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Cannarella R, Crafa A, Curto R, Mongioì LM, Garofalo V, Cannarella V, Condorelli RA, La Vignera S, Calogero AE. Human sperm RNA in male infertility. Nat Rev Urol 2025; 22:92-115. [PMID: 39256514 DOI: 10.1038/s41585-024-00920-9] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 07/10/2024] [Indexed: 09/12/2024]
Abstract
The function and value of specific sperm RNAs in apparently idiopathic male infertility are currently poorly understood. Whether differences exist in the sperm RNA profile between patients with infertility and fertile men needs clarification. Similarly, the utility of sperm RNAs in predicting successful sperm retrieval and assisted reproductive technique (ART) outcome is unknown. Patients with infertility and fertile individuals seem to have differences in the expression of non-coding RNAs that regulate genes controlling spermatogenesis. Several RNAs seem to influence embryo quality and development. Also, RNA types seem to predict successful sperm retrieval in patients with azoospermia. These findings suggest that sperm RNAs could influence decision-making during the management of patients with infertility. This evidence might help to identify possible therapeutic approaches aimed at modulating the expression of dysregulated genes in patients with infertility. Performing prospective studies with large sample sizes is necessary to investigate cost-effective panels consisting of proven molecular targets to ensure that this evidence can be translated to clinical practice.
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Affiliation(s)
- Rossella Cannarella
- Department of Clinical and Experimental Medicine, University of Catania, Catania, Italy.
- Glickman Urological & Kidney Institute, Cleveland Clinic Foundation, Cleveland, OH, USA.
| | - Andrea Crafa
- Department of Clinical and Experimental Medicine, University of Catania, Catania, Italy
| | - Roberto Curto
- Department of Clinical and Experimental Medicine, University of Catania, Catania, Italy
| | - Laura M Mongioì
- Department of Clinical and Experimental Medicine, University of Catania, Catania, Italy
| | - Vincenzo Garofalo
- Department of Clinical and Experimental Medicine, University of Catania, Catania, Italy
| | - Vittorio Cannarella
- Department of Clinical and Experimental Medicine, University of Catania, Catania, Italy
| | - Rosita A Condorelli
- Department of Clinical and Experimental Medicine, University of Catania, Catania, Italy
| | - Sandro La Vignera
- Department of Clinical and Experimental Medicine, University of Catania, Catania, Italy
| | - Aldo E Calogero
- Department of Clinical and Experimental Medicine, University of Catania, Catania, Italy
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2
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Porras-Gómez TJ, Moreno-Mendoza N. Identification of the proliferative activity of germline progenitor cells in the adult ovary of the bat Artibeus jamaicensis. ZYGOTE 2024; 32:366-375. [PMID: 39445446 DOI: 10.1017/s0967199424000364] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/25/2024]
Abstract
Until a few years ago, it was assumed that oocyte renewal did not take place in the ovary of adult organisms; however, the existence of germline progenitor cells (GPCs), which renew the ovarian follicular reserve, has now been documented in mammals. Specifically, in the adult ovary of bats, the presence of cells located in the cortical region with characteristics similar to GPCs, called adult cortical germ cells (ACGC), has been observed. One of the requirements that a GPC must fulfil is to be able to proliferate mitotically, so the evaluation of cell proliferation in ACGC is of utmost importance in order to be able to relate them to a parental lineage. Currently, there are several methods to determine cell proliferation, including BrdU labelling or the use of endogenous proliferation markers. Thus, the aim of this work was to evaluate the proliferative activity of ACGC in the adult ovary of the bat Artibeus jamaicensis, using different proliferation markers and correlating these with the protein expression of the transcription factor Oct4 and the germ line marker Ddx4. We found that the expression pattern of the proliferation markers BrdU, PCNA, Ki-67 and pH3 occurs at different times of the cell cycle, so co-localization of two or more of these markers allows us to identify proliferating cells. This allowed us to identify ACGC with proliferative capacity in the adult ovary of A. jamaicensis, suggesting that GPCs renew the follicle reserve during the adult life of the organism.
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Affiliation(s)
- Tania J Porras-Gómez
- Department of Cell Biology and Phisiology, Instituto de Investigaciones Biomédicas, Universidad Nacional Autónoma de México, Apartado Postal 70228 Ciudad de México04510, Mexico
| | - Norma Moreno-Mendoza
- Department of Cell Biology and Phisiology, Instituto de Investigaciones Biomédicas, Universidad Nacional Autónoma de México, Apartado Postal 70228 Ciudad de México04510, Mexico
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3
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Huang Z, Xie L, Feng H, Lan M, Xu T, Chen D, Pu L, Lu Y. DAZL regulate germline, pluripotency, and proliferation related genes in chicken PGCs and cooperate with DDX4. Theriogenology 2024; 222:22-30. [PMID: 38615433 DOI: 10.1016/j.theriogenology.2024.03.010] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/27/2023] [Revised: 03/15/2024] [Accepted: 03/15/2024] [Indexed: 04/16/2024]
Abstract
Primordial germ cells (PGCs) are the precursors of germ cells and play a crucial role in germline transmission. In chickens, PGCs can be cultured in vitro while maintaining their germline stem cell characteristics. The Deleted in Azoospermia-Like (DAZL) gene, which is highly expressed in PGCs, is essential for germ cell development. Here, through gene knockout experiments, we discovered that the loss of DAZL expression in chicken PGCs led to decreased proliferation and survival. By next employed techniques such as RIP-seq (RNA Binding Protein Immunoprecipitation) and Co-IP-MS/MS (Co-immunoprecipitation Mass Spectrometry), we identified genes directly regulated by DAZL or cooperating with DAZL at the transcriptomic and proteomic levels. DAZL was found to control genes related to germline development, pluripotency, and cell proliferation in PGCs. Additionally, we observed a significant overlap between RNAs and proteins that interact with both DAZL and DDX4, indicating their cooperation in the gene regulation network in chicken PGCs. Our research provides valuable insights into the function of the DAZL gene in germline cells.
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Affiliation(s)
- Zhenwen Huang
- From the Guangxi Key Laboratory of Animal Breeding, Disease Control and Prevention, College of Animal Science and Technology, Guangxi University, Nanning, 530004, China
| | - Long Xie
- From the Guangxi Key Laboratory of Animal Breeding, Disease Control and Prevention, College of Animal Science and Technology, Guangxi University, Nanning, 530004, China
| | - Hu Feng
- From the Agricultural Genomics Institute at Shenzhen Chinese Academy of Agricultural Sciences, Shenzhen, China
| | - Meiyu Lan
- From the Guangxi Key Laboratory of Animal Breeding, Disease Control and Prevention, College of Animal Science and Technology, Guangxi University, Nanning, 530004, China
| | - Tianpeng Xu
- From the Guangxi Key Laboratory of Animal Breeding, Disease Control and Prevention, College of Animal Science and Technology, Guangxi University, Nanning, 530004, China
| | - Dongyang Chen
- From the Guangxi Key Laboratory of Animal Breeding, Disease Control and Prevention, College of Animal Science and Technology, Guangxi University, Nanning, 530004, China
| | - Liping Pu
- From the Guangxi Key Laboratory of Animal Breeding, Disease Control and Prevention, College of Animal Science and Technology, Guangxi University, Nanning, 530004, China
| | - Yangqing Lu
- From the Guangxi Key Laboratory of Animal Breeding, Disease Control and Prevention, College of Animal Science and Technology, Guangxi University, Nanning, 530004, China.
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4
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Jang SW, Kim YR, Han JH, Jang H, Choi HW. Generation of mouse and rat xenogeneic ovaries in vitro for production of mouse oocyte. Anim Cells Syst (Seoul) 2024; 28:303-314. [PMID: 38868077 PMCID: PMC11168328 DOI: 10.1080/19768354.2024.2363601] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/29/2024] [Accepted: 05/30/2024] [Indexed: 06/14/2024] Open
Abstract
The system forming ovarian follicles is developed to investigate in vitro folliculogenesis in a confined environment to obtain functional oocytes. Several studies have reported the successful generation of fully functional oocytes using mouse-induced pluripotent stem cells (iPSCs) and mouse female germline stem cells (fGSCs) as sources of stem cells for in vitro gametogenesis models. In addition, human oogonia have been generated through heterologous co-culture of differentiated human primordial germ cell-like cells (hPGCLCs) with mouse germline somatic cells, although oocyte formation remains challenging. Thus, studies on in vitro ovarian formation in other species are utilized as an introductory approach for in vitro mammalian gametogenesis by understanding the differences in culture systems between species and underlying mechanisms. In this study, we optimized the method of the entire oogenesis process from rat embryonic gonads. We identified well-maturated MII oocytes from rat gonads using our constructed method. Moreover, we generated the first successful in vitro reconstitution of xenogeneic follicles from mouse primordial germ cells (PGCs) and rat somatic cells. We also established an appropriate culture medium and incubation period for xenogeneic follicles. This method will be helpful in studies of xenogeneic follicular development and oocyte generation.
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Affiliation(s)
- Si Won Jang
- Department of Agricultural Convergence Technology, Jeonbuk National University, Jeonju, Republic of Korea
| | - Ye Rim Kim
- Department of Animal Science, Jeonbuk National University, Jeonju, Republic of Korea
| | - Jae Ho Han
- Department of Agricultural Convergence Technology, Jeonbuk National University, Jeonju, Republic of Korea
| | - Hoon Jang
- Department of Life Science, Jeonbuk National University, Jeonju, Republic of Korea
| | - Hyun Woo Choi
- Department of Agricultural Convergence Technology, Jeonbuk National University, Jeonju, Republic of Korea
- Department of Animal Science, Jeonbuk National University, Jeonju, Republic of Korea
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5
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Adashev VE, Kotov AA, Olenina LV. RNA Helicase Vasa as a Multifunctional Conservative Regulator of Gametogenesis in Eukaryotes. Curr Issues Mol Biol 2023; 45:5677-5705. [PMID: 37504274 PMCID: PMC10378496 DOI: 10.3390/cimb45070358] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/18/2023] [Revised: 06/30/2023] [Accepted: 07/03/2023] [Indexed: 07/29/2023] Open
Abstract
Being a conservative marker of germ cells across metazoan species, DEAD box RNA helicase Vasa (DDX4) remains the subject of worldwide investigations thanks to its multiple functional manifestations. Vasa takes part in the preformation of primordial germ cells in a group of organisms and contributes to the maintenance of germline stem cells. Vasa is an essential player in the piRNA-mediated silencing of harmful genomic elements and in the translational regulation of selected mRNAs. Vasa is the top hierarchical protein of germ granules, liquid droplet organelles that compartmentalize RNA processing factors. Here, we survey current advances and problems in the understanding of the multifaceted functions of Vasa proteins in the gametogenesis of different eukaryotic organisms, from nematodes to humans.
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Affiliation(s)
- Vladimir E Adashev
- Department of Molecular Mechanisms for Realization of Genetic Information, Laboratory of Biochemical Genetics of Animals, National Research Center "Kurchatov Institute", Kurchatov Sq. 1, 123182 Moscow, Russia
| | - Alexei A Kotov
- Department of Molecular Mechanisms for Realization of Genetic Information, Laboratory of Biochemical Genetics of Animals, National Research Center "Kurchatov Institute", Kurchatov Sq. 1, 123182 Moscow, Russia
| | - Ludmila V Olenina
- Department of Molecular Mechanisms for Realization of Genetic Information, Laboratory of Biochemical Genetics of Animals, National Research Center "Kurchatov Institute", Kurchatov Sq. 1, 123182 Moscow, Russia
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6
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Yoshimatsu S, Kisu I, Qian E, Noce T. A New Horizon in Reproductive Research with Pluripotent Stem Cells: Successful In Vitro Gametogenesis in Rodents, Its Application to Large Animals, and Future In Vitro Reconstitution of Reproductive Organs Such as “Uteroid” and “Oviductoid”. BIOLOGY 2022; 11:biology11070987. [PMID: 36101367 PMCID: PMC9312112 DOI: 10.3390/biology11070987] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 05/28/2022] [Revised: 06/24/2022] [Accepted: 06/28/2022] [Indexed: 11/16/2022]
Abstract
Simple Summary Functional gametes, such as oocytes and spermatozoa, have been derived from rodent pluripotent stem cells, which can be applied to large animals and ultimately, to humans. In addition to summarizing these topics, we also review additional approaches for in vitro reconstitution of reproductive organs. This review illustrates intensive past efforts and future challenges on stem cell research for in vitro biogenesis in various mammalian models. Abstract Recent success in derivation of functional gametes (oocytes and spermatozoa) from pluripotent stem cells (PSCs) of rodents has made it feasible for future application to large animals including endangered species and to ultimately humans. Here, we summarize backgrounds and recent studies on in vitro gametogenesis from rodent PSCs, and similar approaches using PSCs from large animals, including livestock, nonhuman primates (NHPs), and humans. We also describe additional developing approaches for in vitro reconstitution of reproductive organs, such as the ovary (ovarioid), testis (testisoid), and future challenges in the uterus (uteroid) and oviduct (oviductoid), all of which may be derived from PSCs. Once established, these in vitro systems may serve as a robust platform for elucidating the pathology of infertility-related disorders and ectopic pregnancy, principle of reproduction, and artificial biogenesis. Therefore, these possibilities, especially when using human cells, require consideration of ethical issues, and international agreements and guidelines need to be raised before opening “Pandora’s Box”.
