Control Of Germ Cell Apoptosis Research Articles

Gonadotropin Regulation Testicular RNA Helicase, Two Decades of Studies on Its Structure Function and Regulation From Its Discovery Opens a Window for Development of a Non-hormonal Oral Male Contraceptive.

Gonadotropin Regulated Testicular Helicase (GRTH/DDX25) is member of the DEAD-box family of RNA helicases present in Leydig and germ cells. GRTH is the only family member regulated by hormones, luteinizing hormone, through androgen action. Male mice with knock-out of the GRTH gene are sterile, lack sperm with arrest at round spermatids. GRTH participates on the nuclear export and transport of specific mRNAs, the structural integrity of Chromatoid Bodies of round spermatids, where mRNAs are processed and stored, and in their transit to polyribosomes, where it may regulate translation of relevant genes. GRTH has a central role in the control of germ cell apoptosis and acts as negative regulator of miRNAs which regulate expression of genes involved in the progress of spermatogenesis. In Leydig cells, GRTH gene transcription is regulated by LH via autocrine actions of androgen/androgen receptor and has regulatory effects in steroidogenesis. In germ cells, androgen actions are indirect via receptors in Sertoli cells. Transgenic mice carrying GRTH 5′ flanking region-GFP permitted to discern regions in the gene which directs its expression upstream, in germ cells, and downstream in Leydig cells, and the androgen-regulated transcription at interstitial (autocrine), and germ cell (paracrine) compartments. Further evidence for paracrine actions of androgen/androgen receptor is their transcriptional induction of Germ Cell Nuclear Factor as requisite up-regulator of GRTH gene transcription in round spermatids, linking androgen action to two relevant germ cell genes essential for the progress of spermatogenesis. A missense mutation of R to H at amino acid 242 of GRTH found in 5.8% of a patient population with azoospermia causes loss of the cytoplasmic phospho-GRTH species with preservation of the non-phospho form in transfected cells. Mice with knock-in of the human mutation, lack sperm due to arrest at round spermatids. This model permits to discern the function of phospho-GRTH. The GRTH phospho-site resides at a Threonine structurally adjacent to the mutant site found in patients. Molecular modeling of this site elucidated the amino acids that form the GRTH/PKA interphase and provide the basis for drug design for use as male contraceptive.

ADAM17 As Inducer of Male Germ Cell Apoptosis

Apoptosis during mammalian spermatogenesis is important to control sperm output and to eliminate damaged unwanted cells. A deregulation in apoptosis rate has deleterious consequences and leads to low sperm production. Apoptosis in spermatogenesis has been widely studied, but the mechanism by which it is induced under physiological or pathological conditions has not been clarified. We have recently identified the metalloprotease ADAM17 (TACE) as a putative physiological inducer of germ cell apoptosis. ADAM17 is widely distributed enzyme in many tissues, its main function is to shed (release) its substrates, which leads to the activation of different signaling pathways according to the cellular context. However, in spermatogenesis, ADAM17 is involved in the control or induction of germ cell apoptosis, this seems to be unique and may reflect the complexity of spermatogenesis. In this review I will focus on the published evidence that show how this enzyme involved in the control of germ cell apoptosis during physiological condition and in different insults such as heat stress, DNA damaging agent or endocrine disruptors.

Effect of Antioxidants on Outcome of Testicular Torsion in Rats of Different Ages

We assessed reproductive and testicular function in adult rats after testicular torsion created before, during and after puberty, and with vs without resveratrol or arginine treatment. Age matched rats were divided into groups, including simulated surgery without testicular torsion, 720-degree testicular torsion for 4 hours, testicular torsion with resveratrol treatment and testicular torsion with arginine treatment. To study reproductive function at age 12 weeks each rat mated with 3 females. The males were sacrificed at age 14 weeks. Spermatozoids were collected from the epididymal tail and evaluated for concentration, motility and viability. Testicular samples were collected for morphological analysis. Reproductive function was not altered by testicular torsion but antioxidants improved potency. Compared to sham operated and contralateral samples all spermatozoid parameters from testicular torsion samples were inferior. Resveratrol and arginine did not improve spermatozoid quality or quantity in torsed testes but contralateral samples were improved by each drug. The seminiferous epithelium of rats submitted to testicular torsion during puberty was least affected. Each antioxidant partially to totally prevented the morphological alterations found in rats with untreated testicular torsion. Rats submitted to testicular torsion before puberty that were treated with antioxidants showed the fewest changes. Testicular morphology was altered less in rats when torsion occurred earlier in life, that is during puberty. Treatment with antioxidants improved contralateral spermatozoid production and some fertility parameters. Each antioxidant also prevented testicular morphology alterations after testicular torsion. Prepubertal rats benefited most from antioxidant treatment.

