API

Event: 1688

Key Event Title

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decrease, male anogenital distance

Short name

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short male AGD

Biological Context

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Level of Biological Organization
Tissue


Organ term

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Organ term
perineum


Key Event Components

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Process Object Action
androgen receptor signaling pathway Musculature of male perineum disrupted

Key Event Overview


AOPs Including This Key Event

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Stressors

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Taxonomic Applicability

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Term Scientific Term Evidence Link
human Homo sapiens Moderate NCBI
rat Rattus norvegicus High NCBI
mouse Mus musculus High NCBI

Life Stages

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Life stage Evidence
Foetal High

Sex Applicability

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Term Evidence
Male High

Key Event Description

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The anogenital distance (AGD) refers to the distance between anus and the external genitalia. In rodents and humans, the male AGD is approximately twice the length as the female AGD (Salazar-Martinez et al, 2004; Schwartz et al, 2019). This sexual dimorphisms is a consequence of sex hormone-dependent development of secondary sexual characteristics (Schwartz et al, 2019). In males, it is believed that androgens (primarily DHT) activate AR-positive cells in non-myotic cells in the fetal perineum region to initiate differentiation of the perineal levator ani and bulbocavernosus (LABC) muscle complex (Ipulan et al, 2014). This AR-dependent process occurs within a critical window of development, around gestational days 15-18 in rats (MacLeod et al, 2010). In females, the absence of DHT prevents this masculinization effect from occurring.

The involvement of androgens in masculinization of the male fetus, including the perineum, has been known for a very long time (Jost, 1953), and AGD has historically been used to, for instance, sex newborn kittens. It is now well established that the AGD in newborns is a proxy readout for the intrauterine sex hormone milieu the fetus was developing. Too low androgen levels in XY fetuses makes the male AGD shorter, whereas excess (ectopic) androgen levels in XX fetuses makes the female AGD longer, in humans and rodents (Schwartz et al, 2019).


How It Is Measured or Detected

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The AGD is a morphometric measurement carried out by trained technicians (rodents) or medical staff (humans).

In rodent studies AGD is assessed as the distance between the genital papilla and the anus, and measured using a stereomicroscope with a micrometer eyepiece. The AGD index (AGDi) is often calculated by dividing AGD by the cube root of the body weight.  It is important in statistical analysis to use litter as the statistical unit. This is done when more than one pup from each litter is examined. Statistical analyses is adjusted using litter as an independent, random and nested factor. AGD are analysed using body weight as covariate as recommended in Guidance Document 151 (OECD, 2013).

 


Domain of Applicability

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A short AGD in male offspring is a marker of insufficient androgen action during critical fetal developmental stages (Schwartz et al, 2019; Welsh et al, 2008). A short AGD is thus a sign of undervirilization, which is also associated with a series of male reproductive disorders, including genital malformations and infertility in humans (Juul et al, 2014; Skakkebaek et al, 2001).

There are numerous human epidemiological studies showing associations with intrauterine exposure to anti-androgenic chemicals and short AGD in newborn boys alongside other reproductive disorders (Schwartz et al, 2019). This underscores the human relevance of this AO. However, in reproductive toxicity studies and chemical risk assessment, rodents (rats and mice) are what is tested on. The list of chemicals inducing short male AGD in male rat offspring is extensive, as evidenced by the ‘stressor’ list and reviewed by (Schwartz et al, 2019).


Evidence for Perturbation by Stressor



Butylparaben

Butylparaben has been shown to cause decreased male AGD in rats following intrauterine exposure to 500 and 1000 mg/kg bw/day (Boberg et al, 2016; Zhang et al, 2014). A separate study using 600 mg/kg bw/day did not see an effect on male AGD (Boberg et al, 2008).


p,p'-DDE

p,p,DDE has been shown to cause decreased male AGD in rats following intrauterine exposure to 100-200 mg/kg bw/day (Loeffler & Peterson, 1999; Wolf et al, 1999).


Bis(2-ethylhexyl) phthalate

DEHP has been shown to cause decreased male AGD in rats following intrauterine exposure to 300-1500 mg/kg bw/day (Christiansen et al, 2010; Gray et al, 2000; Howdeshell et al, 2007; Jarfelt et al, 2005; Kita et al, 2016; Li et al, 2013; Lin et al, 2009; Moore et al, 2001; Nardelli et al, 2017; Saillenfait et al, 2009; Wolf et al, 1999).


Dexamethasone

Dexamethasone has been shown to cause decreased male AGD in rats following intrauterine exposure to 0.1 mg/kg bw/day (Van den Driesche et al, 2012).


Fenitrothion

Fenitrothion has been shown to cause decreased male AGD in rats following intrauterine exposure to 25 mg/kg bw/day (Turner et al, 2002).


Finasteride

Finasteride has been shown to cause decreased male AGD in rats following intrauterine exposure to 100 mg/kg bw/day (Bowman et al, 2003).


