Inhibition of testicular germ cell apoptosis and differentiation in mice misexpressing Bcl-2 in spermatogonia

Development ◽  
1996 ◽  
Vol 122 (6) ◽  
pp. 1703-1709 ◽  
Author(s):  
T. Furuchi ◽  
K. Masuko ◽  
Y. Nishimune ◽  
M. Obinata ◽  
Y. Matsui
Keyword(s):  
Transgenic Mice ◽  
Germ Cells ◽  
Molecular Mechanisms ◽  
Elongation Factor ◽  
Chain Elongation ◽  
Seminiferous Tubules ◽  
Testicular Germ Cell ◽  
Abnormal Accumulation ◽  
Normal Spermatogenesis ◽  
Massive Accumulation

During normal spermatogenesis, more than half of the germ cells undergo apoptosis, but the physiological significance and molecular mechanisms of this programmed cell death are largely unknown. Because Bcl-2 functions as a death repressor, we have investigated the effect of misexpressing Bcl-2 in spermatogonia in transgenic mice using the human bcl-2 cDNA under the control of the human polypeptide chain elongation factor 1alpha (EF-1alpha) promoter. In the 2-week-old transgenic testes, exogenous Bcl-2 was expressed in spermatogonia and massive accumulation of spermatogonia was observed in seminiferous tubules by 4 weeks. At this time, only a few spermatocytes were apparent, and the accumulated cells degenerated, leading to vacuolization in some seminiferous tubules by 7 weeks. In older transgenic mice, abnormal accumulation of spermatogonia and degeneration of these germ cells was still observed, but some seminiferous tubules in which the level of Bcl-2 expression was reduced recovered normal spermatogenesis. These observations indicate that spermatogonial apoptosis is part of the normal program of mammalian spermatogenesis and is regulated by a pathway affected by Bcl-2.

scholarly journals Establishment and applications of male germ cell and Sertoli cell lines

Reproduction ◽  
2016 ◽  
Vol 152 (2) ◽  
pp. R31-R40 ◽  
Author(s):  
Hong Wang ◽  
Liping Wen ◽  
Qingqing Yuan ◽  
Min Sun ◽  
Minghui Niu ◽  
...  
Keyword(s):  
Cell Line ◽  
Sertoli Cell ◽  
Cell Lines ◽  
Germ Cells ◽  
Sertoli Cells ◽  
Molecular Mechanisms ◽  
Simian Virus 40 ◽  
T Antigen ◽  
Seminiferous Tubules ◽  
Male Germ Cells

Within the seminiferous tubules there are two major cell types, namely male germ cells and Sertoli cells. Recent studies have demonstrated that male germ cells and Sertoli cells can have significant applications in treating male infertility and other diseases. However, primary male germ cells are hard to proliferatein vitroand the number of spermatogonial stem cells is scarce. Therefore, methods that promote the expansion of these cell populations are essential for their use from the bench to the bed side. Notably, a number of cell lines for rodent spermatogonia, spermatocytes and Sertoli cells have been developed, and significantly we have successfully established a human spermatogonial stem cell line with an unlimited proliferation potential and no tumor formation. This newly developed cell line could provide an abundant source of cells for uncovering molecular mechanisms underlying human spermatogenesis and for their utilization in the field of reproductive and regenerative medicine. In this review, we discuss the methods for establishing spermatogonial, spermatocyte and Sertoli cell lines using various kinds of approaches, including spontaneity, transgenic animals with oncogenes, simian virus 40 (SV40) large T antigen, the gene coding for a temperature-sensitive mutant ofp53, telomerase reverse gene (Tert), and the specific promoter-based selection strategy. We further highlight the essential applications of these cell lines in basic research and translation medicine.

scholarly journals Effect of Cryptorchidism on the Histomorphometry, Proliferation, Apoptosis, and Autophagy in Boar Testes

Animals ◽  
2021 ◽  
Vol 11 (5) ◽  
pp. 1379
Author(s):  
Xiaorui Fan ◽  
Yihui Liu ◽  
Meishan Yue ◽  
Weidong Yue ◽  
Gaoya Ren ◽  
...  
Keyword(s):  
Germ Cells ◽  
Sertoli Cells ◽  
Proliferating Cell Nuclear Antigen ◽  
Molecular Mechanisms ◽  
Inguinal Canal ◽  
The Body ◽  
Nuclear Antigen ◽  
Seminiferous Tubules ◽  
Mrna Levels ◽  
Proliferating Cell

