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Prince Henry’s Institute of Medical Research (L.O., K.M.R., M.E.J., E.R.S.) and Department of Biochemistry (K.M.R.), Monash University, Clayton, 3168, Victoria, Australia
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Abstract |
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 Top Abstract I. IntroductionII. Overview of SpermatogenesisIII. Biosynthesis and Action...IV. ERs, Aromatase, and...V. The Effects of...VI. Estrogen and Spermatogenesis...VII. Estrogen and Sexual...VIII. SummaryReferences |
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and/or ß and aromatase are also compared to
evaluate the physiological role of estrogen in male fertility. The
review focuses on the effects of estrogen administration or
deprivation, primarily in rodents, on the hypothalamo-pituitary-testis
axis, testicular function (including Leydig cell, Sertoli cell, and
germ cell development and function), and in the development and
function of the efferent ductules and epididymis. The requirement for
estrogen in normal male sexual behavior is also reviewed, along with
the somewhat limited data on the fertility of men who lack either the
capacity to produce or respond to estrogen. This review highlights the
ability of exogenous estrogen exposure to perturb spermatogenesis and
male fertility, as well as the emerging physiological
role of estrogens in male fertility, suggesting that, in this local
context, estrogenic substances should also be considered "male
hormones." I. Introduction
II. Overview of Spermatogenesis
A. Germ cell development
B. Regulation
III. Biosynthesis and Action of Estrogen
A. Estrogen biosynthesis
B. Mechanisms of estrogen action
IV. ERs, Aromatase, and Estrogen Production in the Testis
A. ERs and aromatase in the fetal testis
B. ERs and aromatase in the immature testis
C. ERs and aromatase in the adult testis
D. Estrogen production in the testis
V. The Effects of Estrogen Administration or Deprivation on Spermatogenesis
A. Hypothalamo-pituitary-testis axis
B. Efferent ductules and epididymis
C. Testicular descent
D. Leydig cells
E. Sertoli cells
F. Germ cells
G. Comparison of the spermatogenic phenotype of mice with targeted disruptions of ERs or aromatase
VI. Estrogen and Spermatogenesis in Humans
VII. Estrogen and Sexual Behavior
VIII. Summary
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I. Introduction |
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TopAbstractI. IntroductionII. Overview of SpermatogenesisIII. Biosynthesis and Action...IV. ERs, Aromatase, and...V. The Effects of...VI. Estrogen and Spermatogenesis...VII. Estrogen and Sexual...VIII. SummaryReferences |
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(ER)
(3, 4, 5, 6), or aromatase (7, 8, 9), along with the discovery of a second
estrogen receptor-ß (ERß) (10, 11, 12, 13), which is widely expressed in
the male reproductive tract (14). Another important reason for the increased interest in the role of estrogen in male reproduction stems from various reports that exposure to estrogens in the environment may have a detrimental effect on male reproductive development and health and may be related to the reported decreases in sperm counts over the past 50 yr (15, 16, 17). Although controversy exists as to whether male fertility over the past five decades has truly declined (18), and whether the exposure of humans to the relatively low levels of estrogens in the environment would cause significant health problems (19), research into the effects of estrogens on the male reproductive system is clearly warranted (19, 20). In agreement with the notion that estrogen exposure during development may impair male fertility, it has been known for many years that estrogen administration to experimental animals during the neonatal period or adulthood can impair sperm production and maturation (21, 22).
Normal male fertility relies on normal spermatogenesis, i.e., the process by which immature spermatogonia within the testis divide and differentiate into the mature elongated spermatid form that is subsequently released from the seminiferous epithelium. The full fertilizing potential of the released spermatozoan is also dependent on the progression and maturation of sperm through the excurrent duct system and the epididymis.
While the administration of estrogens and xenoestrogens during fetal
and neonatal development has been reported to be associated with a
series of male reproductive disturbances, such as cryptorchidism,
epididymal defects, impaired fertility, and an increased incidence of
testicular cancer (e.g., Refs. 23, 24, 25 and references
therein), an essential physiological role for estrogen in male
fertility was not identified until the early 1990s. The infertility in
mice lacking a functional ER (ERKO) was the first definitive
demonstration that estrogen was required for male fertility (3, 4, 5, 6).
ERKO mice are infertile primarily due to a defect in efferent
ductule development and function (6, 26). Mice lacking a functional
aromatase gene (aromatase knockout, ArKO) are also infertile; however,
this appears to be primarily due to a specific defect in germ cell
development (8). Thus lessons provided by mice transgenic for ER
and/or ERß [i.e., ERKO, ERßKO, and ERßKO mice
(see Ref. 27 for review)] as well as aromatase [ArKO mice (8, 9)],
provide compelling evidence for a role for estrogen in spermatogenesis
and male fertility.
Estrogen is clearly involved in numerous processes in the male, including bone turnover, behavior, and the cardiovascular system (see Ref. 2 for review). However, this review will focus on the effects of estrogen administration and deprivation on the production and function of spermatozoa. This review will address two important questions: 1) can exposure to estrogen or estrogenic substances interfere with spermatogenesis and male fertility? and 2) does estrogen have a physiological role in spermatogenesis and male fertility? Given that spermatogenesis relies on, among other things, the normal development and function of the hypothalamo-pituitary-testis axis, testicular cells (including Leydig cells, Sertoli cells, and germ cells), efferent ductules, and epididymis, the experimental evidence for the effects of, and/or a role for, estrogen in each of these systems will be presented. This review will also provide an overview of estrogen biosynthesis and action in general, and specifically in the testis. A brief overview of the phenotype of humans with mutations in ERs or aromatase will also be presented, as will an overview of a role for estrogen in sexual behavior, given that this is proving to be an important component of estrogen action in male fertility.
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II. Overview of Spermatogenesis |
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TopAbstractI. IntroductionII. Overview of SpermatogenesisIII. Biosynthesis and Action...IV. ERs, Aromatase, and...V. The Effects of...VI. Estrogen and Spermatogenesis...VII. Estrogen and Sexual...VIII. SummaryReferences |
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). When germ cell development is
complete, the mature spermatids are released from the Sertoli cells
into the tubule lumen, and proceed through the excurrent duct system,
known as the rete testis, until they enter the epididymis via the
efferent ducts (Fig. 1). During passage through the epididymis, the
spermatids undergo a series of biochemical changes to become the motile
spermatozoa capable of fertilization.
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), is enclosed by a capsule called the
tunica. The interstitial tissue contains the blood and lymphatic
vessels, which are essential for the movement of hormones and nutrients
into, and out of, the testis. The most frequently encountered cell type
in the interstitium is the Leydig cell (28), which is primarily
involved in the secretion of androgens, notably testosterone, as well
as other steroids including estrogen.
Within the seminiferous tubules, the Sertoli cells reside on a basement
membrane, under which are the lymphatic endothelium and the peritubular
myoid cells (29). The structure of the Sertoli cell is extremely
complex, with numerous cup-shaped processes encompassing the various
germ cell types (30) (Fig. 1). Developing germ cells form intimate
associations with Sertoli cells, with multiple germ cell types in
contact with one Sertoli cell. The various generations of germ cells,
as described below, are not randomly distributed within the
seminiferous epithelium, but are arranged in strictly defined cellular
associations (31). It is the unique associations of these germ cells
with Sertoli cells that constitute the cycle of the seminiferous
epithelium, and each particular association of germ cells is referred
to as a stage. The number of stages of spermatogenesis in a particular
species is thus defined by the number of morphologically recognizable
germ cell associations within the testis; in the mouse there are 12
stages, in the rat there are 14, and in the human there are 6 (32).
A. Germ cell development
Germ cell development involves a complicated series of events, and
the various germ cell types can be distinguished on the basis of
morphology and the differential expression of proteins.
Spermatogonia are present between Sertoli cells, close to the basement membrane of the tubule. They are the most immature germ cells in the testis, and include type A spermatogonia, intermediate spermatogonia (found only in rodents), and type B spermatogonia, the latter of which are considered to be committed to differentiation. The true stem cells of the germ cell population are considered to be a subset of the type A spermatogonial population, although their identity cannot be discerned on the basis of morphology (33). Spermatogonia undergo numerous mitoses to produce a large number of germ cells available for entry into meiosis. Thus, proliferation of the spermatogonial population provides the source for the millions of sperm that are produced per day (32).
After the last mitosis of type B spermatogonia, preleptotene primary spermatocytes are formed (32). These cells replicate their DNA and hence initiate meiosis (33). During the prophase of the first meiotic division, germ cells undergo morphological transitions that can be classified on the basis of nuclear size and morphology (34). In the zygotene phase, pairing of homologous chromosomes occurs, and cells with completely paired chromosomes are termed pachytene spermatocytes. After the pachytene phase, a brief diplotene phase follows in which the chromosome pairs partially separate, and the cells then undergo the first meiotic division to yield secondary spermatocytes. These cells quickly undergo the second meiotic division to yield the haploid round spermatid.
The differentiation of round spermatids into the mature elongated spermatid form takes place, with no further division, during the process known as spermiogenesis. Briefly, spermiogenesis involves formation and development of the acrosome and flagellum, condensation of the chromatin, reshaping and elongation of the nucleus, and removal of the cytoplasm before release of the spermatid during spermiation (31, 32). After commencement of spermatid elongation, the highly condensed spermatid nucleus becomes incapable of transcription, and immature round spermatids transcribe high levels of mRNAs that are subject to translational delay until translation is required during elongation (35). Spermiation is the final step of spermiogenesis and involves the release of the mature elongated spermatid from the Sertoli cell into the lumen of the seminiferous tubule (36).
B. Regulation
Germ cell development relies on a highly coordinated interaction
with the Sertoli cell. Germ cells and Sertoli cells can communicate
directly via ligand/receptor-mediated interactions or paracrine
factors. The production and secretion of many Sertoli cell proteins
involved in germ cell development occur in a stage-dependent manner
(37), reflecting the ability of the Sertoli cell to adapt to the
changing needs of the germ cell. For many years it was presumed that
Sertoli cells were the major controlling factor in the timing of germ
cell development; however, recent studies investigating rat-to-mouse
spermatogonial transplantation clearly demonstrated that rat germ cells
in contact with mouse Sertoli cells develop according to the kinetics
of rat spermatogenesis, thus highlighting the role of germ cells
in controlling their own fate (38).
As well as the production of spermatozoa, the testis is involved in the production of hormones that are required for various functions in the body, including maintenance of secondary sexual functions, and feedback on the hypothalamus and the pituitary to control the secretion of the gonadotropins LH and FSH.
It is well known that the gonadotropins are the major endocrine regulators of spermatogenesis (39, 40, 41). LH targets the Leydig cell to stimulate the secretion of androgens, namely testosterone, which in turn acts on androgen receptors in the seminiferous epithelium to control spermatogenesis. FSH targets receptors within the Sertoli cell to regulate spermatogenesis by stimulating the production of numerous Sertoli cell factors. The roles of testosterone and FSH in the testis have been studied extensively, yet relatively little is known about how these hormones act within the Sertoli cell to stimulate and maintain spermatogenesis (39, 40, 41). Androgens alone have been shown to stimulate all phases of germ cell development in the hypogonadal (hpg) mouse, which is congenitally deficient in GnRH and therefore LH and FSH (42), highlighting the requirement of spermatogenesis for androgen. The question of whether FSH was essential for spermatogenesis in mice was answered by the generation of transgenic mice possessing targeted disruptions of the FSH receptor gene (43) or the FSH ß-subunit gene (44). Males of both transgenic models are fertile and display all stages of germ cell development, as are the androgen-treated hpg mice (42), suggesting that FSH is not an absolute requirement for fertility. However, in both cases the testes are smaller, and less sperm are produced (42, 43, 44), due to the requirement for FSH during the neonatal period of Sertoli cell division (45, 46). More recent quantitative studies on FSH receptor knockout mice also demonstrated defects in sperm development, leading to the production of poor quality sperm (47). Thus while FSH is not essential for spermatogenesis, it is clearly essential for quantitatively normal spermatogenesis and fertility. In terms of the endocrine regulation of spermatogenesis by FSH, LH, and androgens, it is clear that the initiation and maintenance of quantitatively normal spermatogenesis and thus full fertility rely on the delicate balance of the hypothalamo-pituitary-testis axis. The focus of this review is the growing body of evidence that suggests that estrogen should be added to the list of hormones involved in the regulation of spermatogenesis.