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Affiliation(s)
- Sho Yoshimatsu
- Department of Stem Cell Biology and Medicine, Graduate School of Medical Sciences, Kyushu University, Higashi-ku, Fukuoka 812-8582, Japan
- Research Fellow of Japan Society for the Promotion of Science (JSPS), Chiyoda-ku, Tokyo 102-0083, Japan
- Department of Physiology, Keio University School of Medicine, Shinjuku-ku, Tokyo 160-8582, Japan;
- Laboratory for Marmoset Neural Architecture, RIKEN Center for Brain Science, Wako-City 351-0198, Japan;
- Correspondence:
| | - Iori Kisu
- Department of Obstetrics and Gynecology, Keio University School of Medicine, Shinjuku-ku, Tokyo 160-8582, Japan;
| | - Emi Qian
- Department of Physiology, Keio University School of Medicine, Shinjuku-ku, Tokyo 160-8582, Japan;
| | - Toshiaki Noce
- Laboratory for Marmoset Neural Architecture, RIKEN Center for Brain Science, Wako-City 351-0198, Japan;
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7
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Vasa Is a Potential Germ Cell Marker in Leopard Coral Grouper ( Plectropomus leopardus). Genes (Basel) 2022; 13:genes13061077. [PMID: 35741839 PMCID: PMC9222667 DOI: 10.3390/genes13061077] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/23/2022] [Revised: 05/23/2022] [Accepted: 06/13/2022] [Indexed: 11/16/2022] Open
Abstract
Vasa (Ddx4, DEAD box polypeptide 4), an extremely specific marker of germ cells in vivo, is an ATP-dependent RNA helicase that plays an essential role in germ cell development and gametogenesis. However, the expression and function information about this gene in groupers remains lacking. Here, vasa homolog termed Plvasa was isolated and identified Plvasa as a putative germ cell marker in the leopard coral grouper, (Plectropomus leopardus). Results indicated that Plvasa contained 17 exons in the genomic sequence and 9 conserved motifs of the DEAD-box protein by sequence analysis. The sequence comparison, phylogenetic analyses and synteny analyses showed that Plvasa was homologous with other teleosts. Additionally, the expression of Plvasa was significantly higher in gonads than in other tissues in adult individuals (p < 0.05). Further, the distribution of Plvasa revealed that it was only expressed in the germ cells, such as spermatids, germline stem cells and oocytes at different stages, and could not be detected in the somatic cells of gonads. The current study verified that the Plvasa gene is a valuable molecular marker of germ cells in leopard coral grouper, which potentially plays an important role in investigating the genesis and development of teleost germ cells.
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8
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Milani L, Cinelli F, Iannello M, Lazzari M, Franceschini V, Maurizii MG. Immunolocalization of Vasa, PIWI, and TDRKH proteins in male germ cells during spermatogenesis of the teleost fish Poecilia reticulata. Acta Histochem 2022; 124:151870. [PMID: 35218995 DOI: 10.1016/j.acthis.2022.151870] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/15/2021] [Revised: 02/12/2022] [Accepted: 02/14/2022] [Indexed: 11/17/2022]
Abstract
Vasa, PIWI and TDRKH are conserved components of germ granules that in metazoans are involved in germline specification and differentiation, as documented by mutational experiments in some model animals. So far, investigations on PIWI during spermatogenesis of fish has been limited to a few species, and no information is available for TDRKH, another protein involved in the piRNA pathway. In this study, the immunolocalization of these three germline determinants was analyzed in male gonads of the teleost fish Poecilia reticulata to document their localization pattern in the different stages of germ cell differentiation. To analyze their distribution pattern during the different stages of spermatogenesis we performed immunohistochemistry (IHC) and immunofluorescence (IF) assays using primary polyclonal antibodies after testing their specificity with Western Blot. Moreover, sections of testis stained with haematoxylin and eosin clarified the structural organization of P. reticulata testis, while the use of the confocal microscope and the nuclear staining clarified the different stages of germ cell differentiation during spermatogenesis. The results showed that Vasa, PIWI and TDRKH were specifically immunolocalized in the germ cells of P. reticulata, with no specific signal detected in Sertoli cells and in other somatic cells of the gonad. These markers were detected in all stages of differentiation from early spermatogonia to advanced spermatids. Vasa staining was the strongest in spermatogonia, and then decreases throughout differentiation. Instead, both PIWI and TDRKH staining increases during differentiation, and their distribution pattern, similar to what observed in the mouse, suggests their concerted participation in the piRNA pathway also in this fish.
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Affiliation(s)
- L Milani
- Department of Biological, Geological, and Environmental Sciences, University of Bologna, Bologna, Italy.
| | - F Cinelli
- Department of Biological, Geological, and Environmental Sciences, University of Bologna, Bologna, Italy
| | - M Iannello
- Department of Biological, Geological, and Environmental Sciences, University of Bologna, Bologna, Italy
| | - M Lazzari
- Department of Biological, Geological, and Environmental Sciences, University of Bologna, Bologna, Italy
| | - V Franceschini
- Department of Biological, Geological, and Environmental Sciences, University of Bologna, Bologna, Italy
| | - M G Maurizii
- Department of Biological, Geological, and Environmental Sciences, University of Bologna, Bologna, Italy.
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9
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Pou5f1 and Nanog Are Reliable Germ Cell-Specific Genes in Gonad of a Protogynous Hermaphroditic Fish, Orange-Spotted Grouper (Epinephelus coioides). Genes (Basel) 2021; 13:genes13010079. [PMID: 35052423 PMCID: PMC8774525 DOI: 10.3390/genes13010079] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/25/2021] [Revised: 12/12/2021] [Accepted: 12/25/2021] [Indexed: 01/06/2023] Open
Abstract
Pluripotency markers Pou5f1 and Nanog are core transcription factors regulating early embryonic development and maintaining the pluripotency and self-renewal of stem cells. Pou5f1 and Nanog also play important roles in germ cell development and gametogenesis. In this study, Pou5f1 (EcPou5f1) and Nanog (EcNanog) were cloned from orange-spotted grouper, Epinephelus coioides. The full-length cDNAs of EcPou5f1 and EcNanog were 2790 and 1820 bp, and encoded 475 and 432 amino acids, respectively. EcPou5f1 exhibited a specific expression in gonads, whereas EcNanog was expressed highly in gonads and weakly in some somatic tissues. In situ hybridization analyses showed that the mRNA signals of EcNanog and EcPou5f1 were exclusively restricted to germ cells in gonads. Likewise, immunohistofluorescence staining revealed that EcNanog protein was limited to germ cells. Moreover, both EcPou5f1 and EcNanog mRNAs were discovered to be co-localized with Vasa mRNA, a well-known germ cell maker, in male and female germ cells. These results implied that EcPou5f1 and EcNanog could be also regarded as reliable germ cell marker genes. Therefore, the findings of this study would pave the way for elucidating the mechanism whereby EcPou5f1 and EcNanog regulate germ cell development and gametogenesis in grouper fish, and even in other protogynous hermaphroditic species.
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10
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Jang SW, Choi HW. Generation of Miniaturized Ovaries by In Vitro Culture from Mouse Gonads. Dev Reprod 2021; 25:173-183. [PMID: 34950820 PMCID: PMC8670777 DOI: 10.12717/dr.2020.25.3.173] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/16/2021] [Revised: 07/19/2021] [Accepted: 08/17/2021] [Indexed: 11/17/2022]
Abstract
The incidence of infertility among individuals of reproductive age has been
growing due to genetic and environmental factors, and considerable research
efforts are focused on solving this issue. Ovarian development is an overly
complex process in the body, involving the interaction between primordial germ
cells and gonad somatic cells. However, follicles located in the center of the
in vitro ovary are poorly formed owing to ovarian
complexity, nutrient deficiency, and signaling deficiency. In the present study,
we optimized methods for dissociating gonads and culture conditions for the
in vitro generation of miniaturized ovaries. The gonads
from embryos were dissociated into cell masses and cultured on a Transwell-COL
membrane for 3–5 weeks. Approximately 12 follicles were present per
in vitro ovary. We observed that miniaturized ovaries
successfully matured to MII oocytes in vitro from 150 to 100
µm gonad masses. This method will be useful for investigating follicle
development and oocyte production.
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Affiliation(s)
- Si Won Jang
- Dept. of Agricultural Convergence Technology, Jeonbuk National University, Jeonju 54896, Korea
| | - Hyun Woo Choi
- Dept. of Agricultural Convergence Technology, Jeonbuk National University, Jeonju 54896, Korea.,Dept. of Animal Science, Jeonbuk National University, Jeonju 54896, Korea
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11
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Generation of offspring-producing 3D ovarian organoids derived from female germline stem cells and their application in toxicological detection. Biomaterials 2021; 279:121213. [PMID: 34715637 DOI: 10.1016/j.biomaterials.2021.121213] [Citation(s) in RCA: 20] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/26/2020] [Revised: 09/23/2021] [Accepted: 10/20/2021] [Indexed: 12/21/2022]
Abstract
In vitro production of oocytes capable of producing offspring has exciting potential applications in reproductive medicine. Here, we generated and characterized an ovarian organoid model derived from female germline stem cells using a three-dimensional culture system. We show that this model generated normal offspring and detected drug toxicity. The ovarian organoids could produce oocytes and exhibited endocrine functions. Single-cell analysis of ovarian organoids identified six ovarian cell lineages, such as germ, granulosa and theca cells, and produced gene-expression signatures for each cell type. Investigation of the expression patterns of genes related to meiosis and gene ontogeny analysis for germ cell clusters showed that a germ cell population was maintained in the ovarian organoids. Moreover, flow cytometric analysis confirmed that the population of germ cells could be maintained on the organoids and showed that ascorbic acid treatment had a beneficial effect of germ cell population maintenance on the organoids. Furthermore, we demonstrated the successful production of offspring from oocytes derived from ovarian organoids. Finally, we showed the ovarian organoids had the potential to drug toxicological detection. For example, we found that salinomycin impaired the formation of ovarian organoids and germ cell population maintenance by inducing apoptosis. These results indicate that the female germline stem cell-derived ovarian organoids represent a valuable model system for generating oocytes that can yield offspring, and provide a novel model for drug screening and toxicological detection.
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12
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Blondel L, Besse S, Rivard EL, Ylla G, Extavour CG. Evolution of a cytoplasmic determinant: evidence for the biochemical basis of functional evolution of the novel germ line regulator oskar. Mol Biol Evol 2021; 38:5491-5513. [PMID: 34550378 PMCID: PMC8662646 DOI: 10.1093/molbev/msab284] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/30/2022] Open
Abstract
Germ line specification is essential in sexually reproducing organisms. Despite their critical role, the evolutionary history of the genes that specify animal germ cells is heterogeneous and dynamic. In many insects, the gene oskar is required for the specification of the germ line. However, the germ line role of oskar is thought to be a derived role resulting from co-option from an ancestral somatic role. To address how evolutionary changes in protein sequence could have led to changes in the function of Oskar protein that enabled it to regulate germ line specification, we searched for oskar orthologs in 1,565 publicly available insect genomic and transcriptomic data sets. The earliest-diverging lineage in which we identified an oskar ortholog was the order Zygentoma (silverfish and firebrats), suggesting that oskar originated before the origin of winged insects. We noted some order-specific trends in oskar sequence evolution, including whole gene duplications, clade-specific losses, and rapid divergence. An alignment of all known 379 Oskar sequences revealed new highly conserved residues as candidates that promote dimerization of the LOTUS domain. Moreover, we identified regions of the OSK domain with conserved predicted RNA binding potential. Furthermore, we show that despite a low overall amino acid conservation, the LOTUS domain shows higher conservation of predicted secondary structure than the OSK domain. Finally, we suggest new key amino acids in the LOTUS domain that may be involved in the previously reported Oskar−Vasa physical interaction that is required for its germ line role.
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Affiliation(s)
- Leo Blondel
- Department of Molecular and Cellular Biology, Harvard University, Cambridge, MA, USA
| | - Savandara Besse
- Department of Molecular and Cellular Biology, Harvard University, Cambridge, MA, USA
| | - Emily L Rivard
- Department of Molecular and Cellular Biology, Harvard University, Cambridge, MA, USA
| | - Guillem Ylla
- Department of Organismic and Evolutionary Biology, Harvard University, Cambridge, MA, USA
| | - Cassandra G Extavour
- Department of Molecular and Cellular Biology, Harvard University, Cambridge, MA, USA.,Department of Organismic and Evolutionary Biology, Harvard University, Cambridge, MA, USA
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13
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Hansen CL, Pelegri F. Primordial Germ Cell Specification in Vertebrate Embryos: Phylogenetic Distribution and Conserved Molecular Features of Preformation and Induction. Front Cell Dev Biol 2021; 9:730332. [PMID: 34604230 PMCID: PMC8481613 DOI: 10.3389/fcell.2021.730332] [Citation(s) in RCA: 16] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/24/2021] [Accepted: 08/25/2021] [Indexed: 11/24/2022] Open
Abstract
The differentiation of primordial germ cells (PGCs) occurs during early embryonic development and is critical for the survival and fitness of sexually reproducing species. Here, we review the two main mechanisms of PGC specification, induction, and preformation, in the context of four model vertebrate species: mouse, axolotl, Xenopus frogs, and zebrafish. We additionally discuss some notable molecular characteristics shared across PGC specification pathways, including the shared expression of products from three conserved germline gene families, DAZ (Deleted in Azoospermia) genes, nanos-related genes, and DEAD-box RNA helicases. Then, we summarize the current state of knowledge of the distribution of germ cell determination systems across kingdom Animalia, with particular attention to vertebrate species, but include several categories of invertebrates - ranging from the "proto-vertebrate" cephalochordates to arthropods, cnidarians, and ctenophores. We also briefly highlight ongoing investigations and potential lines of inquiry that aim to understand the evolutionary relationships between these modes of specification.
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Affiliation(s)
| | - Francisco Pelegri
- Laboratory of Genetics, University of Wisconsin-Madison, Madison, WI, United States
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Ariyath A, Kanthi JM, Paul-Prasanth B. Differentiation Potential of Cultured Extracellular DEAD-Box Helicase 4+ Oogonial Stem Cells from Adult Human Ovaries into Somatic Lineages. Cells Tissues Organs 2021; 211:577-588. [PMID: 34412061 DOI: 10.1159/000519087] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/27/2021] [Accepted: 08/13/2021] [Indexed: 11/19/2022] Open
Affiliation(s)
- Ajish Ariyath
- Centre for Nanosciences and Molecular Medicine, Amrita Vishwa Vidyapeetham, Kochi, India
| | - Janu Mangala Kanthi
- Department of Gynaecology and Obstetrics, Amrita Institute of Medical Sciences, Amrita Vishwa Vidyapeetham, Kochi, India
| | - Bindhu Paul-Prasanth
- Centre for Nanosciences and Molecular Medicine, Amrita Vishwa Vidyapeetham, Kochi, India
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15
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Li Y, Zhang Y, Liu M. Knockout Gene-Based Evidence for PIWI-Interacting RNA Pathway in Mammals. Front Cell Dev Biol 2021; 9:681188. [PMID: 34336834 PMCID: PMC8317503 DOI: 10.3389/fcell.2021.681188] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/16/2021] [Accepted: 06/08/2021] [Indexed: 01/05/2023] Open
Abstract
The PIWI-interacting RNA (piRNA) pathway mainly consists of evolutionarily conserved protein factors. Intriguingly, many mutations of piRNA pathway factors lead to meiotic arrest during spermatogenesis. The majority of piRNA factor-knockout animals show arrested meiosis in spermatogenesis, and only a few show post-meiosis male germ cell arrest. It is still unclear whether the majority of piRNA factors expressed in spermatids are involved in long interspersed nuclear element-1 repression after meiosis, but future conditional knockout research is expected to resolve this. In addition, recent hamster knockout studies showed that a piRNA factor is necessary for oocytes-in complete contrast to the findings in mice. This species discrepancy allows researchers to reexamine the function of piRNA in female germ cells. This mini-review focuses on the current knowledge of protein factors derived from mammalian knockout studies and summarizes their roles in the biogenesis and function of piRNAs.
