Days 25-90, accompanied by significant changes in plasma inhibin B and testosterone levels. Sertoli cell maturational development in DBP-exposed males, assessed using five protein markers (anti-müllerian hormone, cytokeratin, androgen receptor, CDKN1B, and Nestin), was largely normal, with some evidence of delayed maturation. However, in adulthood, Sertoli cells (SC) in areas lacking germ cells (Sertoli cell-only [SCO] tubules) often exhibited immature features, especially in cryptorchid testes. Sertoli cells in DBP-exposed animals supported fewer germ cells during puberty, but this normalized in scrotal testes by adulthood. Scrotal and especially cryptorchid testes from DBP-exposed animals exhibited abnormalities (SCO tubules, focal dysgenetic areas) at all postnatal ages. Cryptorchid testes from DBP-exposed animals exhibited more Sertoli cell abnormalities at Day 25 compared with scrotal testes, perhaps indicating more severe underlying Sertoli cell malfunction in these testes. Our findings support the concept of altered Sertoli cell development in TDS, especially in cryptorchid testes, but show that maturational defects in Sertoli cells in adulthood most commonly reflect secondary dedifferentiation in absence of germ cells.     

Efficiency of the process of meiosis in scrotal testes of healthy boars and unilateral abdominal cryptorchid boars.

Unilateral abdominal cryptorchidism has usually been correlated with abnormalities in the spermatogenic activity of the scrotal testis. The present study describes the effects of unilateral abdominal cryptorchidism on the meiotic process in scrotal testes from postpubertal boars. The percentage of primary spermatocytes, secondary spermatocytes, and round spermatids was evaluated in testicular smears from scrotal testes of healthy boars and of right-sided unilateral abdominal cryptorchid boars. As compared to the scrotal testes of healthy boars, the scrotal testes of unilateral abdominal cryptorchid boars showed low transformation from primary to secondary spermatocytes (meiosis I), but normal transformation from secondary spermatocytes to round spermatids (meiosis II). The data obtained indicate that spontaneous unilateral abdominal cryptorchidism on the right side induced partial arrest of spermatogenesis at the primary spermatocyte stage that was attributed to anomalies in Sertoli-cell activity. Abnormal paracrine signals from altered Sertoli cells could have resulted in either disturbed mitosis, which led to the formation of spermatocytes with an abnormal DNA content, or abnormalities in the metabolic activity and the organization of the cytoskeleton of primary spermatocytes.     

Cell population indexes of spermatogenic yield and testicular sperm reserves in adult jaguars (Panthera onca)

The intrinsic yield of spermatogenesis and supporting capacity of Sertoli cells are the desirable indicators of sperm production in a species. The objective of the present study was to quantify intrinsic yield and the Sertoli cell index in the spermatogenic process and estimate testicular sperm reserves by histological assessment of fragments obtained by testicular biopsy of five adult jaguars in captivity. The testicular fragments were fixed in 4% glutaric aldehyde, dehydrated at increasing alcohol concentrations, included into hydroxyethyl methacrylate, and were cut into 4 μm thickness. In the seminiferous epithelium of the jaguar, 9.2 primary spermatocytes in pre-leptotene were produced by “A” spermatogonia. During the meiotic divisions only 3.2 spermatids were produced by a primary spermatocyte. The general spermatogenic yield of the jaguar was about 23.4 cells and each Sertoli cell was able to maintain about 19.2 germ cells, 11 of them were round spermatids. In each seminiferous epithelium cycle about 166 million spermatozoa were produced by each gram of testicular tissue. In adult jaguars, the general spermatogenic yield was similar to the yield observed in pumas, greater than that observed for the domestic cat, but less compared to most domestic animals.     

The effects of postnatal FSH substitution on Sertoli cell number and the sperm production capacity of the adult boar