Endocrine Regulation of Male Fertility by the Skeleton

Interactions between bone and the reproductive system have until now been thought to be limited to the regulation of bone remodeling by the gonads. We now show that, in males, bone acts as a regulator of fertility. Using coculture assays, we demonstrate that osteoblasts are able to induce testosterone production by the testes, though they fail to influence estrogen production by the ovaries. Analyses of cell-specific loss- and gain-of-function models reveal that the osteoblast-derived hormone osteocalcin performs this endocrine function. By binding to a G protein-coupled receptor expressed in the Leydig cells of the testes, osteocalcin regulates in a CREB-dependent manner the expression of enzymes that is required for testosterone synthesis, promoting germ cell survival. This study expands the physiological repertoire of osteocalcin and provides the first evidence that the skeleton is an endocrine regulator of reproduction.

Male germ cell apoptosis: regulation and biology

Cellular apoptosis appears to be a constant feature in the adult testis and during early development. This is essential because mammalian spermatogenesis is a complex process that requires precise homeostasis of different cell types. This review discusses the latest information available on male germ cell apoptosis induced by hormones, toxins and temperature in the context of the type of apoptotic pathway either the intrinsic or the extrinsic that may be used under a variety of stimuli. The review also discusses the importance of mechanisms pertaining to cellular apoptosis during testicular development, which is independent of exogenous stimuli. Since instances of germ cell carcinoma have increased over the past few decades, the current status of research on apoptotic pathways in teratocarcinoma cells is included. One other important aspect that is covered in this review is microRNA-mediated control of germ cell apoptosis, a field of research that is going to see intense activity in near future. Since knockout models of various kinds have been used to study many aspects of germ cell development, a comprehensive summary of literature on knockout mice used in reproduction studies is also provided.

Activator protein-1 in human male germ cell apoptosis.

Apoptosis limits germ cell number in the testis, and its dysregulation is associated with male infertility. Here, we evaluated the role of the transcription factor activator protein 1 (AP-1) in male germ cell apoptosis in a culture of human seminiferous tubules. AP-1 DNA-binding activity increased in cultured tubules within 2.5 h, which was earlier than the onset of apoptosis as detected by caspase 3 activation and apoptotic DNA fragmentation. The c-Jun, c-Fos and JunD proteins were detected in the Sertoli cell nuclei, whereas apoptosis occurred in the germ cells. Follicle-stimulating hormone (FSH), whose receptors are expressed in the Sertoli cells, inhibited germ cell apoptosis and concomitantly suppressed AP-1 DNA-binding activity, but had no effect on nuclear factor kappaB (NF-kappaB) activation. These results suggest that AP-1 transcription factors are involved in the Sertoli cell-mediated control of germ cell apoptosis, and that inhibition of germ cell apoptosis by FSH appears to involve suppression of AP-1 activation.

