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Relationship: 2274

Title

A descriptive phrase which clearly defines the two KEs being considered and the sequential relationship between them (i.e., which is upstream, and which is downstream). More help

Impaired, Spermatogenesis leads to impaired, Fertility

Upstream event

The causing Key Event (KE) in a Key Event Relationship (KER). More help

Impaired, Spermatogenesis

Downstream event

The responding Key Event (KE) in a Key Event Relationship (KER). More help

impaired, Fertility

Key Event Relationship Overview

The utility of AOPs for regulatory application is defined, to a large extent, by the confidence and precision with which they facilitate extrapolation of data measured at low levels of biological organisation to predicted outcomes at higher levels of organisation and the extent to which they can link biological effect measurements to their specific causes.Within the AOP framework, the predictive relationships that facilitate extrapolation are represented by the KERs. Consequently, the overall WoE for an AOP is a reflection in part, of the level of confidence in the underlying series of KERs it encompasses. Therefore, describing the KERs in an AOP involves assembling and organising the types of information and evidence that defines the scientific basis for inferring the probable change in, or state of, a downstream KE from the known or measured state of an upstream KE. More help

AOPs Referencing Relationship

AOP Name	Adjacency	Weight of Evidence	Quantitative Understanding	Point of Contact	Author Status	OECD Status
Inhibition of 11β-Hydroxysteroid Dehydrogenase leading to decreased population trajectory	adjacent	High	High	Agnes Aggy (send email)	Under development: Not open for comment. Do not cite	Under Development

Taxonomic Applicability

Latin or common names of a species or broader taxonomic grouping (e.g., class, order, family) that help to define the biological applicability domain of the KER.In general, this will be dictated by the more restrictive of the two KEs being linked together by the KER. More help

Term	Scientific Term	Evidence	Link
rodents	rodents	High	NCBI
teleost fish	teleost fish	High	NCBI

Sex Applicability

An indication of the the relevant sex for this KER. More help

Sex	Evidence
Male	High

Life Stage Applicability

An indication of the the relevant life stage(s) for this KER. More help

Term	Evidence
Adult, reproductively mature	High

Key Event Relationship Description

Provides a concise overview of the information given below as well as addressing details that aren’t inherent in the description of the KEs themselves. More help

Spermatogenesis is a multiphase process of cellular transformation that produces mature male gametes known as sperm for sexual reproduction (Kang et al., 2015). The process of spermatogenesis can be broken down into 3 phases: the mitotic proliferation of spermatogonia, meiosis, and post meiotic differentiation(spermiogenesis) (Boulanger et al., 2015). Male fertility is dependent on the quantity as well as the proper cellular morphology of the sperm formed in the testes (Chen et al., 2020). The fusion of sperm and oocytes is the key step for the beginning of life known as fertilization (Alavi et al., 2019). Impaired spermatogenesis may impact fertility and, consequently, also reduce reproduction.

Evidence Collection Strategy

Include a description of the approach for identification and assembly of the evidence base for the KER. For evidence identification, include, for example, a description of the sources and dates of information consulted including expert knowledge, databases searched and associated search terms/strings. Include also a description of study screening criteria and methodology, study quality assessment considerations, the data extraction strategy and links to any repositories/databases of relevant references.Tabular summaries and links to relevant supporting documentation are encouraged, wherever possible. More help

The majority of papers used in evidence supporting the key event relationship were found through AbstractSifter, a Microsoft Excel-based application that extracts papers from PubMed. AbstractSifter ranks abstracts based on their relevance through key search and filter terms. Initial papers were found through the search engine, Google Scholar, utilizing the search terms “Impaired spermatogenesis male infertility” and “Impaired spermatogenesis male infertility in fish”. These papers were used to help curate search and filter terms used in Abstract Sifter. This search yielded 41600 search results but only papers found on the first page of results were further examined. In AbstractSifter, 3 different searches were done to curate a subset of 40 papers. Search terms for the 3 searches included “spermatogenesis AND fish” and “spermatogenesis AND zebrafish” which yielded an initial set of 1587 and 192 results respectively. Filter terms for the 3 searches included “male, infertility, and reduced”, “male, infertility, and impaired”, and “male and infertil”. The first 2 filter set of words were used for the spermatogenesis and fish search which yielded 9 and 11 papers respectively. The last set of filter terms was used for the spermatogenesis and zebrafish search which yielded a respective 25 papers. Additional sources used towards the weight of evidence were provided through expert knowledge and found through sources in papers initially curated in the AbstractSifter search.

