Elimination Of Germ Cells Research Articles

Higher temperatures directly increase germ cell number, promoting feminization of red-eared slider turtles.

In many reptile species, gonadal sex is affected by environmental temperature during a critical period of embryonic development-a process known as temperature-dependent sex determination (TSD).1 The oviparous red-eared slider turtle, Trachemys scripta, has a warm-female/cool-male TSD system and is among the best-studied members of this group.2 When incubated at low temperatures, the somatic cells of the bipotential gonad differentiate into Sertoli cells, the support cells of the testis, whereas at high temperatures, they differentiate into granulosa cells, the support cells of the ovary.3 Here, we report the unexpected finding that temperature independently affects the number of primordial germ cells (GCs) in the embryonic gonad at a time before somatic cell differentiation has initiated. Specifically, embryos incubated at higher, female-inducing temperatures have more GCs than those incubated at the male-inducing temperature. Furthermore, elimination of GCs in embryos incubating at intermediate temperatures results in a strong shift toward male-biased sex ratios. This is the first evidence that temperature affects GC number and the first evidence that GC number influences sex determination in amniotes. This observation has two important implications. First, it supports a new model in which temperature can impact sex determination in incremental ways through multiple cell types. Second, the findings have important implications for a major unresolved question in the fields of ecology and evolutionary biology-the adaptive significance of TSD. We suggest that linking high GC number with female development improves female reproductive potential and provides an adaptive advantage for TSD.

Global changes in epigenomes during mouse spermatogenesis: possible relation to germ cell apoptosis.

Mammalian spermatogenesis is characterized by disproportionate germ cell apoptosis. The high frequency of apoptosis is considered a safety mechanism that serves to avoid unfavorable transmission of paternal aberrant genetic information to the offspring as well as elimination mechanism for removal of overproduced immature or damaged spermatogenic cells. The molecular mechanisms involved in the induction of germ cell apoptosis include both intrinsic mitochondrial Bcl-2/Bax and extrinsic Fas/FasL pathways. However, little is known about the nuclear trigger of those systems. Recent studies indicate that epigenomes are essential in the regulation of gene expression through remodeling of the chromatin structure, and are genome-like transmission materials that reflect the effects of various environmental factors. In spermatogenesis, epigenetic errors can act as the trigger for elimination of germ cells with abnormal chromatin structure, abnormal gene expression and/or morphological defects (disordered differentiation). In this review, we focus on the relationship between global changes in epigenetic parameters and germ cell apoptosis in mice and other mammals.

The Initiation of Meiotic Sex Chromosome Inactivation Sequesters DNA Damage Signaling from Autosomes in Mouse Spermatogenesis.

Meiotic sex chromosome inactivation (MSCI) is an essential event in the mammalian male germline. MSCI is directed by a DNA damage response (DDR) pathway centered on the phosphorylation of histone variant H2AX at serine 139 (termed γH2AX). The failure to initiate MSCI is linked to complete meiotic arrestand elimination of germ cells; however, the mechanisms underlying this arrest and elimination remain unknown. To address this question, we established a new separation-of-function mouse model for H2ax that shows specific and complete defects in MSCI. The genetic change is a point mutation in which another H2AX amino acid residue important in the DDR, tyrosine 142 (Y142), is converted to alanine (H2ax-Y142A). In H2ax-Y142A meiosis, the establishment of DDR signals on the chromosome-wide domain of the sex chromosomes is impaired. The initiation of MSCI is required for stage progression, which enables crossover formation, suggesting that the establishment of MSCI permits the timely progression of male meiosis. Our results suggest that normal meiotic progression requires the removal of ATR-mediated DDR signaling from autosomes. We propose a novel biological function for MSCI: the initiation of MSCI sequesters DDR factors from autosomes to the sex chromosomes at the onset of the pachytene stage, and the subsequent formation of an isolated XY nuclear compartment-the XY body-sequesters DDR factors to permit meiotic progression from the mid-pachytene stage onward. VIDEO ABSTRACT.