Spermatogenesis and its endocrine regulation

Three major phases compose spermatogenesis: mitotic proliferation of spermatogonia, meiosis of spermatocytes, and spermiogenesis, the restructuring of spermatids into flagellated spermatozoa. The process is fuelled by stem cells that, when dividing, either self-renew or produce spermatogonia that are committed to proliferation, meiosis, and spermiogenesis. During all phases, germ cells are in close contact with and require the structural and functional support of Sertoli cells. In contrast to germ cells, these somatic cells express receptors for sex steroids and follicle-stimulating hormone (FSH), the most important hormones that regulate spermatogenesis. A typical Sertoli cell response to an endocrine stimulus would be to change the release of a growth factor that would then mediate the hormone's effect to the germ cells. Recent studies in the Japanese eel have shown, for example, that in the absence of gonadotropin Sertoli cells produce a growth factor (an orthologue of anti-Mullerian hormone) that restricts stem cell divisions to the self-renewal pathway; also estrogens stimulate stem cell renewal divisions but not spermatogonial proliferation. Gonadotropin or 11-ketotestosterone (11-KT) stimulation, however, induces spermatogonial proliferation, which is in part mimicked by another Sertoli cell-derived growth factor (activin B). Since FSH (besides luteinizing hormone, LH) stimulates steroidogenesis in fish, and since FSH is the only gonadotropin detected in the plasma of sexually immature salmonids, increased FSH signalling may be sufficient to initiate spermatogenesis by activating both Sertoli cell functions and 11-KT production. Another important androgen is testosterone (T), which seems to act via feedback mechanisms that can compromise FSH-dependent signalling or steroidogenesis. The testicular production of T and 11-KT therefore needs to be balanced adequately. Further research is required to elucidate in what way(s) 11-KT stimulates later stages of development, such as entry into meiosis and spermiogenesis. At this period, LH becomes increasingly important for the regulation of androgen production. Results from mammalian models suggest that during the later phases, the control of germ cell apoptosis via Sertoli cell factors is an important regulatory mechanism. In many species, sperm cells cannot fertilize eggs until having passed a maturation process known as capacitation, which includes the acquisition of motility. Progestins that are produced under the influence of LH appear to play an important role in this context, which involves the control of the composition of the seminal plasma (e.g., pH values).

Germ cell suicide: new insights into apoptosis during spermatogenesis.

Mature sperm are the product of a precisely regulated developmental sequence in which germ cell proliferation, differentiation, self-renewal and apoptosis are carefully controlled. The control of germ cell apoptosis during spermatogenesis is especially important. It is mediated by signals derived from the Sertoli cells with which each germ cell is closely associated, as well as by signals originating outside the testis. A greater understanding of these signals is emerging from studies of the spermatogenic defects of genetically modified animals. In particular, the intracellular signaling cascades which ultimately determine germ cell fate are being illuminated by recent studies of the Bcl-2 protein family. This review summarises the crucial role which stringently regulated apoptosis plays in the production of male gametes.

Hormonal and genetic control of germ cell apoptosis in the testis.

Programmed cell death is an evolutionarily conserved cell death process that plays a major role during normal development and homeostasis. In many cases, the ordered execution of this internal death programme leads to typical morphological and biochemical changes that have been termed apoptosis. The crucial role of this mode of cell death in the pathogenesis of diverse human diseases including cancer, acquired immunodeficiency syndrome, neurodegeneratives disorders, atherosclerosis and cardiomyopathy is now supported by a wealth of data. In adult mammals, including humans, germ cell death is conspicuous during normal spermatogenesis and plays a pivotal role in sperm output. Withdrawal of gonadotrophins and testosterone further enhances the degeneration of germ cells in the testis. The availability of a quantitative method for analysing the testicular DNA fragmentation and in situ methods to localize specific germ cells undergoing apoptosis, either spontaneously or in response to a variety of death triggering signals, opens new avenues in the understanding of the significance of germ cell apoptosis during normal and abnormal states of spermatogenesis. A growing body of evidence demonstrates that both spontaneous (during normal spermatogenesis) and accelerated germ cell death triggered by deprivation of the gonadotrophic support or moderately increased scrotal temperature in adult rats occur almost exclusively via apoptosis. Although there has been spectacular progress in the understanding of the molecular mechanisms of apoptosis in various systems other than spermatogenesis, elucidation of the biochemical and molecular mechanisms by which germ cell apoptosis is regulated has only just begun. It is likely that germ cell apoptosis is controlled in a cell-type specific fashion, but the basic elements of the death machinery may be universal. In addition, there is increasing evidence that homozygous disruption of a number of genes in mice results in infertility through accelerated germ cell apoptosis. Manipulation of spermatogenesis by survival factor(s) deprivation or increases in extrinsic death signals in loss-of-function or gain-of-function mouse models provides a basis for further attempts to define the intrinsic regulation of various death-related genes by external death signals. Such information is crucial for effective management of male factor infertility as well as more targeted approaches to male contraception.