Evidence Map 2.0

ID	Experimental Design	Species	Upstream Observation	Downstream Observation	Citation (first author, year)	Notes

Evidence Map

Addresses the scientific evidence supporting KERs in an AOP setting the stage for overall assessment of the AOP. More help

Title	First Author	Biological Plausibility	Dose Concordance	Temporal Concordance	Incidence Concordance

Biological Plausibility

Dose Concordance Evidence

Temporal Concordance Evidence

Incidence Concordance Evidence

Uncertainties and Inconsistencies

Addresses inconsistencies or uncertainties in the relationship including the identification of experimental details that may explain apparent deviations from the expected patterns of concordance. More help

When exposed to 10 and 100 ng/L of EE2 for 62 days leading to spawning, Rainbow trout (Oncorhynchus mykiss) experienced a decrease in GSI and increases in sperm concentration and spermatocrit. However, there were no significant changes to spermatogenesis. Despite this, there was a decrease in viability of embryos. (Schultz et al., 2003)
Male Sprague-Dawley rats (Rattus norvegicus) fed a high fat diet(allowing them to develop Non-alcoholic fatty liver disease) experienced decreased testosterone levels along with reduced sperm number and motility. However, this did not affect fertility of the rats (Li et al., 2013).

Table 4: Concordance Table (T-Z)