The role of FSH for Sertoli cell establishment and sperm production in the boar is not definitely known. In order to elucidate its function FSH was substituted postnatally in male pigs and the resulting effects on testicular histological traits and sperm production capacity were investigated when the boars had reached maturity. Six male piglets received pFSH from 18 to 48 days postnatally. Another six piglets instead received saline and served as controls. Blood samples were drawn to measure FSH, LH, testosterone and estradiol. After 28 weeks, the boars were trained to mount a dummy so that the spermatogenic capacity was tested by increasing the frequency of semen collection at the age of 30 weeks. Libido (latency time) and ejaculate criteria (volume, motility, morphological abnormalities) were determined. Thereafter the boars were killed and their testes analyzed for morphology, number of Sertoli cells, germ cells and Leydig cells as well as the ratio between mitosis and apoptosis in the tubules.FSH concentrations were twofold due to FSH application when compared to the controls. LH was low during the first 2 weeks of FSH treatment. Thereafter concentrations increased in three of the six treated animals but not in controls. Testosterone increased slightly over the application period both in the controls and the treated piglets. Estradiol levels were similar in both groups. Increased ejaculation frequency reduced sperm concentrations and sperm motility in all boars and the percentage of morphologically abnormal sperm increased. Ejaculate volumes and the time of latency were not significantly altered. No differences were observed between the controls and the FSH treated boars. The testicular parameters of both FSH- and control boars were identical for morphology, number of spermatogenic and somatic cells as well as mitosis–apoptosis equilibrium. The data demonstrated that a prolonged postnatal period of FSH concentrations does not influence the sperm production of the adult boar.     

Germ cell transplantation from large domestic animals into mouse testes

Donor-derived spermatogenesis after spermatogonial transplantation to recipient animals could serve as a novel approach to manipulate the male germ line in species where current methods of genetic modification are still inefficient. The objective of the present study was to investigate germ cell transplantation from boars, bulls, and stallions, which are economically important domestic animals, to mouse recipients. Donor testis cells (fresh, cryopreserved, or cultured for 1 month) were transplanted into testes of immunodeficient recipient mice in which endogenous spermatogenesis had been destroyed. Recipient testes were analyzed from 1 to > 12 months after transplantation for the presence of donor germ cells by donor-specific immunohistochemistry. Donor cells were present in most recipient testes with species-dependent differences in pattern and extent of colonization. Porcine donor germ cells formed chains and networks of round cells connected by intercellular bridges but later stages of donor-derived spermatogenesis were not observed. Transplanted bovine testis cells initially appeared similar but then developed predominantly into fibrous tissue within recipient seminiferous tubules. Few equine germ cells proliferated in mouse testes with no obvious difference between cells recovered from a scrotal or a cryptorchid donor testis. The pattern of colonization after transplantation of cultured cells did not resemble spermatogonial proliferation. These results indicate that fresh or cryopreserved germ cells from large animals can colonize the mouse testis but do not differentiate beyond the stage of spermatogonial expansion. Species-specific differences in the compatibility of large animal donors and mouse recipients were detected which cannot be predicted solely on the basis of phylogenetic distance between donor and recipient species.     

Spermatogenesis and sperm transit through the epididymis in mammals with emphasis on pigs

Starting from the period of testis differentiation, the Sertoli cell plays a pivotal role in the development of a functional testis. FSH is the major mitotic factor for Sertoli cells. Because the supporting capacity of Sertoli cells is relatively fixed for each species, their total number per testis, established just before puberty (approximately 4 months in pigs), dictates the potential for sperm production. In contrast to Sertoli cells that are still undifferentiated, mature Leydig cells are already present at birth in pigs. Spermatogenesis lasts from 30 to 75 days in mammals, and this time period is under the control of the germ cell genotype. In boars, each spermatogenic cycle and the entire spermatogenic process lasts 8.6-9.0 and approximately 40 days, respectively. The sperm transit through the epididymis takes approximately 10 days in pigs and this is within the range cited for most mammals. Germ cell loss occurs normally during spermatogenesis, mainly during the spermatogonial and meiotic phases. In pigs, significant germ cell loss also takes place during spermiogenesis. In mammals in general, including pigs, only 2-3 out of a possible 10 spermatozoa are produced from each differentiated type A1 spermatogonium. The high supporting capacity of Sertoli cells and the short duration of the spermatogenic cycle are the main factors responsible for the comparatively high spermatogenic efficiency of pigs.     

Functional importance of palmitoyl protein thioesterase 1 (PPT1) expression by Sertoli cells in mediating cholesterol metabolism and maintenance of sperm quality

Sertoli cells are a type of nurse cell in the seminiferous epithelium that are crucial for sustaining spermatogenesis by extending nutritional and energy support to the developing germ cells. Dysfunction of Sertoli cells could cause disordered spermatogenesis and reduced fertility in males. In this study, we focused on the expression and function of palmitoyl protein thioesterase 1 (PPT1), a lysosomal depalmitoylating enzyme, in Sertoli cells. Here, we show that PPT1 expression in Sertoli cells is responsive to cholesterol treatment and that specific knockout of Ppt1 in Sertoli cells causes male subfertility associated with poor sperm quality and a high ratio of sperm deformity. Specifically, Ppt1 deficiency leads to poor cell variably accompanied with abnormal lysosome accumulation and increased cholesterol levels in Sertoli cells. Further, Ppt1 deficiency results in poor adhesion of developing germ cells to Sertoli cells in the seminiferous epithelium, which is likely to be responsible for the reduced male fertility as a consequence of declines in sperm count and motility as well as a high incidence of sperm head deformity. In summary, PPT1 affects sperm quality and male fertility through regulating lysosomal function and cholesterol metabolism in Sertoli cells.     