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III. Biosynthesis and Action of Estrogen |
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TopAbstractI. IntroductionII. Overview of SpermatogenesisIII. Biosynthesis and Action...IV. ERs, Aromatase, and...V. The Effects of...VI. Estrogen and Spermatogenesis...VII. Estrogen and Sexual...VIII. SummaryReferences |
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The human CYP19 gene was cloned some years ago (50, 51, 52),
when it was shown that the coding region spans 9 exons beginning with
exon II. Upstream of exon II are a number of alternative exons I which
are spliced in the 5'-untranslated region of the transcript in a
tissue-specific fashion (Fig. 2). The
expression of the gene in the ovary and testis utilizes a proximal
promoter, promoter II (53, 54), and thus transcripts in gonadal tissues
contain sequence at their 5'-end that is immediately upstream of
the translational start site. In contrast, placental transcripts
contain at their 5'-end a distal exon, I.1, which is localized 100 kb
upstream from the start of translation in exon II. This is because
placental expression is driven by a powerful distal promoter upstream
of exon I.1 (55). Transcripts in adipose tissue contain yet another
distal exon located 20 kb downstream of exon I.1, exon I.4 (56), and a
number of other untranslated exons have been characterized including
one that is expressed in brain (57). Splicing of these untranslated
exons to form the mature transcript occurs at a common 3'-splice
junction that is upstream of the translational start site. This means
that although transcripts in different tissues have different
5'-termini, the coding region and thus the protein expressed in these
various tissue sites are always the same. However, the promoter regions
upstream of each of the several untranslated first exons have different
cohorts of response elements, and so regulation of aromatase expression
in each tissue that synthesizes estrogens is different. Aromatase
expression in the gonads is regulated by cAMP and gonadotropins due to
an interaction of the gonadal promoter II with the transcription
factors CREB (cAMP response element binding protein) and SP1 (58, 59).
For comparison, the adipose promoter, I.4, is regulated by class I
cytokines such as interleukin-6 (IL-6), interleukin-11 (IL-11), and
oncostatin M, as well as by tumor necrosis factor- (TNF). The
regulation of estrogen biosynthesis in each tissue site of expression
is unique and has been reviewed previously (60).
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. The
two receptors are not isoforms of each other, but rather are the
products of distinct genes located on separate chromosomes.
The ER proteins are each composed of six functional domains labeled
A–F, a signature characteristic of the entire superfamily (Fig. 3). The N'-terminal A/B domain is the
least conserved among all members and demonstrates only 17% homology
between the two ERs. It contains the activation function 1 (AF1)
region, which is one of two regions critical for the transactivation
function of the members of the receptor family. By contrast, the C
domain is the most highly conserved region, being the DNA binding
domain that contains the zinc-finger motifs. The E domain, or
ligand-binding domain, is modestly conserved throughout the superfamily
and confers ligand specificity on the members. Conservation of amino
acid sequence in this region is 60% between the ER and ERß;
however, each binds estradiol with about equal affinity, although the
relative binding of other ligands differs substantially between them
(64, 65). The E domain also contains the major dimerization surface of
the receptors, and the second transactivation function, activation
function 2 (AF-2), is also located in this region of the
C'-terminus.
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gene in vivo leads
predominantly to a single transcript of approximately 6.3 kb, encoding
a protein of 599 amino acids (66). The human ER is slightly shorter
at 595 amino acids (62). The existence of multiple promoter and
regulatory regions in the 5'-untranslated region has been reported, as
well as naturally occurring variants of the transcript in normal and
malignant tissues, but as yet the existence of true protein products of
these variants, as well as their significance, remains controversial
(67). Initial studies of the rodent ERß transcript indicated it was
substantially shorter than the ER, namely 485 amino acids (10, 11).
This is largely due to a significantly shorter N'-terminal region.
Indeed the existence of a functional AF-1 region in the ERß has been
questioned. However, open reading frames initiating upstream from that
first described have now been discovered (68), suggesting the
possibility that an ERß protein of approximately 530 amino acids
might also exist. Unlike the ER gene, Northern blots of rodent
tissues indicate the presence of multiple ERß transcripts, and a
number of variants have been described, including a conserved insertion
of 18 amino acids in a C'-terminal region in the rat, mouse, and human,
the deletion of one or more exons in these same species, and several
isoforms in the extreme C'-terminus of the human transcript (69, 70, 71).
There is considerable tissue specificity in the expression of ER and
ERß. Thus, ER is the dominant species expressed in uterus, liver,
adipose, skeletal muscle, pituitary, and hypothalamus, whereas ERß is
the major form in ovary and prostate, as well as other regions of the
brain including the limbic system, cerebellum, and cerebral cortex (72, 73). The localization of ER and ERß in the testis and epididymis
is discussed in Section IV.
The mechanisms of estrogen and estrogenic ligand action in the control of transcription have been recently reviewed elsewhere (74, 75, 76, 77, 78); however, the following will provide a brief overview of estrogen action on ER via classic pathways, ligand-independent ER activity, and nongenomic actions of estrogen.
The binding of ligand to ER and the events that lead to the regulation of target gene transcription are broadly similar among all steroid receptors, namely that they bind to response elements on the target gene and form a platform to which is recruited the complex of coactivators and transcription factors required for transcriptional activation. In general, estrogen action on ER involves ligand binding, dissociation of chaperone complexes and receptor phosphorylation; receptor dimerization; nuclear translocation; DNA binding and interaction with cofactors; and modulation of transcriptional activity.
Although ER and ERß bind estradiol with similar affinity, there is
considerable selectivity of the different receptor subtypes in terms of
affinities of various ligands (65). In particular, some phytoestrogens,
such as genistein and coumestrol, have a significantly higher affinity
for ERß than ER (64).
ER subtypes form homo- or heterodimers in vitro or in cells.
In general terms, the ER homodimer is more transcriptionally active
than the ERß homodimer in most systems (see Ref. 76 for review). The
ER/ERß heterodimer was recently shown to have similar
transcriptional activity to the ER homodimer under
hormone-saturating conditions, but is less active than the ER
homodimer at subsaturating estradiol concentrations (79).
The fact that there are distinct differences between the ER subtypes
with regard to agonistic/antagonistic effects on transcription is now
well known. For example, the ER agonistic activity of tamoxifen appears
unique to ER (see Ref. 76 for review). The distinct properties
exhibited by ER and ERß have been exploited for therapeutic use by
the development of selective ER modulators (SERMs) (see Refs. 74, 78
for review); however, an understanding of the molecular events
underlying SERM pharmacology has only recently begun to emerge (74, 78, 80). The molecular mechanisms underlying the differences in the
functional characteristics of ER and ERß are well beyond the scope
of this review, but include differences at the level of ligand
binding/affinity, cofactor recruitment, and activity of the AF-1 and
AF-2 domains (which are influenced by cell and promoter context) (see
Refs. 74, 75, 76 for review). For example, the relative expression levels
of the respective ER subtypes within a cell are key determinants of
transcriptional activity in response to agonists and antagonists (79).
Another way in which ER subtypes exhibit functional differences is in
their ability to recruit coactivators and corepressors (74, 75, 76, 80, 81). Examples of coactivators of ERs are the steroid receptor
coactivators (SRCs) (82) (see Ref. 83 for review). Often, coactivators
such as SRC1 facilitate transcription via histone acetylation activity,
which results in decondensation of chromatin and hence increases the
ability of the transcriptional machinery to interact with the promoter
(see Ref. 76 for review). In contrast to coactivators, corepressors
generally bind to ER in the absence of ligand or to antagonist-occupied
ER and repress transcriptional activity. These frequently possess, or
else activate, histone deacetylase activity. Examples of corepressors
include nuclear receptor corepressor (NCoR) (84) and short heterodimer
partner (SHP) (85, 86). The differences in transcriptional activation
of ER compared with ERß by agonists and antagonists is due, at
least in part, to differences in the ability of the ligand-receptor
complex to recruit coactivators, which is related to the conformational
changes induced in the helix H12 of the ligand binding cavity of each
receptor by the agonist/antagonist (80). In summary, the ability of an
ER subtype to stimulate transcription is dependent on promoter context,
the nature of the ligand bound, the expression of coactivator and
corepressor proteins, and the relative expression levels of ER subtypes
in a given cell. It is also worthwhile to note that the ability of a
ligand for the ER to influence transcription is determined by its
structure, its affinity for ER subtypes, and the recruitment of
coactivators and corepressors that is, in turn, dependent on cell and
promoter context (see Ref. 78 for review).
ERs can also associate with target gene promoters in a manner that is
not dependent on direct DNA binding, but involves the binding of ER to
proteins within a preformed transcriptional complex. For example,
ligand-activated ER can positively regulate transcription by
associating with the activating protein 1 (AP1) transcription factor
complex in target cells (see Ref. 77 for review). Similar interactions
of ER with other transcriptional complexes exist (see Ref. 74 for
review) indicating that ER can interact directly or indirectly with DNA
to modulate transcription. ER and ERß exhibit different properties
in both DNA binding-dependent and -independent mechanisms (see Ref.
76).
There is now also increasing evidence for ligand-independent
transcriptional activation of ERs (87, 88). Epidermal growth factor
(EGF)-induced phosphorylation of ER results in ligand-independent
transcriptional activity, a pathway that has been shown to have
functional significance (87, 89, 90). Growth factor-induced
phosphorylation of ER is dependent on the mitogen-activated protein
kinase (MAPK) pathway (see Ref. 91 for review). MAPK
phosphorylates the AF-1 region of ER (92) and ERß (93) and in the
latter case has been shown to promote the recruitment of SRC1 in a
ligand- independent manner (93). Other factors, such as cyclin D1,
can activate ER-mediated transcription in the absence of estradiol (94, 95).
In addition to the classic genomic pathway of ligand- occupied ER
interaction with target genes, estrogen can also induce extremely rapid
(within seconds to minutes) increases in the concentration of calcium
or cAMP second messengers (96, 97) in an apparently nongenomic
mechanism of action (see Ref. 98 for review), presumably via receptors
on the plasma membrane. Also, physiological concentrations of estradiol
can induce a rapid release of nitric oxide in endothelial cells via
membrane-bound receptors (99, 100). Nongenomic actions of estrogen
appear to be of particular importance in cardiovascular (101) and
neuronal (102) tissues. Evidence has been presented that in some
systems estrogen appears to act on a membrane-associated ER that is
immunologically related to ER (103, 104, 105, 106, 107). ERß transcripts can also
produce membrane-associated forms (104). However, in other systems,
nongenomic effects of estrogen do not appear to be via
membrane-associated forms of ER or ERß (108, 109, 110, 111). For example,
nongenomic actions of estrogen on pancreatic ß-cells appear to be via
a membrane binding site with the pharmacological profile of the
-adrenergic receptor (110). Therefore, the potential exists for
estrogen action on plasma membrane receptors/complexes unrelated to
ER or ERß.
In summary, ligand-bound ERs act as transcription factors and are
capable of modulating target gene transcription by both DNA
binding-dependent and -independent means. Molecular mechanisms of
estrogen action are now known to be more complex than originally
thought, with ligand-independent pathways that mediate cross-talk with
growth factor signaling pathways, as well as rapid nongenomic actions.
Understanding how estrogen will act within a cell is further
complicated by the fact that ER and ERß differ in their functional
characteristics and in their tissue localization. Thus, estrogenic
ligand-dependent and/or ER-dependent regulation of target gene
expression depends on the nature of the ligand, the relative levels of
ER subtypes and cofactors, the molecular pathway of action and, in
particular, the cell and promoter context.
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IV. ERs, Aromatase, and Estrogen Production in the Testis |
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TopAbstractI. IntroductionII. Overview of SpermatogenesisIII. Biosynthesis and Action...IV. ERs, Aromatase, and...V. The Effects of...VI. Estrogen and Spermatogenesis...VII. Estrogen and Sexual...VIII. SummaryReferences |
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, ERß, and aromatase mRNA and protein in the
testis from the fetal stage through to adulthood will be presented. The
reader is also directed to other relevant reviews (113, 114, 115, 116) for the
discussion of ER and aromatase localization in the male reproductive
tract.
A. ERs and aromatase in the fetal testis
ERs and aromatase are found at all stages of testicular
development in the rodent. For more detailed information of male
gonadal differentiation, see Refs. 117, 118, 119 .
Immunohistochemical studies using antibodies that recognize ER show
that this protein is present in the mouse undifferentiated gonad at day
10 (120) and day 11.5 (121), suggesting that estrogen may have a role
very early in the differentiation process. Leydig cells within the
rodent fetal testis contain ER until birth (120, 121, 122, 123, 124, 125). In fact, ERs
are expressed in the Leydig cells at a stage in development when the
androgen receptor is not yet expressed (126), highlighting a role for
estrogen at this stage (see Ref. 125). Although one study reports that
ER is present within the seminiferous tubules of the fetal testis
(120), other studies do not (see Table 1). ER is also present in the
developing efferent ductules and epididymis (120, 121, 125).
