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.     

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.     

Fate of bone marrow stem cells transplanted into the testis: potential implication for men with testicular failure

To assess adult stem cell differentiation in the testis, we injected bone marrow cells from adult green fluorescent protein (GFP) transgenic mice into the seminiferous tubules and the testicular interstitium of busulfan-treated wild-type or c-kit mutant (W/W(v)) mice. Ten to 12 weeks after transplantation, we examined the fate of the transplanted bone marrow cells and found that they survived in recipient testes. In both the busulfan-treated and W/W(v) mice, some of the GFP-positive donor cells had a Sertoli cell appearance and expressed follicle-stimulating hormone receptor within the seminiferous tubules. In addition, GFP-positive donor cells were found in the interstitium of recipient testes, and they expressed the cytochrome P450 side chain cleavage enzyme (P450scc). In the seminiferous tubules of busulfan-treated mice, GFP-positive donor cells had the appearance of spermatogonia or spermatocytes and expressed VASA. However, this was not found in the seminiferous tubules of W/W(v) mice. We conclude that adult bone marrow cells, in a favorable testicular environment, differentiate into somatic and germ cell lineages. The resident neighboring cells in the recipient testis may control site-appropriate stem cell differentiation. This clinically relevant finding raises the possibility for treatment of male infertility and testosterone deficiency through the therapeutic use of stem 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.     

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.     

Toxic effects of anatoxin-a on testes and sperm counts of male mice

Anatoxin-a, a potent neurotoxin, is one of a number of toxins produced by cyanobacteria especially some strains of Anabaena. Toxic cyanobacteria are found worldwide in inland and coastal water environments. The present study was performed to evaluate the toxicity of anatoxin-a on testes and sperm counts of male mice. The animals of the treatment groups were administered with 50, 100 and 150microg/kg/day anatoxin-a for seven consecutive days by intraperitoneal (i.p.) route. Although there were no significant changes in body weight gain, and absolute and relative testes weights, absolute and relative weights of cauda epididymis reduced significantly in the 100 and 150microg/kg groups when compared with control group. The number of sperm count in cauda epididymis was reduced dose dependently in all treatment groups compared with control animals. Anatoxin-a caused dose-dependent histopathological changes in the testes of mice such as degenerations in seminiferous tubules, intercellular disassociation of spermatogenetic cell lines, sloughing of germ cells into tubular lumen, vacuolisation in Sertoli cells and loss of germ cells. The epithelial thickness of seminiferous tubules decreased significantly in all treatment groups in a dose-dependent manner.     

Experimental allergic orchitis in mice

Summary Experimental allergic orchids was induced in (C57BL/6J × A/J)F₁ mice by two injections of syngeneic testicular homogenate emulsified with adjuvants immediately followed by intravenous injection of pertussis vaccine, at a 2 week interval.Histologically, in the initial stage there was occasional focal degeneration and desquamation of both spermatogonia and Sertoli cells within limited parts of the seminiferous tubules, in the peripheral region of the testis. No inflammatory change was present. In some cases, however, inflammatory reaction in the rete testis and focal lymphocytic infiltration in the interstitium were also observed. Subsequently, marked infiltration of lymphocytes, monocytes, and polymorphs were found not only in the testes, but also in rete testis and epididymis. In later stages the inflammatory reaction gradually subsided, but the testes became atrophic due to progression of spermatogenic arrest. Many tubules were lined only with monolayers of Sertoli cells, surrounded by hyperplastic Leydig cells in the interstitium. At 5 months after the 2nd immunization, there was still variable depression of spermatogenesis and hyperplasia of Leydig cells with scattered fibrous scars in the seminiferous tubules, although good regeneration of germ cells appeared in some tubules.Immunological studies revealed that lymphocytes obtained from mice bearing developed orchitis showed a significantly enhanced response in the mixed culture with syngeneic testicular cells, and suggest that cellular immunity plays an important role in the induction of experimental allergic orchitis in mice.     