Development of germ cell transplants: morphometric and ultrastructural studies.

Mouse-to-mouse transplants were studied at 10 min, 9 h, 24 h, 1 week, 1 month, 2 months, and 3 months post-transplantation. Data from a previous light microscope study were confirmed and extended using morphometric and ultrastructural techniques. As soon as 10 min after introduction of the germ cells from one mouse into the tubule lumen of a recipient mouse they developed relationships with small Sertoli cell processes. The extent of this surface-to-surface relationship increased in animals sacrificed up to 1 week post-transplantation. Most transplanted germ cells retained the characteristics of the donor germ cells after they had been isolated and pelleted. Nearly all transplanted cells eventually underwent phagocytosis by the recipient Sertoli cells. The presence of small apparent clones of germ cells after 1 week of transplantation indicated that some germ cells may divide and survive for short periods within the epithelium. No discernible qualitative subcellular changes in the host Sertoli cell accompanying the development of transplant spermatogenesis were noted. Macrophages were present in the region of the boundary tissue between myoid cells and appeared to increase in number in the peritubular tissue of transplanted testes. Images suggest that they migrated into the tubule to gain entrance to the lumen and there take on the form of activated macrophages. Some macrophages phagocytose sperm at 2 months and 3 months post-transplantation. A testis weight increase previously demonstrate to occur at 24 h post-introduction of germ cells was found to be due to an increase in the volume of the tubular lumen. The increase of lumen size at 24 h was not related to the volume of the injected material. It is suggested that the presence of injected cells, likely germ cells, in the tubule lumen stimulated increased secretion by the Sertoli cell.     

Role of epithelial cells and programmed cell death in Hydra spermatogenesis

Spermatogenesis in higher animals is a tightly regulated process, in which survival and death of sperm precursor cells depends on the presence of somatic cells in gonads. In the basal metazoan Hydra spermatogenesis takes place in anatomically simple testes and in the absence of accessory structures. Hydra sperm precursors are derived from interstitial stem cells. Here we show that large numbers of sperm precursors in testes of Hydra vulgaris undergo programmed cell death (apoptosis) and that ectodermal epithelial cells phagocytose the apoptotic sperm precursors. This is surprising since so far no evidence has been reported that epithelial cells are directly involved in germ cell differentiation in Hydra. We propose that, similar to Sertoli cells in mammals, in Hydra epithelial cells support and perhaps even control spermatogenesis.     

Long-term (6-wk) hindlimb suspension inhibits spermatogenesis in adult male rats.

The International Space Station will allow extended habitation in space and long-term exposure to microgravity (microG). A concern is the impact of long-term microG exposure on the ability of species to reproduce. The model often used to simulate microG is rat hindlimb suspension (HLS), where the hindlimbs are elevated above the cage floor with a tail harness. Experiments described here are the first to examine the effect of long-term HLS on testicular function in adult male rats. Free-roaming (controls), animals with only the tail harnessed but hindlimbs in contact with the cage floor (TO), and HLS animals were tested for 6 wk. Cryptorchidism was prevented in TO and HLS animals by partial constriction of the inguinal canal with sutures. All parameters were compared at the end of the 6-wk experiment. Testicular weights and spermatogenesis were significantly reduced by HLS, such that no spermatogenic cells beyond round spermatids were present and epididymides were devoid of mature sperm. In many tubules, loss of all germ cells, except a few spermatogonia, resulting in histopathology similar to the Sertoli cell, was observed. Spermatogenesis appeared unaffected in control and TO animals. Sertoli and Leydig cell appearance, testosterone, luteinizing hormone, and follicle-stimulating hormone levels, and epididymal and seminal vesicle weight were unchanged by HLS. Cortisone was not elevated by HLS; thus stress may not be a factor. These results demonstrate that spermatogenesis is severely inhibited by long-term HLS, whereas testicular androgen production is not. These results have significant implications regarding serious effects of long-term exposure to microG on the reproductive capability of scrotal mammals, including humans.     