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Affiliation(s)
- Yinuo Li
- State Key Laboratory of Reproductive Medicine, Department of Histology and Embryology, School of Basic Medical Sciences, Nanjing Medical University, Nanjing, China
| | - Yue Zhang
- State Key Laboratory of Reproductive Medicine, Clinical Center of Reproductive Medicine, The First Affiliated Hospital of Nanjing Medical University, Nanjing, China
| | - Mingxi Liu
- State Key Laboratory of Reproductive Medicine, Department of Histology and Embryology, School of Basic Medical Sciences, Nanjing Medical University, Nanjing, China
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Cai YT, Xiong CL, Liu TS, Shen SL, Rao JP, Qiu F. Secretions released from mesenchymal stem cells improve spermatogenesis restoration of cytotoxic treatment with busulfan in azoospermia mice. Andrologia 2021; 53:e14144. [PMID: 34143903 DOI: 10.1111/and.14144] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/08/2021] [Revised: 05/21/2021] [Accepted: 05/23/2021] [Indexed: 12/25/2022] Open
Abstract
This study aimed at the efficacy of sequential treatment of bone marrow-derived mesenchymal stem cell secretion for busulfan-treated azoospermia in mice. The conditioned media (CM) was obtained from bone marrow mesenchymal stem cells (MSCs) or 293 cells. Chemically induced azoospermia mice received 200 μl MSC-CM or 293-CM twice a week intravenously for three consecutive weeks. The histological assessment of spermatogenic recovery quantifying the expression of meiosis-associated genes, and Sertoli cell barrier functional factors were assessed. The characteristics of TM4 cells (Sertoli cell line) after pre-incubation of MSC-CM in vitro were also obtained. The MSC-CM group had the most spermatogenic colonies among the three groups (p < .05), but no spermatids were seen. Expressions of the meiosis-associated genes Dazl, Vasa, Miwi, Stra8, CyclinA1, Pgk2 and Scp3 in MSC-CM testis were remarkably higher compared with 293-CM and busulfan groups respectively (p < .05). The levels of Sertoli cell barrier functional factors, for example ICAM-1 and N-cadherin, were significantly increased during MSC-CM treatment (p < .05). Moreover, pre-incubation of MSC-CM particularly accelerated the CD54 (ICAM-1) and CD44 expressions of TM4 cells and promoted cell inherent adhesion. MSC-CM treatment can significantly improve the short-term restoration of spermatogonial structures of chemically induced azoospermia related to facilitating Sertoli cell adhesion integrity.
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Affiliation(s)
- Yi-Ting Cai
- Department of Reproductive Medicine, The Second Affiliated Hospital of Zhejiang University School of Medicine, Hangzhou, China
| | - Cheng-Liang Xiong
- Center for Reproductive Medicine, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China
| | - Tian-Shu Liu
- Department of Thoracic Surgery, The Second Affiliated Hospital of Zhejiang University School of Medicine, Hangzhou, China
| | - Shi-Liang Shen
- Center for Reproductive Medicine, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China
| | - Jin-Peng Rao
- Department of Reproductive Medicine, The Second Affiliated Hospital of Zhejiang University School of Medicine, Hangzhou, China
| | - Feng Qiu
- Department of Reproductive Medicine, The Second Affiliated Hospital of Zhejiang University School of Medicine, Hangzhou, China
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Liu X, Zhu Y, Zhao Y, Wang Y, Li W, Hong X, Yu L, Chen C, Xu H, Zhu X. Vasa expression is associated with sex differentiation in the Asian yellow pond turtle, Mauremys mutica. JOURNAL OF EXPERIMENTAL ZOOLOGY PART B-MOLECULAR AND DEVELOPMENTAL EVOLUTION 2021; 336:431-442. [PMID: 34101984 DOI: 10.1002/jez.b.23064] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 10/05/2020] [Revised: 04/10/2021] [Accepted: 05/18/2021] [Indexed: 01/04/2023]
Abstract
Vasa, one of the best-studied germ cell markers plays a critical role in germ cell development and differentiation in animals. Vasa deficiency would lead to male-specific sterility in most vertebrates, but female sterility in the fly. However, the role of the vasa gene involved in germ cell differentiation is largely elusive. Here, we first characterized the expression profile of vasa products in the Asian yellow pond turtle by quantitative reverse-transcription polymerase chain reaction and fluorescence immunostaining. The results showed that vasa messenger RNA (mRNA) is initially detected in embryos at stage 16, and then dramatically increased in embryos at stage 19. In particular, like the sex-related genes, vasa mRNA exhibited differential expression in embryos between the male-producing temperature (MPT, 25°C) and the female-producing temperature (FPT, 33°C), whereas there was no difference in methylation levels of vasa promoter detected between FPT and MPT. In contrast, in the adult Asian yellow pond, the level of vasa mRNA was much higher in the testis than ovary. Moreover, the immunostaining on testicular sections and cells showed that Vasa protein was exclusively expressed in germ cells: Weak but detectable in spermatogonia, highest in spermatocytes, moderate and concentrated in chromatid bodies in spermatids and spermatozoa, and bare in somatic cells. The expression profile of Vasa protein is similar in turtle species studied so far but distinct from those in fish species in this study. The findings of this study would provide new insights into our understanding of the conservation and divergence of the vasa gene, even other germ cell genes across phyla.
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Affiliation(s)
- Xiaoli Liu
- Key Laboratory of Tropical & Subtropical Fishery Resource Application & Cultivation of Ministry of Agriculture and Rural Affairs, Pearl River Fisheries Research Institute, Chinese Academy of Fishery Sciences, Guangzhou, China
| | - Yanyu Zhu
- Key Laboratory of Tropical & Subtropical Fishery Resource Application & Cultivation of Ministry of Agriculture and Rural Affairs, Pearl River Fisheries Research Institute, Chinese Academy of Fishery Sciences, Guangzhou, China.,Shanghai Ocean University, Shanghai, China
| | - Yanyan Zhao
- Key Laboratory of Tropical & Subtropical Fishery Resource Application & Cultivation of Ministry of Agriculture and Rural Affairs, Pearl River Fisheries Research Institute, Chinese Academy of Fishery Sciences, Guangzhou, China
| | - Yakun Wang
- Key Laboratory of Tropical & Subtropical Fishery Resource Application & Cultivation of Ministry of Agriculture and Rural Affairs, Pearl River Fisheries Research Institute, Chinese Academy of Fishery Sciences, Guangzhou, China
| | - Wei Li
- Key Laboratory of Tropical & Subtropical Fishery Resource Application & Cultivation of Ministry of Agriculture and Rural Affairs, Pearl River Fisheries Research Institute, Chinese Academy of Fishery Sciences, Guangzhou, China
| | - Xiaoyou Hong
- Key Laboratory of Tropical & Subtropical Fishery Resource Application & Cultivation of Ministry of Agriculture and Rural Affairs, Pearl River Fisheries Research Institute, Chinese Academy of Fishery Sciences, Guangzhou, China
| | - Lingyun Yu
- Key Laboratory of Tropical & Subtropical Fishery Resource Application & Cultivation of Ministry of Agriculture and Rural Affairs, Pearl River Fisheries Research Institute, Chinese Academy of Fishery Sciences, Guangzhou, China
| | - Chen Chen
- Key Laboratory of Tropical & Subtropical Fishery Resource Application & Cultivation of Ministry of Agriculture and Rural Affairs, Pearl River Fisheries Research Institute, Chinese Academy of Fishery Sciences, Guangzhou, China
| | - Hongyan Xu
- Key Laboratory of Freshwater Fish Reproduction and Development, Ministry of Education, Key Laboratory of Aquatic Sciences of Chongqing, College of Fisheries, Southwest University, Chongqing, China
| | - Xinping Zhu
- Key Laboratory of Tropical & Subtropical Fishery Resource Application & Cultivation of Ministry of Agriculture and Rural Affairs, Pearl River Fisheries Research Institute, Chinese Academy of Fishery Sciences, Guangzhou, China.,Shanghai Ocean University, Shanghai, China
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Mokdadi M, Abdelkrim YZ, Banroques J, Huvelle E, Oualha R, Yeter-Alat H, Guizani I, Barhoumi M, Tanner NK. The In Silico Identification of Potential Members of the Ded1/DDX3 Subfamily of DEAD-Box RNA Helicases from the Protozoan Parasite Leishmania infantum and Their Analyses in Yeast. Genes (Basel) 2021; 12:212. [PMID: 33535521 PMCID: PMC7912733 DOI: 10.3390/genes12020212] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/27/2020] [Revised: 01/27/2021] [Accepted: 01/29/2021] [Indexed: 12/14/2022] Open
Abstract
DEAD-box RNA helicases are ubiquitous proteins found in all kingdoms of life and that are associated with all processes involving RNA. Their central roles in biology make these proteins potential targets for therapeutic or prophylactic drugs. The Ded1/DDX3 subfamily of DEAD-box proteins is of particular interest because of their important role(s) in translation. In this paper, we identified and aligned the protein sequences of 28 different DEAD-box proteins from the kinetoplast-protozoan parasite Leishmania infantum, which is the cause of the visceral form of leishmaniasis that is often lethal if left untreated, and compared them with the consensus sequence derived from DEAD-box proteins in general, and from the Ded1/DDX3 subfamily in particular, from a wide variety of other organisms. We identified three potential homologs of the Ded1/DDX3 subfamily and the equivalent proteins from the related protozoan parasite Trypanosoma brucei, which is the causative agent of sleeping sickness. We subsequently tested these proteins for their ability to complement a yeast strain deleted for the essential DED1 gene. We found that the DEAD-box proteins from Trypanosomatids are highly divergent from other eukaryotes, and consequently they are suitable targets for protein-specific drugs.
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Affiliation(s)
- Molka Mokdadi
- Expression Génétique Microbienne, UMR8261 CNRS, Université de Paris, Institut de Biologie Physico-Chimique, 13 rue Pierre et Marie Curie, 75005 Paris, France; (M.M.); (Y.Z.A.); (J.B.); (E.H.); (H.Y.-A.)
- PSL Research University, 75005 Paris, France
- Laboratory of Molecular Epidemiology and Experimental Pathology (LR16IPT04), Institut Pasteur de Tunis, Université de Tunis El Manar, 13 Place Pasteur, BP74 Tunis-Belvédère 1002, Tunisia; (R.O.); (I.G.)
- Institut National des Sciences Appliquées et Technologies, Université de Carthage, CEDEX, Tunis 1080, Tunisia
| | - Yosser Zina Abdelkrim
- Expression Génétique Microbienne, UMR8261 CNRS, Université de Paris, Institut de Biologie Physico-Chimique, 13 rue Pierre et Marie Curie, 75005 Paris, France; (M.M.); (Y.Z.A.); (J.B.); (E.H.); (H.Y.-A.)
- Laboratory of Molecular Epidemiology and Experimental Pathology (LR16IPT04), Institut Pasteur de Tunis, Université de Tunis El Manar, 13 Place Pasteur, BP74 Tunis-Belvédère 1002, Tunisia; (R.O.); (I.G.)
| | - Josette Banroques
- Expression Génétique Microbienne, UMR8261 CNRS, Université de Paris, Institut de Biologie Physico-Chimique, 13 rue Pierre et Marie Curie, 75005 Paris, France; (M.M.); (Y.Z.A.); (J.B.); (E.H.); (H.Y.-A.)
- PSL Research University, 75005 Paris, France
| | - Emmeline Huvelle
- Expression Génétique Microbienne, UMR8261 CNRS, Université de Paris, Institut de Biologie Physico-Chimique, 13 rue Pierre et Marie Curie, 75005 Paris, France; (M.M.); (Y.Z.A.); (J.B.); (E.H.); (H.Y.-A.)
- PSL Research University, 75005 Paris, France
| | - Rafeh Oualha
- Laboratory of Molecular Epidemiology and Experimental Pathology (LR16IPT04), Institut Pasteur de Tunis, Université de Tunis El Manar, 13 Place Pasteur, BP74 Tunis-Belvédère 1002, Tunisia; (R.O.); (I.G.)
| | - Hilal Yeter-Alat
- Expression Génétique Microbienne, UMR8261 CNRS, Université de Paris, Institut de Biologie Physico-Chimique, 13 rue Pierre et Marie Curie, 75005 Paris, France; (M.M.); (Y.Z.A.); (J.B.); (E.H.); (H.Y.-A.)
- PSL Research University, 75005 Paris, France
| | - Ikram Guizani
- Laboratory of Molecular Epidemiology and Experimental Pathology (LR16IPT04), Institut Pasteur de Tunis, Université de Tunis El Manar, 13 Place Pasteur, BP74 Tunis-Belvédère 1002, Tunisia; (R.O.); (I.G.)
| | - Mourad Barhoumi
- Laboratory of Molecular Epidemiology and Experimental Pathology (LR16IPT04), Institut Pasteur de Tunis, Université de Tunis El Manar, 13 Place Pasteur, BP74 Tunis-Belvédère 1002, Tunisia; (R.O.); (I.G.)
| | - N. Kyle Tanner
- Expression Génétique Microbienne, UMR8261 CNRS, Université de Paris, Institut de Biologie Physico-Chimique, 13 rue Pierre et Marie Curie, 75005 Paris, France; (M.M.); (Y.Z.A.); (J.B.); (E.H.); (H.Y.-A.)
