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Event: 1890

Key Event Title

The KE title should describe a discrete biological change that can be measured. It should generally define the biological object or process being measured and whether it is increased, decreased, or otherwise definably altered relative to a control state. For example “enzyme activity, decreased”, “hormone concentration, increased”, or “growth rate, decreased”, where the specific enzyme or hormone being measured is defined. More help

Decrease (loss of) fetal male germ cells

Short name
The KE short name should be a reasonable abbreviation of the KE title and is used in labelling this object throughout the AOP-Wiki. The short name should be less than 80 characters in length. More help
Germ cell loss, male

Biological Context

Structured terms, selected from a drop-down menu, are used to identify the level of biological organization for each KE. Note, KEs should be defined within a particular level of biological organization. Only KERs should be used to transition from one level of organization to another. Selection of the level of biological organization defines which structured terms will be available to select when defining the Event Components (below). More help
Level of Biological Organization
Cellular

Cell term

Further information on Event Components and Biological Context may be viewed on the attached pdf.The biological context describes the location/biological environment in which the event takes place.  For molecular/cellular events this would include the cellular context (if known), organ context, and species/life stage/sex for which the event is relevant. For tissue/organ events cellular context is not applicable.  For individual/population events, the organ context is not applicable. More help

Organ term

Further information on Event Components and Biological Context may be viewed on the attached pdf.The biological context describes the location/biological environment in which the event takes place.  For molecular/cellular events this would include the cellular context (if known), organ context, and species/life stage/sex for which the event is relevant. For tissue/organ events cellular context is not applicable.  For individual/population events, the organ context is not applicable. More help

Key Event Components

Further information on Event Components and Biological Context may be viewed on the attached pdf.Because one of the aims of the AOP-KB is to facilitate de facto construction of AOP networks through the use of shared KE and KER elements, authors are also asked to define their KEs using a set of structured ontology terms (Event Components). In the absence of structured terms, the same KE can readily be defined using a number of synonymous titles (read by a computer as character strings). In order to make these synonymous KEs more machine-readable, KEs should also be defined by one or more “event components” consisting of a biological process, object, and action with each term originating from one of 22 biological ontologies (Ives, et al., 2017; See List). Biological process describes dynamics of the underlying biological system (e.g., receptor signalling). The biological object is the subject of the perturbation (e.g., a specific biological receptor that is activated or inhibited). Action represents the direction of perturbation of this system (generally increased or decreased; e.g., ‘decreased’ in the case of a receptor that is inhibited to indicate a decrease in the signalling by that receptor).Note that when editing Event Components, clicking an existing Event Component from the Suggestions menu will autopopulate these fields, along with their source ID and description. To clear any fields before submitting the event component, use the 'Clear process,' 'Clear object,' or 'Clear action' buttons. If a desired term does not exist, a new term request may be made via Term Requests. Event components may not be edited; to edit an event component, remove the existing event component and create a new one using the terms that you wish to add. More help

Key Event Overview

AOPs Including This Key Event

All of the AOPs that are linked to this KE will automatically be listed in this subsection. This table can be particularly useful for derivation of AOP networks including the KE. Clicking on the name of the AOP will bring you to the individual page for that AOP. More help
AOP Name Role of event in AOP Point of Contact Author Status OECD Status
Ectopic ATRA in fetal testis leads to reduced spem count KeyEvent Arthur Author (send email) Under development: Not open for comment. Do not cite

Stressors

This is a structured field used to identify specific agents (generally chemicals) that can trigger the KE. Stressors identified in this field will be linked to the KE in a machine-readable manner, such that, for example, a stressor search would identify this as an event the stressor can trigger. NOTE: intermediate or downstream KEs in one AOP may function as MIEs in other AOPs, meaning that stressor information may be added to the KE description, even if it is a downstream KE in the pathway currently under development.Information concerning the stressors that may trigger an MIE can be defined using a combination of structured and unstructured (free-text) fields. For example, structured fields may be used to indicate specific chemicals for which there is evidence of an interaction relevant to this MIE. By linking the KE description to a structured chemical name, it will be increasingly possible to link the MIE to other sources of chemical data and information, enhancing searchability and inter-operability among different data-sources and knowledgebases. The free-text section “Evidence for perturbation of this MIE by stressor” can be used both to identify the supporting evidence for specific stressors triggering the MIE as well as to define broad chemical categories or other properties that classify the stressors able to trigger the MIE for which specific structured terms may not exist. More help