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), and this receptor subtype appears to remain in these cells until
birth (122, 123, 124, 127). However, it is the gonocytes that express ERß
in higher abundance than the other testicular cells (123, 127),
possibly suggesting a direct role in gonocyte maturation. ERß is also
present in rat associated ducts, which will eventually differentiate
into the efferent ductules and adjoining epididymis (124, 125). The fetal rat testis also has aromatase activity (128, 129, 130), which is first expressed by day 19 (129). The cell type responsible may be Sertoli cells, as estrogen production by fetal testes in culture is stimulated by FSH (131). Interestingly, this is close to the time when FSH receptors first appear on the Sertoli cells (132). There is also evidence that fetal Leydig cells have the capacity for LH-stimulated aromatization of androgens (133).
Thus the demonstration that rodent fetal testicular cells synthesize
estrogen and express both ER and -ß suggests a role for estrogen
in the development of the fetal male reproductive tract and in gonocyte
differentiation.
B. ERs and aromatase in the immature testis
Around the time of birth, the testis continues to express both ER
isoforms and aromatase (Table 1). Most reports suggest that ERß
protein and mRNA, rather than ER, appear to be localized to the rat
seminiferous epithelium, in both Sertoli cells and developing germ
cells (123, 127) (Table 1). In the mouse, the testis at this time also
continues to abundantly express ERß mRNA, with immunocytochemical
studies localizing this ER to the germ cells (122).
ER expression is restricted to the cells that lie outside of the
seminiferous epithelium shortly after birth (121, 125), with ER
(121, 122, 123, 124, 125), along with ERß (123, 127), continuing to be found in the
Leydig cells of mice and rats during the neonatal period. ER protein
is also found in the rodent rete testis (125), efferent ductules (121, 125), and the epididymis (121, 124, 125), where it has remained since
fetal development. In fact, ER is expressed in much higher levels in
the efferent ducts than the cauda epididymis (121, 125). ERß is
also found in the epididymis during development (122, 124, 134).
At this stage of development, basal aromatase activity is found in both the immature Leydig cells and Sertoli cells (135, 136, 137); however, activity is significantly induced by FSH in the Sertoli cells (135). In fact at this age, Sertoli cells were more active in producing estrogen than neonatal Leydig cells and adult Sertoli cells, suggesting that these cells are an important source of estrogen in the postnatal testis (137). At this age, germ cells have been reported not to contain detectable aromatase (138).
During days 10–26 in the immature rat, Leydig cells and Sertoli cells
are dividing and undergoing functional maturation (see Ref. 139 for
review). Again, ER is absent from the seminiferous epithelium with
only ERß prominent in the epithelium of the immature rodent (Table 1). Specifically in the tubules of the rat, ERß mRNA and protein are
present in relatively low levels in spermatogonia (123, 127) and
immature Sertoli cells (123, 127), and by day 21 ERß is abundantly
expressed in pachytene spermatocytes (123). Other germ cells do not
show ERß staining at this time (127). By day 12 in the mouse, ERß
is also specifically immunolocalized to the spermatocytes; however, its
expression appears to decrease and was undetectable by day 26 (122).
Again, rat and mouse Leydig cells express ER at this time (122, 123, 124, 125)
(Table 1).
During the neonatal and pubertal period of development, ER is
prominent in the rete testis and efferent ductules (125, 134) and is
present in the mouse epididymis (122). Recent studies on the
immunoexpression of ERs in the rat described the specific cellular and
regional localization of ER and ERß in the developing rat efferent
ductules, epididymis, and vas deferens and showed that the localization
of ER in the epididymis is confined to a relatively short window of
development (134). ERß is also present in the epididymis at this time
(122, 134).
Aromatase appears to have an age-dependent pattern of expression. As the animal matures, the Leydig cells appear to significantly supplement estrogen production from the Sertoli cells (136), with Leydig cell basal aromatase activity increasing 3- to 4-fold (137) and now stimulated by LH (135). However, the Sertoli cells do continue to express aromatase during their maturation (135).
C. ERs and aromatase in the adult testis
The presence of ER, ERß, and aromatase in the adult testis
has been the subject of numerous recent studies. An intense effort has
focused on the expression and localization of ER subtypes in the adult
testis of mice (73, 140), rats (14, 123, 125, 127, 141, 142), primates
(125, 143, 144), and humans (13, 145, 146, 147), primarily due to the fact
that there has been a considerable amount of conflicting data.
Various evidence indicates that Leydig cells of rats and mice express
ER (125, 142, 146) (see Table 1); however, the localization of ER
in Leydig cells of primates and humans is more controversial; studies
have shown immunostaining in some primate Leydig cells (125), and in
human Leydig cells (147); however, other studies could not detect ER
immunostaining in Leydig cells from primates (144, 146) or humans (146, 147). In the adult mouse ERß protein is expressed in the
Leydig cells (140), but this does not seem to be the case in the adult
rat (127, 142). Again, the localization of ERß in Leydig cells from
primates and humans is controversial with some studies showing no ERß
in Leydig cells from primates (143) or humans (13), whereas other
studies showed immunoexpression of ERß in Leydig cells from two
primate species (146) as well as humans (146, 147, 148).
By adulthood, rodent Leydig cells express a high level of aromatase (115, 138, 149, 150, 151), which is stimulated by LH (136, 152, 153) and steroids (153). In fact, aromatase activity is higher in the adult than at any other age (136) and is higher in the adult Leydig cells than in the Sertoli cells (149). The decrease in aromatase activity during Sertoli cell maturation into the adult form (see Ref. 154 for review) may be related to the control of Sertoli cell division and adult cell function (see Section V.E). The presence of aromatase in the Leydig cells of primates and humans is well established (see Ref. 115 for review).
Rat Sertoli cells contain both ERß mRNA and protein from the fetus to
adulthood (Table 1 and Section IV.A and B) (14, 123, 127, 141, 142). Immunohistochemical studies suggest that ERß is
not stage dependent in the rat (14, 123). In contrast to the rat, there
are no reports of ERß in mouse Sertoli cells (Table 1). Sertoli cells
in primates (146) and humans contain ERß (146, 147, 148) but not ER
(125, 146, 147).
There is now considerable evidence that germ cells contain both ERs and
aromatase (see Fig. 4 for a summary of
the proposed localization). In general, ERß is the predominant, and
potentially the only, ER in germ cells (see Table 1). One study in rats
found immunoexpression of ER in spermatocytes and spermatids (142);
however, there are no other reports of ER in rodent germ cells (see
Table 1). ER immunoexpression in human germ cells within
seminiferous tubule fragments in culture has been suggested by one
study (145), whereas other studies did not show ER protein in
primate or human germ cells (146, 147). ERß is present in
rat type A spermatogonia (127) as well as in intermediate and type B
spermatogonia in some studies (123), but not others (127) and is also
seen in spermatogonia in monkeys and humans (146, 148).
ERß is found in pachytene spermatocytes and round spermatids, but not
elongating spermatids in rats (123, 127, 141) and in primates and
humans (13, 143, 146, 148), although one study suggested ERß in human
elongating spermatids (145). However, one study in mice found ERß
only in elongated spermatids and not earlier germ cells (140) (see
Table 1). Other studies in rats and humans found ERß in Sertoli cells
but not in germ cells (142, 147). A further complexity to the
testicular localization of ERß is that one study in mice could not
detect ERß mRNA at all in mouse testis by RNase protection assay
(73). Therefore, considerable conflict in the literature exists in
terms of ERß localization in the testis (see Table 1 and below).
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). Aromatase localization is
observed to move from the Golgi apparatus to the cytoplasm during
spermatid development (150). When round spermatids begin the
morphological transformation into elongated spermatids, aromatase
continues to be found in these cells (Table 1) and is immunolocalized
to the flagella of the developing spermatid (150). Aromatase appears to
be present in higher levels in mature spermatids of the rat than in
earlier germ cells (149, 155). Aromatase mRNA and activity was higher
in the germ cells of the mouse and rat when compared with Leydig cells
(150, 155, 157), suggesting that the germ cells are an important source
of estrogen in the testis.
When elongated spermatids are released from the epithelium, during the
process of spermiation, aromatase remains in the residual body that is
subsequently phagocytosed by the Sertoli cell (150, 157). However, not
all the cytoplasm is phagocytosed, and aromatase activity remains in
the cytoplasmic droplet that is still attached to the flagellum as the
sperm make their way through the epididymis (151, 158). Thus it appears
as if mature sperm are able to synthesize their own estrogen, as they
traverse the efferent ducts (116, 151, 158) (see Fig. 4). The ability
to synthesize estrogen gradually decreases as the droplet slowly moves
to the end of the tail during epididymal transit until it is finally
lost (158). The demonstration of aromatase in sperm is important as it
suggests that the sperm themselves could control the levels of estrogen
present in the luminal fluid, directly modulating functions such as the
reabsorption of fluid from the efferent ductules (116).
A very high level of expression of ER is seen in the efferent
ductules of the rat (125, 159). In fact, it has been found that it is
the efferent ductules that possess the highest level of ER
immunostaining, relative to the testis, excurrent ducts, and
epididymis, throughout life (125). In addition, the efferent ductules
appear to be the first male reproductive structure to express the ER in
fetal development (160), suggesting a role for estrogen in the
development of this tissue. ER is present in the rat and mouse
epididymis (73, 124, 159), although in low levels, while another report
could not detect it in rats (125). Again, similar to earlier ages, both
ERß mRNA and protein are present from the efferent ducts to the
epididymis of the mouse (73, 140) and the rat (124, 159). A very recent
study of the localization of ER and ERß in the rat efferent
ductules and epididymis showed that in adult rats ER was strongly
immunoexpressed in the epithelial cells of the efferent ductules but
was not found in the epididymis, whereas ERß was found in epithelial
cells and some periductal cells throughout the efferent ductules and
epididymis (134). ER is found in the nonciliated epithelial cells of
the efferent ductules in primates and humans (146), although another
study in humans did not observe ER immunoexpression in the efferent
ductules (147). ER was rarely detected in the epididymis of primates
or humans (146, 147). In contrast, ERß is found in both stromal and
epithelial cells throughout the efferent ductules and epididymis in
these species (146).
The studies reviewed in this section highlight the fact that there is
considerable conflict in the literature regarding the localization of
ER and ERß, particularly in the testis. The conflict usually
arises when comparing studies utilizing immunohistochemical techniques.
In the studies cited in Table 1 and above, there is considerable
variation in the antibodies used. The antibodies have been raised
against different regions of the ER molecules, and thus detect
different epitopes (see Ref. 161 for a recent review on ER and ERß
antibodies, including some of those used in the studies cited here).
Most studies cited here have used antigen retrieval methods and have
presented some evidence of antibody specificity, usually by
preabsorption controls. Even though various studies used commercially
available antibodies, it is clear that a range of commercial antibodies
to ERs are not well characterized (see Ref. 161 for review). In
addition, whether the antibody used detects ligand-occupied and/or
inactive receptor, monomeric or dimerized receptors, an ER in
heterodimer form, receptors bound to DNA, etc., is usually not known. A
further complexity in the immunohistochemical studies cited in this
review is that different methods of tissue fixation, antigen retrieval,
antibody dilution, and antibody detection methods were used, all of
which can affect the ability of an antibody to bind to its antigen.
Thus at present it is almost impossible to reconcile the differences in
the various reports on ER and ERß localization. Clarification of
the exact localization of ER subtypes in the testis would be
facilitated by the comparison of a panel of very well characterized
antibodies, in which the exact nature of the protein detected is known.
In summary, the studies reviewed above suggest that the testis is
capable of synthesizing and responding to estrogens throughout all
stages of development. The localization of ER, ERß, and aromatase
in the adult testis is summarized in Fig. 4 in an effort to bring
together the literature in this area. The localization of ER, ERß,
and aromatase demonstrates that estrogen action is likely to be
important for Leydig cell, Sertoli cell, and germ cell development and
function, as well as in the development and function of the efferent
ductules and epididymis. In particular, germ cells are capable of local
estrogen synthesis and response, via ERß, suggesting that paracrine
and intracrine actions of estrogens may be important in male germ cell
development. The localization of aromatase in sperm in the testis, and
as they traverse the efferent ductules and epididymis, together with
the demonstration of high levels of ER and ERß in the efferent
ductules, support the hypothesis that estrogen in sperm acts on ER in
the efferent ductules (see Section V.B).
D. Estrogen production in the testis
The synthesis of estrogens in the testis has been reviewed
extensively elsewhere (154, 162); therefore, only a few relevant
details will be considered here. There is a high concentration of
estrogen in rete testis fluid (163) and, in the rat, the concentration
of estrogen in the caput epididymis is approximately 25 times the level
measured in plasma (164). It is clear that the concentrations of
estrogen in the testis and rete testis fluid far exceeds the
concentration in male serum in various species (see Ref. 162 for
review) thus suggesting a central role for estrogen in testicular and
epididymal function.