Flutamide

Flutamide has been shown to cause decreased male AGD in rats following intrauterine exposure to doses between 16-100 mg/kg bw/day (Foster & Harris, 2005; Hass et al, 2007; Kita et al, 2016; McIntyre et al, 2001; Mylchreest et al, 1999; Scott et al, 2007; Welsh et al, 2007).


Ketoconazole

Ketoconazole has been shown to cause decreased male AGD in rats following intrauterine exposure to 50 mg/kg bw/day in one study (Taxvig et al, 2008), but no effect in another study using same dose (Wolf et al, 1999).


Linuron

Linuron has been shown to cause decreased male AGD in rats following intrauterine exposure to 50-100 mg/kg bw/day (Hotchkiss et al, 2004; McIntyre et al, 2002; Wolf et al, 1999).


Prochloraz

Prochloraz has been shown to cause decreased male AGD in rats following intrauterine exposure to 150-250 mg/kg bw/day (Laier et al, 2006; Noriega et al, 2005).


Procymidone

Procymidone has been shown to cause decreased male AGD in rats following intrauterine exposure to doses between 50-150 mg/kg bw/day (Hass et al, 2012; Hass et al, 2007; Wolf et al, 1999).


Triticonazole

Triticonazole has been shown to cause decreased male AGD in rats following intrauterine exposure to 150 and 450 mg/kg bw/day (Draskau et al, 2019).


Vinclozolin

Vinclozolin has been shown to cause decreased male AGD in rats following intrauterine exposure to doses between 50-200 mg/kg bw/day (Christiansen et al, 2009; Gray et al, 1994; Hass et al, 2007; Matsuura et al, 2005; Ostby et al, 1999; Schneider et al, 2011; Wolf et al, 2004).


di-n-hexyl phthalate

DnHP has been shown to cause decreased male AGD in rats following intrauterine exposure to 500-750 mg/kg bw/day (Saillenfait et al, 2009a; Saillenfait et al, 2009b).


Dicyclohexyl phthalate

DCHP has been shown to cause decreased male AGD in rats following intrauterine exposure to 350-750 mg/kg bw/day (Aydoğan Ahbab & Barlas, 2015; Hoshino et al, 2005; Saillenfait et al, 2009a).


butyl benzyl phthalate

BBP has been shown to cause decreased male AGD in rats following intrauterine exposure to 500-1000 mg/kg bw/day (Ema & Miyawaki, 2002; Gray et al, 2000; Hotchkiss et al, 2004; Nagao et al, 2000; Tyl et al, 2004).


monobenzyl phthalate

MBeP has been shown to cause decreased male AGD in rats following intrauterine exposure to 375 mg/kg bw/day (Ema et al, 2003).


di-n-heptyl phthalate

DHPP has been shown to cause decreased male AGD in rats following intrauterine exposure to 1000 mg/kg bw/day (Saillenfait et al, 2011).


Regulatory Significance of the Adverse Outcome

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In regulatory toxicology, the AGD is mandatory inclusions in OECD test guidelines used to test for developmental and reproductive toxicity of chemicals. Guidelines include ‘TG 443 extended one-generation study’, ‘TG 421/422 reproductive toxicity screening studies’ and ‘TG 414 developmental toxicity study’.


References

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Aydoğan Ahbab M, Barlas N (2015) Influence of in utero di-n-hexyl phthalate and dicyclohexyl phthalate on fetal testicular development in rats. Toxicol Lett 233: 125-137

Boberg J, Axelstad M, Svingen T, Mandrup K, Christiansen S, Vinggaard AM, Hass U (2016) Multiple endocrine disrupting effects in rats perinatally exposed to butylparaben. Toxicol Sci 152: 244-256

Boberg J, Metzdorff S, Wortziger R, Axelstad M, Brokken L, Vinggaard AM, Dalgaard M, Nellemann C (2008) Impact of diisobutyl phthalate and other PPAR agonists on steroidogenesis and plasma insulin and leptin levels in fetal rats. Toxicology 250: 75-81

Bowman CJ, Barlow NJ, Turner KJ, Wallace DG, Foster PM (2003) Effects of in utero exposure to finasteride on androgen-dependent reproductive development in the male rat. Toxicol Sci 74: 393-406

Christiansen S, Boberg J, Axelstad M, Dalgaard M, Vinggaard AM, Metzdorff SB, Hass U (2010) Low-dose perinatal exposure to di(2-ethylhexyl) phthalate induces anti-androgenic effects in male rats. Reprod Toxicol 30: 313-321

Christiansen S, Scholze M, Dalgaard M, Vinggaard AM, Axelstad M, Kortenkamp A, Hass U (2009) Synergistic disruption of external male sex organ development by a mixture of four antiandrogens. Environ Health Perspect 117: 1839-1846

Draskau MK, Boberg J, Taxvig C, Pedersen M, Frandsen HL, Christiansen S, Svingen T (2019) In vitro and in vivo endocrine disrupting effects of the azole fungicides triticonazole and flusilazole. Environ Pollut 255: 113309