Taxonomic Applicability

Latin or common names of a species or broader taxonomic grouping (e.g., class, order, family) can be selected from an ontology. In many cases, individual species identified in these structured fields will be those for which the strongest evidence used in constructing the AOP was available in relation to this KE. More help
Term Scientific Term Evidence Link
human, mouse, rat human, mouse, rat Moderate NCBI

Life Stages

The structured ontology terms for life-stage are more comprehensive than those for taxa, but may still require further description/development and explanation in the free text section. More help
Life stage Evidence
Fetal High

Sex Applicability

The authors must select from one of the following: Male, female, mixed, asexual, third gender, hermaphrodite, or unspecific. More help
Term Evidence
Male High

Key Event Description

A description of the biological state being observed or measured, the biological compartment in which it is measured, and its general role in the biology should be provided. For example, the biological state being measured could be the activity of an enzyme, the expression of a gene or abundance of an mRNA transcript, the concentration of a hormone or protein, neuronal activity, heart rate, etc. The biological compartment may be a particular cell type, tissue, organ, fluid (e.g., plasma, cerebrospinal fluid), etc. The role in the biology could describe the reaction that an enzyme catalyses and the role of that reaction within a given metabolic pathway; the protein that a gene or mRNA transcript codes for and the function of that protein; the function of a hormone in a given target tissue, physiological function of an organ, etc. Careful attention should be taken to avoid reference to other KEs, KERs or AOPs. Only describe this KE as a single isolated measurable event/state. This will ensure that the KE is modular and can be used by other AOPs, thereby facilitating construction of AOP networks. More help

Male germ cell apoptosis in the fetal testis

In the fetal testis, apoptosis of XY germ cells (pro-spermatogonia) takes place early during gonad differentiation (Coucouvanis et al, 1993; Nguyen et al, 2020; Rucker 3rd et al, 2000; Wang et al, 1998) and is required to adjust overall germ cell numbers to Sertoli cells within the testis cords (Aitken et al, 2011). Later in development, spermatogonia that have been damaged by, for instance by chemical exposures, are also eliminated by apoptosis (Aitken et al, 2011; Wang et al, 2007). Hence, the process of germ cell apoptosis in integral to reproductive development and a failure to eliminate damaged and excess spermatogonia can result in sterility (Knudson et al, 1995; Rodriguez et al, 1997).  Nonetheless, it stands to reason that abnormally high levels of apoptosis during fetal life will result in a smaller spermatogonial stem cell pool, and that this will likely result in diminished reproductive potential (Aitken et al, 2011).

Fetal germ cell loss as Key Event

Although it is normal that a large number of pro-spermatogonia are eliminated by apoptosis during development, excessive loss during the prenatal period would be expected to have a direct consequence for fertility later in life. If all or the majority of pro-spermatogoia are lost, the spermatogonial stem cell pool will be either depleted and/or be of lower quality, and therefore the efficiency of spermatogenesis in the adult testis will be compromised. Hence, loss of germ cells during fetal life, in excess of what is normally ‘programmed’, would be expected to negatively impact adult fertility. It is relevant that spermatogenesis is relatively robust in rodents, compared with humans; in the latter, the number of sperm per ejaculate is only 2 – 4 fold higher than the number at which fertility is significantly reduced (Rahban & Nef, 2020; Working, 1988).

How It Is Measured or Detected

One of the primary considerations in evaluating AOPs is the relevance and reliability of the methods with which the KEs can be measured. The aim of this section of the KE description is not to provide detailed protocols, but rather to capture, in a sentence or two, per method, the type(s) of measurements that can be employed to evaluate the KE and the relative level of scientific confidence in those measurements. Methods that can be used to detect or measure the biological state represented in the KE should be briefly described and/or cited. These can range from citation of specific validated test guidelines, citation of specific methods published in the peer reviewed literature, or outlines of a general protocol or approach (e.g., a protein may be measured by ELISA).Key considerations regarding scientific confidence in the measurement approach include whether the assay is fit for purpose, whether it provides a direct or indirect measure of the biological state in question, whether it is repeatable and reproducible, and the extent to which it is accepted in the scientific and/or regulatory community. Information can be obtained from the OECD Test Guidelines website and the EURL ECVAM Database Service on Alternative Methods to Animal Experimentation (DB-ALM). ?