Hormonal and genetic control of germ cell apoptosis in the testis

Programmed cell death is an evolutionarily conserved cell death process that plays a major role during normal development and homeostasis. In many cases, the ordered execution of this internal death programme leads to typical morphological and biochemical changes that have been termed apoptosis. The crucial role of this mode of cell death in the pathogenesis of diverse human diseases including cancer, acquired immunodeficiency syndrome, neurodegeneratives disorders, atherosclerosis and cardiomyopathy is now supported by a wealth of data. In adult mammals, including humans, germ cell death is conspicuous during normal spermatogenesis and plays a pivotal role in sperm output. Withdrawal of gonadotrophins and testosterone further enhances the degeneration of germ cells in the testis. The availability of a quantitative method for analysing the testicular DNA fragmentation and in situ methods to localize specific germ cells undergoing apoptosis, either spontaneously or in response to a variety of death triggering signals, opens new avenues in the understanding of the significance of germ cell apoptosis during normal and abnormal states of spermatogenesis. A growing body of evidence demonstrates that both spontaneous (during normal spermatogenesis) and accelerated germ cell death triggered by deprivation of the gonadotrophic support or moderately increased scrotal temperature in adult rats occur almost exclusively via apoptosis. Although there has been spectacular progress in the understanding of the molecular mechanisms of apoptosis in various systems other than spermatogenesis, elucidation of the biochemical and molecular mechanisms by which germ cell apoptosis is regulated has only just begun. It is likely that germ cell apoptosis is controlled in a cell-type specific fashion, but the basic elements of the death machinery may be universal. In addition, there is increasing evidence that homozygous disruption of a number of genes in mice results in infertility through accelerated germ cell apoptosis. Manipulation of spermatogenesis by survival factor(s) deprivation or increases in extrinsic death signals in loss-of-function or gain-of-function mouse models provides a basis for further attempts to define the intrinsic regulation of various death-related genes by external death signals. Such information is crucial for effective management of male factor infertility as well as more targeted approaches to male contraception.

Expression of Fas system-related genes in the testis during development and after toxicant exposure

The Fas system has been identified as a key regulator of testicular germ cell apoptosis. The goal of these experiments was to explore the expression of Fas system-related genes in the testis during development and after toxicant exposure. Both Fas ligand (FasL) and Fas receptor (Fas) were expressed postnatally in rat testis with peak expression associated with the high levels of germ cell apoptosis found during the first wave of spermatogenesis. The testicular expression of RIP and FAP-1, components of the Fas activating complex, increased after exposure to mono-(2-ethylhexyl)phthalate (MEHP), a Sertoli cell toxicant which induces massive germ cell death. Finally, the expression of additional apoptosis-inducing genes, including tumor necrosis factor receptor (TNFR), FADD, TRAIL, and DR5, was detected in mammalian testis. These results provide additional support for the following concepts: (1) Sertoli-germ cell interactions are important in the control of germ cell apoptosis; and (2) the Fas system and similar paracrine systems are important modulators of testicular homeostasis.

A matter of death and life: the significance of germ cell death during spermatogenesis.

The significance of cell death occurring during spermatogenesis is a subject of interest because of its potential medical importance. Unfortunately, the field has been difficult for andrologists to penetrate, in part because of the difficulties of studying germ cells in vitro and the complexity of designing suitable models in which to dissect the molecular signalling pathways involved in control of germ cell apoptosis. As a result, the reasons for these deaths remain unclear despite considerable investigative effort. As developments which have occurred over the last few years in understanding of apoptosis can shed light on this important topic, this review focuses on what is currently known about germ cell apoptosis and outlines the emerging picture of what might be the causes and biological role of germ cell deaths in spermatogenesis.