Species	Experimental design	Evidence of Impaired Spermatogenesis (IS)	Evidence of Impaired Fertility (IR)	IS observed?	IR observed?	Citation	Notes
Zebrafish (Danio rerio)	TALEN mRNAs injected into one-cell stage zebrafish embryos to generate parent gen that was outcrossed with WT for F1 F1 was then self-crossed for mutants Targeted the androgen receptor (AR)	Upregulation of amh and gsdf Downregulation igf3 Reduced # of sperm Reduction in # of germ cells observed in AR mutant fish Increased proportion of pre-spermatids sperm cells	Reduced in vitro fertilization (20%) Failed to induce spawning	Yes	Yes	Tang et al., 2018	Androgen receptor
Mice	Disruption of Protein C inhibitor (PCI) through combining mutant embryonic stem cells with swiss morula embryos to create mutants F1 heterozygous mice were then bred to create an F2 that was subsequently used in the study	Morphologically abnormal sperm Reduced motility (12.5%) compared to control (51.5%) Apoptotic spermatocytes likely due to destruction of Sertoli cells	Reduced in vivo fertilization rate (n=416 blastocysts) (0.5%) vs control (n= 420 blastocysts) (94%) In vivo fertilization rate: only 0.5% (n=416) oocytes were fertilized and developed into the blastocyst stage compared to PCI+/+ or PCI+/– males (92% (n = 415) and 94% (n = 420), respectively) Infertile under standard breeding despite showing signs of normal sexual activity	Yes	Yes	Uhrin et al., 2000	PCI - inhibitor of anticoagulant serine protease activated protein C and a variety of proteases PCI is largely present in seminal plasma and is responsible for inhibiting acrosin
Zebrafish (Danio rerio)	Adults exposed to 0.5 mg DEHP kg-1 (body weight) for 10 days via intraperitoneal injection	No large observable difference	No significant decrease in fertilization success	No	No	Uren-Webster et al., 2010	DEHP is phthalate which is a plasticizer in many mass-produced products
	Adults exposed to 50 mg DEHP kg-1 for 10 days via intraperitoneal injection	Reduction in the proportion of spermatozoa and an increase in the proportion of spermatocytes	No significant decrease in fertilization success	Yes	No
	Adults exposed to 5000 mg DEHP kg-1 for 10 days via intraperitoneal injection	Reduction in the proportion of spermatozoa and an increase in the proportion of spermatocytes	Reduction of fertilization success of oocytes spawned by females especially during the second 5-day period	Yes	Yes
Marine medaka (Oryzias melastigma)	Multi-generational study evaluating hypoxia; initial exposure sexually mature fish (F0) for 1 month; F1H and F2H reared in hypoxic conditions; F1T and FT2 reared in normoxia	Significant decrease in curvilinear velocity, straight line velocity and average path velocity of sperm (FO, F1H and F2H) Decrease in percentage of spermatids, number of spermatids, and increases in interstitial space of the testis (F1H, F2T, F3H)	Percentage of fertilization success significantly reduced in F1	Yes	Yes	Wang et al., 2016
Mice (C57BL/6)	Knockout of BRD7 was done through Cre/loxP and flp/FRT recombination and embryonic cells to create a positive clone that was then used to create BRD7-deficient mice	Irregular head shape Deformed acrosome Post-meiotic development of elongating spermatids disruption Increased proportion of abnormal spermatids (49.95 ± 7.13% of round spermatids, 67.84 ± 3.51% of elongating spermatids, 80.65 ± 5.8 % of condensing spermatids and 100% of condensed spermatids) Downregulation of various spermatogenic markers	Infertile under standard breeding despite showing signs of normal sexual activity	Yes	Yes	Wang et al., 2016	BRD7 is a bromodomain gene that inhibits cell growth and cell cycle progression and is a co-factor for p53 BRD7 has high expression in mice testes
Flies (Drosophila)	CRISPR/Cas9-mediated genome editing to SNP locus	64% of cysts contained germ cells and only 25% of those progressed to spermatocyte stage in mutants Lacked mature sperm Lack of mei-p26 protein which facilitates transition from amplifying to differentiating spermatogonia Accumulation of mitotic spermatogonia that fail to differentiate into spermatocytes	Reduced fertility rate (70%) Some were completely sterile	Yes	Yes	Wu et al., 2016
Zebrafish (Danio rerio)	TALEN target of the 1st exon of mettl3 that is injected into WT zebrafish embryos which are raised and outcrossed with WT to obtain F1 F1 outcrossed again for F2 then F2 self-crossed.	Sperm motility reduced (average path velocity, curvilinear velocity, and straight-line velocity) Decreased male GSI (0.62%) compared to WT (1.02%) Reduced # of mature sperm Increased proportions of spermatogonia (24.4% vs 7.5% in WT) and spermatocytes (56.1% vs 26.7% in WT) Decreased proportion of spermatozoa (10.4% vs 50.1% in WT)	Decreased standard breeding rates (48.8.% vs 91.4% in WT) 8.1% of mutant male x WT female spawned successfully vs 94.4% in WT	Yes	Yes	Xia et al., 2018	MEttl3 - multicomponent methyltransferase complex
Zebrafish (Danio rerio)	CRISPR/Cas9 gene targeting of E2f5	Reduced # of spermatozoa compared to WT Increased % of spermatocytes at leptotene and zygotene stages compared to WT Suggests arrest at zygotene stage due to strong y-H2AX staining during mid-leptotene stage Arrested during prophase I Increased apoptosis	Decreased fertilization rates (3% vs 94% in WT) under standard breeding conditions	Yes	Yes	Xie et al., 2020	E2f5 is a transcriptional repressor during cell-cycle progression
Marine medaka (Oryzias melastigma)	0.1 mg/L of DEHP for 6 months from larval stage	Increased GSI vs control (1.18 vs 0.81) Contained mostly spermatocytes (Sp) and spermatids (Sd) with few spermatozoa especially in this treatment	Decreased fertilization success vs control (84.12% vs 94.21%)	Yes	Yes	Ye et al., 2014	DEHP - phthalate MEHP - active metabolite of DEHP
	0.5 mg/L of DEHP for 6 months from larval stage	Increased GSI vs control (1.05 vs 0.81) Contained mostly Sp and Sd with few spermatozoa	Decreased fertilization success vs control (81.61% vs 94.21%)	Yes	Yes
	0.1 mg/L of MEHP for 6 months from larval stage	Increased GSI vs control (1.09 vs 0.81) Contained mostly Sp and Sd with few spermatozoa	Decreased fertilization success vs control (87.46% vs 94.21%)	Yes	Yes
	0.5 mg/L of MEHP for 6 months from larval stage	Increased GSI vs control (1.03 vs 0.81) Contained mostly Sp and Sd with few spermatozoa	Decreased fertilization success vs control (82.16% vs 94.21%)	Yes	Yes
Zebrafish (Danio rerio)	fshr mutant zebrafish line created using TALEN-induced gene knockout	At 40 dpf, control (heterozygotes) could induce spermatogenesis whereas mutants remained undeveloped At 45 and 65 dpf, testes in mutant males showed significant retardation in growth with delayed spermatogenesis and arrest at spermatogonial stage vs control who had mature spermatozoa At 90 dpf, testes of most mutant males were normal and could induce normal fertilization Testes of mutant males that failed to spawn showed retarded spermatogenesis, fewer spermatocytes and more abundant spermatogonia	At 90 dpf, 7 out of 10 fshr mutant males could induce spawning with normal fertilization	Yes	No	Zhang et al., 2015	fshr mutant zebrafish develop as all-male population and the mutants who could not induce spawning are believed to be females who had follicles arrested 105-d fshr mutant zebrafish well developed testis with degenerating oocytes
	lhcgr mutant zebrafish line created using TALEN-induced gene knockout	No observable differences between control and mutants	No observable differences between control and mutants	No	No
	Fshr and lhcgr double mutant zebrafish line created using TALEN-induced gene knockout	Arrest of spermatogonial stage with apoptotic oocyte-like germ cells early on Underdeveloped testis and retarded spermatogenesis at adult stage	At 90 dpf, double knockout completely infertile	Yes	Yes