- PSL Research University, 75005 Paris, France
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19
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In vitro cytotoxicity of zinc oxide nanoparticles in mouse ovarian germ cells. Toxicol In Vitro 2021; 70:105032. [DOI: 10.1016/j.tiv.2020.105032] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/23/2020] [Revised: 09/28/2020] [Accepted: 10/15/2020] [Indexed: 01/19/2023]
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20
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Anbazhagan R, Kavarthapu R, Coon SL, Dufau ML. Role of Phosphorylated Gonadotropin-Regulated Testicular RNA Helicase (GRTH/DDX25) in the Regulation of Germ Cell Specific mRNAs in Chromatoid Bodies During Spermatogenesis. Front Cell Dev Biol 2020; 8:580019. [PMID: 33425888 PMCID: PMC7786181 DOI: 10.3389/fcell.2020.580019] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/04/2020] [Accepted: 11/11/2020] [Indexed: 12/13/2022] Open
Abstract
GRTH/DDX25 is a member of the DEAD-box family of RNA helicases that play an essential role in spermatogenesis. GRTH knock-in (KI) mice with the human mutant GRTH gene (R242H) show loss of the phospho-species from cytoplasm with preservation of the non-phospho form in the cytoplasm and nucleus. GRTH KI mice are sterile and lack elongated spermatids and spermatozoa, with spermatogenic arrest at step 8 of round spermatids which contain chromatoid body (CB) markedly reduced in size. We observed an absence of phospho-GRTH in CB of GRTH KI mice. RNA-Seq analysis of mRNA isolated from CB revealed that 1,421 genes show differential abundance, of which 947 genes showed a decrease in abundance and 474 genes showed an increase in abundance in GRTH KI mice. The transcripts related to spermatid development, differentiation, and chromatin remodeling (Tnp1/2, Prm1/2/3, Spem1/2, Tssk 2/3/6, Grth, tAce, and Upf2) were reduced, and the transcripts encoding for factors involved in RNA transport, regulation, and surveillance and transcriptional and translational regulation (Eef1a1, Ppp1cc, Pabpc1, Ybx3, Tent5b, H2al1m, Dctn2, and Dync1h1) were increased in the CB of KI mice and were further validated by qPCR. In the round spermatids of wild-type mice, mRNAs of Tnp2, Prm2, and Grth were abundantly co-localized with MVH protein in the CB, while in GRTH KI mice these were minimally present. In addition, GRTH binding to Tnp1/2, Prm1/2, Grth, and Tssk6 mRNAs was found to be markedly decreased in KI. These results demonstrate the importance of phospho-GRTH in the maintenance of the structure of CB and its role in the storage and stability of germ cell-specific mRNAs during spermiogenesis.
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Affiliation(s)
- Rajakumar Anbazhagan
- Section on Molecular Endocrinology, Division of Developmental Biology, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD, United States
| | - Raghuveer Kavarthapu
- Section on Molecular Endocrinology, Division of Developmental Biology, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD, United States
| | - Steven L Coon
- Molecular Genomics Core, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD, United States
| | - Maria L Dufau
- Section on Molecular Endocrinology, Division of Developmental Biology, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD, United States
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21
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Seyoum A, Pradhan A, Jass J, Olsson PE. Perfluorinated alkyl substances impede growth, reproduction, lipid metabolism and lifespan in Daphnia magna. THE SCIENCE OF THE TOTAL ENVIRONMENT 2020; 737:139682. [PMID: 32521362 DOI: 10.1016/j.scitotenv.2020.139682] [Citation(s) in RCA: 51] [Impact Index Per Article: 10.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 12/17/2019] [Revised: 05/18/2020] [Accepted: 05/22/2020] [Indexed: 05/15/2023]
Abstract
Per- and polyfluorinated alkyl substances (PFASs) are synthetic organofluorine compounds with unique stability accompanied with hydrophobic and lipophobic properties. Perfluorooctane sulfonate (PFOS) and Perfluorooctanoic acid (PFOA) are of high concern due to their wide application in consumer and industrial products, extreme persistence, abundant occurrence in the environment and their toxic effect to humans and animals. However, knowledge on the molecular mechanisms of toxicity and the effects on reproduction output remain scarce. In this study, we analyzed the effects of PFOS and PFOA on Daphnia magna. Acute toxicity, development, reproduction, lipid metabolism (lipid-accumulation) and lifespan was investigated, as well as the expression of genes related to these endpoints. Exposure of PFOS and PFOA at 1, 10 and 25 μM did not cause acute lethality. Hatching was reduced following exposure to both compounds, and lifespan was decreased following exposure to 25 μM PFOS. Body length of Daphnia magna was reduced significantly by 25 μM PFOS following 7 days exposure. Lipid staining revealed that all PFAS exposures increased lipid accumulation. qRT-PCR analysis of genes involved in lipid metabolism suggests that the increase in lipid content could be due to inhibition of genes involved on absorption and catabolism of fatty acids. Exposure to both PFOA and PFOS reduced the fecundity significantly. Downregulation of genes involved in development and reproductive process, including vtg2, vasa, EcRA, EcRB, usp, jhe, HR3, ftz-F1, E74 and E75 were observed. The alterations in developmental and reproductive genes as well as the disturbed lipid metabolism provides mechanistic insight into the possible causes for decreased fecundity and lifespan observed following exposure to both PFOS and PFOA.
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Affiliation(s)
- Asmerom Seyoum
- The Life Science Center-Biology, School of Science and Technology, Örebro University, SE-701 82 Örebro, Sweden
| | - Ajay Pradhan
- The Life Science Center-Biology, School of Science and Technology, Örebro University, SE-701 82 Örebro, Sweden
| | - Jana Jass
- The Life Science Center-Biology, School of Science and Technology, Örebro University, SE-701 82 Örebro, Sweden
| | - Per-Erik Olsson
- The Life Science Center-Biology, School of Science and Technology, Örebro University, SE-701 82 Örebro, Sweden.
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Lee S, Kim S, Ahn J, Park J, Ryu BY, Park JY. Membrane-bottomed microwell array added to Transwell insert to facilitate non-contact co-culture of spermatogonial stem cell and STO feeder cell. Biofabrication 2020; 12:045031. [DOI: 10.1088/1758-5090/abb529] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/31/2022]
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Melo LH, Melo RMC, Luz RK, Bazzoli N, Rizzo E. Expression of Vasa, Nanos2 and Sox9 during initial testicular development in Nile tilapia (Oreochromis niloticus) submitted to sex reversal. Reprod Fertil Dev 2020; 31:1637-1646. [PMID: 31097079 DOI: 10.1071/rd18488] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/01/2018] [Accepted: 04/28/2019] [Indexed: 11/23/2022] Open
Abstract
Sexual differentiation and early gonadal development are critical events in vertebrate reproduction. In this study, the initial testis development and expression of the Vasa, Nanos2 and Sox9 proteins were examined in Nile tilapia Oreochromis niloticus submitted to induced sex reversal. To that end, 150O. niloticus larvae at 5 days post-hatching (dph) were kept in nurseries with no hormonal addition (control group) and 150 larvae were kept with feed containing 17α-methyltestosterone to induce male sex reversal (treated group). Morphological sexual differentiation of Nile tilapia occurred between 21 and 25 dph and sex reversal resulted in 94% males, whereas the control group presented 53% males. During sexual differentiation, gonocytes (Gon) were the predominant germ cells, which decreased and disappeared after that stage in both groups. Undifferentiated spermatogonia (Aund) were identified at 21 dph in the control group and at 23 dph in the treated group. Differentiated spermatogonia (Adiff) were found at 23 dph in both groups. Vasa and Nanos2 occurred in Gon, Aund and Adiff and there were no significant differences between groups. Vasa-labelled Adiff increased at 50 dph in both groups and Nanos2 presented a high proportion of labelled germ cells during sampling. Sertoli cells expressed Sox9 throughout the experiment and its expression was significantly greater during sexual differentiation in the control group. The results indicate that hormonal treatment did not alter initial testis development and expression of Vasa and Nanos2 in Nile tilapia, although lower expression of Sox9 and a delay in sexual differentiation was detected in the treated group.
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Affiliation(s)
- Luis H Melo
- Departamento de Morfologia, Instituto de Ciências Biológicas, Universidade Federal de Minas Gerais, UFMG, Av. Antônio Carlos 6627, 31270-901 Belo Horizonte, Minas Gerais, Brazil
| | - Rafael M C Melo
- Departamento de Morfologia, Instituto de Ciências Biológicas, Universidade Federal de Minas Gerais, UFMG, Av. Antônio Carlos 6627, 31270-901 Belo Horizonte, Minas Gerais, Brazil
| | - Ronald K Luz
- Laboratório de Aquacultura, Escola de Veterinária, Universidade Federal de Minas Gerais, Av. Antônio Carlos 6627, 31270-901 Belo Horizonte, MG, Brazil
| | - Nilo Bazzoli
- Programa de Pós-Graduação em Biologia de Vertebrados, Pontifícia Universidade Católica de Minas Gerais, PUC Minas, Av. Dom José Gaspar 500, 30535-610 Belo Horizonte, Minas Gerais, Brazil
| | - Elizete Rizzo
- Departamento de Morfologia, Instituto de Ciências Biológicas, Universidade Federal de Minas Gerais, UFMG, Av. Antônio Carlos 6627, 31270-901 Belo Horizonte, Minas Gerais, Brazil; and Corresponding author.
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24
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Javadi A, Mokhtari S, Moraveji SF, Sayahpour FA, Farzaneh M, Gourabi H, Esfandiari F. Short time exposure to low concentration of zinc oxide nanoparticles up-regulates self-renewal and spermatogenesis-related gene expression. Int J Biochem Cell Biol 2020; 127:105822. [PMID: 32771442 DOI: 10.1016/j.biocel.2020.105822] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/13/2020] [Revised: 07/21/2020] [Accepted: 07/31/2020] [Indexed: 12/16/2022]
Abstract
Extensive application of zinc oxide (ZnO) nanoparticles (NPs) in everyday life results in increased exposure to these NPs. Spermatogonial stem cells (SSCs) guarantee sperm production throughout the male reproductive life by providing a balance between self-renewal and differentiation. We used an in vitro platform to investigate the ZnO NPs effects on SSCs. We successfully synthesized ZnO NPs. In order to investigate these NPs, we isolated SSCs from mouse testes and cultured them in vitro. Our results confirmed the uptake of ZnO NPs by the cultured SSCs. We observed a dose- and time-dependent decrease in SSC viability. Both spherical and nanosheet ZnO NPs had the same cytotoxic effects on the SSCs, irrespective of their shapes. Moreover, we have shown that short time (one day) exposure of SSCs to a low concentration of ZnO NPs (10 μg/mL) promoted expressions of specific genes (Plzf, Gfr α1 and Bcl6b) for SSC self-renewal and differentiation genes (Vasa, Dazl, C-kit and Sycp3) expressed by spermatogonia during spermatogenesis. Our study provides the first insight into ZnO NPs function in SSCs and suggests a new function for ZnO NPs in the male reproductive system. We demonstrated that ZnO NPs might promote spermatogenesis via upregulation of gene expression related to SSC self-renewal and differentiation at low concentrations. Additional research should clarify the possible effect of ZnO NPs on the SSC genome and its effects on human SSCs.
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Affiliation(s)
- Azam Javadi
- Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran; Department of Genetics, Faculty of Basic Science and Advanced Technologies, University of Science and Culture, Tehran, Iran
| | - Saadat Mokhtari
- Department of Physics, Shahid Beheshti University, Tehran, Iran
| | - Seyedeh-Faezeh Moraveji
- Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran
| | - Forough-Azam Sayahpour
- Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran
| | - Maryam Farzaneh
- Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran; Department of Molecular and Cellular Biology, Faculty of Basic Science and Advanced Technologies, University of Science and Culture, Tehran, Iran
| | - Hamid Gourabi
- Department of Genetics, Reproductive Biomedicine Research Center, Royan Institute for Reproductive Biomedicine, ACECR, Tehran, Iran.
| | - Fereshteh Esfandiari
- Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran.
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Jin K, Li D, Jin J, Song J, Zhang Y, Chang G, Chen G, Li B. C1EIP Functions as an Activator of ENO1 to Promote Chicken PGCs Formation via Inhibition of the Notch Signaling Pathway. Front Genet 2020; 11:751. [PMID: 32849782 PMCID: PMC7396672 DOI: 10.3389/fgene.2020.00751] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/23/2020] [Accepted: 06/23/2020] [Indexed: 12/14/2022] Open
Abstract
The production of germ cells, especially primordial germ cells (PGCs), is important for avian stem cells and reproduction biology. However, key factors involved in the regulation of PGCs remain unknown. Here, we report a PGC-related marker gene: C1EIP (Chromosome 1 Expression in PGCs), whose activation and expression are regulated by the transcription factor STAT3 (signal transducer and activator of transcription 3), histone acetylation, and promoter methylation. C1EIP regulates PGCs formation by mediating the expression of PGC-associated genes, such as CVH (Chicken Vasa Homologous) and CKIT (Chicken KIT proto-oncogene). C1EIP knockdown during embryonic development reduces PGC generation efficiency both in vitro and in ovo. Conversely, C1EIP overexpression increases the formation efficiency of PGCs. C1EIP encodes a cytoplasmic protein that interacts with ENO1 (Enolase 1) in the cytoplasm, inhibits the Notch signaling pathway, and positively regulates PGC generation. Collectively, our findings demonstrate C1EIP as a novel gene involved in PGC formation, which regulates genes involved in embryonic stem cell differentiation through interaction with ENO1 and subsequent inhibition of the Notch signaling pathway by the impression of Myc (MYC proto-oncogene).