Spontaneous unilateral cryptorchid boars have one testis in the abdomen or inguinal canal, causing its temperature to be at or near the body temperature, which impairs spermatogenesis, although the histomorphometry and molecular mechanisms underlying this process remain unclear. The aim of the present study was to determine the histomorphometry, proliferation, apoptosis, and autophagy alterations in spermatogonia and Sertoli cells in unilateral cryptorchid, scrotal (contrascrotal), and preweaning piglet (preweaning) testes. Histomorphometrical analysis of cryptorchid testes showed that the seminiferous tubules contained only Sertoli cells and a few spermatogonia, but did not contain post-meiotic germ cells. The number of spermatogonia markedly decreased, and the number of Sertoli cells did not change remarkably in cryptorchid testes. TUNEL assay results showed that apoptosis signals were predominantly observed in spermatogonia. In cryptorchid and contrascrotal testes, proliferating cell nuclear antigen (PCNA) and LC3 were located in spermatogonia. The number of PCNA-positive, TUNEL-positive, and LC3-positive germ cells was low, and the protein and mRNA levels of PCNA and LC3 were significantly decreased in cryptorchid testes. Taken together, the number of Sertoli cells did not change remarkably, whereas the number of germ cells decreased in the cryptorchid testes, compared with that in the contrascrotal testes. Insufficient proliferation, excessive apoptosis, and autophagy were involved in the regulation of the decrease in spermatogonia in cryptorchid boar testes.

scholarly journals Knockdown of Dehydrodolichyl Diphosphate Synthase in the Drosophila Retina Leads to a Unique Pattern of Retinal Degeneration

2021 ◽  
Vol 14 ◽  
Author(s):  
Tal Brandwine ◽  
Reut Ifrah ◽  
Tzofia Bialistoky ◽  
Rachel Zaguri ◽  
Elisheva Rhodes-Mordov ◽  
...  
Keyword(s):  
Western Blot ◽  
Retinal Degeneration ◽  
Blot Analysis ◽  
Western Blot Analysis ◽  
Molecular Mechanisms ◽  
Retinal Disease ◽  
Chain Elongation ◽  
Missense Mutations ◽  
Unique Pattern ◽  
Abnormal Accumulation

Dehydrodolichyl diphosphate synthase (DHDDS) is a ubiquitously expressed enzyme that catalyzes cis-prenyl chain elongation to produce the poly-prenyl backbone of dolichol. It appears in all tissues including the nervous system and it is a highly conserved enzyme that can be found in all animal species. Individuals who have biallelic missense mutations in the DHDDS gene are presented with non-syndromic retinitis pigmentosa with unknown underlying mechanism. We have used the Drosophila model to compromise DHDDS ortholog gene (CG10778) in order to look for cellular and molecular mechanisms that, when defective, might be responsible for this retinal disease. The Gal4/UAS system was used to suppress the expression of CG10778 via RNAi-mediated-knockdown in various tissues. The resulting phenotypes were assessed using q-RT-PCR, transmission-electron-microscopy (TEM), electroretinogram, antibody staining and Western blot analysis. Targeted knockdown of CG10778-mRNA in the early embryo using the actin promoter or in the developing wings using the nub promoter resulted in lethality, or wings loss, respectively. Targeted expression of CG10778-RNAi using the glass multiple reporter (GMR)-Gal4 driver (GMR-DHDDS-RNAi) in the larva eye disc and pupal retina resulted in a complex phenotype: (a) TEM retinal sections revealed a unique pattern of retinal-degeneration, where photoreceptors R2 and R5 exhibited a nearly normal structure of their signaling-compartment (rhabdomere), but only at the region of the nucleus, while all other photoreceptors showed retinal degeneration at all regions. (b) Western blot analysis revealed a drastic reduction in rhodopsin levels in GMR-DHDDS-RNAi-flies and TEM sections showed an abnormal accumulation of endoplasmic reticulum (ER). To conclude, compromising DHDDS in the developing retina, while allowing formation of the retina, resulted in a unique pattern of retinal degeneration, characterized by a dramatic reduction in rhodopsin protein level and an abnormal accumulation of ER membranes in the photoreceptors cells, thus indicating that DHDDS is essential for normal retinal formation.

scholarly journals Ectopic FGF23 production induces mineral changes, osteogenic transdifferentiation, and cancer associated microcalcifications

2020 ◽  
Author(s):  
Ida Marie Boisen ◽  
John Erik Nielsen ◽  
Ireen Kooij ◽  
Jovana Kaludjerovic ◽  
Peter J. O’Shaughnessy ◽  
...  
Keyword(s):  
Alkaline Phosphatase ◽  
Germ Cell ◽  
Phosphatase Activity ◽  
Sertoli Cell ◽  
Germ Cells ◽  
Alkaline Phosphatase Activity ◽  
Cell Cancer ◽  
Seminiferous Tubules ◽  
Testicular Germ Cell ◽  
Increased Risk