Unspecific histological and hematological alterations in anadromous and resident Salvelinus malma induced by volcanogenic pollution

The streams draining Kamchatkan volcanic areas are perfect natural models producing the long-lasting effects similar to inorganic anthropogenic pollution. The most abundant local fish species Salvelinus malma is able to reproduce under highly unfavorable conditions: contamination with heavy metals and mineral suspensions. The aim of this study was to assess the adaptability of the landlocked and anadromous populations in adverse conditions. Here, we examined the liver toxicant accumulation in the overwintered S. malma juveniles, as well as aberrations in gills, internal organs, and peripheral blood. Compared to the anadromous populations the landlocked populations demonstrate poor ability to develop the mechanisms of decreasing toxicants’ bioaccumulation. The sets of histological and hematological alterations demonstrated a striking similarity for both ecotypes, and the difference in occurrence of tissue alterations was mostly insignificant. Extreme negative effects of the heavily polluted environment were: the secondary lamellas’ fusion, necrosis of liver and spleen tissues, erythrocyte morphological abnormalities, and elimination of germ cells. We suppose that the resistance to organs’ malfunctions is not the main adaptive vector in S. malma populations in the polluted streams, and expect the same long-term pattern of injury in the resident fish populations under anthropogenic pollution.

Stress and Cell Death in Testicular Cells

The success of male reproduction requires the production of a large number of spermatozoa by a unique process known as spermatogenesis. Spermatogenesis is carried out in close association with the Sertoli cells, the only somatic cells of the seminiferous epithelium which are responsible for providing structural, nutritional and endocrine support to the developing germ cells. The seminiferous epithelium of the testes is a rapid proliferation tissue, where the germinal cells, through a large number of mitotic and meiotic divisions prior to their differentiation culminate with the structural and functional formation of spermatozoa. The number of germ cells that Sertoli cells can sustain is maintained by apoptosis, which fulfills the elimination of germ cells with genetic errors, damage to DNA or excess cell production. Apoptosis can also be activated by external factors such as stress, causing alterations in spermatogenesis and testicular involution, which compromises fertility. However, death in testicular cells is not attributed only to apoptosis, as cells use different mechanisms to activate their self-elimination, such as anoikis and autophagy. All of these mechanisms are discussed.

CHARACTERISTICS ЩА REPRODUCTIVE ORGANS OF CASPIAN SEAL (PHOCA CASPICA, GMELIN, 1788) IN THE CONDITIONS OF ANTHROPOGENIC PRESSING

Abstract. Aim. To study the morpho-functional state of reproductive organs of different-age specimens of the Caspian seal. Methods. We have analyzed the organs of the reproductive system of 26 females and 19 males of the Caspian seal, 29 of which are sexually mature specimens and 18 specimens are not sexually mature. The biological material was sampled during specialized research expeditions to the pre-winter herds in October-November 2012, 2014, 2015. The treatment of samples was carried out in accordance with the standard histology techniques. Results. In organs of the reproductive system of investigated fishes scientists have revealed various pathologies. Sexually maturity of males in testicles had the next violations: hyalinosis of the vessel walls: the absence of the layer of spermatogone: pyknosis of kernels of sperm. Pathologies in tests of immature males were not essential. Sexually maturity females had the signs of glandular endometrial hyperplasia. It is noted the signs of sclerous-cystous ovaries. In the uterus of immature females were revealed pathological changes due to inflammatory reaction; in the feeding substance of the ovaries it was discovered a significant number of interstitial glands. Such violations were registered by a number of authors under chronic intoxication (ecotoxicity). Main conclusions. Immature individuals have not expressed pathological changes, which are less expressed than in older age groups. Increased elimination of germ cells is a result of the damaging effects on the reproductive system of animals. Arduous duty of functioning may cause of the violation of the reproductive potential of the population,that on the background of ecological problem can lead to further reduction in the number of Caspian seal.

Bim Mediates Germ Cell Death During Ovarian Development.