Apoptosis is most routinely detected by DNA ladder assay, TUNEL assay or Comet assay (Majtnerová & Roušar, 2018).

Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay, which detects apoptotic DNA fragmentations (Gorczyca et al, 1992) is available commercially from numerous companies using various staining technologies.

DNA laddering can be used to measure apoptosis at later stages only and is used to detect apoptosis of many cells, as it involves separation of DNA by agarose gel electrophoresis (Gong et al, 1994)

Comet assay, or single cell gel electrophoresis assay, can detect DNA damage at single-cell resolution (Singh et al, 1988). The alkaline Comet Assay is part of OECD Test Guideline 489 (OECD, 2016).

Direct measurements of total germ cell number in animal models can be performed with using various probes and antibodies to germ cell markers that are commercially available and reporter assays using germ cell specific promoter elements driving expression of reporter proteins. These reporter assays can detect the presence of germ cells in a quantitative manner. Examples include reporter mouse line OG2 (Szabó et al, 2002).

Domain of Applicability

This free text section should be used to elaborate on the scientific basis for the indicated domains of applicability and the WoE calls (if provided). While structured terms may be selected to define the taxonomic, life stage and sex applicability (see structured applicability terms, above) of the KE, the structured terms may not adequately reflect or capture the overall biological applicability domain (particularly with regard to taxa). Likewise, the structured terms do not provide an explanation or rationale for the selection. The free-text section on evidence for taxonomic, life stage, and sex applicability can be used to elaborate on why the specific structured terms were selected, and provide supporting references and background information.  More help

Fetal male germ cells must enter cell cycle quiescence during fetal life (McLaren, 2001). This process is conserved between mice, rats and humans (Francavilla et al, 1990)

Evidence for Perturbation by Stressor

Bis(2-ethylhexyl) phthalate

Di(2-ethylhexyl) phthalate (DEHP) exposure induces apoptosis to gonocytes in rat testis after intrauterine exposure to 500 and 750 mg/kg bw/day (Ryu et al, 2007).

all-trans-Retinoic acid

Exposure to 1 µM retinoic acid (RA) to explanted human fetal testes culture results in reduced number of gonocytes (Jørgensen et al, 2015).

Ectopic atRA in the fetal mouse testis can induce germ cell apoptosis (Cupp et al, 1999; Livera et al, 2000; Marinos et al, 1995; Trautmann et al, 2008), also seen in Cyp26b1-/- mice which are incapable of eliminating atRA from the fetal testes (MacLean et al, 2007)

References

List of the literature that was cited for this KE description. Ideally, the list of references, should conform, to the extent possible, with the OECD Style Guide (https://www.oecd.org/about/publishing/OECD-Style-Guide-Third-Edition.pdf) (OECD, 2015). More help

Aitken RJ, Findlay JK, Hutt KJ, Kerr JB (2011) Apoptosis in the germ line. Reproduction 141: 139-150

Coucouvanis EC, Sherwood SW, Carswell-Crumpton C, Spack EG, Jones PP (1993) Evidence that the mechanism of prenatal germ cell death in the mouse is apoptosis. Exp Cell Res 209: 238-247

Cupp AS, Dufour JM, Kim G, Skinner MK, Kim KH (1999) Action of retinoids on embryonic and early postnatal testis development. Endocrinology 140: 2343-2352

Francavilla S, Cordeschi G, Properzi G, Concordia N, Cappa F, Pozzi V (1990) Ultrastructure of fetal human gonad before sexual differentiation and during early testicular and ovarian development. J Submicrosc Cytol Pathol 22: 389-400

Gong J, Traganos F, Darzynkiewicz Z (1994) A selective procedure for DNA extraction from apoptotic cells applicable for gel electrophoresis and flow cytometry. Anal Biochem 218: 314-319

Gorczyca W, Bruno S, Darzynkiewicz R, Gong J, Darzynkiewicz Z (1992) DNA strand breaks occurring during apoptosis - their early insitu detection by the terminal deoxynucleotidyl transferase and nick translation assays and prevention by serine protease inhibitors. Int J Oncol 1: 639-648