AbstractTesticular microcalcifications consist of hydroxyapatite and their demonstration by ultrasound has been associated with increased risk of testicular germ cell cancer (TGCT). Here, we show that fibroblast growth factor 23 (FGF23), a bone-specific regulator of phosphate homeostasis, is expressed in testicular germ cell neoplasia in situ (GCNIS), embryonal carcinoma (EC), and human embryonic stem cells. FGF23 is not glycosylated in TGCTs and thus rapidly cleaved into a C-terminal fragment that serves as a competitive antagonist for full-length FGF23. High levels of C-terminal FGF23 occupy the receptor formed by Klotho and FGF receptor 1 (FGFR1) in the germ cells facilitating a shift in the expression of phosphate transport proteins from SLC34A2 to SLC34A1 in seminiferous tubules adjacent to GCNIS. Fgf23 knockout mice have a marked epididymal deposition of hydroxyapatite, while the testicular phenotype is milder with spermatogenic arrest and focal germ-cell-specific expression of the bone-like markers runt-related transcription factor 2 (RUNX2) and bone gamma-carboxyglutamic acid-containing protein (BGLAP). In accordance, mice with no functional androgen receptor and lack of circulating gonadotropins develop microcalcifications in 94% of cases and have lower Slc34a2, and higher Slc34a1 and Bglap expression. In accordance, human testicular specimens with microcalcifications also have lower SLC34A2, and focally germ cells express SLC34A1, BGLAP, and RUNX2. Importantly, calcium or phosphate induced osteogenic transdifferentiation of a spermatogonial cell line in vitro demonstrated by induction of alkaline phosphatase activity and deposition of hydroxyapatite, which could be fully rescued by pyrophosphate (PPi). Severe microcalcifications were also found in a mouse model with Sertoli-cell ablation particularly when Sertoli-ablation was conducted prepubertally where the germ cells retain stem cell potential. In conclusion, cancer-related microcalcifications may arise secondary to gonadal mineral alterations, which in combination with impaired Sertoli cell function and reduced PPi due to high alkaline phosphatase activity in GCNIS and TGCTs, facilitates osteogenic transdifferentiation of testicular germ cells and deposition of hydroxyapatite.

scholarly journals Mammalian gonocyte and spermatogonia differentiation: recent advances and remaining challenges

Reproduction ◽  
2015 ◽  
Vol 149 (3) ◽  
pp. R139-R157 ◽  
Author(s):  
Gurpreet Manku ◽  
Martine Culty
Keyword(s):  
Germ Cell ◽  
Molecular Mechanisms ◽  
Germ Cell Tumors ◽  
Adult Life ◽  
Seminiferous Tubules ◽  
Testicular Germ Cell Tumors ◽  
Testicular Germ Cell ◽  
Self Renewal ◽  
Proliferation And Differentiation ◽  
Developmental Phases

The production of spermatozoa relies on a pool of spermatogonial stem cells (SSCs), formed in infancy from the differentiation of their precursor cells, the gonocytes. Throughout adult life, SSCs will either self-renew or differentiate, in order to maintain a stem cell reserve while providing cells to the spermatogenic cycle. By contrast, gonocytes represent a transient and finite phase of development leading to the formation of SSCs or spermatogonia of the first spermatogenic wave. Gonocyte development involves phases of quiescence, cell proliferation, migration, and differentiation. Spermatogonia, on the other hand, remain located at the basement membrane of the seminiferous tubules throughout their successive phases of proliferation and differentiation. Apoptosis is an integral part of both developmental phases, allowing for the removal of defective cells and the maintenance of proper germ–Sertoli cell ratios. While gonocytes and spermatogonia mitosis are regulated by distinct factors, they both undergo differentiation in response to retinoic acid. In contrast to postpubertal spermatogenesis, the early steps of germ cell development have only recently attracted attention, unveiling genes and pathways regulating SSC self-renewal and proliferation. Yet, less is known on the mechanisms regulating differentiation. The processes leading from gonocytes to spermatogonia have been seldom investigated. While the formation of abnormal gonocytes or SSCs could lead to infertility, defective gonocyte differentiation might be at the origin of testicular germ cell tumors. Thus, it is important to better understand the molecular mechanisms regulating these processes. This review summarizes and compares the present knowledge on the mechanisms regulating mammalian gonocyte and spermatogonial differentiation.

Apoptotic germ cells regulate Sertoli cell lipid storage and fatty acid oxidation

Reproduction ◽  
2018 ◽  
Author(s):  
Mariana Regueira ◽  
Agostina Gorga ◽  
Gustavo Marcelo Rindone ◽  
Eliana Herminia Pellizzari ◽  
Selva Cigorraga ◽  
...  
Keyword(s):  
Fatty Acid ◽  
Sertoli Cell ◽  
Germ Cells ◽  
Energy Homeostasis ◽  
Molecular Mechanisms ◽  
Lipid Storage ◽  
Seminiferous Tubules ◽  
Acid Oxidation ◽  
Mrna Levels ◽  
Metabolism Regulation