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Affiliation(s)
- Kai Jin
- Key Laboratory of Animal Breeding Reproduction and Molecular Design for Jiangsu Province, College of Animal Science and Technology, Yangzhou University, Yangzhou, China.,Institutes of Agricultural Science and Technology Development, Yangzhou University, Yangzhou, China.,Joint International Research Laboratory of Agriculture and Agri-Product Safety of Ministry of Education of China, Yangzhou University, Yangzhou, China
| | - Dong Li
- Reproductive Medicine Center, The Affiliated Drum Tower Hospital of Nanjing University, Medical School, Nanjing, China
| | - Jing Jin
- Key Laboratory of Animal Breeding Reproduction and Molecular Design for Jiangsu Province, College of Animal Science and Technology, Yangzhou University, Yangzhou, China.,Institutes of Agricultural Science and Technology Development, Yangzhou University, Yangzhou, China.,Joint International Research Laboratory of Agriculture and Agri-Product Safety of Ministry of Education of China, Yangzhou University, Yangzhou, China
| | - Jiuzhou Song
- Animal & Avian Sciences, University of Maryland, College Park, College Park, MD, United States
| | - Yani Zhang
- Key Laboratory of Animal Breeding Reproduction and Molecular Design for Jiangsu Province, College of Animal Science and Technology, Yangzhou University, Yangzhou, China.,Institutes of Agricultural Science and Technology Development, Yangzhou University, Yangzhou, China.,Joint International Research Laboratory of Agriculture and Agri-Product Safety of Ministry of Education of China, Yangzhou University, Yangzhou, China
| | - Guobing Chang
- Key Laboratory of Animal Breeding Reproduction and Molecular Design for Jiangsu Province, College of Animal Science and Technology, Yangzhou University, Yangzhou, China.,Institutes of Agricultural Science and Technology Development, Yangzhou University, Yangzhou, China.,Joint International Research Laboratory of Agriculture and Agri-Product Safety of Ministry of Education of China, Yangzhou University, Yangzhou, China
| | - Guohong Chen
- Key Laboratory of Animal Breeding Reproduction and Molecular Design for Jiangsu Province, College of Animal Science and Technology, Yangzhou University, Yangzhou, China.,Institutes of Agricultural Science and Technology Development, Yangzhou University, Yangzhou, China.,Joint International Research Laboratory of Agriculture and Agri-Product Safety of Ministry of Education of China, Yangzhou University, Yangzhou, China
| | - Bichun Li
- Key Laboratory of Animal Breeding Reproduction and Molecular Design for Jiangsu Province, College of Animal Science and Technology, Yangzhou University, Yangzhou, China.,Institutes of Agricultural Science and Technology Development, Yangzhou University, Yangzhou, China.,Joint International Research Laboratory of Agriculture and Agri-Product Safety of Ministry of Education of China, Yangzhou University, Yangzhou, China
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Malik HN, Singhal DK, Saini S, Malakar D. Derivation of oocyte-like cells from putative embryonic stem cells and parthenogenetically activated into blastocysts in goat. Sci Rep 2020; 10:10086. [PMID: 32572061 PMCID: PMC7308273 DOI: 10.1038/s41598-020-66609-2] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/22/2019] [Accepted: 05/21/2020] [Indexed: 02/02/2023] Open
Abstract
Germ cells are responsible for the propagation of live animals from generation to generation, but to surprise, a steep increase in infertile problems among livestock poses great threat for economic development of human race. An alternative and robust approach is essential to combat these ailments. Here, we demonstrate that goat putative embryonic stem cells (ESCs) were successfully in vitro differentiated into primordial germ cells and oocyte-like cells using bone morphogenetic protein-4 (BMP-4) and trans-retinoic acid (RA). Oocyte-like cells having distinct zonapellucida recruited adjacent somatic cells in differentiating culture to form cumulus-oocyte complexes (COCs). The putative COCs were found to express the zonapellucida specific (ZP1 and ZP2) and oocyte-specific markers. Primordial germ cell-specific markers VASA, DAZL, STELLA, and PUM1 were detected at protein and mRNA level. In addition to that, the surface architecture of these putative COCs was thoroughly visualized by the scanning electron microscope. The putative COCs were further parthenogenetically activated to develop into healthy morula, blastocysts and hatched blastocyst stage like embryos. Our findings may contribute to the fundamental understanding of mammalian germ cell biology and may provide clinical insights regarding infertility ailments.
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Affiliation(s)
- Hruda Nanda Malik
- Animal Biotechnology Centre, National Dairy Research Institute, Karnal, 32001, India
| | - Dinesh Kumar Singhal
- Animal Biotechnology Centre, National Dairy Research Institute, Karnal, 32001, India
| | - Sikander Saini
- Animal Biotechnology Centre, National Dairy Research Institute, Karnal, 32001, India
| | - Dhruba Malakar
- Animal Biotechnology Centre, National Dairy Research Institute, Karnal, 32001, India.
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Sheng X, Tian C, Liu L, Wang L, Ye X, Li J, Zeng M, Liu L. Characterization of oogonia stem cells in mice by Fragilis. Protein Cell 2020; 10:825-831. [PMID: 31559571 PMCID: PMC6834537 DOI: 10.1007/s13238-019-00654-0] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/26/2022] Open
Affiliation(s)
- Xiaoyan Sheng
- State Key Laboratory of Medicinal Chemical Biology, College of Life Sciences, Nankai University, Tianjin, 300071, China
- Department of Cell Biology and Genetics, College of Life Sciences, Nankai University, Tianjin, 300071, China
| | - Chenglei Tian
- State Key Laboratory of Medicinal Chemical Biology, College of Life Sciences, Nankai University, Tianjin, 300071, China
- Department of Cell Biology and Genetics, College of Life Sciences, Nankai University, Tianjin, 300071, China
| | - Linlin Liu
- State Key Laboratory of Medicinal Chemical Biology, College of Life Sciences, Nankai University, Tianjin, 300071, China
- Department of Cell Biology and Genetics, College of Life Sciences, Nankai University, Tianjin, 300071, China
| | - Lingling Wang
- State Key Laboratory of Medicinal Chemical Biology, College of Life Sciences, Nankai University, Tianjin, 300071, China
| | - Xiaoying Ye
- State Key Laboratory of Medicinal Chemical Biology, College of Life Sciences, Nankai University, Tianjin, 300071, China
- Department of Cell Biology and Genetics, College of Life Sciences, Nankai University, Tianjin, 300071, China
| | - Jie Li
- State Key Laboratory of Medicinal Chemical Biology, College of Life Sciences, Nankai University, Tianjin, 300071, China
- Department of Cell Biology and Genetics, College of Life Sciences, Nankai University, Tianjin, 300071, China
| | - Ming Zeng
- State Key Laboratory of Medicinal Chemical Biology, College of Life Sciences, Nankai University, Tianjin, 300071, China
| | - Lin Liu
- State Key Laboratory of Medicinal Chemical Biology, College of Life Sciences, Nankai University, Tianjin, 300071, China.
- Department of Cell Biology and Genetics, College of Life Sciences, Nankai University, Tianjin, 300071, China.
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28
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Wagner M, Yoshihara M, Douagi I, Damdimopoulos A, Panula S, Petropoulos S, Lu H, Pettersson K, Palm K, Katayama S, Hovatta O, Kere J, Lanner F, Damdimopoulou P. Single-cell analysis of human ovarian cortex identifies distinct cell populations but no oogonial stem cells. Nat Commun 2020; 11:1147. [PMID: 32123174 PMCID: PMC7052271 DOI: 10.1038/s41467-020-14936-3] [Citation(s) in RCA: 207] [Impact Index Per Article: 41.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/22/2019] [Accepted: 02/10/2020] [Indexed: 01/05/2023] Open
Abstract
The human ovary orchestrates sex hormone production and undergoes monthly structural changes to release mature oocytes. The outer lining of the ovary (cortex) has a key role in defining fertility in women as it harbors the ovarian reserve. It has been postulated that putative oogonial stem cells exist in the ovarian cortex and that these can be captured by DDX4 antibody isolation. Here, we report single-cell transcriptomes and cell surface antigen profiles of over 24,000 cells from high quality ovarian cortex samples from 21 patients. Our data identify transcriptional profiles of six main cell types; oocytes, granulosa cells, immune cells, endothelial cells, perivascular cells, and stromal cells. Cells captured by DDX4 antibody are perivascular cells, not oogonial stem cells. Our data do not support the existence of germline stem cells in adult human ovaries, thereby reinforcing the dogma of a limited ovarian reserve.
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Affiliation(s)
- Magdalena Wagner
- Division of Obstetrics and Gynecology, Department of Clinical Science, Intervention and Technology, Karolinska Institutet and Karolinska University Hospital, Stockholm, Sweden
| | - Masahito Yoshihara
- Department of Biosciences and Nutrition, Karolinska Institutet, Stockholm, Sweden
| | - Iyadh Douagi
- Center of Hematology and Regenerative Medicine, Department of Medicine, Karolinska Institutet, Stockholm, Sweden.,Flow Cytometry Section, Research Technologies Branch, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD, USA
| | | | - Sarita Panula
- Division of Obstetrics and Gynecology, Department of Clinical Science, Intervention and Technology, Karolinska Institutet and Karolinska University Hospital, Stockholm, Sweden
| | - Sophie Petropoulos
- Division of Obstetrics and Gynecology, Department of Clinical Science, Intervention and Technology, Karolinska Institutet and Karolinska University Hospital, Stockholm, Sweden.,Department of Medicine, Centre de recherche du CHUM, University of Montreal, Montreal, Canada
| | - Haojiang Lu
- Division of Obstetrics and Gynecology, Department of Clinical Science, Intervention and Technology, Karolinska Institutet and Karolinska University Hospital, Stockholm, Sweden
| | - Karin Pettersson
- Division of Obstetrics and Gynecology, Department of Clinical Science, Intervention and Technology, Karolinska Institutet and Karolinska University Hospital, Stockholm, Sweden
| | - Kerstin Palm
- Division of Obstetrics and Gynecology, Department of Clinical Science, Intervention and Technology, Karolinska Institutet and Karolinska University Hospital, Stockholm, Sweden.,Department of Gynecology and Obstetrics, Visby hospital, Visby, Sweden
| | - Shintaro Katayama
- Department of Biosciences and Nutrition, Karolinska Institutet, Stockholm, Sweden
| | - Outi Hovatta
- Division of Obstetrics and Gynecology, Department of Clinical Science, Intervention and Technology, Karolinska Institutet and Karolinska University Hospital, Stockholm, Sweden
| | - Juha Kere
- Department of Biosciences and Nutrition, Karolinska Institutet, Stockholm, Sweden.,Folkhälsan Research Institute, Helsinki, and Stem Cells and Metabolism Research Program, University of Helsinki, Helsinki, Finland
| | - Fredrik Lanner
- Division of Obstetrics and Gynecology, Department of Clinical Science, Intervention and Technology, Karolinska Institutet and Karolinska University Hospital, Stockholm, Sweden. .,Ming Wai Lau Center for Reparative Medicine, Stockholm node, Karolinska Institutet, Stockholm, Sweden.
| | - Pauliina Damdimopoulou
- Division of Obstetrics and Gynecology, Department of Clinical Science, Intervention and Technology, Karolinska Institutet and Karolinska University Hospital, Stockholm, Sweden.
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Lejong M, Choa-Duterre M, Vanmuylder N, Louryan S. Is Vasa such a highly specific marker for primordial germ cells? A comparison of VASA and HSP90 proteins expression in young chicken embryos. Morphologie 2020; 104:20-26. [PMID: 32057659 DOI: 10.1016/j.morpho.2020.01.003] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/25/2019] [Revised: 01/20/2020] [Accepted: 01/20/2020] [Indexed: 01/21/2023]
Abstract
INTRODUCTION Primordial germ cells (PGCs) have been studied since the 19th century with several different methods. The earliest works were based on the morphological criteria of these cells associated or not with a particular staining. Different markers have been proposed in immunohistochemistry among which we can quote the Stage-specific embryonic antigene-1 (SSEA-1), the embryonic mouse antigen-1 (EMA-1) or the heat shock protein 90. Unfortunately, none of them are germline specific. The VASA protein is considered as one of the most reliable marker for PGCs by some authors with its expression being considered to limited to the germ cells. However, other studies have reported its expression in somatic cells. Here, we described the expression of the heat shock protein, HSP90, and the VASA protein in the early chick embryo. MATERIAL AND METHODS Embryos from stages Hamburger-Hamilton (HH) 19, 21 and 28 were collected. Embryos were dissected and fixed in Serra's medium. Sections were placed on slides for PAS staining and for double immunohistochemistry with HSP90 and VASA. RESULTS VASA and HSP90 expression have been observed in germ cells but as well in other cell lineages with a spatio-temporal gradient in respect to the characteristics of development of each organ. The conclusion is that VASA expression is not limited to the germ line in chick embryo.
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Affiliation(s)
- M Lejong
- Laboratoire d'anatomie, biomécanique et organogenèse, faculté de médecine, université Libre de Bruxelles, route de Lennik, 808, 1070 Bruxelles, Belgium
| | - M Choa-Duterre
- Laboratoire d'anatomie, biomécanique et organogenèse, faculté de médecine, université Libre de Bruxelles, route de Lennik, 808, 1070 Bruxelles, Belgium
| | - N Vanmuylder
- Laboratoire d'anatomie, biomécanique et organogenèse, faculté de médecine, université Libre de Bruxelles, route de Lennik, 808, 1070 Bruxelles, Belgium
| | - S Louryan
- Laboratoire d'anatomie, biomécanique et organogenèse, faculté de médecine, université Libre de Bruxelles, route de Lennik, 808, 1070 Bruxelles, Belgium.
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Disruption of Spermatogenesis and Infertility in Ataxia with Oculomotor Apraxia Type 2 (AOA2). THE CEREBELLUM 2019; 18:448-456. [PMID: 30778901 DOI: 10.1007/s12311-019-01012-w] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
Abstract
Ataxia with oculomotor apraxia type 2 (AOA2) is a rare autosomal recessive cerebellar ataxia characterized by onset between 10 and 20 years of age and a range of neurological features that include progressive cerebellar atrophy, axonal sensorimotor neuropathy, oculomotor apraxia in a majority of patients, and elevated serum alpha-fetoprotein (AFP). AOA2 is caused by mutation of the SETX gene which encodes senataxin, a DNA/RNA helicase involved in transcription regulation, RNA processing, and DNA maintenance. Disruption of senataxin in rodents led to defective spermatogenesis and sterility in males uncovering a key role for senataxin in male germ cell survival. Here, we report the first clinical and cellular evidence of impaired spermatogenesis in AOA2 patients. We assessed sperm production in three AOA2 patients and testicular pathology in one patient and compared the findings to those of Setx-knockout mice. Sperm production was impaired in all patients assessed (3/3, 100%). Analyses of testicular biopsies from an AOA2 patient recapitulate features of the histology seen in Setx-knockout mice, strongly suggesting an underlying mechanism centering on DNA-damage-mediated germ cell apoptosis. These findings support a role for senataxin in human reproductive function and highlight a novel clinical feature of AOA2 that extends the extra-neurological roles of senataxin. This raises an important reproductive counseling issue for clinicians, and fertility specialists should be aware of SETX mutations as a possible diagnosis in young male patients presenting with oligospermia or azoospermia since infertility may presage the later onset of neurological manifestations in some individuals.