Analysis of RhoE expression in the testis,epididymis and ductus deferens,and the effects of its deficiency in mice

Rho proteins are a large family of GTPases involved in the control of actin cytoskeleton dynamics, proliferation and survival. Rnd1, Rnd2 and RhoE/Rnd3 form a subfamily of Rho proteins characterized by being constitutively active. The role of these proteins has been studied during the last years in several systems; however, little is known about their expression and functions in the reproductive organs. In this work we analysed the localization and the effect of RhoE deficiency in the testes using mice lacking RhoE expression (RhoE gt/gt), and our research shows some unexpected and relevant results. First, we have observed that RhoE is only expressed in Leydig cells within the testicular parenchyma and it is absent of seminiferous tubules. In addition, RhoE is expressed in the excurrent ducts of the testis, including the ductuli efferentes, epididymis and ductus deferens. Moreover, the testes of postnatal 15‐day‐old RhoE null mice are smaller, both in absolute values and in relation to the body weight. Furthermore, the dimensions of their seminiferous tubules are also reduced compared with wild‐types. In order to study the role of RhoE in the adult, we analysed heterozygous animals as RhoE null mice die early postnatally. Our results show that the testes of adult RhoE heterozygous mice are also smaller than those of the wild‐types, with a 17% decrease in the ratio testis weight/body weight. In addition, their seminiferous tubules have reduced tubular diameter (12%) and a thinner epithelial wall (33%) that appears disorganized and with a swollen lumen. Finally, and probably as a consequence of those alterations, the sperm concentration of heterozygous animals was found to be lower than in the wild‐types. These results indicate that accurate levels of RhoE in the testes are necessary for a correct development and function of male gonads, and suggest novel and unexpected roles of Rnd GTPases in the reproductive physiology.     

Early testicular changes after vasectomy and vasovasostomy in Lewis rats

The testes of Lewis rats were studied at intervals from 2 weeks to 3 months after bilateral vasectomy, vasectomy followed 1 month later by vasovasostomy, or sham operations. Aims were to determine the nature of early alterations after vasectomy, and to determine whether vasovasostomy after 1 month would result in reversal of vasectomy-induced changes. Approximately one-fourth of the testes in the vasectomy and vasovasostomy groups displayed histological changes, which consisted mainly of depletion of germ cells. The extent of the depletion varied greatly in different seminiferous tubules. In testes altered in this way, no abnormal infiltrations of lymphocytes, macrophages, or other cells were observed in the seminiferous epithelium or in the interstitium. The rete testis and straight tubules were normal in testes with altered seminiferous epithelium. A few testes in the vasectomy and vasovasostomy groups had necrotic centers. The results suggest that depletion of germ cells occurred as a result of shedding from the seminiferous epithelium into the lumen of the tubules. A cellular immune response, such as occurs in experimental allergic orchitis in other species, did not appear to be responsible for the observed loss of germ cells. This suggests a possible role for humoral antibody in this model, since there is an association between testicular changes and serum antisperm antibodies at longer intervals after vasectomy. Testicular alterations were not reversed by performance of a vasovasostomy 1 month after vasectomy.     

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.     

Characterization, cryopreservation, and ablation of spermatogonial stem cells in adult rhesus macaques

Spermatogonial stem cells (SSCs) are at the foundation of mammalian spermatogenesis. Whereas rare A(single) spermatogonia comprise the rodent SSC pool, primate spermatogenesis arises from more abundant A(dark) and A(pale) spermatogonia, and the identity of the stem cell is subject to debate. The fundamental differences between these models highlight the need to investigate the biology of primate SSCs, which have greater relevance to human physiology. The alkylating chemotherapeutic agent, busulfan, ablates spermatogenesis in rodents and causes infertility in humans. We treated adult rhesus macaques with busulfan to gain insights about its effects on SSCs and spermatogenesis. Busulfan treatment caused acute declines in testis volume and sperm counts, indicating a disruption of spermatogenesis. One year following high-dose busulfan treatment, sperm counts remained undetectable, and testes were depleted of germ cells. Similar to rodents, rhesus spermatogonia expressed markers of germ cells (VASA, DAZL) and stem/progenitor spermatogonia (PLZF and GFRalpha1), and cells expressing these markers were depleted following high-dose busulfan treatment. Furthermore, fresh or cryopreserved germ cells from normal rhesus testes produced colonies of spermatogonia, which persisted as chains on the basement membrane of mouse seminiferous tubules in the primate to nude mouse xenotransplant assay. In contrast, testis cells from animals that received high-dose busulfan produced no colonies. These studies provide basic information about rhesus SSC activity and the impact of busulfan on the stem cell pool. In addition, the germ cell-depleted testis model will enable autologous/homologous transplantation to study stem cell/niche interactions in nonhuman primate testes.     