In dissected testicular tissue from adult rats, the concentration of estrogen in interstitial tissue was 9 times higher than that in the seminiferous tubules (165). However, it is now becoming clear that the level of aromatase activity in germ cells of the adult rodent is equal to or higher than the aromatase activity in Leydig cells (149, 150) (see above), suggesting that while Leydig cells have previously been considered to be the primary source of estrogen in the testis (166), germ cells must now be considered to have an important role also (see Refs. 114, 115, 116 for review, and Section IV.C). Thus the source of the high concentration of estrogen in fluid leaving the testis may be due largely to the high levels of aromatase mRNA, protein, and activity in testicular germ cells and particularly in spermatids (149, 150, 151, 157, 158, 167, 168) (see Refs. 114, 115, 116 for review).
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V. The Effects of Estrogen Administration or Deprivation on Spermatogenesis |
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TopAbstractI. IntroductionII. Overview of SpermatogenesisIII. Biosynthesis and Action...IV. ERs, Aromatase, and...V. The Effects of...VI. Estrogen and Spermatogenesis...VII. Estrogen and Sexual...VIII. SummaryReferences |
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A. Hypothalamo-pituitary-testis axis
As mentioned in Section II.B, the initiation and
maintenance of spermatogenesis require the secretion of gonadotropins
from the pituitary and thus is dependent on the balance of the
hypothalamo-pituitary-testis axis. The negative feedback effect of
testosterone on both the hypothalamus and the pituitary to regulate
gonadotropin secretion is well known. In humans, the negative feedback
of testosterone on the hypothalamo-pituitary axis to inhibit secretion
of both LH and FSH is the basis for the current approach to male
contraception (169). It is now becoming clear that a major component of
the negative feedback action of androgens on gonadotropin secretion is
mediated via aromatization to estrogen (170, 171, 172, 173). In particular,
studies in humans showed that administration of estradiol could further
enhance gonadotropin suppression that was induced by a
testosterone-based contraceptive (174), further indicating estrogen’s
role as a negative feedback regulator of gonadotropin secretion. The
demonstration of ER and ERß in the rodent hypothalamus and
pituitary (73, 142, 175, 176), of ERß in the monkey pituitary (143),
together with the immunolocalization of aromatase in the brain and in
particular the hypothalamus (177), indicates that estrogen has an
important role in these tissues.
During pubertal development in the rodent, numerous studies have shown that neonatal exposure to either estrogens or estrogen-like compounds promotes changes in gonadotropin secretion (178, 179, 180) and can, in fact, alter the organization of the hypothalamo-pituitary-testis axis so that changes persist into adulthood (181). A single high dose of estradiol benzoate to 1-day-old male rats causes a reduction in both GnRH secretion and pituitary responsiveness to GnRH (182), as well as the profound suppression of circulating FSH, LH and, consequently, testosterone levels (180). Interestingly, recent studies administering low doses of estrogenic compounds during the neonatal period in rats could actually stimulate serum FSH levels during puberty, an effect that could not be explained by changes in inhibin B (178). Given that the appropriate concentrations of LH and FSH, as well as a tightly regulated onset of secretion during the neonatal and pubertal periods, is fundamental to whether normal spermatogenesis proceeds (183, 184), neonatal estrogen exposure can have important long-term effects on the hypothalamo-pituitary-testis axis and thus spermatogenesis. Interestingly, whether or not estrogen administration to juvenile mice will interfere with the hypothalamo-pituitary-gonadal axis appears to be strain-dependent (185), which could lead to confusion when interpreting the literature on the interaction between estrogen, the regulation of pituitary hormone production, and fertility.
In the adult, there are many examples of a role for estrogen in the regulation of gonadotropin secretion. Adult male rats given increasing doses of estradiol for 10 days showed significant decreases in circulating concentrations of FSH and LH, which leads to subsequent reductions in serum and testicular testosterone levels (186). Surprisingly, the authors of this study also noted a stimulatory effect of low doses of estradiol on FSH, as was demonstrated by studies in the neonate (178), and in adult hypogonadal (hpg) mice given reasonably physiological doses of estradiol (187), indicating that estrogen can participate in both negative and positive effects on the pituitary in the male. Some of the effects of estrogen on FSH secretion may be mediated by its ability to promote changes in the in vitro Sertoli cell production of inhibin B (188), which is an important mediator of FSH secretion in males (Ref. 189 and references therein). Interestingly, estrogen has also been shown to increase the expression of the ßB-subunit of inhibin B in breast cancer cells (190). Therefore, estrogen may play a role in the regulation of this peptide that is primarily involved in mediating pituitary gonadotropin secretion, but may also have other roles in the testis (see Ref. 191 for review).
While estrogen administration clearly causes decreases in circulating gonadotropin levels, the administration of aromatase inhibitors causes increases in serum LH and testosterone in adult dogs (192) and rats (193), and in serum testosterone and the responsiveness of Leydig cells to a bolus injection of LH in monkeys (194). An increase in the circulating concentration of FSH was also seen in rats treated with an aromatase inhibitor (193). Administration of an aromatase inhibitor to men causes increases in the circulating concentrations of LH, FSH, and testosterone (172, 195). An interesting study in which the effects of administration of an aromatase inhibitor was compared in normal vs. GnRH-treated hypogonadotropic-hypogonadal men demonstrated that estrogen acts at the hypothalamus to decrease GnRH pulse frequency and at the pituitary to decrease responsiveness to GnRH (173). Thus, in the human, it is clear that estrogen is an important regulator of the hypothalamo- pituitary-gonadal axis in both sexes.
Investigation into the role of estrogen in the control of the
hypothalamo-pituitary-testis axis has been facilitated by transgenic
mouse models (see Table 2). As would be
predicted from aromatase inhibitor studies, male mice that lack a
functional aromatase gene (ArKO) have increased levels of serum LH and
testosterone yet normal levels of FSH (8, 196). Although ER is
present in the mouse hypothalamus and is the only ER in the mouse
pituitary (73), male mice deficient in ER (ERKO) showed
surprisingly little change in LH and FSH levels, although serum
testosterone levels were higher (5). A later study with a larger number
of animals showed that ERKO males had a 2-fold increase in both
circulating LH and in the pituitary content of LH ß mRNA, but
confirmed that ERKO animals had no change in FSH (197). An elegant
series of experiments performed by Lindzey and colleagues (197), and
reviewed in detail elsewhere (27), suggested that estrogen is likely to
be involved in facilitating the negative feedback effects of
testosterone on the male mouse hypothalamo-pituitary-testis axis and
further demonstrated that ERKO animals may have an increased
sensitivity to androgens, suggesting an altered organization of the
hypothalamo-pituitary-testis axis. The endocrine profile of the ERßKO
has not been detailed; however, the fact that ERßKO males are fully
fertile suggests no endocrine disruption (198).
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B. Efferent ductules and epididymis
After the release of mature spermatids from the Sertoli cell
during spermiation, spermatids via the seminiferous tubule fluid
proceed into the rete testis. Arising from the rete testis are a series
of tubules known as the efferent ductules, which connect the rete
testis to the initial segment at the head of the epididymis (see Ref.
199 for review). The primary function of the efferent ductules is to
resorb water, ions, and proteins, and various studies have shown that
approximately 90% of rete testis fluid is resorbed within these
ductules, so that spermatozoa become concentrated as they enter the
epididymis, thereby ensuring that a large number of spermatozoa are
released upon ejaculation (Refs. 162, 199, 200, 201 and references
therein). The passage of sperm through the initial segment, caput,
corpus, and cauda epididymis is essential for the final maturation of
sperm. The epididymis synthesizes and secretes numerous proteins and
also actively endocytoses substances from the epididymal lumen to
remove secreted proteins that are no longer required (see Refs. 202, 203 for review). Thus, the resorption of fluid through the efferent
ductules, as well as the attainment of various morphological,
biochemical, and motile properties during passage through the
epididymis, is fundamental for adequate sperm content of the ejaculate
and for full fertilizing capabilities.
Whereas a role for androgen in the regulation of epididymal function is
well known (204, 205, 206, 207), studies in the 1970s showed that there is a high
concentration of estrogen-binding sites in the immature and adult
epididymis (208, 209, 210), suggesting a role for estrogens in sperm
maturation and male reproduction. The concentration of estrogen leaving
the testis is far higher than in the circulation (see Section
IV.D and Ref. 162 for review), and estrogen receptors (ER and
ß) are present in the efferent ductules and epididymis (73, 125, 134, 140, 159, 160, 211, 212, 213) (see Section IV). In the rat
efferent ductules, ER is expressed at 3.5 times the level seen in
uterus (159). The presence of abundant estrogen and ER in the efferent
ductules and epididymis points to a role for estrogen in the regulation
of these tissues and thus in modifying sperm maturation and function.
A study by McLachlan and colleagues in 1975 (24) showed that prenatal exposure of mice to diethylstilbestrol (DES) caused epididymal granulomas, suggesting an action of exogenous estrogen on epididymal development. Numerous studies since have shown that neonatal estrogen exposure causes an impairment of efferent ductule and epididymal development and function that can lead to deleterious effects on fertility in adulthood (e.g., Refs. 134, 214, 215, 216, 217, 218, 219, 220). While at least part of these effects could be attributed to changes in the hypothalamo-pituitary-gonadal axis, a direct action on the epididymis to mediate these changes is likely given the high levels of ER expression.
Although the above studies indicate that estrogen administration can
affect the development of efferent ductule and epididymal function and
lead to an impairment of male reproductive function, the first major
insights into the mechanism of estrogen action in the efferent ductules
and epididymis, and the requirement for estrogen in male fertility,
were gained from the ERKO mouse. The ERKO male mouse is
infertile, due to disruptions to spermatogenesis, reduced
epididymal sperm content, reduced sperm motility and fertilizing
ability, as well as defects in reproductive behavior (4, 5, 221) (see
Ref. 27 for review and Table 2 and Section V.G). Studies by
Hess and colleagues (6) clearly demonstrated that dysfunction of the
efferent ductules of the ERKO mice contributed to the impairment of
fertility. ERKO testis weights initially increase during
postpubertal testicular development, due to an accumulation of fluid in
the lumen of the seminiferous tubules (5, 6), suggesting either an
excess of fluid secreted by the testis and/or a failure of fluid
resorption by the efferent ductules. In vivo efferent
ductule ligation experiments showed that the first possibility did not
occur, since ERKO testes secreted less fluid than wild-type animals
(6). Isolated efferent ductules were then cultured and ligated at
either end in vitro to analyze the removal of luminal fluids
by the efferent ductule epithelium. While ligated wild-type ductules
efficiently remove luminal fluid and collapse in vitro, the
efferent ductules of the ERKO mice were shown to swell, indicating
that fluid resorption by the efferent ductules in the ERKO mice was
impaired (6). Interestingly, efferent ductules isolated from wild-type
animals treated in vivo with the dual ER antagonist ICI
182,780, did not swell to the same extent as ERKO tissues,
suggesting that estrogen acting via ERß may contribute to efferent
ductule function (6). There have been no reports of efferent ductule
and epididymal dysfunction in ERßKO mice that exhibit normal
fertility (27, 198). However, more recent studies in which the effects
of the dual ER/ERß antagonist ICI 182,780 on adult efferent
ductule function in vivo were compared with the phenotype of
ERKO animals showed that both ICI-treated and ERKO animals
displayed similar disruptions to the rete testis and efferent ductules,
suggesting that ER, rather than ERß, plays a major role in these
tissues (26). The fact that ERßKO mice appear to have a similar
phenotype to ERKO mice (222) could be interpreted to mean that ERß
may not have a major role. However, detailed comparisons of the
efferent ductule and epididymal morphology and function in ERKO,
ERßKO, and ERßKO animals are required to confirm the involvement
of ERß in these tissues.
Recent studies on the morphology of the efferent ductules of the
ERKO mouse indicated a multitude of defects including decreases in
efferent ductule epithelial cell height and number of cilia, and a loss
of endocytotic apparatus, whereas examination of the epididymis also
demonstrated abnormalities of epithelial cells and sperm granulomas
(223). These observations highlight the importance of ER in the
adult function of the efferent ductules and epididymis but in
particular point to a physiological role for estrogen in the
development of these tissues. A second important study compared ER
antagonist administration to postpubertal mice with ERKO mice at a
similar age to demonstrate that the development of blind-ending
efferent tubules and an unusual growth of initial segment epithelial
cells was a consequence of a congenital absence of estrogen action,
while the dysfunction of efferent ductule epithelial cells is primarily
due to a lack of estrogen action in the adult (26). Thus, the
developmental abnormalities in efferent ductules and epididymal tissues
in ERKO mice demonstrate a physiological role for estrogen in the
development of these tissues.