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31
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Azizi H, Ranjbar M, Rahaiee S, Govahi M, Skutella T. Investigation of VASA Gene and Protein Expression in Neonate and Adult Testicular Germ Cells in Mice In Vivo and In Vitro. CELL JOURNAL 2019; 22:171-177. [PMID: 31721531 PMCID: PMC6874794 DOI: 10.22074/cellj.2020.6619] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 12/16/2018] [Accepted: 04/07/2019] [Indexed: 12/14/2022]
Abstract
Objective We aimed to examine the expression levels of the VASA gene and protein in testis sections of neonate and adult mice as well as testicular cell cultures. Materials and Methods In this experimental study, in order to investigate the expression of this germ cell marker gene in more detail, we analyzed the expression of VASA by immunocytochemistry, immunohistochemistry and fluidigm reverse transcription-polymerase chain reaction (RT-PCR). Results The immunohistochemical assays showed that the VASA protein was exclusively expressed in germ cells in the seminiferous tubules of the neonate and adult testis and not in somatic cells. VASA was not detectable in PLZF positive spermatogonial stem cells (SSCs), was weakly expressed in proliferating spermatogonia, and became abundant in spermatocytes and round spermatozoa. Counting VASA-positive cells in the seminiferous tubules of the neonate and adult testis depicted significant higher expression (P<0.05) of VASA in the adult testis in comparison to its neonate counterpart. SSC colonies were established in vitro after digestion of the testis and characterized by immunocytochemistry for CD90 and stage-specific embryonic antigens 3 (SSEA3). Immunocytochemistry confirmed that in contrast to the not detectable signal in vivo, VASA protein was strongly localized in the cytoplasm of both neonate and adult mouse SSCs under in vitro conditions. The results of Fluidigm RT-PCR revealed a significant higher expression of the germ cell gene VASA in adult SSCs in comparison to neonate SSCs in cell culture (P<0.05). Conclusion The VASA protein is, therefore, an extremely specific marker of testicular germ cell differentiation in vivo and mostly expressed in the adult testis in spermatocytes and round spermatids. The immunohistochemical signal in spermatogonia is very low. So, PLZF positive SSCs are negative for VASA in vivo, while in contrast, once isolated from the testicular niche VASA is also strongly expressed in SSCs under in vitro conditions.
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Affiliation(s)
- Hossein Azizi
- Faculty of Biotechnology, Amol University of Special Modern Technologies, Amol, Iran. Electronic Address:
| | - Mojtaba Ranjbar
- Faculty of Biotechnology, Amol University of Special Modern Technologies, Amol, Iran
| | - Somayeh Rahaiee
- Faculty of Biotechnology, Amol University of Special Modern Technologies, Amol, Iran
| | - Mostafa Govahi
- Faculty of Biotechnology, Amol University of Special Modern Technologies, Amol, Iran
| | - Thomas Skutella
- Institute for Anatomy and Cell Biology, Medical Faculty, University of Heidelberg, Im Neuenheimer Feld, Heidelberg, Germany
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A versatile toolbox for knock-in gene targeting based on the Multisite Gateway technology. PLoS One 2019; 14:e0221164. [PMID: 31454364 PMCID: PMC6711506 DOI: 10.1371/journal.pone.0221164] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/11/2019] [Accepted: 07/31/2019] [Indexed: 12/20/2022] Open
Abstract
Knock-in (KI) gene targeting can be employed for a wide range of applications in stem cell research. However, vectors for KI require multiple complicated processes for construction, including multiple times of digestion/ligation steps and extensive restriction mapping, which has imposed limitations for the robust applicability of KI gene targeting. To circumvent this issue, here we introduce versatile and systematic methods for generating KI vectors by molecular cloning. In this approach, we employed the Multisite Gateway technology, an efficient in vitro DNA recombination system using proprietary sequences and enzymes. KI vector construction exploiting these methods requires only efficient steps, such as PCR and recombination, enabling robust KI gene targeting. We show that combinatorial usage of the KI vectors generated using this method and site-specific nucleases enabled the precise integration of fluorescent protein genes in multiple loci of human and common marmoset (marmoset; Callithrix jacchus) pluripotent stem cells. The methods described here will facilitate the usage of KI technology and ultimately help to accelerate stem cell research.
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Jiang Y, Zhang Z, Cha L, Li L, Zhu D, Fang Z, He Z, Huang J, Pan Z. Resveratrol Plays a Protective Role against Premature Ovarian Failure and Prompts Female Germline Stem Cell Survival. Int J Mol Sci 2019; 20:ijms20143605. [PMID: 31340581 PMCID: PMC6678805 DOI: 10.3390/ijms20143605] [Citation(s) in RCA: 45] [Impact Index Per Article: 7.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/12/2019] [Revised: 07/09/2019] [Accepted: 07/17/2019] [Indexed: 12/12/2022] Open
Abstract
This study was designed to investigate the protective effect of resveratrol (RES) on premature ovarian failure (POF) and the proliferation of female germline stem cells (FGSCs) at the tissue and cell levels. POF mice were lavaged with RES, and POF ovaries were co-cultured with RES and/or GANT61 in vitro. FGSCs were pretreated with Busulfan and RES and/or GANT61 and co-cultured with M1 macrophages, which were pretreated with RES. The weights of mice and their ovaries, as well as their follicle number, were measured. Ovarian function, antioxidative stress, inflammation, and FGSCs survival were evaluated. RES significantly increased the weights of POF mice and their ovaries as well as the number of follicles, while it decreased the atresia rate of follicles. Higher levels of Mvh, Oct4, SOD2, GPx, and CAT were detected after treatment with RES in vivo and in vitro. RES treatment resulted in significantly lower TNF-α and IL-6 concentrations and an obviously higher IL-10 concentration in the ovaries. In FGSCs, higher Mvh, Oct4, and SOD2 concentrations and lower TNF-α, IL-6, and MDA concentrations were measured in the RES group. Blockage of the Hh signaling pathway reversed the protective effect of RES on FGSCs. In conclusion, RES effectively improved the ovarian function of the POF model and the productive capacity of FGSCs via relieving oxidative stress and inflammation and a mechanism involving the Hh signaling pathway, suggesting that RES is a potential agent against POF and can aid in the survival of FGSCs.
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Affiliation(s)
- Yu Jiang
- Medical College, Nanchang University, Nanchang 330006, Jiangxi Province, China
| | - Zhaoyuan Zhang
- Fuzhou Medical College of Nanchang University, Nanchang 344000, Jiangxi Province, China
| | - Lijun Cha
- Medical College, Nanchang University, Nanchang 330006, Jiangxi Province, China
| | - Lili Li
- Medical College, Nanchang University, Nanchang 330006, Jiangxi Province, China
| | - Dantian Zhu
- Medical College, Nanchang University, Nanchang 330006, Jiangxi Province, China
| | - Zhi Fang
- Medical College, Nanchang University, Nanchang 330006, Jiangxi Province, China
| | - Zhiqiang He
- Medical College, Nanchang University, Nanchang 330006, Jiangxi Province, China
| | - Jian Huang
- The Key Laboratory of Reproductive Physiology and Pathology of Jiangxi Provincial, Nanchang University, Nanchang 330031, Jiangxi Province, China
| | - Zezheng Pan
- Medical College, Nanchang University, Nanchang 330006, Jiangxi Province, China.
- Faculty of Basic Medical Science, Nanchang University, Nanchang 330006, Jiangxi Province, China.
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Jiang Y, Zhu D, Liu W, Qin Q, Fang Z, Pan Z. Hedgehog pathway inhibition causes primary follicle atresia and decreases female germline stem cell proliferation capacity or stemness. Stem Cell Res Ther 2019; 10:198. [PMID: 31277696 PMCID: PMC6612207 DOI: 10.1186/s13287-019-1299-5] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/04/2019] [Revised: 05/23/2019] [Accepted: 06/07/2019] [Indexed: 02/07/2023] Open
Abstract
Background Follicle depletion is one of the causes of premature ovarian failure (POF) and primary ovarian insufficiency (POI). Hence, maintenance of a certain number of female germline stem cells (FGSCs) is optimal to produce oocytes and replenish the primordial follicle pool. The mechanism that regulates proliferation or stemness of FGSCs could contribute to restoring ovarian function, but it remains uncharacterized in postnatal mammalian ovaries. This study aims to investigate the mechanism by which inhibiting the activity of the hedgehog (Hh) signaling pathway regulates follicle development and FGSC proliferation. Methods and results To understand the role of the Hh pathway in ovarian aging, we measured Hh signaling activity at different reproductive ages and the correlation between them in physiological and pathological mice. Furthermore, we evaluated the follicle number and development and the changes in FGSC proliferation or stemness after blocking the Hh pathway in vitro and in vivo. In addition, we aimed to explain one of the mechanisms for the FGSC phenotype changes induced by treatment with the Hh pathway-specific inhibitor GANT61 via oxidative stress and apoptosis. The results show that the activity of Hh signaling is decreased in the ovaries in physiological aging and POF models, which is consistent with the trend of expression levels of the germline stem cell markers Mvh and Oct4. In vitro, blocking the Hh pathway causes follicular developmental disorders and depletes ovarian germ cells and FGSCs after treating ovaries with GANT61. The proliferation or stemness of cultured primary FGSCs is reduced when Hh activity is blocked. Our results show that the antioxidative enzyme level and the ratio of Bcl-2/Bax decrease, the expression level of caspase 3 increases, the mitochondrial membrane potential is abnormal, and ROS accumulate in this system. Conclusions We observed that the inhibition of the Hh signaling pathway with GANT61 could reduce primordial follicle number and decrease FGSC reproductive capacity or stemness through oxidative damage and apoptosis. Electronic supplementary material The online version of this article (10.1186/s13287-019-1299-5) contains supplementary material, which is available to authorized users.
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Affiliation(s)
- Yu Jiang
- Medical College, Nanchang University, Nanchang, Jiangxi Province, China
| | - Dantian Zhu
- Medical College, Nanchang University, Nanchang, Jiangxi Province, China
| | - Wenfeng Liu
- Medical College, Nanchang University, Nanchang, Jiangxi Province, China
| | - Qiushi Qin
- Medical College, Nanchang University, Nanchang, Jiangxi Province, China
| | - Zhi Fang
- Medical College, Nanchang University, Nanchang, Jiangxi Province, China
| | - Zezheng Pan
- Faculty of Basic Medical Science, Jiangxi Medical College, Nanchang University, Nanchang, 330006, China. .,Medical College, Nanchang University, Nanchang, Jiangxi Province, China.
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Peng Z, Xueb G, Chen W, Xia S. Environmental inhibitors of the expression of cytochrome P450 17A1 in mammals. ENVIRONMENTAL TOXICOLOGY AND PHARMACOLOGY 2019; 69:16-25. [PMID: 30921671 DOI: 10.1016/j.etap.2019.02.007] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 10/03/2018] [Revised: 02/09/2019] [Accepted: 02/14/2019] [Indexed: 05/23/2023]
Abstract
Cytochrome P450 17A1 (CYP17A1; EC: 1.14.14.19) is a critically important bifunctional enzyme with nicotinamide adenine dinucleotide phosphate (NADPH) as its cofactor that catalyzes the formation of all endogenous androgens. Its hydroxylase activity catalyzes the 17α-hydroxylation of pregnenolone (PREG)/progesterone (P4) to 17α-OH-pregnenolone/17α-OH-progesterone, and its 17,20-lyase activity converts 17α-OH-pregnenolone/17α-OH-progesterone to dehydroepiandrosterone/androstenedione. Androgens are required for male reproductive development, so androgen deficiency resulting from CYP17A1 inhibition may lead to reproductive disorders. There has been some advances on the study of environmental chemicals inhibiting mammalian CYP17A1 expression but no related review was available so we think it now necessary to review their characteristics and inhibiting properties.
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Affiliation(s)
- Zhiheng Peng
- Department of Clinical Laboratory Center, The second Hospital of Lanzhou University, Lanzhou, Gansu 730000, China.
| | - Guoqiang Xueb
- Second Provincial People's Hospital of Gansu, Lanzou, Gansu 730000, China.
| | - Wenci Chen
- Wenzhou Hospital of Integrated Traditional Chinese and Western Medicine, Wenzhou, Zhejiang, 32500, China.
| | - Shenglong Xia
- The Second Affiliated Hospital and Yuying Children's Hospital of Wenzhou Medical University, Wenzhou, Zhejiang, 32500, China.
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36
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Ddx4 + Oogonial Stem Cells in Postmenopausal Women's Ovaries: A Controversial, Undefined Role. Cells 2019; 8:cells8070650. [PMID: 31261822 PMCID: PMC6678385 DOI: 10.3390/cells8070650] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/09/2019] [Revised: 06/13/2019] [Accepted: 06/20/2019] [Indexed: 12/18/2022] Open
Abstract
Recent studies support the existence of oogonial stem cells (OSCs) in the ovarian cortex of different mammals, including women.These cells are characterized by small size, membrane expression of DEAD(Asp-Glu-Ala-Asp)-box polypeptide-4 (Ddx4), and stemness properties (such as self-renewal and clonal expansion) as well as the ability to differentiate in vitro into oocyte-like cells. However, the discovery of OSCs contrasts with the popular theory that there is a numerically defined oocyte pool for female fertility which undergoes exhaustion with menopause. Indeed, in the ovarian cortex of postmenopausal women OSCs have been detected that possess both viability and capability to differentiate into oocytes, which is similar to those observed in younger patients. The pathophysiological role of this cell population in aged women is still debated since OSCs, under appropriate stimuli, differentiate into somatic cells, and the occurrence of Ddx4+ cells in ovarian tumor samples also suggests their potential involvement in carcinogenesis. Although further investigation into these observations is needed to clarify OSC function in ovary physiology, clinical investigators and researchers studying female infertility are presently focusing on OSCs as a novel opportunity to restore ovarian reserve in both young women undergoing early ovarian failure and cancer survivors experiencing iatrogenic menopause.