Normal ovarian development in mice is characterized by the initial expansion of germ cells followed by the death of more than two thirds of their maximum number. The final result of germ cell mitosis and apoptosis during embryogenesis is a fixed number of oocytes stored within the ovary as primordial follicles just after birth, from which all ovulated oocytes are derived. Thus, apoptosis plays a critical role in determining the number of oocytes endowed within the ovary and the fertile lifespan of the female. In this study, we investigated the role of Bim (Bcl-2 interacting mediator of cell death), a pro-apoptotic protein belonging to the BH3-only subgroup of the Bcl-2 family of apoptotic regulators, in mediating cell death during the establishment and maintenance of the primordial follicle pool. Immunohistochemical (IHC) staining revealed that Bim protein is expressed within a small subset of mouse oocytes in meiotic prophase at embryonic day (E) 15.5 to E17.5, with minimal expression detected in oocytes once primordial follicle assembly is complete at post natal (PN) day 5. Analysis of follicle numbers at PN10 showed that bim-/- mice had significantly more primordial follicles than WT (2.76 ± 0.25 vs 1.51 ± 0.23 follicles/104μm2, P<0.05), supporting an important role for Bim in determining the initial size of the primordial follicle pool. Furthermore, while quantitative real-time polymerase chain reaction (QRT-PCR), in situ hybridisation (ISH) and IHC analysis revealed the induction of Bim expression within primordial follicle oocytes following ?-irradiation-induced DNA damage, Bim deficiency could not rescue oocytes from ?-irradiation induced apoptosis, probably due to the presence of other BH3-only family members, most likely Puma and Noxa. In summary, this study supports a non-redundant and essential role for pro-apoptotic Bim in the elimination of germ cells during ovarian development and in the establishment of the primordial follicle reserve. This work was supported by the NHMRC of Australia (Program Grants #494802 and #257502, Fellowships JKF (#441101), KJH (#494836), CLS (#406675), AS (#461299); Victorian Government's Operational Infrastructure Support Program.

Protective effect of Selenium on the testes and spermatogenesis in acute and long–term cadmium treated male Swiss mice

The protective effect of selenium chloride (SeCl2) against cadmium chloride (CdCl2), induced toxicity on sperms parameters, testes architecture and antioxidant enzymes in male Swiss mice was investigated. Intraperitoneal injection of CdCl2 (0.15 mg) dose in mice reduced total sperm count and significantly associated with increased sperm abnormality and death sperm percentage when compared with controls. The effect of Cd-treated mice showed a significant decrease in weight of testes and epididymides as compared with control groups. The degree of organ weight loss (testes and epididymides) were less severe in long-term treatment. Long-term exposure of both cadmium-injected mice had similar effect as acute exposure but with lower efficacy. Recovery period showed restoring effects in both exposure periods. The histopathological testes changes indicated by the reduction of seminiferous tubule diameter and testes architecture appearance due to the epithelial sloughing and elimination of germ cells were more pronounced at 72h/ Cd treatment. Substantially proves the ongoing damage effect of cadmium on developing germ cells. Long-term exposure of a batch of cadmium –injected mice had a similar effect as acute exposure but with lower efficacy. In addition, Intraperitoneal injection of CdCl2 (0.15 mg) also stimulate lipid peroxidation in testicular tissues, indicated by significant decrease in antioxidant enzymatic activity such as peroxidase and catalase in the experimental mice group. Administration of SeCl2 in combination with equal volume CdCl2 at the above mentioned doses led to partially organs weight resorted, elevated sperm count and reduction in the percentage of abnormal and dead sperm population along with significant increase in antioxidant enzymatic activities. The protective action and the antagonistic effect of Se at different periods against cadmium –induce toxicity are discussed.