Although a congenital absence of ER leads to permanent defects in
efferent ductule development and function, so too does neonatal
exposure to highly estrogenic compounds (134, 215, 216). Male rats
exposed to a high dose of diethylstilbestrol (DES) during the
first few days of life show dilation of the rete testis and an
accumulation of fluid in the testis during development (215, 216),
eventually leading to a deleterious effect on the seminiferous tubules
in adulthood (216). Fisher and colleagues (215) showed that this
treatment causes a permanent reduction in the water channel protein
aquaporin-1, which is found at high levels in the ER-bearing
nonciliated epithelial cells lining the efferent ductules. They
suggested that the efferent ductule dysfunction induced by neonatal DES
exposure is related to the absence of aquaporin-1, which may be
required for fluid movement across the efferent ductule epithelium.
Importantly, the effects of DES treatment were compared with neonatal
treatment with a GnRH antagonist to show that the effect of DES on the
development and function of the efferent ductules is via an estrogenic
action, rather than decreases in LH, FSH, or androgens (215). Further
studies showed that neonatal exposure to other estrogenic compounds has
similar effects on the development and function of the efferent
ductules, and the magnitude of the effect of each compound was
comparable to its in vitro estrogenic activity (214). More
recent studies show that neonatal exposure to DES caused various
histological changes as well as changes in the cell- and
region-specific expression of ER protein in the efferent ductules as
well as in the epididymis and vas deferens (134), highlighting the fact
that the development of these tissues, and indeed the pattern of ER
expression, is a target for exogenous estrogen action. While neonatal
estrogen exposure produces developmental defects in the efferent
ductules and epididymis, short-term treatment of adult rats with
estradiol benzoate also disturbs efferent ductule function (224).
It is perhaps surprising that a congenital absence of ER and
neonatal, or indeed adult, exposure to high levels of estrogens produce
similar effects on the development and function of the efferent
ductules. However, the fact that both models involving contrasting
changes in estrogen during the neonatal period produce the same
dysfunction in adulthood underscores the importance of estrogen during
the development of these tissues. One proposition for the similar
effects of neonatal estrogen exposure and deficiency is that
inappropriate estrogen exposure may lead to down-regulation of ERs
resulting in an estrogen deficiency syndrome similar to that seen in
the ER null mice. Such a proposition is supported by studies in the
uterus, in which developmental exposure to DES clearly leads to
decreases in ER levels in adulthood, and hence the uterus is
permanently unresponsive to estrogen (225, 226). Similarly, studies in
the testis suggest that neonatal DES exposure leads to down-regulation
of ER and androgen receptor, but an increase in ERß (180), perhaps
suggesting a permanent change in either estrogen responsiveness, or in
estrogen-dependent gene expression. The proposition that ERs in the
efferent ductules and epididymis are down-regulated by neonatal DES
treatment, however, is not supported by immunolocalization studies in
DES-treated rats. One study has shown that ER immunostaining intensity
in the efferent ductules is increased by neonatal DES treatment (220),
perhaps suggesting that the epithelial cells would become
hypersensitive to the already highly potent agonistic action of DES. A
second study showed that the morphological abnormalities in epididymal
development induced by neonatal DES treatment were associated with
changes in ER, but not ERß, immunoexpression, such that ER
became more widespread in its expression pattern (134). Indeed,
neonatal DES treatment leads to reduced androgen receptor expression in
male rat reproductive tracts, an effect that was shown to be directly
attributed to DES, rather than changes in androgen (227). This study
showed that DES-induced changes in the efferent ductules and epididymis
may be due, in part, to an altered androgen-estrogen balance (227).
Perhaps the observations on efferent ductule development and function
in both the neonatal estrogen exposure and ER null mice models will
be reconciled when a better understanding of the relative expression of
the ER subtypes during the development of these tissues, and of the
downstream targets of estrogen action in these tissues, is gained.
In vitro studies on the interactions of estrogenic ligands
and ERs show clearly that the relative expression of both ER and
ERß in a given cell type will determine the sensitivity of that cell
to estrogen as well as determine how the cell responds to receptor
agonists and antagonists (79).
A further complexity to the above account of the involvement of
estrogen in efferent ductule and epididymal function is that the ArKO
mouse did not show obvious fluid retention and disturbance of fluid
resorption (8), as would be predicted by the ERKO phenotype. This
could be due to the fact that germ cell development is compromised in
the ArKO (8), and therefore perhaps less fluid is being secreted by the
seminiferous tubules in these animals. The demonstration that wild-type
efferent ductules treated with a dual ER and ERß antagonist
in vitro did not swell like ERKO tubules (6) predicts
that the lack of estrogen action on both ER subtypes would lead to a
less pronounced effect on efferent ductule fluid resorption compared
with ER alone, and thus these observations could be extended to the
interpretation of the ArKO phenotype. In vivo ER antagonist
studies showed that while the effect on efferent ductules and rete
testis was similar to ERKO animals, most parameters measured showed
a less profound effect in ER antagonist-treated animals compared with
ERKO (26). It is thus possible that the blockade of ERß action
along with ER may cause less pronounced effects on the efferent
ductules; however, the possibility that the less profound effects of
the antagonist are due to incomplete blockade of estrogen action in
these tissues cannot be ruled out. Also the difference between the
ERKO and the antagonist-treated mice (26) could also be related to
the lack of ER during development compared with a lack of ER in
adulthood. Since ArKOs would have access to maternal estrogen during
development, perhaps the lack of estrogen in adulthood would cause less
profound effects on efferent ductule function than a lack of estrogen
(or ER) during development. An alternative explanation for the lack
of obvious efferent ductule dysfunction in the ArKOs is that ERs in
these tissues may be activated by ligand-independent means, or else
that alternate ligands for these receptors could lead to continued
fluid resorption by the efferent ductules (see Section V.G).
There are also studies to show that disruption of estrogen in adulthood can compromise efferent ductule and/or epididymal function and fertility. The administration of an aromatase inhibitor to adult male monkeys caused apparent defects in epididymal sperm maturation since sperm had compromised motility and condensation (194). Chronic aromatase inhibitor treatment in adult rats, however, did not compromise fertility, although a few rats showed evidence of a disturbance in efferent ductule fluid resorption (193). Interestingly, adult mice that had been treated for 35 days with an ER antagonist remained fertile, despite compromised efferent ductule function (26). While estrogen deficiency in the adult promotes efferent ductule and epididymal dysfunction, studies by Meistrich and colleagues (22) showed that estradiol benzoate given to adult mice for 50 days had negligible effects on sperm number in the testis but clearly reduced the sperm content of the epididymis and increased the time taken for sperm to transit through the epididymis. The authors demonstrated a decrease in sperm maturation and suggested that estradiol administration increases epididymal transport rate, with the result that sperm spend less time in the epididymis and cannot fully mature.
In summary, this section has highlighted the importance of the normal
development and function of the efferent ductules and epididymis for
spermatogenesis and fertility, and in particular the influence of
estrogen on these tissues. Estrogen overexposure during neonatal
development and in adulthood leads to changes in efferent ductule and
epididymal tissues that can produce permanent defects in their function
and hence have deleterious effects on male fertility. Importantly,
studies from ER null mice demonstrate that estrogen plays a crucial
role in the development and normal functioning of these tissues that
has profound implications for male fertility.
C. Testicular descent
Testicular descent is an important aspect of male sexual
development, and fetal Leydig cells produce factors, notably androgens,
that are necessary for this process. The relocation of the testes from
the urogenital ridge to the inguinal abdominal wall (transabdominal
descent) and the subsequent migration of the testis into the scrotum
(inguinoscrotal descent) is a hormonally regulated process (see Ref.
228 for review).
The undescended testes and resultant infertility in mice lacking
functional androgen receptors or type 2 5-reductase underscores the
importance of androgens in testicular descent (228, 229, 230, 231); however, it
is well known that in utero exposure of male fetuses to high
levels of estrogens can also interfere with this process. For example,
the treatment of pregnant women with DES is associated with
cryptorchidism of the male offspring (25).
Animal models have been used to study estrogen-induced cryptorchidism, and several studies have suggested that estrogen exposure may interfere with the fetal hypothalamo-pituitary-testis axis, leading to an inhibition of fetal Leydig cell androgen production, thus interfering with testicular descent (see Ref. 113 for review). However, ERs are present in fetal Leydig cells of the rat (see Section IV) and thus a direct action of estrogen on these cells is possible. Insight into the mechanism by which in utero estrogen exposure causes cryptorchidism comes from recent studies on the insulin-3 (Insl3) gene. Targeted disruption of this gene, which is specifically expressed in fetal Leydig cells, causes bilateral cryptorchidism (232, 233). Since both in utero exposure to estrogen and lack of a functional Insl3 gene causes cryptorchidism, it is possible that Insl3 plays a role in estrogen-induced cryptorchidism. Indeed, two recent studies have clearly demonstrated that in utero exposure to 17ß-estradiol or DES causes a specific down-regulation of Insl3 transcription in fetal Leydig cells (234, 235). Steroidogenic factor-1 (SF-1) is an important transcriptional activator of Insl3 in fetal Leydig cells (236), and therefore estrogen may down-regulate Insl3 transcription via SF-1. While one study has shown that in utero estrogen exposure decreases testicular SF-1 expression (237), other studies have shown that the estrogen-induced decrease in Insl3 was not coincident with a decrease in SF-1 (234, 235), and therefore the exact mechanism by which estrogen regulates fetal Leydig cell Insl3 transcription is unclear. Thus an important consequence of estrogen overexposure during the prenatal period is a decrease in transcription of Insl3 in fetal Leydig cells, leading to cryptorchidism and deleterious effects on fertility. Given that cryptorchidism is also associated with an increased risk of testicular cancer (228), inappropriate estrogen exposure is an important consideration for the etiology of this disease. Indeed, overexpression of aromatase in a mouse model has been shown to result in testicular Leydig cell tumors (238).
Estrogen deprivation during fetal development is also associated with
problems with testicular descent. Although ERKO mice have descended
testes, defects in cremaster muscle development was noted in these
animals, indicating a role for ER in some aspects of male
reproductive tract development and testicular descent (239). A male
patient deficient in ER had bilaterally descended testes (240).
Aromatase-deficient mice also do not appear to have defects in
testicular descent (8, 196); however, it is likely that these mice are
subjected to maternal estrogens in utero. There are no
reports of undescended testes in three male patients identified with
aromatase deficiency (241, 242, 243); however, preliminary reports from our
laboratory of a recently identified aromatase-deficient man indicate
that this patient is cryptorchid (our unpublished data).
D. Leydig cells
Leydig cells first appear in the testis during day 15 of embryonic
development in the rat (see Ref. 244 for review). These fetal Leydig
cells secrete high concentrations of androgens that are required for
Wolffian duct development and subsequent male sexual development. The
fetal Leydig cells present at birth are not progenitors of the adult
Leydig cell population; rather they remain present in low numbers in
the mature testis, presumably in a quiescent state (245). During the
prepubertal period, there is a rapid growth of Leydig cells, which
arise from mesenchymal precursor cells, while after about day 28 in the
rat, morphologically recognizable Leydig cells divide to produce the
adult Leydig cell population (246) (see Ref. 139 for review). Leydig
cells through pre- and postnatal development differ in their morphology
as well as function (see Refs. 28, 139, 247 for review). In the
adult, perhaps the most notable function of the Leydig cell is to
produce androgens that are necessary for spermatogenesis and the
maintenance of secondary sexual functions. The following section will
briefly review the evidence for a role for estrogen in Leydig cell
function. The reader is also directed to another relevant review (113).
The proliferation of precursor and adult-type Leydig cells during a defined period of pubertal development is important for the establishment of the adult complement of Leydig cells. Estrogen appears to play an inhibitory role in this process and therefore may be important in controlling the steroidogenic capacity of the adult testis. Neonatal estrogen exposure can interfere with Leydig cell development and proliferation during puberty (see Ref. 113 for review). Using ethane dimethansulfonate (EDS) to cause the destruction of Leydig cells as a model to study Leydig cell regeneration and development, estrogen was shown to block Leydig cell regeneration, probably via the inhibition of the rapid phase of Leydig cell proliferation (248). The fact that estrogen treatment between days 5–16 after EDS treatment was most effective at inhibiting Leydig cell regeneration suggests that estrogen can act on precursor Leydig cells (248). It has been suggested that, while LH is clearly important in regulating Leydig cell development, locally produced factors also appear to be involved (249, 250). These studies, together with the fact that more mature Leydig cells have higher levels of aromatase activity (251), have led to the hypothesis that mature Leydig cells may produce estrogens that inhibit precursor Leydig cell development (248). Consistent with this hypothesis is the fact that precursor Leydig cells have 20 times the level of ER mRNA than do mature Leydig cells (252). Also of interest is the fact that neonatal estrogen exposure, at a time when Leydig cells are developing, results in permanently decreased serum testosterone levels in the adult rat, despite no decreases in LH, suggesting that inappropriate exposure to estrogens during Leydig cell development can cause permanent changes to Leydig cell function (181). Taken together, these studies lend support to the hypothesis that estrogen may act in a paracrine fashion in the testis to control Leydig cell development (see Ref. 113 for review).