Ema M, Miyawaki E (2002) Effects on development of the reproductive system in male offspring of rats given butyl benzyl phthalate during late pregnancy. Reprod Toxicol 16: 71-76

Ema M, Miyawaki E, Hirose A, Kamata E (2003) Decreased anogenital distance and increased incidence of undescended testes in fetuses of rats given monobenzyl phthalate, a major metabolite of butyl benzyl phthalate. Reprod Toxicol 17: 407-412

Foster PM, Harris MW (2005) Changes in androgen-mediated reproductive development in male rat offspring following exposure to a single oral dose of flutamide at different gestational ages. Toxicol Sci 85: 1024-1032

Gray LE, Jr., Ostby J, Furr J, Price M, Veeramachaneni DN, Parks L (2000) Perinatal exposure to the phthalates DEHP, BBP, and DINP, but not DEP, DMP, or DOTP, alters sexual differentiation of the male rat. Toxicol Sci 58: 350-365

Gray LEJ, Ostby JS, Kelce WR (1994) Developmental effects of an environmental antiandrogen: the fungicide vinclozolin alters sex differentiation of the male rat. Toxicol Appl Pharmacol 129: 46-52

Hass U, Boberg J, Christiansen S, Jacobsen PR, Vinggaard AM, Taxvig C, Poulsen ME, Herrmann SS, Jensen BH, Petersen A, Clemmensen LH, Axelstad M (2012) Adverse effects on sexual development in rat offspring after low dose exposure to a mixture of endocrine disrupting pesticides. Reprod Toxicol 34: 261-274

Hass U, Scholze M, Christiansen S, Dalgaard M, Vinggaard AM, Axelstad M, Metzdorff SB, Kortenkamp A (2007) Combined exposure to anti-androgens exacerbates disruption of sexual differentiation in the rat. Environ Health Perspect 115 Suppl. 1: 122-128

Hoshino N, Iwai M, Okazaki Y (2005) A two-generation reproductive toxicity study of dicyclohexyl phthalate in rats. J Toxicol Sci 30 Spec No: 79-96

Hotchkiss AK, Parks-Saldutti LG, Ostby JS, Lambright C, Furr J, Vandenbergh JG, Gray LEJ (2004) A mixture of the "antiandrogens" linuron and butyl benzyl phthalate alters sexual differentiation of the male rat in a cumulative fashion. Biol Reprod 71: 1852-1861

Howdeshell KL, Furr J, Lambright CR, Rider CV, Wilson VS, Gray LE, Jr. (2007) Cumulative effects of dibutyl phthalate and diethylhexyl phthalate on male rat reproductive tract development: altered fetal steroid hormones and genes. Toxicol Sci 99: 190-202

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Jost A (1953) Problems of fetal endocrinology: The gonadal and hypophyseal hormones. Recent Prog Horm Res 8: 379-418

Juul A, Almstrup K, Andersson AM, Jensen TK, Jorgensen N, Main KM, Rajpert-De Meyts E, Toppari J, Skakkebaek NE (2014) Possible fetal determinants of male infertility. Nat Rev Endocrinol 10: 553-562

Kita DH, Meyer KB, Venturelli AC, Adams R, Machado DL, Morais RN, Swan SH, Gennings C, Martino-Andrade AJ (2016) Manipulation of pre and postnatal androgen environments and anogenital distance in rats. Toxicology 368-369: 152-161

Laier P, Metzdorff SB, Borch J, Hagen ML, Hass U, Christiansen S, Axelstad M, Kledal T, Dalgaard M, McKinnell C, Brokken LJ, Vinggaard AM (2006) Mechanisms of action underlying the antiandrogenic effects of the fungicide prochloraz. Toxicol Appl Pharmacol 213: 2

Li M, Qiu L, Zhang Y, Hua Y, Tu S, He Y, Wen S, Wang Q, Wei G (2013) Dose-related effect by maternal exposure to di-(2-ethylhexyl) phthalate plasticizer on inducing hypospadiac male rats. Environ Toxicol Pharmacol 35: 55-60

Lin H, Lian QQ, Hu GX, Jin Y, Zhang Y, Hardy DO, Chen GR, Lu ZQ, Sottas CM, Hardy MP, Ge RS (2009) In utero and lactational exposures to diethylhexyl-phthalate affect two populations of Leydig cells in male Long-Evans rats. Biol Reprod 80: 882-888

Loeffler IK, Peterson RE (1999) Interactive effects of TCDD and p,p'-DDE on male reproductive tract development in in utero and lactationally exposed rats. Toxicol Appl Pharmacol 154: 28-39

MacLeod DJ, Sharpe RM, Welsh M, Fisken M, Scott HM, Hutchison GR, Drake AJ, van den Driesche S (2010) Androgen action in the masculinization programming window and development of male reproductive organs. Int J Androl 33: 279-287

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