The influence of vasovasostomy on testicular alterations after vasectomy in Lewis rats

The occurrence of alterations in testicular weight and morphology after vasectomy and vasectomy reversal by vasovasostomy was studied in Lewis rats. Animals were studied 3, 4, and 7 months after bilateral vasectomy or a vasectomy followed 3 months later by vasovasostomy. Other rats served as sham-operated controls. The weights of the testes in vasectomy and vasovasostomy animals fell into two groups-small testes weighing less than 0.88 g and normal-sized testes of 1.2 g or more. When the extent of testicular alterations was estimated in sections for light microscopy by use of a semiquantitative testicular biopsy score count (TBSC), the morphology of the testes corresponded closely to the testis weight (r = .94), small testes having correspondingly low TBSC scores. In severely altered small testes, the seminiferous tubules were narrower than in sham-operated rats, and numbers of germ cells were greatly depleted. Many tubules contained only Sertoli cells and spermatogonia, although spermatocytes were present in a minority of tubules. A few seminiferous tubules contained multinucleate spermatids. Electron microscopy of severely altered tubules revealed closely apposed processes of Sertoli cells, which contained filaments, microtubules, and endoplasmic reticulum. In contrast, testes with normal weight in vasectomy and vasovasostomy groups resembled those of the sham-operated animals. Comparison of distributions of testicular biopsy score counts demonstrated differences between vasectomy and vasovasostomy groups as time after operation increased. At the 3-4-month intervals, approximately one-third of the testes were severely altered in both vasectomy and vasovasostomy groups.(ABSTRACT TRUNCATED AT 250 WORDS)     

细辛对D-半乳糖所致衰老小鼠睾丸的形态学研究

本文研究了细辛对D－半乳糖所致衰老小鼠的抗衰老作用，应用光镜电镜技术测定小鼠睾丸的重量、生精小管直径、生精上皮细胞数、随龄变化的间质细胞数，同时观察了细辛对上述指标的影响。结果 1．随增龄、睾丸重量减轻，生精小管直径缩小。衰老时租精细胞缺如，仅有支持细胞或散在分布，间质细胞随龄递减。2．细辛可以使小鼠的曲细精管增粗，生精过程活跃，生精细胞增多，间质细胞增多。结论 细辛有一定抗衰老作用。     

Incipient germ cell tumor in Sertoli-cell-only syndrome testis, accompanied with retroperitoneal teratocarcinoma and widespread metastases

Incipient germ cell tumor in Sertoli-cell-only syndrome testis was examined in an autopsy case of retroperitoneal teratocarcinoma with widespread metastases. Although both testes of a 28-year-old man had clinically been small and free from tumor mass to palpation, histopathological examinations revealed a malignancy in the right testis with the appearance of Sertoli-cell-only syndrome. The left testis showed solely the histology of Sertoli-cell-only syndrome. The testicular malignancy consisted of undifferentiated, atypical germ cells mainly confined within approximately one-tenth of seminiferous tubules, and only one small cartilage nodule. Some tubules showed intratubular growth pattern suggestive of seminoma. A few syncytiotrophoblast-like giant cells occurred in the tubules. These findings seem to furnish substantial evidence to the concept that atypical germ cells are the origin of testicular germ cell tumors of different types.     

Quantitative assessment of the biological effects of follicle regulatory protein on dihydrotestosterone-maintained spermatogenesis in hypophysectomized rat

Follicle regulatory protein (FRP) isolated from porcine ovarian follicles influences folliculogenesis through a paracrine mechanism. A similar protein has been found in the testes and seems to have some inhibitory effects on spermatogenesis when administered to intact male experimental animals. On the basis of female and male studies, it has been ascertained that the effects of FRP are at the level of gonads and not the pituitary or the hypothalamus. In the studies with intact males it was not possible to determine the exact site of FRP action on the testes. Dihydrotestosterone (DHT) has been shown to maintain spermatogenesis in hypophysectomized rats. In order to determine if the inhibitory effects of FRP are at steps prior to the formation of DHT, FRP was administered to hypophysectomized rats that were injected with DHT. Groups of adult rats were hypophysectomized and treated daily with FRP, DHT, FRP + DHT, or vehicle alone for 30 days. At necropsy, body, testes, prostate glands, and seminal vesicle weights were recorded. One testis and sexual accessory glands were fixed for histological evaluation. The contralateral testis was decapsulated, six 2 mm segments of seminiferous tubules, representing defined stages of spermatogenesis, were isolated by transillumination-assisted microdissection, and spermatogenic cells were quantified by DNA flow cytometry. Histologically, the seminiferous tubules of vehicle-treated hypophysectomized controls showed advanced regression. Rats treated with FRP alone showed similar degeneration. On the other hand, rats treated with DHT showed maintenance of spermatogenesis comparable to normal controls. The testes of rats treated with FRP + DHT were indistinguishable from those treated with DHT only. Flow cytometric quantification of germinal cells from all groups confirmed the histological findings. In this study FRP did not exert deleterious effects on DHT-maintained spermatogenesis. This finding suggests that the inhibitory effects of FRP on spermatogenesis in intact animals may not be a direct effect on spermatogenic cells but may impair androgen action or production or DHT formation.     