In addition to the potential role for estrogen in controlling Leydig
cell development, there is also evidence for estrogen acting as a
paracrine factor in the control of adult Leydig cell steroidogenesis.
Although LH is the primary driving force for testosterone production,
the theory that intratesticular factors are required to modulate Leydig
cell steroidogenesis, such as by mediating LH responsiveness, does have
merit (see Ref. 253 for review). Estrogen has been shown to inhibit
Leydig cell steroidogenic enzymes that are required for testosterone
biosynthesis. For example, estrogen has been shown to inhibit P450
17-hydroxylase/C17,20 lyase activity in both
neonatal (254, 255) and postpubertal (256) testes. DES can decrease
testosterone production in adult rats in the absence of changes in LH
(257). Estrogens and xenoestrogens have also been shown to inhibit
androgen production by testicular tissue from the Atlantic croaker
amphibian, and this effect appears to be predominantly via a nongenomic
mechanism (258). In humans, estrogen can directly inhibit testicular
steroidogenesis, and at least part of this action may be by altering
Leydig cell LH receptors (259). Indeed, there is a suggestion that
estrogen may be involved in mediating Leydig cell responsiveness to LH
(260) (see Ref. 113 for review), which may partially explain the
reported increase in Leydig cell responsiveness to LH in monkeys given
an aromatase inhibitor (194). Given that estrogen appears to have an
inhibitory action on adult Leydig cell steroidogenesis, it is
interesting to note that androgens, produced in high concentrations by
adult Leydig cells, have been shown to regulate Leydig cell production
of estrogen sulfotransferase, which is an enzyme involved in the
sulfurylation and inactivation of estrogens (261). Finally, it should
be mentioned that ArKO mice show evidence of Leydig cell
hyperplasia/hypertrophy after 18 weeks of age (8); however, this is
likely to be a consequence of the elevated LH levels in these
animals.
Thus, normal Leydig cell function and development are important for male sexual development, testicular steroidogenesis during puberty and adulthood, and hence normal fertility. The demonstration of estrogen modulation of, and action at, each stage of Leydig cell development suggests that estrogen exposure could have important consequences for Leydig cell function and hence for male fertility.
E. Sertoli cells
The proliferation of Sertoli cells occurs from day 16 of fetal
life in the rat and reaches a maximum 2 days before birth (262).
Approximately 1 million Sertoli cells are present in the rat testis at
birth and, with the continued proliferative activity of these cells,
albeit at a declining rate (262), the numbers increase to a maximum of
around 40 million at day 15 of postnatal life (263). After postnatal
day 15, proliferation ceases (262), differentiation commences (see Ref.
139 for review), and the number of the Sertoli cells in the testis
remains stable throughout adulthood (263) (see Ref. 264 for review). It
is well known that this period of Sertoli cell proliferation, and the
postproliferative differentiation and maturation of these cells, is
essential for the full spermatogenic potential of the adult. This is
perhaps best illustrated by experiments showing changes in the
testicular size and spermatogenic potential of the adult after the
alteration during puberty of factors that either potentiate Sertoli
cell proliferation (265, 266) or interfere with or delay their
maturation (179, 267).
In the following section, a role for estrogen in the control of Sertoli cell proliferation and differentiation will be discussed, and therefore it is pertinent to briefly review the hormonal control of these processes. Sertoli cell proliferation is likely controlled by numerous factors, including pituitary hormones and intratesticular factors (see Ref. 264 for review). The importance of FSH in this phase of development was demonstrated by the administration of human recombinant FSH to rats during the neonatal period, resulting in an increased number of Sertoli cells and an increased spermatogenic potential of the adult (266), and by the fact that neonatal administration of human recombinant FSH to hpg mice also stimulated Sertoli cell numbers (46). While FSH is thus a mitogenic factor for neonatal Sertoli cells (see Ref. 264 for review), thyroid hormone is thought to inhibit Sertoli cell division but promote differentiation. This knowledge comes from the observation that transient neonatal hypothyroidism results in enlarged testes and enhanced production of sperm (268). The mechanism behind this observation is now known to be via a direct action of thyroid hormone on Sertoli cells to inhibit Sertoli cell proliferation and stimulate differentiation (265, 269). Thyroid hormone acts directly on its receptor, which is expressed at high levels in proliferating Sertoli cells, and then declines toward the end of the proliferative period (270). Thus it is clear that endocrine signals control Sertoli cell division and differentiation.
Sertoli cells produce considerable amounts of estrogen during the period of division, leading to the suggestion that estrogen is involved in this process. Aromatase activity is highest in Sertoli cells from prepubertal rats, declines as Sertoli cells mature, and is hormonally regulated, principally by FSH (see Ref. 154 for review). Indeed, the FSH-induced aromatase activity and the measurement of estradiol produced by primary cultures of Sertoli cells from immature rats is the basis of a bioassay for FSH (271). ERs are present in Sertoli cells throughout development and appear to be primarily of the ß-form (123, 127, 142, 143) (see Section IV). Dorrington and colleagues (154, 272) have proposed that estradiol, along with FSH, may be a mitogen for Sertoli cell division and have demonstrated that in granulosa cells of the ovary, estrogen induced TGFß, which in turn stimulated DNA synthesis. This suggests that in these somatic cells of the gonads, FSH may induce aromatase activity and hence estrogen production, which stimulates TGFß, which then, along with FSH, promotes cell division (272). The fact that similar control mechanisms exist in Sertoli cells led these authors to hypothesize that estrogen may participate in the FSH-mediated mitogenic activity on Sertoli cells via induction of TGFß (154, 272).
Confirmation of whether estrogen has an action on Sertoli cell division
may be facilitated by ER and aromatase null mice models. There are no
descriptions of Sertoli cell numbers in ERKO, ERßKO, or ERßKO
animals. Sertoli cell numbers in ArKO mice were not different from
wild-type animals in our initial published studies (8) although very
recent preliminary data on larger numbers of animals suggest an
increase in Sertoli cell numbers in ArKO animals (our
unpublished data), thus lending support to the hypothesis that estrogen
may act to control the neonatal period of Sertoli cell division and
differentiation.
Estrogen may also be involved in the postproliferative period of Sertoli cell maturation. Although studying the direct effects of exogenous estrogen treatment on Sertoli cells is complicated by disruptions to circulating hormones (see Section V.A), some studies have shown a specific action of estrogen in Sertoli cell development. Sharpe and colleagues (179) compared the effect of neonatal exposure to DES to a GnRH antagonist administered for the same period to dissect out the specific actions of inappropriate estrogen exposure during days 2–12 of life in the rat. These studies showed that while DES and GnRH antagonist decreased FSH levels and Sertoli cell numbers, DES caused a more profound delay of Sertoli cell maturation, as evidenced by the immunolocalization of Sertoli cell proteins, and permanent defects in spermatogenesis and testicular histology in adulthood (179), suggesting that exogenous estrogen has an inhibitory role in Sertoli cell maturation.
The theories put forward by Sharpe et al. (179), Dorrington
and Khan (154), and others that estrogen has a stimulatory effect on
Sertoli cell division yet a negative effect on Sertoli cell
differentiation and development is supported by various observations:
1) the demonstration that estrogen production is high in proliferating
Sertoli cells yet is lower in Sertoli cells with a more differentiated
morphology (135, 154, 273); 2) toward the end of the period of Sertoli
cell proliferation, FSH-induced aromatase activity starts to decline,
partly due to a decreased Sertoli cell responsiveness to FSH (see Refs.
154, 272 for review); 3) coincident with the fall in estrogen
production and Sertoli cell mitotic activity, TGFß, which may
stimulate Sertoli cell mitosis in response to estrogen, also declines
(154); 4) thyroid hormone, which stimulates Sertoli cell
differentiation (265, 269), decreases aromatase activity in prepubertal
Sertoli cells (274); 5) the capacity of Leydig cells to produce
testosterone increases from about day 14 postpartum (247), and androgen
has been shown to inhibit Sertoli cell aromatase activity either
directly (275) or via androgen-mediated effects on peritubular cells
(276), suggesting that androgens from the maturing Leydig cells may
participate in the down-regulation of aromatase during the switch from
Sertoli cell division to differentiation; 6) various growth factors
produced by the Sertoli cell can control Sertoli cell estrogen
production and responsiveness to FSH and could therefore be involved in
switching off aromatase during the period between division and
differentiation (154); 7) germ cells, which are starting to develop
during the switch between Sertoli cell division and differentiation,
decrease Sertoli cell aromatase activity (277). Finally, perhaps the
most interesting observation that supports an inhibitory role of
estrogen in Sertoli cell differentiation is the fact that cells with
the morphological features of a differentiated Sertoli cell are found
in the ovaries of mice lacking both functional ER and ERß
(ERßKO mice) (222, 278) as well as in ovaries from ArKO mice
(279). This observation suggests that the removal of estrogen or
estrogen-induced factors is required for granulosa cell survival and
differentiation, and the loss of estrogen results in an environment in
which Sertoli cells are able to differentiate, presumably in this case
from granulosa cells.
Finally, it is of interest to note that estrogen is thought to have a role in regulating the expression of the cell adhesion molecule neural cadherin (NCad) in the immature mouse testis (280) and in cultured mouse Sertoli cells (281). Given that NCad is thought to be important for cell-cell interactions in the testis, particularly between germ cells and Sertoli cells (282, 283), this may be one way in which estrogen is involved in establishing and maintaining the seminiferous epithelium.
In summary, the normal proliferation, differentiation, and functional maturation of Sertoli cells is essential for the initiation of spermatogenesis and the full spermatogenic potential of the adult. Estrogen administration studies show that Sertoli cell proliferation and function can be affected by exogenous estrogens and estrogen-like substances, leading to permanent defects in reproductive function in adulthood.
F. Germ cells
Germ cell development involves a series of mitotic and meiotic
divisions and differentiation from the immature spermatogonia into the
specialized elongated spermatid. The development of germ cells is well
known to be dependent on the action of FSH and testosterone on the
Sertoli cell, and both of these hormones have been shown to prevent
germ cell apoptosis as well as to potentiate division and/or
differentiation (see Refs. 39, 40, 41 for review). In addition to the well
documented hormonal control of spermatogenesis by androgens and FSH,
evidence for a direct role for estrogen in mediating germ cell
proliferation, viability, and function is now emerging.
While it is known that neonatal estrogen exposure leads to permanent
defects in germ cell development in adult rats (179), it is possible
that this is due to permanent defects in Sertoli cell function rather
than an effect on germ cells themselves. A direct action of estrogen on
germ cells, however, is entirely possible given the localization of
aromatase and ERs during various stages of germ cell development (see
Section IV). Therefore, a paracrine action of estrogen from
the Sertoli cells on the germ cells is possible, as is an action of
estrogen that is produced within germ cells (see Fig. 4). Although the
actions of estrogen on Leydig and Sertoli cells appear to be mainly
inhibitory (see Sections V.D and E), there is
accumulating evidence that estrogen has a predominantly stimulatory
effect on germ cells.
Studies in which estrogen was administered to rats during the neonatal period between days 5 and 11 showed that the numbers of undifferentiated and differentiating type A spermatogonia were increased at day 15 of life (284). Although these studies suggest a stimulatory role for estrogen in spermatogonial division, it is difficult to conclude whether this effect is direct, or via perturbation of the hormonal signals from the pituitary (see Section V.A). However, in vivo and in vitro studies in Japanese eels suggest a direct stimulatory effect of estrogen on these cells, since spermatogonial renewal in this species was stimulated by estrogen but blocked by the estrogen antagonist tamoxifen (285). Studies using rat gonocytes (or prespermatogonial cells) in culture clearly demonstrated an effect of estrogen in stimulating gonocyte proliferation (286). The proliferation of these spermatogonial precursor cells, as evidenced by 5-bromo-2'-deoxyuridine (BrDU) incorporation, was stimulated by a 1 µM dose of estradiol but not by higher doses, and the stimulatory effect was blocked by the ER antagonist ICI 164,384. The effect of estradiol was mirrored by that of platelet-derived growth factor (PDGF) and, since the addition of estradiol and PDGF in combination did not have additive effects, the authors postulated that estradiol and PDGF acted via a similar mechanism to stimulate gonocyte proliferation (286). Given that aromatase activity in the Sertoli cells is high during the neonatal period when gonocytes are proliferating and differentiating into spermatogonia, and that gonocytes and differentiating spermatogonia during the early neonatal period have been shown to contain ERß (122, 123, 127) (see Section IV), a direct action of estrogen in stimulating precursor germ cell mitosis is possible.