Transgene-mediated cosuppression and RNA interference enhance germ-line apoptosis in Caenorhabditis elegans

Introduction of multiple copies of a germ-line-expressed gene elicits silencing of the corresponding endogenous gene during Caenorhabditis elegans oogenesis; this process is referred to as germ-line cosuppression. Transformed plasmids assemble into extrachromosomal arrays resembling extra minichromosomes with repetitive structures. Loss of the transgene extrachromosomal array leads to reversion of the silencing phenomenon. Cosuppression and RNAi depend upon some of the same genes. In the C. elegans germ line, about half the cells undergo a physiological programmed cell death that shares most genetic requirements with somatic apoptosis. In addition, apoptosis is stimulated by DNA damage and synaptic failure mediated through different apoptotic checkpoints. We found that both germ-line cosuppression and RNAi of germ-line-expressed genes enhance apoptosis during C. elegans oogenesis. In contrast, apoptosis is not enhanced by extrachromosomal arrays carrying genes not driven by germ-line-specific promoters that thus do not elicit transgene-mediated cosuppression/silencing. Similarly, introduction of doubled-stranded RNA that shares no homology with endogenous genes has no effect on apoptosis. "Silencing-induced apoptosis" is dependent upon sir-2.1 and cep-1 (the worm p53 ortholog), and is accompanied by a rise in RAD-51 foci, a marker for ongoing DNA repair, indicating induction of DNA double-strand breaks. This finding suggests that the DNA damage-response pathway is involved. RNAi and cosuppression have been postulated as defense mechanisms against genomic intruders. We speculate that the mechanism here described may trigger the elimination of germ cells that have undergone viral infection or transposon activation.

Simultaneous Accumulation of Hyaluronan Binding Protein 1 (HABP1/p32/gC1qR) and Apoptotic Induction of Germ Cells in Cryptorchid Testis

In the experimentally cryptorchid rat, spermatogenic arrest is associated with the formation of multinuclear giant cells, leading to large-scale apoptosis and elimination of germ cells from the seminiferous epithelium. Using this model, the role of Hyaluronan Binding Protein 1 (HABP1), which expresses a stage specifically in post-meiotic cells during spermatogenesis, was examined. Cryptorchidism induced complete arrest of spermatogenesis by 2 days, and by 3-5 days many large and small multinucleated giant cells populated the affected tubules. Ultrastructure of the giant cells revealed both single and multiple chromatin aggregation, with some less compact and distorted, and others broken down into tiny fragments. These cells along with other germ cells were stained terminal deoxynucleotidyl transferase dUTP nick-end labeling positive, demonstrating strong expression of Bax and Heat Shock Protein 70. Simultaneously, there was an up-regulation of the proprotein form of HABP1 in these cells and a decrease in the mature form of protein. The above findings indicate a possible role for HABP1 proprotein in apoptosis induction of germ cells in the cryptorchid testes.

Apoptotic Changes in the Epithelium Germinativum of the Cat (Felis catus s. domestica, L. 1758) at Different Ages and Breeding Seasons

Apoptosis (programmed cell death) could be considered as a physiological process that takes part in a healthy organism, which helps to maintain organism homeostasis. The visible deterioration of semen quality and the number of germ cells is accompanied by a seasonal decrease of the reproductive activity in some species. This post-effect cascade is caused by apoptosis, which is the primary mechanism responsible for the elimination of germ cells during spermatogenesis. The aim of our study was to assess apoptotic changes in the epithelium germinativum in cat testes at different ages. One hundred and two pairs of testes were obtained from domestic cats aged between 4 months and 10 years. The paraffin-embedded tissue sections were labelled using the Oncogene and Calbiochem Research Products DNA Fragmentation Detection Kit (Cat# QIA21; Darmstadt, Germany), which allows the recognition of apoptotic nuclei in tissue sections with Fragment End Labelling (FragEL) of DNA. The activity of apoptotic processes in cat testes collected from the spring-summer period compared with the autumn-winter season revealed that, 59.42% and 51.51%, respectively, males testes were characterized by insignificant changes. The obtained data revealed a distinctive apoptotic changes in the young animal testes before spermatogenesis onset. An intensification of programmed death cells in the epithelium germinativum in the elder cats (between 3-6 and 6-10 years) was not observed. Apoptotic changes slightly intensified in cats aged between 12 and 36 months.

