Decreased ALDH1A (RALDH) activity leading to decreased fertility via disrupted meiotic initiation of fetal oogonia

• Terje Svingen

• You Song

Revision dates for related pages

• Sex: This AOP applies to females. atRA is also involved in meiosis of testicular gonocytes, but this occurs postnatally. In the female ovaries, atRA induces meiosis of oocytes during gestation, thus the spatiotemporal expression of atRA in the ovaries are tightly controlled. Finally, as this AOP is concerned with establishing the ovarian reserve/follicle pool through mechanisms that are unique to ovaries, restricting the AOP to female only is appropriate.

• Life stages: This AOP spans the period from mid- to late-gestation in mammals, all the way to adulthood where fertility is manifested. The upstream event pertains to fetal/neonatal life stages, whereas the downstream events pertain to adult reproductive life stages. 

• Taxonomy: Strongest evidence for the role of atRA in regulating oocyte entry into meiosis stems from mouse studies, so the taxonomic applicability is strongest for this animal model. Studies have also been done in rats. Evidence for the same mechanisms in humans is less substantiated (Li & Clagett-Dame, 2009; Griswold et al, 2012; Spiller & Bowles, 2022; Jørgensen & Rajpert-De Meyts, 2014).

The essentiality of each key event (KE) was evaluated, meaning that if an upstream KE is blocked or does not occur, subsequent downstream KEs or the adverse outcome (AO) are prevented or altered. Both direct and indirect evidence of essentiality were assessed according to the OECD developer’s handbook (see Appendix B, 1t3n0mxigk_App_B_Essentiality_2025.pdf) with a summary provided in Table 1.

Table 1: Essentiality assessment of KEs for AOP 398

Retrieved literature and overall assessments are detailed in Appendix B. ****substantial evidence, ***good evidence, **moderate evidence, *some evidence.

Uncertainties, inconsistencies and data gaps

In mice, there is strong evidence to support the view that atRA is important for initiating meiosis in germ cells (Bowles et al, 2016; Spiller et al, 2017; Teletin et al, 2017). Some studies suggest that atAR is not critical but important for meiotic entry under normal physiological conditions by evidencing meiosis in Aldh1a1, Aldh1a2 and Aldh1a3 ablated mice, individually and in tandem (Bellutti et al, 2019; Chassot et al, 2020; Kumar et al, 2011); however, additional studies have shown redundant roles between all three Aldha isoforms which can compensate for deletion of one or two (Bowles et al, 2016). More specifically, both double (Aldh1a2/3) and triple (Aldh1a1/2/3) knockout mouse models display reduced Stra8 expression in oocytes, yet oocytes eventually go through meiosis, which could suggest a redundant role for atRA for meiosis in the ovaries (Chassot et al, 2020; Kumar et al, 2011). A similar phenotype with reduced Stra8 expression but eventual meiotic initiation is seen for deletion of atRA receptors RAR-α, -β, -γ) in mice (Vernet et al, 2020). But, although RAR knockouts were also capable of producing offspring, it remains unclear if any of the above-mentioned mouse models display impaired fertility or whether the size of their oocyte pools are affected.

Biological Plausibility, coherence, and consistency of the experimental evidence

The role for ALDH1A2 in the synthesis of atRA is well established as an essential component of regulating regional expression of retinoid species during development. It is also well established that atRA is an inducer of meiosis in germ cells in mice; however, there is some debate about the essentiality of atRA in this process in human fetal ovaries. The requirement for oocytes to enter the first phase of meiosis during fetal development is also well established, hence the biological plausibility linking meiotic failure with loss of oocytes at later developmental stages is strong.