Recent evidence for a direct role for estrogen in preventing germ cell apoptosis was gained from studies using human adult seminiferous tubules cultured in vitro (145). When seminiferous tubules were cultured in the absence of serum and survival factors, spermatocyte and spermatid apoptosis was induced. The apoptosis of these cells could be prevented by low doses of estradiol, or higher doses of dihydrotestosterone, suggesting that estradiol is a potent inhibitor of germ cell apoptosis. The effect of estradiol was rapid, within 4 h, leading the authors to speculate that at least part of the effect may be mediated by a nongenomic action of estrogen; however, the authors used immunocytochemistry to show that ERs were present within these cells (145).
The fact that round spermatid apoptosis has been shown to occur in the seminiferous tubules of older ArKO mice (8) also highlights a role for estrogen in acting as a spermatid survival factor. In these mice, the numbers of spermatogonia and spermatocytes did not differ compared with wild-type animals; however, significant decreases in round and elongated spermatid numbers were seen after 18 weeks of age. In situ detection of apoptotic cells suggested that numerous round spermatids undergo apoptosis, leading to decreases in the spermatid numbers (8). In agreement with this finding, adult monkeys treated with an aromatase inhibitor showed a decrease in the conversion of round to elongated spermatids as evidenced by flow cytometry, and a decrease in sperm output from the testis, also suggesting that estrogen is important for spermatid differentiation (194, 287). The fact that aromatase deficiency leads to defects in spermatid differentiation is also supported by earlier studies in which a granulosa cell-secreted factor, purified to homogeneity and shown to have aromatase-inhibitory activity, induced round spermatid degeneration and a decrease in mature spermatids in adult male rats (288, 289).
A surprising finding in ArKO mice was that round spermatids that did
not undergo apoptosis early in spermiogenesis had acrosomal dysgenesis
(8). The acrosome is a vesicle associated with the postmeiotic
spermatid nucleus and contains a number of hydrolytic enzymes that are
required for the sperm’s penetration of the zona pellucida of the
ovum. The acrosome arises from the Golgi complex and, during acrosome
biogenesis, proacrosomic granules in the Golgi coalesce to form a
single large vesicle that becomes closely associated with the spermatid
nucleus in the early stages of spermiogenesis (see Ref. 139 for
review). The acrosomal vesicle gradually spreads out and flattens over
the nucleus, in a process that involves anterograde and retrograde
vesicular trafficking between the Golgi and the developing acrosome
(290). The observation of abnormal acrosome development in the ArKO
mouse suggests that acrosome biogenesis could be an estrogen-dependent
process. This hypothesis is supported by the immunolocalization of high
levels of aromatase in the Golgi complex of the developing spermatid
(150), as well as the presence of ERß in spermatids (see
Section IV and Table 1).
The stimulatory effect of estrogen on spermatogenesis was recently investigated in detail by the administration of estradiol to hpg mice (187), which lack FSH and LH due to a congenital deficiency of GnRH. The testes of the postpubertal hpg mouse are underdeveloped with spermatogenesis arrested at the early stages of germ cell development; however, the administration of estradiol-filled SILASTIC implants to these mice for a period of 70 days was able to induce all stages of spermatogenic cell development (187). This somewhat surprising finding strongly suggests that estrogen is capable of inducing spermatogenesis. Although the literature reviewed above suggests that estrogen can have a direct action on germ cells, the estrogenic induction of spermatogenesis in the hpg mouse may also have been due to an indirect effect via the stimulation of low levels of FSH. The relatively physiological levels of estrogen present in these mice after 35–70 days of treatment were associated with a significant stimulation of FSH, presumably by a direct action at the pituitary. Although the levels of FSH were approximately one-third of the levels in wild-type animals, an effect of these low levels of circulating FSH on stimulating germ cell development cannot be ruled out (187). Despite the obvious caution needed when studying the hormonal regulation of spermatogenesis in a mouse congenitally deficient in all gonadotrophic stimulus, the induction of spermatogenesis by estrogen in this model provides evidence that estrogen can stimulate male germ cell development.
Recent studies have suggested that estrogen is involved in the function
of mature spermatozoa. The incubation of human spermatozoa in the
presence of estrogen is known to stimulate various sperm functions
including motility and lactate production (see Ref. 98 for review).
Evidence has been presented that the stimulatory actions of estrogen on
human spermatozoa are via a membrane-associated ER, which is a 29-kDa
protein that is recognized by an antibody to the ligand-binding domain
of the human genomic ER (107). The action of estradiol on this
receptor is apparently nongenomic, as the effect was rapid (within
minutes) and involved a rapid influx of calcium. Estradiol action via
this receptor causes changes in tyrosine phosphorylation of various
proteins and inhibited the nongenomic actions of progesterone such as
the progesterone-induced acrosome reaction (107).
Although the above studies suggest a role for estrogen in germ cell
development, it should be pointed out that recent reports show that the
administration of a dual ER/ERß antagonist ICI 182,780 to
wild-type mice for 35 days did not produce observable changes in the
morphology of the seminiferous epithelium, apart from the expected
distention of the rete testis (26). However the ability of the ICI
antagonist to cross the blood-testis barrier and enter the adluminal
compartment of the seminiferous epithelium to locally block estrogen
action has not been demonstrated, nor has the ability of the antagonist
to fully block the high levels of estrogen that are present in the
testis.
Therefore, the studies presented above provide evidence of a stimulatory role for estrogen in germ cell development including spermatogonial division, germ cell viability and differentiation, acrosome biogenesis, and function of spermatozoa.
G. Comparison of the spermatogenic phenotype of mice with targeted
disruptions of ERs or aromatase
Previously published studies have described the fertility and/or
testicular phenotypes of the ERKO (5, 6, 27, 278, 291), ERßKO
(198, 278), ERßKO (222, 278), and ArKO (7, 8, 9, 196) mice.
Although the phenotypes of these animals have been mentioned above, a
more direct comparison between the phenotypes of animals in which
aromatase or the ERs are inactivated is of interest when reviewing
the evidence for a role for estrogen in spermatogenesis (see Table 2).
Although spermatogenesis is clearly disrupted in the ERKO mouse (5),
the lack of germ cell development and the reduction in mature
spermatids in the epididymis can be primarily attributed to compromised
fluid resorption due to defective efferent ductule function (5, 6, 27, 223) (see Section V.B). Efferent ductule ligation
experiments show that the seminiferous epithelium in the ERKO
secretes significantly less fluid than wild-type animals (6), and this
could be attributed to either fluid build-up causing Sertoli cell
dysfunction or a direct effect due to the lack of ERKO in the
testis. It seems likely that the spermatogenic phenotype in the ERKO
mice is primarily due to an indirect effect via efferent ductule
dysfunction, rather than a direct effect on spermatogenesis (6, 27).
Studies by Mahato and colleagues (292) showed that the infertility in
the ERKO mice is not due to a defect within the ER null germ
cells themselves, since transplantation of ERKO germ cells to
wild-type mice depleted of germ cells demonstrated that ER-deficient
germ cells can develop in an environment in which aromatase and ERs are
present. The fact that mouse germ cells do not require ER for
development is perhaps not surprising because mouse germ cells do not
appear to contain ER, nor do germ cells in other species (see Table 1 and Section IV).
The lack of any apparent spermatogenic phenotype in mice lacking a
functional ERß (198, 278), however, is surprising, given the fact
that ERß appears to be the only ER present in the germ cells, and the
predominant, if only, subtype in the Sertoli cell (see Section
IV). Indeed in most species, ERß appears to be the most
predominant and more widely expressed ER in the testis. The absence of
spermatogenic disruption in these mice is not readily explicable, but
could perhaps be due, in part, to a compensation by ER, since ER
expression and localization in ERßKO testes have not been explored.
In the efferent ductules of the ERKO mice, ERß has been shown to
have a different subcellular localization compared to wild-type tissues
(140). Thus, perhaps there is a permanent change in the localization of
ER in the ERßKO testis. The fact that germ cells from ERKO mice
can develop normally when transplanted in wild-type mice (292),
together with the fact that ERßKO animals do not show direct
disruptions to spermatogenesis (see Ref. 27 for review; see Table 2),
raises questions as to the role of ERs in germ cells. However, the
potential action of estrogen in germ cells via nonclassical receptors
cannot be ruled out. As discussed above, nongenomic actions of estrogen
have been implicated in Leydig cell function (258), germ cell viability
(145), and sperm function (107). The nongenomic action of estrogen on
plasma membrane-associated receptors has been documented in various
reproductive systems (see Ref. 98 for review, and Section
III.B). The estrogen-dependent initiation of this nongenomic
pathway can include a rapid increase in cAMP (e.g., Ref.
293), or intracellular calcium (e.g., Ref. 107), or the MAPK
pathway (e.g., Ref. 294). Consequently, it is possible that
estrogens have actions independent of ER and ERß, and thus the
potential for estrogen acting in the testes of mice with inactivated
ER and/or ß cannot be discounted.
To date, three laboratories have generated ArKO mice (7, 8, 9). The
transgenic lines from each of the three laboratories all show defects
in sexual behavior (see Table 2); however, fertility in these mice is
variable (see Ref. 7 for a direct comparison of the three phenotypes).
All mice show normal spermatogenesis at 14 weeks of age (7, 8, 9). One
line of ArKO mice shows progressive disruptions to spermatogenesis
until by 1 yr of age, all animals show evidence of spermatogenic
disruption (8). In contrast, ArKO mice generated recently in another
laboratory show no disruptions to spermatogenesis at 14 weeks to 10
months of age, although there is a significant reduction in
seminiferous epithelial height in these animals (7). Testicular
histology in older mice from a third line of ArKO mice has not been
described (9). The reason for the heterogeneity in the phenotypes of
different ArKO mice is unclear. Diet can provide variable levels of
phytoestrogens that may contribute to the heterogeneity. Another source
of variation could be in the extent of in utero exposure of
male ArKO pups to estrogen from the maternal circulation as well as
that produced by wild-type and heterozygous littermates. Also pups are
presumably exposed to maternal estrogens during the lactational period,
which may also be affected by the phytoestrogen content of the
mother’s diet. Thus variations in estrogen exposure of ArKO males
in utero, before weaning, and during adulthood are likely to
contribute to the variable spermatogenic phenotype.
In our laboratory, we noticed a decrease in fertility of ArKO mice after the age of 18 weeks (8), and this was related to the decrease in the numbers of round and elongated spermatids in the testes, despite no changes in earlier germ cell numbers. The fact that round spermatids undergo apoptosis and/or have disruptions to acrosome biogenesis in the ArKO mice (8) and contain both aromatase and ERß (see Section IV) suggests local estrogen action in these cells. A surprising finding in these ArKO mice was that the spermatogenic phenotype was age dependent, developing only after 18 weeks of age, leading to the speculation that dietary phytoestrogens could allow the maintenance of spermatogenesis in younger animals (8). Whether the disruptions to spermatogenesis have an earlier onset and are more profound in ArKO animals raised on a phytoestrogen-free diet, and thus whether dietary estrogens could have a modulatory role in the testis of ArKO mice, is currently under examination in our laboratory.
A second unexpected finding in our ArKO mice (8) and in other ArKO mice
(7, 9) was the absence of seminiferous tubule dysfunction related to
dysfunction of the efferent ductules as is seen in the ERKO (6, 26, 27, 223, 291) (see Table 2). Thus while spermatogenesis is disrupted in
ArKO mice (8), this does not seem to be primarily related to failure of
the efferent ductules to resorb fluid, since no significant increase in
either testis weight or seminiferous tubule luminal volume was seen
(8). It is possible that efferent ductule morphology and function could
be impaired in the ArKO, yet there may be reduced fluid secretion by
the seminiferous tubules and thus fluid buildup may not be obvious. A
second proposition is that ArKO animals, which will be exposed to
maternal estrogens in utero, may have normal development of
the efferent ductules, which allows these tissues to function normally
in adulthood.
The lack of an effect on efferent ductules and the late onset phenotype
in the ArKOs (8) leads to the speculation that there is
ligand-independent activation of ERs within the ArKO reproductive
tract. There is evidence for ligand (estrogen)-independent activation
of ERs in various systems, e.g., by cyclin D1 (94, 95) or by
growth factors (see Ref. 87 for review; see Section III
B). An alternative explanation to account for these
observations is that, despite the absence of aromatase products in ArKO
animals (196), there is the potential for other endogenous estrogenic
ligands to activate ER-mediated transcription. In vitro
relative binding affinity studies show that ER and ERß can
interact with a variety of non-aromatase-derived ligands such as
5-androstene-3ß,17ß-diol and 5-androstane-3ß,17ß-diol (65).
If ERs in ArKO mice do interact with alternate ligands such as those
derived from androgens, presumably there would be a gradual
disappearance of such ligand(s) in the testis after pubertal
development leading to the onset of spermatogenic dis- ruption (8).