The Fas System is a Key Regulator of Germ Cell Apoptosis in the Testis

Apoptosis occurs in the testis as an important physiological mechanism to limit the number of germ cells in the seminiferous epithelium. Sertoli cells, which tightly regulate germ cell proliferation and differentiation, are implicated in the control of germ cell apoptosis. Fas (APO-1, CD95), a transmembrane receptor protein, transmits an apoptotic signal within cells when bound by Fas ligand (FasL). The Fas system has been implicated in immune regulation, including cytotoxic T cell-mediated cytotoxicity, activation-induced suicide of T cells, and control of immune-privileged sites. Here we propose the Fas system as a key regulator of spermatogenesis. In this model, FasL expressed by Sertoli cells initiates the apoptotic death of germ cells expressing Fas. Using immunohistochemistry, we localized Fas to germ cells and FasL to Sertoli cells. The expression of these genes was dramatically up-regulated after exposure to mono-(2-ethylhexyl) phthalate and 2,5-hexanedione, two widely studied Sertoli cell toxicants known to induce germ cell apoptosis. Mouse germ cells in vitro were susceptible to anti-Fas antibody-induced death, and the survival of rat germ cells was increased after disruption of FasL by antisense oligonucleotide treatment. Unlike its expression in other tissues, testicular expression of Fas in the lpr mouse, a spontaneous mutant of the Fas gene, is similar to that in the normal mouse, arguing for the importance of the Fas system in maintaining testicular homeostasis. These data implicate the Sertoli cell in the paracrine control of germ cell output during spermatogenesis by a Fas-mediated pathway.

The Fas System Is a Key Regulator of Germ Cell Apoptosis in the Testis*

Apoptosis occurs in the testis as an important physiological mechanism to limit the number of germ cells in the seminiferous epithelium. Sertoli cells, which tightly regulate germ cell proliferation and differentiation, are implicated in the control of germ cell apoptosis. Fas (APO-1, CD95), a transmembrane receptor protein, transmits an apoptotic signal within cells when bound by Fas ligand (FasL). The Fas system has been implicated in immune regulation, including cytotoxic T cell-mediated cytotoxicity, activation-induced suicide of T cells, and control of immune-privileged sites. Here we propose the Fas system as a key regulator of spermatogenesis. In this model, FasL expressed by Sertoli cells initiates the apoptotic death of germ cells expressing Fas. Using immunohistochemistry, we localized Fas to germ cells and FasL to Sertoli cells. The expression of these genes was dramatically up-regulated after exposure to mono-(2-ethylhexyl) phthalate and 2,5-hexanedione, two widely studied Sertoli cell toxicants known to induce germ cell apoptosis. Mouse germ cells in vitro were susceptible to anti-Fas antibody-induced death, and the survival of rat germ cells was increased after disruption of FasL by antisense oligonucleotide treatment. Unlike its expression in other tissues, testicular expression of Fas in the lpr mouse, a spontaneous mutant of the Fas gene, is similar to that in the normal mouse, arguing for the importance of the Fas system in maintaining testicular homeostasis. These data implicate the Sertoli cell in the paracrine control of germ cell output during spermatogenesis by a Fas-mediated pathway.

The Fas System Is a Key Regulator of Germ Cell Apoptosis in the Testis

Apoptosis occurs in the testis as an important physiological mechanism to limit the number of germ cells in the seminiferous epithelium. Sertoli cells, which tightly regulate germ cell proliferation and differentiation, are implicated in the control of germ cell apoptosis. Fas (APO-1, CD95), a transmembrane receptor protein, transmits an apoptotic signal within cells when bound by Fas ligand (FasL). The Fas system has been implicated in immune regulation, including cytotoxic T cell-mediated cytotoxicity, activation-induced suicide of T cells, and control of immune-privileged sites. Here we propose the Fas system as a key regulator of spermatogenesis. In this model, FasL expressed by Sertoli cells initiates the apoptotic death of germ cells expressing Fas. Using immunohistochemistry, we localized Fas to germ cells and FasL to Sertoli cells. The expression of these genes was dramatically up-regulated after exposure to mono-(2-ethylhexyl) phthalate and 2,5-hexanedione, two widely studied Sertoli cell toxicants known to induce germ cell apoptosis. Mouse germ cells in vitro were susceptible to anti-Fas antibody-induced death, and the survival of rat germ cells was increased after disruption of FasL by antisense oligonucleotide treatment. Unlike its expression in other tissues, testicular expression of Fas in the lpr mouse, a spontaneous mutant of the Fas gene, is similar to that in the normal mouse, arguing for the importance of the Fas system in maintaining testicular homeostasis. These data implicate the Sertoli cell in the paracrine control of germ cell output during spermatogenesis by a Fas-mediated pathway.