Jørgensen A, Nielsen JE, Perlman S, Lundvall L, Mitchell RT, Juul A, Rajpert-De Meyts E (2015) Ex vivo culture of human fetal gonads: manipulation of meiosis signalling by retinoic acid treatment disrupts testis development. Hum Reprod 30: 2351-2363

Knudson CM, Tung KS, Tourtellotte WG, Brown GA, Korsmeyer SJ (1995) Bax-deficient mice with lymphoid hyperplasia and male germ cell death. Science 270: 96-99

Livera G, Rouiller-Fabre V, Durand P, Habert R (2000) Multiple effects of retinoids on the development of Sertoli, germ, and Leydig cells of fetal and neonatal rat testis in culture. Biol Reprod 62: 1303-1314

MacLean G, Li H, Metzger D, Chambon P, Petkovich M (2007) Apoptotic extinction of germ cells in testes of Cyp26b1 knockout mice. Endocrinology 148: 4560-4567

Majtnerová P, Roušar T (2018) An overview of apoptosis assays detecting DNA fragmentation. Mol Biol Rep 45: 1469-1478

Marinos E, Kulukussa M, Zotos A, Kittas C (1995) Retinoic acid affects basement membrane formation of the seminiferous cords in 14-day male rat gonads in vitro. Differentiation 59: 87-94

McLaren A (2001) Mammalian germ cells: birth, sex, and immortality. Cell Struct Funct 26: 119-122

Nguyen DH, Soygur B, Peng SP, Malki S, Hu G, Laird DJ (2020) Apoptosis in the fetal testis eliminates developmentally defective germ cell clones. Nat Cell Biol 22: 1423-1435

OECD. (2016) Test No. 489: In Vivo Mammalian Alkaline Comet Assay. OECD Guidelines for the Testing of Chemicals, Section 4. OECD Publishing, Paris.

Rahban R, Nef S (2020) Regional difference in semen quality of young men: a review on the implication of environmental and lifestyle factors during fetal life and adulthood. Basic Clin Androl 30: 16

Rodriguez I, Ody C, Araki K, Garcia I, Vassalli P (1997) An early and massive wave of germinal cell apoptosis is required for the development of functional spermatogenesis. EMBO J 16: 2262-2270

Rucker 3rd EB, Dierisseau P, Wagner KU, Garrett L, Wynshaw-Boris A, Flaws JA, Hennighausen L (2000) Bcl-x and Bax regulate mouse primordial germ cell survival and apoptosis during embryogenesis. Mol Endocrinol 14: 1038-1052

Ryu JY, Whang J, Park H, Im JY, Kim J, Ahn MY, Lee J, Kim HS, Lee BM, Yoo SD, Kwack SJ, Oh JH, Park KL, Han SY, Kim SH (2007) Di(2-ethylhexyl) phthalate induces apoptosis through peroxisome proliferators-activated receptor-gamma and ERK 1/2 activation in testis of Sprague-Dawley rats. J Toxicol Environ Health A 70: 1296-1303

Singh NP, McCoy MT, Tice RR, Schneider EL (1988) A simple technique for quantitation of low levels of DNA damage in individual cells. Exp Cell Res 175: 184-191

Szabó PE, Hübner K, Schöler H, Mann JR (2002) Allele-specific expression of imprinted genes in mouse migratory primordial germ cells. Mech Dev 115: 157-160

Trautmann E, Guerquin MJ, Duquenne C, Lahaye JB, Habert R, Livera G (2008) Retinoic acid prevents germ cell mitotic arrest in mouse fetal testes. Cell Cycle 7: 656-664

Wang C, Cui YG, Wang XH, Jia Y, Hikim AS, Lue YH, Tong JS, Qian LX, Sha JH, Zhou ZM, Hull L, Leung A, Swerdloff RS (2007) transient scrotal hyperthermia and levonorgestrel enhance testosterone-induced spermatogenesis suppression in men through increased germ cell apoptosis. J Clin Endocrinol Metab 92: 3292-3304

Wang RA, Nakane PK, Koji T (1998) Autonomous cell death of mouse male germ cells during fetal and postnatal period. Biol Reprod 58: 1250-1256

Working PK (1988) Male reproductive toxicology: comparison of the human to animal models. Environ Health Perspect 77: 37-44