Known modulating factors

This table captures specific information on the MF, its properties, how it affects the KER and respective references.1.) What is the modulating factor? Name the factor for which solid evidence exists that it influences this KER. Examples: age, sex, genotype, diet 2.) Details of this modulating factor. Specify which features of this MF are relevant for this KER. Examples: a specific age range or a specific biological age (defined by...); a specific gene mutation or variant, a specific nutrient (deficit or surplus); a sex-specific homone; a certain threshold value (e.g. serum levels of a chemical above...) 3.) Description of how this modulating factor affects this KER. Describe the provable modification of the KER (also quantitatively, if known). Examples: increase or decrease of the magnitude of effect (by a factor of...); change of the time-course of the effect (onset delay by...); alteration of the probability of the effect; increase or decrease of the sensitivity of the downstream effect (by a factor of...) 4.) Provision of supporting scientific evidence for an effect of this MF on this KER. Give a list of references. More help

Fertilization success in Chinook salmon (Oncorhynchus tshawytscha) was significantly biased towards the male whose sperm swam fastest in the female’s ovarian fluid (Rosengrave et al., 2016).
Seminal plasma pH(R2=0.525) is positively correlated with fertilization rate in Rainbow Trout (Oncorhynchus mykiss) and African catfish (Clarias gariepinus) (Lahnsteiner et al., 1998, Mansour et al., 2005).

Quantitative Understanding of the Linkage

Captures information that helps to define how much change in the upstream KE, and/or for how long, is needed to elicit a detectable and defined change in the downstream KE. More help

Quantitative understanding is shown bellow.

Response-response Relationship

Provides sources of data that define the response-response relationships between the KEs. More help

Lahnsteiner et al.(1998) determined that fertilization rate in Rainbow Trout (Oncorhynchus mykiss) can be described by sperm motility rate(y=0.72x * 25.99 where y is fertilization rate and x is sperm motility rate, R=0.594, P < 0.001), seminal plasma pH(R²=0.525, P < 0.001), and spermatozoal respiration activation(R²=0.554, P < 0.001). They found a positive correlation between % of motile spermatozoa and total swimming velocity with fertilization rate (P < 0.001) and % of immotile spermatozoa inversely. The 2 parameters accounted for 65% of total variance in fertilization rate.
Relative sperm velocity(p=0.008) and longevity (p < 0.0001) showed significant association with sperm competition success in Atlantic salmon (Salmo salar). Males with faster spermatozoa achieved greater fertilization success. (Gage et al., 2004)
Highly significant correlations were found between sperm motility (R=0.932, p < 0.001) and fertilization rate in Rainbow Trout (Oncorhynchus mykiss) (Ciereszko and Dabrowski, 1993).

Time-scale

Information regarding the approximate time-scale of the changes in KEdownstream relative to changes in KEupstream (i.e., do effects on KEdownstream lag those on KEupstream by seconds, minutes, hours, or days?). More help

The duration of spermatogenesis in humans (Homo sapiens) is reported to be 74 days (Griswold, M.D, 2016). Consequently, effects on spermatogenesis may not manifest as observable impacts on fertility until perhaps 74 days after impacts on spermatogenesis began. This may vary depending on the stage(s) of spermatogenesis that are impacted by the stressor.
The duration of the meiotic and spermiogenic phases in zebrafish (Danio rerio) is reported to be 6 days which means there could be a delay of at least 6 days before signs of impaired fertility may be detected (Leal et al., 2009).

Known Feedforward/Feedback loops influencing this KER

Define whether there are known positive or negative feedback mechanisms involved and what is understood about their time-course and homeostatic limits. More help

Feedforward/feedback loops haven’t been evaluated yet. However, given that fertilization pertains to the interaction between sperm and oocyte, it seems unlikely that fertilization rates (external to the male) would feedback on and impact spermatogenesis.

Domain of Applicability

A free-text section of the KER description that the developers can use to explain their rationale for the taxonomic, life stage, or sex applicability structured terms. More help

Taxonomic Applicability: Spermatogenesis is one of the most conserved biological processes from Drosophila to humans (Wu et al., 2016). As a result, animals who utilize sexual reproduction as their way to produce offspring are heavily reliant on spermatogenesis being effective and normal (Kang et al., 2015). There are studies on reproduction and spermatogenesis across a multitude of taxas.
Sex Applicability: Spermatogenesis is a male-specific process (Tang et al., 2018, Wu et al., 2015, Kang et al., 2015, Wang et al., 2015). Thus, the present relationship is only relevant for males.
Life Stage Applicability: Spermatogenesis and reproduction are only relevant for sexually-mature adults.