Although non-meiotic oocytes can survive in germ cell nests and during nest breakdown, they will ultimately be eliminated from the oocyte pool of competent follicles. There is therefore a direct link between meiotic entry and fertility during adulthood. Thus, this AOP provides a plausible chain of events linking reduced atRA during fetal life with reduced ovarian reserve and fertility during reproductive age. The strength of the downstream KEs and KER – reduced ovarian reserve and reduced fertility – is very well documented and thus the biological plausibility is very strong. Evidence for a direct link between the AO and perturbed atRA synthesis, or reduced atRA levels, during early development comes mainly from mouse studies; yet the relationship is regarded biologically plausible also in humans, but with weight of evidence not being as strong. 

The overall evidence assessment scores for each KER is summarized in the below Table:

Table 2: Biological plausibility of key event relationships of AOP-398. 

Empirical support

Concordance of dose-response relationships

The quantitative understanding of dose-response relationships in this AOP is limited. Whilst the relative levels of endogenous atRA produced by the ovary (for any species) remains unknown, similarly, the quantitative relationship between atRA levels and induction of meiosis also remains unclear. Nevertheless, it is has been conclusively shown that low levels of exogenous atRA can induce mouse and rat germ cells to enter meiosis both in vitro and ex vivo (Bowles et al, 2006; Livera et al, 2000). Likewise, atRA is necessary to achieve meiosis in in vitro-derived oocytes via PGCLCs (Miyauchi et al, 2017).

Temporal concordance among the key events and the adverse outcome

This AOP bridges two different life stages: fetal/perinatal and adult/reproductive age. The adverse outcome is the result of perturbation taking place during early stages of ovary development. In mice, rats and humans, the oocytes must enter meiosis prophase in order to establish the follicle pool/ovarian reserve postnatally. Thus, the AOP focusses on chemical perturbations during fetal life, which occurs around E13-E16 in mice and E15-E18 is rats, or first trimester in humans (Peters, 1970), but the adverse outcome does not manifest until adulthood. 

There is strong temporal concordance between the various key events, from inhibition of ALDH1A2 (RALDH2) that leads to reduced atRA synthesis. In turn, atRA must be present in the fetal ovaries at the time when oocytes are supposed to enter meiosis mid-gestation in mice (or first trimester in human). With a significant reduction in available atRA the oocytes will not enter meiosis, ultimately leading to the downstream key event of loss of oocytes beyond what is normal. The number of oocytes, or the oocyte pool/ovarian reserve, in turn will affect ovary function and fertility at reproductive stages, hence the temporal sequence of events is rational based on the biological process.

Strength, consistency, and specificity of association of adverse effect and initiating event

In mice, there is strong evidence to support the view that atRA is an inducer of meiosis in germ cells, with consistent results from in vitro (PGCLCs), ex vivo (ovary cultures) and in vivo studies as listed under KE 2477. There is strong evidence showing the importance of RA for female fertility, but this relates to many aspects of reproductive development and function from fetal life to adulthood, including maintaining pregnancy (Clagett-Dame & Knutson, 2011). Thus, it can be difficult to distill exactly how atRA-controlled meiotic entry of oocytes directly link to reduced fertility. Nevertheless, a direct relationship is strongly supported by the fact that Stra8-depleted mice are infertile with small ovaries lacking oocytes (Baltus et al, 2006) and that Stra8 induction in germ cells is controlled by atRA in mice, rats and humans (Bowles et al, 2006; Childs et al, 2011; Koubova et al, 2006; Livera et al, 2000). Furthermore, vitamin A-deficient (VAD) mice display delayed or failed meiotic entry of fetal oocytes depending on level of Vitamin A deficiency (Li & Clagett-Dame, 2009).

More detailed descriptions and supporting references are found at respective KER pages on AOPwiki. Overall assessments are summarized in Table 3.

Table 3: Empirical evidence for key event relationships of AOP-398. 

This AOP is still largely qualitative, as the quantitative understanding between chemical potency and perturbation of KEs are insufficient. This relates to the dose-response relationship between concentrations of atRA in the ovary relative to meiotic initiation of oocytes. It also relates to the relationship between number of lost oocytes during development relative to the oocyte pool/ovarian reserve, as there naturally is a large loss of oocytes during development.