Deletion of Genes Implicated in Protecting the Integrity of Male Germ Cells Has Differential Effects on the Incidence of DNA Breaks and Germ Cell Loss

BackgroundInfertility affects ∼20% of couples in Europe and in 50% of cases the problem lies with the male partner. The impact of damaged DNA originating in the male germ line on infertility is poorly understood but may increase miscarriage. Mouse models allow us to investigate how deficiencies in DNA repair/damage response pathways impact on formation and function of male germ cells. We have investigated mice with deletions of ERCC1 (excision repair cross-complementing gene 1), MSH2 (MutS homolog 2, involved in mismatch repair pathway), and p53 (tumour suppressor gene implicated in elimination of germ cells with DNA damage).Principal FindingsWe demonstrate for the first time that depletion of ERCC1 or p53 from germ cells results in an increased incidence of unrepaired DNA breaks in pachytene spermatocytes and increased numbers of caspase-3 positive (apoptotic) germ cells. Sertoli cell-only tubules were detected in testes from mice lacking expression of ERCC1 or MSH2 but not p53. The number of sperm recovered from epididymes was significantly reduced in mice lacking testicular ERCC1 and 40% of sperm contained DNA breaks whereas the numbers of sperm were not different to controls in adult Msh2 −/− or p53 −/− mice nor did they have significantly compromised DNA.ConclusionsThese data have demonstrated that deletion of Ercc1, Msh2 and p53 can have differential but overlapping affects on germ cell function and sperm production. These findings increase our understanding of the ways in which gene mutations can have an impact on male fertility.

Switch from BAX-dependent to BAX-independent germ cell loss during the development of fetal mouse ovaries

Female reproductive life is limited by the oocyte/follicle pool, which has been determined by the number of germ cells to enter meiosis and subsequent loss of oocytes. It has been suggested that apoptosis accounts for the elimination of germ cells throughout oogenesis. However, female germ cells are lost continuously while they undergo distinct cell cycles in fetal and neonatal life. No convincing evidence has yet been provided to show apoptotic death of oocytes during meiotic prophase in vivo. In this study, we examined the change in the germ cell population in mice deficient of BAX, a key proapoptotic molecule. The number of germ cells, identified by GCNA1 immunolabeling, approximately doubled in ovaries of Bax(-/-) mice compared with ovaries of heterozygous Bax(+/-) mice and wild-type Bax(+/+) mice by 14.5 days post coitum (d.p.c.) and remained higher up to 24.5 d.p.c. However, there was a rapid loss of germ cells in Bax(-/-) ovaries, paralleling that in Bax(+/-) and Bax(+/+) ovaries from 14.5-24.5 d.p.c., a period in which most germ cells entered and progressed in meiotic prophase. These results suggest that, while progressing through meiotic prophase, oocytes are eliminated by a BAX-independent mechanism.

From spermatocytes to spermatozoa in an infertile XYY male

Sex chromosome distribution and aneuploidy as well as germ cell degeneration were evaluated in meiotic and post-meiotic cells from an infertile XYY male. Sex chromosome distribution was assessed by multicolour fluorescence in situ hybridization on meiotic preparations. Post-meiotic cell aneuploidy was characterized by a method combining multicolour fluorescence in situ hybridization and immunocytochemistry using the proacrosin-specific monoclonal antibody (mAb 4D4). TUNEL assay was carried out on seminiferous tubules to evaluate germ cell degeneration. At the prophase stage of the first meiotic division, 63.64% of cells at the pachytene stage carried three sex chromosomes. The ratio of X-bearing to Y-bearing spermatids and spermatozoa differed significantly from 1 : 1 with an excess of Y-bearing spermatids and spermatozoa. The frequency of hyperhaploid XY spermatids was increased in the XYY male, as well as the incidence of YY, XY and disomic 18 ejaculated spermatozoa. A preferential elimination of germ cells by apoptosis occurred in spermatocytes I. The persistence of the extra Y chromosome during meiosis of an XYY male is associated with a high rate of spermatocyte I degeneration and a low rate of aneuploid spermatozoa.

