To the extent possible under law, AOP-Wiki has waived all copyright and related or neighboring rights to KE:1890
Key Event Title
Decrease (loss of) fetal male germ cells
|Level of Biological Organization|
Key Event Components
Key Event Overview
AOPs Including This Key Event
|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||Terje Svingen (send email)||Under development: Not open for comment. Do not cite|
|human, mouse, rat||human, mouse, rat||Moderate||NCBI|
Key Event Description
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
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
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)
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