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Clarkson YL, Weatherall E, Waterfall M, McLaughlin M, Lu H, Skehel PA, Anderson RA, Telfer EE. Extracellular Localisation of the C-Terminus of DDX4 Confirmed by Immunocytochemistry and Fluorescence-Activated Cell Sorting. Cells 2019; 8:cells8060578. [PMID: 31212843 PMCID: PMC6627596 DOI: 10.3390/cells8060578] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/16/2019] [Revised: 06/06/2019] [Accepted: 06/09/2019] [Indexed: 02/03/2023] Open
Abstract
Putative oogonial stem cells (OSCs) have been isolated by fluorescence-activated cell sorting (FACS) from adult human ovarian tissue using an antibody against DEAD-box helicase 4 (DDX4). DDX4 has been reported to be germ cell specific within the gonads and localised intracellularly. White et al. (2012) hypothesised that the C-terminus of DDX4 is localised on the surface of putative OSCs but is internalised during the process of oogenesis. This hypothesis is controversial since it is assumed that RNA helicases function intracellularly with no extracellular expression. To determine whether the C-terminus of DDX4 could be expressed on the cell surface, we generated a novel expression construct to express full-length DDX4 as a DsRed2 fusion protein with unique C- and N-terminal epitope tags. DDX4 and the C-terminal myc tag were detected at the cell surface by immunocytochemistry and FACS of non-permeabilised human embryonic kidney HEK 293T cells transfected with the DDX4 construct. DDX4 mRNA expression was detected in the DDX4-positive sorted cells by RT-PCR. This study clearly demonstrates that the C-terminus of DDX4 can be expressed on the cell surface despite its lack of a conventional membrane-targeting or secretory sequence. These results validate the use of antibody-based FACS to isolate DDX4-positive putative OSCs.
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Affiliation(s)
- Yvonne L Clarkson
- Institute of Cell Biology, University of Edinburgh, Edinburgh EH9 3FF, UK.
- Centre for Discovery Brain Sciences, University of Edinburgh, Edinburgh EH8 9XD, UK.
- Ashworth Laboratories, School of Biological Sciences, University of Edinburgh, Edinburgh EH9 3JR, UK.
| | - Emma Weatherall
- Institute of Cell Biology, University of Edinburgh, Edinburgh EH9 3FF, UK.
- Centre for Discovery Brain Sciences, University of Edinburgh, Edinburgh EH8 9XD, UK.
- Ashworth Laboratories, School of Biological Sciences, University of Edinburgh, Edinburgh EH9 3JR, UK.
| | - Martin Waterfall
- Ashworth Laboratories, School of Biological Sciences, University of Edinburgh, Edinburgh EH9 3JR, UK.
| | - Marie McLaughlin
- Institute of Cell Biology, University of Edinburgh, Edinburgh EH9 3FF, UK.
- Centre for Discovery Brain Sciences, University of Edinburgh, Edinburgh EH8 9XD, UK.
- Ashworth Laboratories, School of Biological Sciences, University of Edinburgh, Edinburgh EH9 3JR, UK.
| | - Haojiang Lu
- Institute of Cell Biology, University of Edinburgh, Edinburgh EH9 3FF, UK.
- Centre for Discovery Brain Sciences, University of Edinburgh, Edinburgh EH8 9XD, UK.
- Ashworth Laboratories, School of Biological Sciences, University of Edinburgh, Edinburgh EH9 3JR, UK.
| | - Paul A Skehel
- Centre for Discovery Brain Sciences, University of Edinburgh, Edinburgh EH8 9XD, UK.
| | - Richard A Anderson
- MRC Centre for Reproductive Health, Queens Medical Research Institute, University of Edinburgh, Edinburgh EH16 4TJ, UK.
| | - Evelyn E Telfer
- Institute of Cell Biology, University of Edinburgh, Edinburgh EH9 3FF, UK.
- Centre for Discovery Brain Sciences, University of Edinburgh, Edinburgh EH8 9XD, UK.
- Ashworth Laboratories, School of Biological Sciences, University of Edinburgh, Edinburgh EH9 3JR, UK.
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38
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Milani L, Maurizii MG. Insights into Germline Development and Differentiation in Molluscs and Reptiles: The Use of Molecular Markers in the Study of Non-model Animals. Results Probl Cell Differ 2019; 68:321-353. [PMID: 31598863 DOI: 10.1007/978-3-030-23459-1_14] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022]
Abstract
When shifting research focus from model to non-model species, many differences in the working approach should be taken into account and usually methodological modifications are required because of the lack of genetics/genomics and developmental information for the vast majority of organisms. This lack of data accounts for the largely incomplete understanding of how the two components-genes and developmental programs-are intermingled in the process of evolution. A deeper level of knowledge was reached for a few model animals, making it possible to understand some of the processes that guide developmental changes during evolutionary time. However, it is often difficult to transfer the obtained information to other, even closely related, animals. In this chapter, we present and discuss some examples, such as the choice of molecular markers to be used to characterize differentiation and developmental processes. The chosen examples pertain to the study of germline in molluscs, reptiles, and other non-model animals.
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Affiliation(s)
- Liliana Milani
- Department of Biological, Geological and Environmental Sciences (BiGeA), University of Bologna, Bologna, Italy.
| | - Maria Gabriella Maurizii
- Department of Biological, Geological and Environmental Sciences (BiGeA), University of Bologna, Bologna, Italy
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39
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Dufau ML, Kavarthapu R. 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. Front Endocrinol (Lausanne) 2019; 10:576. [PMID: 31555207 PMCID: PMC6727037 DOI: 10.3389/fendo.2019.00576] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/12/2019] [Accepted: 08/07/2019] [Indexed: 12/22/2022] Open
Abstract
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.
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40
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Ahlawat S, Sharma R, Arora R, Kumari N, Mishra AK, Tantia MS. Promoter methylation and expression analysis of Bvh gene in bulls with varying semen motility parameters. Theriogenology 2018; 125:152-156. [PMID: 30447494 DOI: 10.1016/j.theriogenology.2018.11.001] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/25/2018] [Revised: 10/09/2018] [Accepted: 11/04/2018] [Indexed: 12/18/2022]
Abstract
Crossbreds of low-producing indigenous cattle and high-producing exotic dairy bulls (Holstein Friesian and Jersey) have contributed in ensuring that India continues to be the world's top milk-producing country. However, subfertility observed in crossbred male progenies has been a major obstacle in exploitation of heterosis due to crossbreeding. There is sufficient scientific evidence in support of genetic and epigenetic regulation of key physiological processes including spermatogenesis. Bovine Vasa Homology (Bvh) is considered a molecular marker for the study of gametogenesis. Significant negative correlation between DNA methylation and gene expression has been reported in cattle-yaks hybrids and their parents. The present study analyzed promoter methylation status and expression profile of Bvh gene in spermatozoa from exotic Holstein Friesian cattle, indigenous Sahiwal cattle and their crossbreds with varying semen motility parameters. The degree of methylation of the Bvh promoter region was significantly higher in poor motility crossbred bulls (13.3%) as compared to good motility crossbreds (5.3%), Sahiwal (3%) and Holstein Friesian bulls (1%) (P < 0.05). Gene expression analysis revealed significantly higher mRNA abundance of Bvh in purebreds (Holstein Friesian and Sahiwal) as compared to crossbred counterparts (P < 0.001). Inverse correlation observed in this study between promoter methylation and gene expression of Bvh gene in spermatozoa from crossbred bulls with poor motility phenotype as compared to purebred parents provides an important insight into understanding the graded fertility of crossbred bulls.
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Affiliation(s)
- Sonika Ahlawat
- ICAR-National Bureau of Animal Genetic Resources, Karnal, India.
| | - Rekha Sharma
- ICAR-National Bureau of Animal Genetic Resources, Karnal, India
| | - Reena Arora
- ICAR-National Bureau of Animal Genetic Resources, Karnal, India
| | - Namita Kumari
- ICAR-National Bureau of Animal Genetic Resources, Karnal, India
| | - A K Mishra
- ICAR-National Bureau of Animal Genetic Resources, Karnal, India
| | - M S Tantia
- ICAR-National Bureau of Animal Genetic Resources, Karnal, India
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41
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Gaur M, Ramathal C, Reijo Pera RA, Turek PJ, John CM. Isolation of human testicular cells and co-culture with embryonic stem cells. Reproduction 2018; 155:153-166. [PMID: 29326135 DOI: 10.1530/rep-17-0346] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/07/2017] [Revised: 10/17/2017] [Accepted: 11/13/2017] [Indexed: 01/17/2023]
Abstract
Our overall goal is to create a three-dimensional human cell-based testicular model for toxicological and spermatogenesis studies. Methods to purify the major somatic testicular cells, namely Leydig cells (LCs), peritubular myoid cells (PCs) and Sertoli cells (SCs), from rats, mice and guinea pigs have been reported. In humans, the isolation of populations enriched for primary LCs, PCs or SCs also have described. One objective of this study was to determine if populations of cells enriched for all three of these cell types can be isolated from testes of single human donors, and we were successful in doing so from testes of three donors. Testes tissues were enzymatically digested, gravity sedimented and Percoll filtered to isolate populations enriched for LCs, PCs and SCs. LCs and PCs were identified by colorimetric detection of the expression of prototypical enzymes. Division of PCs and SCs in culture has been reported. We observed that primary human LCs could divide in culture by incorporation of 5-ethynyl-2'-deoxyuridine. SCs were identified and their functionality was demonstrated by the formation of tight junctions as shown by the expression of tight junction proteins, increased transepithelial electrical resistance, polarized secretion of biomolecules and inhibition of lucifer yellow penetration. Furthermore, we found that human SC feeder layers could facilitate germ cell progression of human embryonic stem cells (hESCs) by microarray analysis of gene expression.
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Affiliation(s)
| | - Cyril Ramathal
- Institute for Stem Cell BiologyStanford University, Stanford, California, USA
| | - Renee A Reijo Pera
- Institute for Stem Cell BiologyStanford University, Stanford, California, USA.,Stanford University School of MedicineStanford, California, USA
| | - Paul J Turek
- MandalMedInc., San Francisco, California, USA.,The Turek ClinicSan Francisco, California, USA
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42
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Li X, Ao J, Wu J. Systematic identification and comparison of expressed profiles of lncRNAs and circRNAs with associated co-expression and ceRNA networks in mouse germline stem cells. Oncotarget 2018; 8:26573-26590. [PMID: 28404936 PMCID: PMC5432280 DOI: 10.18632/oncotarget.15719] [Citation(s) in RCA: 47] [Impact Index Per Article: 6.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/19/2016] [Accepted: 01/22/2017] [Indexed: 01/01/2023] Open
Abstract
Accumulating evidence indicates that long noncoding RNAs (lncRNAs) and circular RNAs (circRNAs) involve in germ cell development. However, little is known about the functions and mechanisms of lncRNAs and circRNAs in self-renewal and differentiation of germline stem cells. Therefore, we explored the expression profiles of mRNAs, lncRNAs, and circRNAs in male and female mouse germline stem cells by high-throughput sequencing. We identified 18573 novel lncRNAs and 18822 circRNAs in the germline stem cells and further confirmed the existence of these lncRNAs and circRNAs by RT-PCR. The results showed that male and female germline stem cells had similar GDNF signaling mechanism. Subsequently, 8115 mRNAs, 3996 lncRNAs, and 921 circRNAs exhibited sex-biased expression that may be associated with germline stem cell acquisition of the sex-specific properties required for differentiation into gametes. Gene Ontology (GO) and KEGG pathway enrichment analyses revealed different functions for these sex-biased lncRNAs and circRNAs. We further constructed correlated expression networks including coding–noncoding co-expression and competing endogenous RNAs with bioinformatics. Co-expression analysis showed hundreds of lncRNAs were correlated with sex differences in mouse germline stem cells, including lncRNA Gm11851, lncRNA Gm12840, lncRNA 4930405O22Rik, and lncRNA Atp10d. CeRNA network inferred that lncRNA Meg3 and cirRNA Igf1r could bind competitively with miRNA-15a-5p increasing target gene Inha, Acsl3, Kif21b, and Igfbp2 expressions. These findings provide novel perspectives on lncRNAs and circRNAs and lay a foundation for future research into the regulating mechanisms of lncRNAs and circRNAs in germline stem cells.
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Affiliation(s)
- Xiaoyong Li
- Renji Hospital, Key Laboratory for the Genetics of Developmental and Neuropsychiatric Disorders (Ministry of Education), Bio-X Institutes, School of Medicine, Shanghai Jiao Tong University, Shanghai, 200240, China
| | - Junping Ao
- State Key Laboratory of Oncogenes and Related Genes, Shanghai Cancer Institute, Renji Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, 200032, China
| | - Ji Wu
- Renji Hospital, Key Laboratory for the Genetics of Developmental and Neuropsychiatric Disorders (Ministry of Education), Bio-X Institutes, School of Medicine, Shanghai Jiao Tong University, Shanghai, 200240, China.,Key Laboratory of Fertility Preservation and Maintenance of Ministry of Education, Ningxia Medical University, Yinchuan, 750004, China.,Shanghai Key Laboratory of Reproduction Medicine, Shanghai, 200025, China
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43
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Li Q, Yang H, He L, Wang Q. Characterization of the Es -DDX52 involved in the spermatogonial mitosis and spermatid differentiation in Chinese mitten crab ( Eriocheir sinensis ). Gene 2018; 646:106-119. [DOI: 10.1016/j.gene.2017.12.044] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/22/2017] [Revised: 12/14/2017] [Accepted: 12/20/2017] [Indexed: 11/26/2022]
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44
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Matsuura T, Miyazaki S, Miyazaki T, Tashiro F, Miyazaki JI. Zfp296 negatively regulates H3K9 methylation in embryonic development as a component of heterochromatin. Sci Rep 2017; 7:12462. [PMID: 28963472 PMCID: PMC5622089 DOI: 10.1038/s41598-017-12772-y] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/12/2017] [Accepted: 09/14/2017] [Indexed: 12/20/2022] Open
Abstract
The Cys2/His2-type zinc finger protein Zfp296 has been implicated in stem cell pluripotency and tumor pathogenesis. However, its mechanisms remain elusive. Here, we demonstrated that a Zfp296 deficiency in mice impairs germ-cell development and embryonic growth. Zfp296 was intracellularly localized to heterochromatin in embryos. A GST-Zfp296 pull-down experiment using ES cell nuclear extract followed by LC-MS/MS showed that Zfp296 interacts with component proteins of heterochromatin (such as HP1, Dnmt1, Dnmt3b, and ATRX) and the NuRD complex. We focused on H3K9 methylation as a hallmark of heterochromatin, and found that Zfp296 overexpression in cultured cells reduces the Suv39h1-mediated H3K9 methylation. Consistent with this finding, in Zfp296 -/- mouse embryos, we observed a global increase in H3K9 methylation in a developmental stage-dependent manner, and showed, by ChIP-qPCR, that the H3K9me3 levels at major satellite repeats were elevated in Zfp296 -/- embryos. Our results demonstrate that Zfp296 is a component of heterochromatin that affects embryonic development by negatively regulating H3K9 methylation.