The p53 protein influences the sensitivity of testicular germ cells to mono-(2-ethylhexyl) phthalate-induced apoptosis by increasing the membrane levels of Fas and DR5 and decreasing the intracellular amount of c-FLIP.

The consequence of mono-(2-ethylhexyl) phthalate (MEHP)-induced injury of testicular Sertoli cells is the Fas-dependent apoptotic elimination of germ cells. In addition to the well-known ability of p53 to regulate the transcription of various apoptosis-associated proteins, p53 also has been implicated in mediating the localization of Fas to the plasma membrane of various cell types in a transcription-independent manner. To resolve the role of p53 in MEHP-mediated testicular toxicity, we used wild-type (p53(+/+)) and p53 knockout (p53(-/-)) mice. A significantly lower incidence of TUNEL-positive germ cells was observed in p53(-/-) mice compared to p53(+/+) mice at 1, 1.5, and 24 h after MEHP exposure. In these same mice, an induction of Fas and death receptor-5 (DR5) in testicular membrane preparations was observed only in p53(+/+) mice. Analyses of mRNA levels in testes of p53(+/+) and p53(-/-) mice by reverse transcription-polymerase chain reaction revealed that increases in membrane levels of Fas occurred in the absence of their transcriptional up-regulation. Processing of procaspase-8 was observed only in MEHP-treated p53(+/+) mice, and this correlated with the observed incidence of germ cell apoptosis. Interestingly, the p53 status of mice also influenced the stability of c-FLIP (L), a caspase-8 inhibitory protein, that was measured at levels approximately two- to fivefold higher in p53(-/-) mice after MEHP-exposure compared to those in p53(+/+) mice. Taken together, these data suggest that MEHP-induced germ cell apoptosis is dependent, in part, on the p53 protein and on its abilities to increase the localization of Fas and DR5 on the germ cell membrane as well as to decrease the cellular levels of c-FLIP (L).

Delivery of a cyclic adenosine 3',5'-monophosphate response element-binding protein (creb) mutant to seminiferous tubules results in impaired spermatogenesis.

FSH binding to Sertoli cells is required for optimal production of sperm in mammals. The cAMP response element-binding protein (CREB) is a major mediator of FSH-induced changes in gene expression. To determine whether CREB is required for spermatogenesis, an adenovirus encoding a phosphorylation-defective CREB mutant (AdCREBm1) was used to inhibit CREB activity in Sertoli cells. Addition of AdCREBm1 to primary rat Sertoli cell cultures completely abolished induction of the CREB-regulated c-fos gene. Injection of an adenovirus encoding ss-galactosidase into the rat testis seminiferous tubules in vivo demonstrated that predominately Sertoli cells were infected by adenovirus. AdCREBm1-directed expression of CREBm1 in seminiferous tubules did not affect Sertoli cell viability, but resulted in the apoptosis of meiotic spermatocyte germ cells within 4 days of adenovirus injection into seminiferous tubules. Disrupted spermatogenesis, defined by at least a 75% reduction of round spermatids, was observed in 42 +/- 5.8% of seminiferous tubules 14 days after AdCREBm1 infection, whereas using this criteria, testes injected with a control adenovirus did not display disrupted spermatogenesis. These data demonstrate that AdCREBm1 causes apoptosis and elimination of germ cells and suggest that CREB is required to produce a Sertoli cell-derived factor that is critical for germ cell survival.

The relevance of spontaneous- and chemically-induced alterations in testicular germ cell apoptosis to toxicology

Elimination of germ cells via apoptosis occurs spontaneously under normal physiologic conditions and is often heightened after chemical-induced testicular injury. Though many different apoptosis-related elements have been identified in the testis, the molecular and cellular mechanisms that regulate germ cell apoptosis are not thoroughly understood. In this review, the role of germ cell apoptosis in spermatogenesis and possible key regulators of apoptosis is described. The involvement of the Fas-signaling pathway between Sertoli cells and germ cells is highlighted as a crucial paracrine-signaling mechanism that responds to both physiologic- or toxicant-induced declines in the supportive capacity of the testis and reduces the germ cell population accordingly.