The presence of lipid droplets (LD) and the utilization of fatty acids (FA) as a source of energy are Sertoli cell (SC) putative characteristics. It is well known that SC can phagocyte and degrade apoptotic germ cells (AGC) resulting in increasing lipid content and ATP levels. A relationship between the regulation of lipid storage and of lipid oxidation in SC might be envisaged. The aim of this study was to analyze whether AGC and FA are able to simultaneously regulate molecular mechanisms involved in lipid storage and FA oxidation in SC. The experimental model utilized in this study consisted in SC cultures obtained from 20-day-old rats that were co-cultured with AGC or treated with palmitic acid (PA, 500μM) for 24 and 48 hours. AGC and PA increase LD, triacylglycerol (TAG) content, and mRNA levels of Plin1, Plin2, Plin3 (proteins involved in TAG storage). Simultaneously, AGC and PA rise the extent of FA oxidation and mRNA levels of Cpt1, Lcad (proteins involved in FA degradation). PPARs are transcription factors which participate in lipid metabolism regulation. Results show that AGC and PA treatment increase PPAR transcriptional activity in SC. Additionally, the presence of a PPARg antagonist decreases the up-regulation of LD content and Plin1 expression. Similarly, the presence of a PPARb/d antagonist reduces the increase in FA oxidation and Cpt1 mRNA levels. Altogether these results suggest that AGC and FA, which probably generates PPARs ligands, regulate lipid storage and fatty acid utilization, contributing to the energy homeostasis in the seminiferous tubules.

Regulation of meiotic entry and gonadal sex differentiation in the human: normal and disrupted signaling

2014 ◽  
Vol 5 (4) ◽  
pp. 331-341 ◽  
Author(s):  
Anne Jørgensen ◽  
Ewa Rajpert-De Meyts
Keyword(s):  
Germ Cell ◽  
Germ Cells ◽  
Fetal Development ◽  
Molecular Mechanisms ◽  
Current Knowledge ◽  
Sex Differentiation ◽  
Cell Cancer ◽  
Testicular Germ Cell ◽  
Unique Type ◽  
Germ Cell Differentiation

AbstractMeiosis is a unique type of cell division that is performed only by germ cells to form haploid gametes. The switch from mitosis to meiosis exhibits a distinct sex-specific difference in timing, with female germ cells entering meiosis during fetal development and male germ cells at puberty when spermatogenesis is initiated. During early fetal development, bipotential primordial germ cells migrate to the forming gonad where they remain sexually indifferent until the sex-specific differentiation of germ cells is initiated by cues from the somatic cells. This irreversible step in gonadal sex differentiation involves the initiation of meiosis in fetal ovaries and prevention of meiosis in the germ cells of fetal testes. During the last decade, major advances in the understanding of meiosis regulation have been accomplished, with the discovery of retinoic acid as an inducer of meiosis being the most prominent finding. Knowledge about the molecular mechanisms regulating meiosis signaling has mainly been established by studies in rodents, while this has not yet been extensively investigated in humans. In this review, the current knowledge about the regulation of meiosis signaling is summarized and placed in the context of fetal gonad development and germ cell differentiation, with emphasis on results obtained in humans. Furthermore, the consequences of dysregulated meiosis signaling in humans are briefly discussed in the context of selected pathologies, including testicular germ cell cancer and some forms of male infertility.

Molecular Mechanisms of Resistance in Testicular Germ Cell Tumors - clinical Implications

2018 ◽  
Vol 18 (10) ◽  
pp. 967-978 ◽  
Author(s):  
Katarina Kalavska ◽  
Vincenza Conteduca ◽  
Ugo De Giorgi ◽  
Michal Mego
Keyword(s):  
Germ Cell ◽  
Molecular Mechanisms ◽  
Cisplatin Resistance ◽  
Apoptotic Cell ◽  
Germ Cell Tumors ◽  
Treatment Resistance ◽  
Experimental Models ◽  
Testicular Germ Cell Tumors ◽  
Testicular Germ Cell ◽  
Repair Capacity

Testicular germ cell tumors (TGCTs) represent the most common malignancy in men aged 15-35. Due to these tumors’ biological and clinical characteristics, they can serve as an appropriate system for studying molecular mechanisms associated with cisplatin-based treatment resistance. This review describes treatment resistance from clinical and molecular viewpoints. Cisplatin resistance is determined by various biological mechanisms, including the modulation of the DNA repair capacity of cancer cells, alterations to apoptotic cell death pathways, deregulation of gene expression pathways, epigenetic alterations and insufficient DNA binding. Moreover, this review describes TGCTs as a model system that enables the study of the cellular features of cancer stem cells in metastatic process and describes experimental models that can be used to study treatment resistance in TGCTs. All of the abovementioned aspects may help to elucidate the molecular mechanisms underlying cisplatin resistance and may help to identify promising new therapeutic targets.

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