Development and Morphogenesis of Testis in Human Fetuses

Development and maturation of prenatal testis was studied on 104 human male fetuses. At 9 weeks, the testes were seen as elongated yellowish tissue superolateral to the developing urinary bladder and medial to mesonephros. The testis assumed ellipsoidal shape at 13 weeks. At about 16 weeks, the testes developed convexity on its anterior aspect. Epididymis and testis were at the same level until 20 weeks thereafter the upper testicular pole was encroached upon by the epididymis, with complete encroachment till its anterior aspect at 24 week. The sinus was also distinctly present between the two. The testis assumed miniature adult testis at term, but its size was approximately 1/15th of the adult. Cytoarchitecture of the testis at 9 weeks revealed radially disposed sex cords. At 13 weeks, tunica albuginea,tubular organization in the parenchyma and the Leydig cells were identifiable. At 17 weeks, tunica vasculosa and incomplete lobules were apparent. At 24 weeks, testis was marked by more but solid seminiferous tubules lined by 4–5 germ layers. Leydig cells were also identifiable till 24 weeks. Spermatogenic cells, both pale and dark type were distinguishable although pale spermatogonia were more numerous than dark cells. 28 weeks onwards tunica vaginalis and complete septa in the testicular parenchyma were evident. Moreover, the seminiferous tubules developed central vacuolation. By 30 weeks, seminiferous tubules became more complex as evident by their increase in number and coiled and tortuousity. At term, the fetal testis had not yet attained the cytoarchitecture of the adult testis suggesting that testicular maturation continues postnatally.     

Intratubular germ cell neoplasia of unclassified type occupying the whole testis accompanied by a small mature teratoma and metastatic choriocarcinoma and Sertoli cell-only tubules in the other testis

A 19-year-old man with mild mental retardation was diagnosed as having metastatic choriocarcinoma and a testicular tumor. Histopathological examination of the resected testis revealed the presence of a small lesion of mature teratoma but no trace of choriocarcinoma. The remaining seminiferous tubules were atrophic and lined by large atypical germ cells, which were diagnosed as intratubular germ cell neoplasia of the unclassified type (IGCNU). A small area with prominent tubules was also observed. Within this lesion, the tubules were dilated and contained several layers of cells with central necrosis. Immunohistological comparison of staining for several biological markers (Ki-67, c-kit and placental alkaline phosphatase) between cells in the atrophic tubules and those in the dilated tubules indicated a progression of the latter cells to cells with a more proliferative ability. In the opposite testis, examined at autopsy after death due to metastatic choriocarcinoma, all seminiferous tubules were lined by Sertoli cells only. It was therefore assumed that the germ cell tumor of the combined histological type had primarily arisen in the background of IGCNU, and that choriocarcinoma had spontaneously regressed. The early onset of these testicular neoplastic lesions strongly indicates their occurrence under the genetic background of gonadal dysplasia, the Sertoli cell-only syndrome. The possible relation of gonadal disease to mental retardation in this patient is also discussed.     

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.     

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.     

猕猴输精管内注射高分子聚合物HFMC后对睾丸组织结构的影响

本实验选用具有生育力成年雄性猕猴７只，在直视下行双侧ＨＦＭＣ输精管内注射，每侧剂量分别为３０ｍｇ１只，６０ｍｇ和１００ｍｇ各３只；于注射后２．５年和３．５年分别处死动物，取睾丸组织进行光镜和电镜观察．结果发现：猕猴注射ＨＦＭＣ２．５年后，睾丸光镜大部分曲细精管生精上皮结构完整，排列整齐。仅见局部少数管腔生精上皮层数减少，上皮细胞轻度水样变性等病理改变。电镜下曲细精管内除支持细胞内脂褐素增多，轻度基底膜增厚和精母细胞内质网扩张外，各级生精细胞，支持细胞及细胞间连接复合体等超微结构未见明显异常。注射ＨＦＭＣ３．５年后猕猴的光镜、电镜结果与注射后２．５年结果相似，但局部改变较２．５年组轻。上述结果表明：猕猴输精管内注射一定剂量ＨＦＭＣ节育不会引起睾丸组织的严重病理改变。但是，由于注射ＨＦＭＣ后，ＨＦＭＣ释放Ｈ＋及其对输精管的暂时阻塞，改变了精子生存的内环境，使睾丸出现局部轻度病理改变，随着ＨＦＭＣ逐渐溶解排出，睾丸功能相继恢复正常，配对产仔。为ＨＦＭＣ应用提供了安全性依据。