Is Leydig cell steroidogenic function affected by the germ cell content of the seminiferous tubules?

The effect of testicular germ cell content on Leydig cell steroidogenic function in vivo in adult rats was examined. Three experimental paradigms were used to effect germ cell changes. First, a vitamin A-depletion/repletion regimen was used to achieve synchrony at different stages of the cycle of the seminiferous epithelium and thus produce testes with widely differing germ cell contents. Second, long term vitamin A depletion was used to effect germ cell, but not Leydig cell, loss. Third, Leydig cells and germ cells first were eliminated from the testes of adult rats by the administration of ethane 1,2-dimethane sulfonate (EDS) along with testosterone- and estradiol-filled Silastic capsules; Leydig cells were then restored to the germ cell-depleted testes by removal of the luteiniging hormone (LH)-suppressing capsules. Serum, interstitial fluid, and seminiferous tubule fluid testosterone concentrations did not differ between rats in which at least 70% of the seminiferous tubules contained germ cells at stages VII-VIII or at stages XII-III of the cycle. The capacity of the testes of these rats to produce testosterone, assessed by their in vitro perfusion with maximally stimulating LH, also showed no differences despite the differences in germ cell content. Elimination of germ cells throughout the testes by long term vitamin A depletion also did not affect the steroidogenic function of the testes. Finally, the steroidogenic function of Leydig cells restored to germ cell-depleted testes was indistinguishable from that of germ cell-containing controls. These results, taken together, provide evidence supporting the contention that the germ cell content of the testis has little or no effect on testicular steroidogenic function.

THE INTER-SERTOLI CELL TIGHT JUNCTIONS IN GERM CELL-FREE SEMINIFEROUS TUBULES FROM PRENATALLY IRRADIATED RATS: A FREEZE-FRACTURE STUDY

The tight junctions between Sertoli cells were examined by freeze-fracture in 3-month-old prenatally irradiated rats, whose seminiferous tubules are devoid of germ cells. The replicas from irradiated tubules show elaborate interdigitations of the lateral membranes of Sertoli cells and very extensive tight junctions. These junctions are characterized by a great number of continuous parallel or complex interweaving strands of intramembranous particles, preferentially associated with E fracture faces. The presence of highly cross-linked tight junctional strands is compatible with an epithelium deprived of germ cells, with a reduced need for flexibility. Anomalous ectoplasmic specializations, consisting of groups of cisternae arranged perpendicularly to the lateral surface, are found in the irradiated tubules. These structures may be involved in a storage mechanism of redundant lateral membrane resulting from the elimination of germ cells. Typical gap junctions, intercalated between the tight junctional strands, are larger and more frequently found in treated animals than in controls. These findings indicate that a very tight permeability barrier seems to be established in the irradiated testis even in the absence of germ cells. Thus, the formation and maintenance of Sertoli tight junctions do not appear to be directly dependent on the presence of germ cells. Nevertheless, the alterations detected in the tight junction architecture and in the ectoplasmic specializations indicate that maturing germ cells probably contribute to the functional organization of the blood–testis barrier in the normal testis.

Elimination of germ cells during differentiation of the human ovary: an electron microscopic study

Observations by scanning and transmission electron microscopy during ovarian development (from 7 wk until term of gestation) showed numerous germ cells within the superficial epithelial layer covering the ovary. In early stages of differentiation (between 7 and 11 wk), germ cells appeared to be actively migrating to the surface of the ovary by ameboid-like movements. Later, the transfer of the germ cells to the surface epithelium was also occurring passively, as a consequence of an intense proliferation of germ and somatic cells — arranged in clusters and cords — migrating towards the most superficial areas of the ovary. The present evidence suggests that a number of primordial germ cells — as well as oogonia and oocytes — which lie in the superficial epithelium can leave the ovary and ultimately rest freely upon its surface. Elimination of germ cells during differentiation of the ovary in humans was always paralleled by necrosis of oogonia and oocytes and atretic alteration of primitive follicles. The significance of these processes is discussed in relation to the reduction of the pool of oocytes at birth.