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Affiliation(s)
- Takumi Matsuura
- Division of Stem Cell Regulation Research, Osaka University Graduate School of Medicine, 2-2 Yamadaoka, Suita, 565-0871, Osaka, Japan
| | - Satsuki Miyazaki
- Division of Stem Cell Regulation Research, Osaka University Graduate School of Medicine, 2-2 Yamadaoka, Suita, 565-0871, Osaka, Japan
| | - Tatsushi Miyazaki
- Division of Stem Cell Regulation Research, Osaka University Graduate School of Medicine, 2-2 Yamadaoka, Suita, 565-0871, Osaka, Japan
| | - Fumi Tashiro
- Division of Stem Cell Regulation Research, Osaka University Graduate School of Medicine, 2-2 Yamadaoka, Suita, 565-0871, Osaka, Japan
| | - Jun-Ichi Miyazaki
- Division of Stem Cell Regulation Research, Osaka University Graduate School of Medicine, 2-2 Yamadaoka, Suita, 565-0871, Osaka, Japan.
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45
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Not All H3K4 Methylations Are Created Equal: Mll2/COMPASS Dependency in Primordial Germ Cell Specification. Mol Cell 2017; 65:460-475.e6. [PMID: 28157506 DOI: 10.1016/j.molcel.2017.01.013] [Citation(s) in RCA: 72] [Impact Index Per Article: 9.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/10/2016] [Revised: 07/16/2016] [Accepted: 01/04/2017] [Indexed: 11/20/2022]
Abstract
The spatiotemporal regulation of gene expression is central for cell-lineage specification during embryonic development and is achieved through the combinatorial action of transcription factors/co-factors and epigenetic states at cis-regulatory elements. Here, we show that in addition to implementing H3K4me3 at promoters of bivalent genes, Mll2 (KMT2B)/COMPASS can also implement H3K4me3 at a subset of non-TSS regulatory elements, a subset of which shares epigenetic signatures of active enhancers. Our mechanistic studies reveal that association of Mll2's CXXC domain with CpG-rich regions plays an instrumental role for chromatin targeting and subsequent implementation of H3K4me3. Although Mll2/COMPASS is required for H3K4me3 implementation on thousands of loci, generation of catalytically mutant MLL2/COMPASS demonstrated that H3K4me3 implemented by this enzyme was essential for expression of a subset of genes, including those functioning in the control of transcriptional programs during embryonic development. Our findings suggest that not all H3K4 trimethylations implemented by MLL2/COMPASS are functionally equivalent.
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46
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Dosage compensation in the process of inactivation/reactivation during both germ cell development and early embryogenesis in mouse. Sci Rep 2017. [PMID: 28623283 PMCID: PMC5473838 DOI: 10.1038/s41598-017-03829-z] [Citation(s) in RCA: 17] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/20/2022] Open
Abstract
Ohno proposed that dosage compensation in mammals evolved as a two-step mechanism involving X-inactivation and X-upregulation. While X-inactivation is well characterized, it remains to further analysis whether upregulation of the single activated X chromosome in mammals occurs. We obtained RNA-seq data, including single-cell RNA-seq data, from cells undergoing inactivation/reactivation in both germ cell development and early embryogenesis stages in mouse and calculated the X: A ratio from the gene expression. Our results showed that the X: A ratio is always 1, regardless of the number of X chromosomes being transcribed for expressed genes. Furthermore, the single-cell RNA-seq data across individual cells of mouse preimplantation embryos of mixed backgrounds indicated that strain-specific SNPs could be used to distinguish transcription from maternal and paternal chromosomes and further showed that when the paternal was inactivated, the average gene dosage of the active maternal X chromosome was increased to restore the balance between the X chromosome and autosomes. In conclusion, our analysis of RNA-seq data (particularly single-cell RNA-seq) from cells undergoing the process of inactivation/reactivation provides direct evidence that the average gene dosage of the single active X chromosome is upregulated to achieve a similar level to that of two active X chromosomes and autosomes present in two copies.
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47
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Leopardo NP, Vitullo AD. Early embryonic development and spatiotemporal localization of mammalian primordial germ cell-associated proteins in the basal rodent Lagostomus maximus. Sci Rep 2017; 7:594. [PMID: 28377629 PMCID: PMC5429608 DOI: 10.1038/s41598-017-00723-6] [Citation(s) in RCA: 17] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/20/2016] [Accepted: 03/09/2017] [Indexed: 11/22/2022] Open
Abstract
The gene network controlling primordial germ cell (PGC) specification in eutherian mammals has been exhaustively investigated in mice. The egg-cylinder morphology of the mouse embryo is the key event enabling inductive signals from the extra-embryonic ectoderm (ExE) to specify epiblast cells as PGCs early on. We investigated the embryonic development and the spatiotemporal localization of PGC-associated proteins in the basal Hystricognathi rodent Lagostomus maximus. L. maximus develops through a flat-disc epiblast far apart from the ExE. In the primitive streak stage, OCT4-positive cells are detected in the posterior pole of the embryo disc in the mesoderm of the proximal epiblast. In the neural plate stage, a reduced 8 to 12 OCT4-positive cell population transiently expresses FRAGILIS, STELLA and SOX17 in the posterior streak. Soon after translocation to the hindgut, pluripotent OCT4 cells start expressing VASA, and then, STELLA and FRAGILIS are turned on during migration toward the genital ridge. L. maximus shows a spatiotemporal pattern of PGC-associated markers divergent from the early PGC restriction model seen in mice. This pattern conforms to alternative models that are based on a pluripotent population in the embryonic axis, where PGCs are specified later during development.
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Affiliation(s)
- Noelia P Leopardo
- Departamento de Ciencias Biomédicas y Biotecnológicas, Centro de Estudios Biomédicos, Biotecnológicos, Ambientales y Diagnóstico -CEBBAD-, Universidad Maimónides, Hidalgo 775, C1405BCK, Buenos Aires, Argentina.,Consejo Nacional de Investigaciones Científicas y Técnicas, CONICET, Buenos Aires, Argentina
| | - Alfredo D Vitullo
- Departamento de Ciencias Biomédicas y Biotecnológicas, Centro de Estudios Biomédicos, Biotecnológicos, Ambientales y Diagnóstico -CEBBAD-, Universidad Maimónides, Hidalgo 775, C1405BCK, Buenos Aires, Argentina. .,Consejo Nacional de Investigaciones Científicas y Técnicas, CONICET, Buenos Aires, Argentina.
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48
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Liu WS, Zhao Y, Lu C, Ning G, Ma Y, Diaz F, O'Connor M. A novel testis-specific protein, PRAMEY, is involved in spermatogenesis in cattle. Reproduction 2017; 153:847-863. [PMID: 28356500 DOI: 10.1530/rep-17-0013] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/09/2017] [Revised: 03/06/2017] [Accepted: 03/29/2017] [Indexed: 11/08/2022]
Abstract
Preferentially expressed antigen in melanoma (PRAME) is a cancer/testis antigen that is predominantly expressed in normal testicular tissues and a variety of tumors. The function of the PRAME family in spermatogenesis remains unknown. This study was designed to characterize the Y-linked PRAME (PRAMEY) protein during spermatogenesis in cattle. We found that PRAMEY is a novel male germ cell-specific, and a germinal granule-associated protein that is expressed in spermatogenic cells during spermatogenesis. The intact PRAMEY protein (58 kDa) was detected in different ages of testes but not in epididymal spermatozoa. A PRAMEY isoform (30 kDa) was highly expressed only in testes after puberty and in epididymal spermatozoa. This isoform interacts with PP1γ2 and is likely the mature protein present in the testes and sperm. Immunofluorescent staining demonstrated that PRAMEY was located predominantly in the acrosome granule of spermatids, and in acrosome and flagellum of spermatozoa. Immunogold electron microscopy further localized the PRAMEY protein complex to the nucleus and several cytoplasmic organelles, including the rough endoplasmic reticulum, some small vesicles, the intermitochondrial cement, the chromatoid body and the centrioles, in spermatogonia, spermatocytes, spermatids and/or spermatozoa. PRAMEY was highly enriched in and structurally associated with the matrix of the acrosomal granule (AG) in round spermatids, and migrated with the expansion of the AG during acrosomal biogenesis. While the function of PRAMEY remains unclear during spermatogenesis, our results suggest that PRAMEY may play an essential role in acrosome biogenesis and spermatogenesis.Free Chinese abstract: A Chinese translation of this abstract is freely available at http://www.reproduction-online.org/content/153/6/847/suppl/DC1.FreeSpanish abstract: A Spanish translation of this abstract is freely available at http://www.reproduction-online.org/content/153/6/847/suppl/DC2.
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Affiliation(s)
- Wan-Sheng Liu
- Department of Animal ScienceCenter for Reproductive Biology and Health (CRBH), College of Agricultural Sciences
| | - Yaqi Zhao
- Department of Animal ScienceCenter for Reproductive Biology and Health (CRBH), College of Agricultural Sciences
| | - Chen Lu
- Department of Animal ScienceCenter for Reproductive Biology and Health (CRBH), College of Agricultural Sciences
| | - Gang Ning
- Microscopy and Cytometry FacilityThe Huck Institutes of the Life Sciences, The Pennsylvania State University, University Park, Pennsylvania, USA
| | - Yun Ma
- Department of Animal ScienceCenter for Reproductive Biology and Health (CRBH), College of Agricultural Sciences.,College of Life ScienceXinyang Normal University, Xinyang, Henan, China
| | - Francisco Diaz
- Department of Animal ScienceCenter for Reproductive Biology and Health (CRBH), College of Agricultural Sciences
| | - Michael O'Connor
- Department of Animal ScienceCenter for Reproductive Biology and Health (CRBH), College of Agricultural Sciences
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Kawashima I, Kawamura K. Disorganization of the germ cell pool leads to primary ovarian insufficiency. Reproduction 2017; 153:R205-R213. [PMID: 28289071 DOI: 10.1530/rep-17-0015] [Citation(s) in RCA: 17] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/08/2017] [Revised: 02/10/2017] [Accepted: 03/13/2017] [Indexed: 01/26/2023]
Abstract
The mammalian ovary is an organ that controls female germ cell development, storing them and releasing mature oocytes for transporting to the oviduct. During the fetal stage, female germ cells change from a proliferative state to meiosis before forming follicles with the potential for the growth of surrounding somatic cells. Understanding of molecular and physiological bases of germ cell development in the fetal ovary contributed not only to the elucidation of genetic disorders in primary ovarian insufficiency (POI), but also to the advancement of novel treatments for patients with POI. Accumulating evidence indicates that mutations in NOBOX, DAZL and FIGLAgenes are associated with POI. In addition, cell biology studies revealed the important roles of these genes as essential translational factors for germ cell development. Recent insights into the role of the PI3K (phosphatidylinositol 3-kinase)-Akt signaling pathway in primordial follicle activation allowed the development of a new infertility treatment, IVA (in vitro activation), leading to successful pregnancy/delivery in POI patients. Furthermore, elucidation of genetic dynamics underlying female germ cell development could allow regeneration of oocytes from ES (embryonic stem)/iPS (induced pluripotent stem) cells in mammals. The purpose of this review is to summarize basic findings related to female germ cell development and potential clinical implications, especially focusing on POI etiologies. We also summarize evolving new POI therapies based on IVA as well as oocyte regeneration.
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Affiliation(s)
- Ikko Kawashima
- Department of Advanced Reproductive MedicineSt. Marianna University School of Medicine, Kawasaki City, Kanagawa, Japan
| | - Kazuhiro Kawamura
- Department of Advanced Reproductive MedicineSt. Marianna University School of Medicine, Kawasaki City, Kanagawa, Japan
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Li W, Zhang P, Wu X, Zhu X, Xu H. A Novel Dynamic Expression of vasa in Male Germ Cells during Spermatogenesis in the Chinese Soft-Shell Turtle (Pelidiscus sinensis
). JOURNAL OF EXPERIMENTAL ZOOLOGY PART B-MOLECULAR AND DEVELOPMENTAL EVOLUTION 2017; 328:230-239. [DOI: 10.1002/jez.b.22728] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Received: 11/04/2016] [Revised: 12/17/2016] [Accepted: 01/07/2017] [Indexed: 11/06/2022]
Affiliation(s)
- Wei Li
- Pearl River Fisheries Research Institute; Chinese Academic of Fisheries Sciences; Guangzhou People's Republic of China
| | - Piaoyi Zhang
- Pearl River Fisheries Research Institute; Chinese Academic of Fisheries Sciences; Guangzhou People's Republic of China
- College of Fisheries and Life Science; Shanghai Ocean University; Shanghai People's Republic of China
| | - Xuling Wu
- Pearl River Fisheries Research Institute; Chinese Academic of Fisheries Sciences; Guangzhou People's Republic of China
- College of Fisheries and Life Science; Shanghai Ocean University; Shanghai People's Republic of China
| | - Xinping Zhu
- Pearl River Fisheries Research Institute; Chinese Academic of Fisheries Sciences; Guangzhou People's Republic of China
| | - Hongyan Xu
- Pearl River Fisheries Research Institute; Chinese Academic of Fisheries Sciences; Guangzhou People's Republic of China
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