Ultrastructure of germ cells, the Leydig cells, and Sertoli cells during spermatogenesis in Boleophthalmus pectinirostris (Teleostei, Perciformes, Gobiidae)

The ultrastructures of germ cells, Leydig cells, and Sertoli cells during spermatogenesis in male Boleophthalmus pectinirostris were investigated by electron microscopic observations. During the period of maturation divisions, well-developed Leydig cells have three major morphological characteristics: a vesicular nucleus, mitochondria with tubular cristae, and a number of smooth endoplasmic reticulum. Based on cytoplasmic features, it appears that Leydig cells are responsible for the synthesis of male sex steroids. Although no clear evidence of steroidogenesis was found in the Sertoli cells, they were found to perform a phagocytic function in the seminiferous lobules. Most Sertoli cells contain granules thought to represent deposited glycogen or lipid but there is no indication of a transfer of nutrients to the spermatids. During the period of germ cell degeneration, several characteristics of phagocytosis appear in the cytoplasm of the Sertoli cells. In particular, it is assumed that the Sertoli cells are involved in the degeneration and resorption of undischarged spermatids after spermiation. No acrosome of the sperm is formed. The structure of the spermatozoon in B. pectinirostris is very similar and closely resembles to those of suborder Gobioidei (perciform type teleosts). The flagellum or sperm tail shows the typical 9+2 array of microtubules.     

Necrosis Is the Dominant Cell Death Pathway in Uropathogenic Escherichia coli Elicited Epididymo-Orchitis and Is Responsible for Damage of Rat Testis

Male infertility is a frequent medical condition, compromising approximately one in twenty men, with infections of the reproductive tract constituting a major etiological factor. Bacterial epididymo-orchitis results in acute inflammation most often caused by ascending canalicular infections from the urethra via the continuous male excurrent ductal system. Uropathogenic Escherichia coli (UPEC) represent a relevant pathogen in urogenital tract infections. To explore how bacteria can cause damage and cell loss and thus impair fertility, an in vivo epididymo-orchitis model was employed in rats by injecting UPEC strain CFT073 into the vas deference in close proximity to the epididymis. Seven days post infection bacteria were found predominantly in the testicular interstitial space. UPEC infection resulted in severe impairment of spermatogenesis by germ cell loss, damage of testicular somatic cells, a decrease in sperm numbers and a significant increase in TUNEL (+) cells. Activation of caspase-8 (extrinsic apoptotic pathway), caspase-3/−6 (intrinsic apoptotic pathway), caspase-1 (pyroptosis pathway) and the presence of 180 bp DNA fragments, all of which serve as indicators of the classical apoptotic pathway, were not observed in infected testis. Notably, electron microscopical examination revealed degenerative features of Sertoli cells (SC) in UPEC infected testis. Furthermore, the passive release of high mobility group protein B1 (HMGB1), as an indication of necrosis, was observed in vivo in infected testis. Thus, necrosis appears to be the dominant cell death pathway in UPEC infected testis. Substantial necrotic changes seen in Sertoli cells will contribute to impaired spermatogenesis by loss of function in supporting the dependent germ cells.     

Inducible nitric oxide synthase and nitrotyrosine in the central nervous system of mice chronically infected with Trypanosoma brucei brucei

Human African trypanosomiasis, or sleeping sickness, evolves toward a meningoencephalitic stage, with a breakage in the blood-brain barrier, perivascular infiltrates, and astrocytosis. The involvement of nitric oxide (NO) has been evoked in the pathogenic development of the illness, since NO was found to be increased in the brain of animals infected with Trypanosoma brucei (T. b.) brucei. An excessive NO production can lead to alterations of neuronal signaling and to cell damage through the cytotoxicity of NO and its derivatives, especially peroxynitrites. In African trypanosomiasis, the sites of NO production and its role in the pathogenicity of lesions in the central nervous system (CNS) are unknown. In a chronic model of African trypanosomiasis (mice infected with T. b. brucei surviving with episodic suramin administration), NADPH-diaphorase staining of brain slides revealed that NO synthase (NOS) activity is located not only in endothelial cells, choroid plexus ependymal cells, and neurons as in control mice but also in mononuclear inflammatory cells located in perivascular and parenchyma infiltrates. An immunohistochemical study showed that the mononuclear inflammatory cells expressed an inducible NOS activity. Furthermore, the presence of nitrotyrosine in inflammatory lesions demonstrated an increased NO production and the intermediate formation of peroxynitrites. The detection of extensive formation of nitrotyrosine in the CNS parenchyma was observed in mice having shown neurological disorders, suggesting the role of peroxynitrites in the appearance of neurological troubles. In conclusion, this study confirmed the increased NO synthesis in the CNS of mice infected with T. b. brucei and suggests a deleterious role for NO, through the formation of peroxynitrites, in the pathogenesis of African CNS trypanosomiasis.     

Increased germ cell degeneration and reduced germ cell:Sertoli cell ratio in stallions with low sperm production

To determine the relationship between germ cell degeneration or germ cell:Sertoli cell ratio and daily sperm production, testes were obtained during the months of May to July (breeding season) and November to January (nonbreeding season) from adult (4 to 20-yr-old) stallions with either high (n = 15) or low (n = 15) sperm production. Serum was assayed for concentrations of LH, FSH and testosterone. Testes were assayed for testosterone content and for the number of elongated spermatids, after which parenchymal samples were prepared for histologic assessment. Using morphometric procedures, the types and numbers of spermatogonia, germ cells and Sertoli cells were determined. High sperm producing stallions had greater serum testosterone concentration, total intratesticular testosterone content, testicular parenchymal weight, seminiferous epithelial height, diameter of seminiferous tubules, numbers of A and B spermatogonia per testis, number of Sertoli cells per testis, and number of B spermatogonia, late primary spermatocytes, round spermatids and elongated spermatids per Sertoli cell than low sperm producing stallions (P < 0.05). The number of germ cells (total number of all spermatocytes and spermatids in Stage VIII tubules) accommodated by Sertoli cells was reduced in low sperm producing stallions (18.6 +/- 1.3 germ cells/Sertoli cell) compared with that of high sperm producing stallions (25.4 +/- 1.3 germ cells/Sertoli cell; P < 0.001). The conversion from (yield between) early to late primary spermatocytes and round to elongated spermatids was less efficient for the low sperm producing stallions (P < 0.05). Increased germ cell degeneration during early meiosis and spermiogenesis and reduced germ cell:Sertoli cell ratio was associated with low daily sperm production. These findings can be explained either by a compromised ability of the Sertoli cells to support germ cell division and/or maturation or the presence of defects in germ cells that predisposed them to degeneration.     

Arylsulfatase A deficiency causes seminolipid accumulation and a lysosomal storage disorder in Sertoli cells

Sulfogalactosylglycerolipid (SGG) is the major sulfoglycolipid of male germ cells. During spermatogenesis, apoptosis occurs in >50% of total germ cells. Sertoli cells phagocytose these apoptotic germ cells and degrade their components using lysosomal enzymes. Here we demonstrated that SGG was a physiological substrate of Sertoli lysosomal arylsulfatase A (ARSA). SGG accumulated in Sertoli cells of Arsa(-/-) mice, and at 8 months of age, this buildup led to lysosomal swelling and other cellular abnormalities typical of a lysosomal storage disorder. This disorder likely compromised Sertoli cell functions, manifesting as impaired spermatogenesis and production of sperm with near-zero fertilizing ability in vitro. Fecundity of Arsa(-/-) males was thus reduced when they were older than 5 months. Sperm SGG is known for its roles in fertilization. Therefore, the minimal sperm fertilizing ability of 8-month-old Arsa(-/-) males may be explained by the 50% reduction of their sperm SGG levels, a result that was also observed in testicular germ cells. These unexpected decreases in SGG levels might be partly due to depletion of the backbone lipid palmitylpalmitoylglycerol that is generated from the SGG degradation pathway in Sertoli cells and normally recycled to new generations of primary spermatocytes for SGG synthesis.     

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.     

Developmental regulation of the bcl-2 family during spermatogenesis: insights into the sterility of bcl-w-/- male mice

Expression of bcl-w, a close relative of bcl-2 is essential for male fertility in mice. Although the initial wave of spermatogenesis in bcl-w -/- mice proceeds normally until 3-4 weeks of age, adults fail to produce sperm. To clarify why bcl-w is essential for adult but not juvenile spermatogenesis, we investigated the expression pattern of eight bcl-2 family members. We found that both the level and pattern of expression varied in different cell types during juvenile and adult spermatogenesis. Anti-apoptotic genes bcl-w, bcl-2 and bcl-xL were all expressed in spermatogonia during juvenile spermatogenesis, but only bcl-w was detected in spermatogonia of adult mice. A similar shift was evident in Sertoli cells. This developmental regulation may co-ordinate physiological germ cell apoptosis in wild type mice and account for the time of onset for pathological germ cell apoptosis in bcl-w -/- animals.