Finally, it is also worthwhile to note that recent reports on the
infertility of mice deficient in vitamin D receptors indicate that the
testes of these mice have disrupted estrogen biosynthesis due to a
reduction in aromatase expression (295). While the authors report that
the spermatogenic phenotype is similar to both the ERKO and ArKO
phenotypes, further analysis of this phenotype is required to ascertain
whether the observed reduction in sperm count and disruption to
seminiferous epithelial morphology are due to a direct effect on germ
cell development, such as in the ArKO, and/or due to dysfunction of the
efferent ductules, such as in the ERKO.
Studies on the spermatogenic phenotypes in ER (reviewed in Ref. 27) and
aromatase null mice clearly provide evidence that estrogen and ERs are
required for spermatogenesis and normal male fertility (see Table 2).
Studies in the ER null mouse have shown that one of the most
important estrogen, or at least, ER-regulated events is the
development and function of the efferent ductules, and that a lack of
ER in this tissue has profound implications for male fertility. The
observations in ArKO mice suggest that aromatase and estrogen are
important for germ cell development. However, since ERß is apparently
the only ER present in germ cells, and ERßKO mice are fully fertile,
it is unclear how estrogen is acting in germ cells. Observations in
isolated germ cells have provided evidence for plasma-membrane ER
and/or nongenomic actions of estrogen; a mechanism that may not be
revealed in ERßKO mice. Discrepancies between the ERKO,
ERßKO, and ArKO phenotypes (see Table 2) perhaps suggest the
presence of novel genes encoding ERs and/or aromatase and/or
ligand-independent activation of ERs. Thus, while the studies in ER and
aromatase null mice indicate that ER is essential for normal male
fertility, as is aromatase, the discrepancies between the phenotypes
demonstrate that a more comprehensive understanding of estrogen action
in the testis is required.
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VI. Estrogen and Spermatogenesis in Humans |
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KO, ERßKO,
ERßKO, and ArKO mice, have been especially useful in elucidating
the spermatogenic consequences of removing the global and local actions
of estrogen. Naturally occurring mutations in humans, which render them
devoid of estrogen or resistant to its actions, can similarly provide
us with information specific to the role of estrogen in human
spermatogenesis. Such mutations have been extremely rare, and indeed
were once thought to be lethal. Only a single case has been reported
for a mutation in the ER (240), whereas two cases have been described
resulting from aromatase deficiency in men (242, 243) and one in a male
infant (241). It is valuable for the purposes of this article, however,
to review the salient features of the reproductive phenotypes of these
men, to better understand the direct and indirect impact estrogen has
upon human spermatogenesis and fertility.
The first aromatase-deficient adult male presented with scant estradiol
(<7 pg/ml) and elevated testosterone, 5-dihydrotestosterone,
androstenedione, FSH, and LH levels (243). Semen analysis was not
performed on this patient and at the time of reporting, he was a
virgin; hence, no conclusions can be drawn about the effect of his
aromatase deficiency upon his fertility. Morishima and colleagues (243)
report that the volume of both testes was greater than 25 ml and that
the consistency of the testes was normal. At 24 yr of age, the patient
exhibited Tanner stage 5 pubic hair and genital development.
Behaviorally, he was heterosexually oriented and reported a normal
pattern of nocturnal emissions and ejaculations. The second
aromatase-deficient male studied also had undetectable levels of
estradiol; however, he had normal serum concentrations of testosterone
and androstenedione, with only slightly elevated levels of FSH and LH
(242). Semen analyses of this patient revealed a more than 20-fold
decrease in sperm count and all sperm were immotile. A testicular
biopsy showed hypospermatogenesis and germ cell arrest, mainly at the
level of primary spermatocytes. Consequently, this man was infertile,
one of the symptoms for which he initially sought therapy. Treatment
with human menopausal and chorionic gonadotropins, estradiol, or
testosterone, did not restore the sperm count (242). Testis volume was
subnormal in this male, the volume of each testis being only 8 ml at 29
yr of age, although other sexual parameters of pubertal development
were normal. His sexual identity and psychosexual orientation were
heterosexual. It should be noted that azoospermia and infertility were
also reported in a brother of this man who did not have an
aromatase deficiency; therefore, the infertility of this
estrogen-deficient male may not be related to a lack of
estrogen (242).
In the single reported case of an adult male with a mutation in the
ER gene, a premature stop codon resulted from the replacement of
cytosine with thymine at codon 157, and thus the translated receptor
would lack the DNA binding and ligand binding domains and be
functionally inert (240). Like the aromatase-deficient males,
gonadotropin levels in this man were elevated, and circulating
testosterone was normal [similar to the patient described by Carani
and colleagues (242)], but serum estrogens were elevated (119 pg/ml).
This male experienced normal onset of puberty and was normally
masculinized, each testis with a volume of 20–25 ml. Semen analysis
revealed a normal sperm density (25 x
106/ml) but with a decreased sperm viability of
only 18% (normal = 50%). Treatment with high-dose ethinyl
estradiol did not restore the hormone profile of this patient to within
normal parameters, despite a 10-fold increase in the serum free
estradiol concentration (240). Behaviorally, this male expressed no
gender identity disorder and was heterosexual, and his sexual function
was apparently normal.
Evidently, few conclusions about the action(s) of estrogen on human spermatogenesis may be drawn from this small number of examples. The available data, although scant, certainly imply that estrogen is required for human spermatogenesis to proceed normally, and for full male fertility.
Other, nonreproductive similarities exist between the case of male estrogen insensitivity and those described for aromatase-deficient men. These include significant skeletal aberrations, such as osteopenia, osteoporosis, increased bone turnover, and delayed or absent epiphyseal fusion, and compromised circulating lipid levels and insulin resistance. While beyond the scope of the current paper, these facets of the phenotypes were reviewed by Faustini-Fustini et al. (296) and Grumbach and Auchus (297).
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VII. Estrogen and Sexual Behavior |
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TopAbstractI. IntroductionII. Overview of SpermatogenesisIII. Biosynthesis and Action...IV. ERs, Aromatase, and...V. The Effects of...VI. Estrogen and Spermatogenesis...VII. Estrogen and Sexual...VIII. SummaryReferences |
|---|
). While an analysis of the estrogen control of behavior clearly
extends beyond this review, a brief overview is warranted.
Perturbations have been reported in the sexual behavior of ER null
mice, disturbances that by themselves (to the exclusion of the
spermatogenic aberrations) would render the mice either infertile or
with diminished fertility (see Ref. 27 for review; see Table 2).
ERKO mice, while motivated to engage in sexual activity in terms of
numbers of mounts, display a severely reduced number of intromissions
and ejaculations (221). In contrast, sexual behavior of ERßKO male
mice is reportedly not perturbed (298), suggesting that ER is the
receptor required to transduce normal sexual behavior in males (27).
Sexual behavior has recently been reported as being completely
abolished in ERßKO mice (299), highlighting the fact that estrogen
action is required for normal sexual behavior. Of particular interest
is the fact that ERßKO mice do not mount receptive females,
whereas ERKO and ERßKO mice display normal mounting behavior (see
Table 2), indicating that expression of either ER is sufficient for
mounting behavior and suggesting that ERß plays a role in maintaining
sexual behavior in the brain, but that lack of ERß (in ERßKO mice)
can be compensated for by ER (299). The information from ER null
mice complements evidence from Honda and colleagues (9), Toda and
colleagues (7), and our own laboratory (our unpublished data) (see
Table 2), that ArKO mice display disturbed sexual behavior, either not
mounting receptive females at all, or taking longer to mount and
mounting less often than wild-type control males.
A single study has been performed to assess the impact of estrogen on human male sexual behavior (300). By restoring estrogen to the second aromatase-deficient patient that was described in the literature, significant modifications of this man’s sexual behavior occurred. Although originally reported as having a normal libido (242), the administration of estradiol to this patient increased libido, frequency of sexual intercourse, masturbation, and erotic fantasies (300). Thus the evidence for an involvement of estrogen in sexual behavior gained from mouse models may have parallels in the human.
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VIII. Summary |
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TopAbstractI. IntroductionII. Overview of SpermatogenesisIII. Biosynthesis and Action...IV. ERs, Aromatase, and...V. The Effects of...VI. Estrogen and Spermatogenesis...VII. Estrogen and Sexual...VIII. SummaryReferences |
|---|
That estrogen can influence testicular and epididymal function is not
unexpected, given the evidence presented that estrogen biosynthesis,
via the aromatase enzyme, and action on its receptors ( and/or ß),
occurs in these tissues. Estrogen is produced by the testis from the
fetal period throughout adulthood and, similarly, ER and ß are
found in the testis at all ages. While some cells express both ER
and ß, such as the Leydig cells, the cells in the seminiferous
epithelium appear to predominantly contain ERß. Nongenomic actions of
estrogen can also occur in the testis, although whether this involves
the "classic" ERs remains uncertain.
Evidence has been presented that estrogens act at multiple levels to control, or interfere with, spermatogenesis. Estrogen is clearly involved in the negative feedback effects of testosterone on the brain to control pituitary gonadotropin secretion, and hence an absence of, or inappropriate exposure to, estrogens leads to disturbances in the delicate balance of the hypothalamo-pituitary-testis axis in both mice and men. In view of the fact that the development and spermatogenic potential of the testis is reliant upon this axis, such disturbances are likely to have a deleterious effect on spermatogenesis and fertility.
We and others (162) have reviewed the evidence for an essential role
for estrogen in the development and maintenance of the efferent
ductules and epididymis. An absence of ER causes defects in efferent
ductule development, resulting in disturbed function, particularly in
terms of fluid resorption, causing a reduced number of sperm to enter
the epididymis. A consequence of the disturbed fluid dynamics is a
buildup of fluid in the testis, resulting in seminiferous epithelial
damage and impaired germ cell development. The fact that overexposure
to estrogens during neonatal development can also produce similar
defects in efferent ductule function highlights the need for a tightly
coordinated series of estrogen-dependent events in this tissue.
There is compelling evidence for a role for estrogen in testicular function. Notable roles for estrogen in Leydig cells include the coordinated regulation of progenitor Leydig cells into immature and adult forms. Estrogen also plays a role in testicular descent, at least in part, by the regulation of fetal Leydig cell gene expression. Sertoli cells are influenced by estrogen overexposure, which is probably related to the fact that estrogen appears to play an inhibitory role, acting as an internal control mechanism in Sertoli cell proliferation, development, and function. A compelling body of evidence has also been presented that estrogen has a functional role in germ cells. Germ cells contain ERs as well as aromatase, and thus it is possible that estrogen acts in an intracrine manner in these cells, to control viability/apoptosis and, potentially, acrosome biogenesis. In addition, nongenomic actions of estrogen on sperm function can now be considered along with the well documented nongenomic actions of progesterone (98).
The models of gene disruption and estrogen administration studies
reviewed in this manuscript and the deleterious effects of such
situations on male fertility have answered many questions as to the
role of, and sites of action for, estrogen in several aspects of male
fertility, from testicular function to sexual behavior. However, a
large amount of conflicting data also exist, such as the comparison of
the phenotypes of mice deficient in ER and/or ERß and aromatase.
Such conflicting data highlight the complexity of the molecular
mechanisms of estrogen action in the male, and serve to remind us that
the study of the role of estrogen in male fertility is far from
complete.
|
Footnotes |
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1 Supported by USPHS Grant R37-AG-08174, Program Grant 983212 from the
National Health and Medical Research Council of Australia (NH&MRC), and
a Wellcome Trust Research Training Fellowship in Reproductive Biology
050387. 
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References |
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TopAbstractI. IntroductionII. Overview of SpermatogenesisIII. Biosynthesis and Action...IV. ERs, Aromatase, and...V. The Effects of...VI. Estrogen and Spermatogenesis...VII. Estrogen and Sexual...VIII. SummaryReferences |
|---|
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L. Aksglaede, A. M. Wikstrom, E. R.-D. Meyts, L. Dunkel, N. E. Skakkebaek, and A. Juul Natural history of seminiferous tubule degeneration in Klinefelter syndrome Hum. Reprod. Update, January 1, 2006; 12(1): 39 - 48. [Abstract] [Full Text] [PDF]
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S. Solakidi, A-M.G. Psarra, S. Nikolaropoulos, and C.E. Sekeris Estrogen receptors {alpha} and {beta} (ER{alpha} and ER{beta}) and androgen receptor (AR) in human sperm: localization of ER{beta} and AR in mitochondria of the midpiece Hum. Reprod., December 1, 2005; 20(12): 3481 - 3487. [Abstract] [Full Text] [PDF]
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H. Baines, M. O Nwagwu, E. C Furneaux, J. Stewart, J. B Kerr, T. M Mayhew, and F. J P Ebling Estrogenic induction of spermatogenesis in the hypogonadal (hpg) mouse: role of androgens Reproduction, November 1, 2005; 130(5): 643 - 654. [Abstract] [Full Text] [PDF]
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 S. Ramaswamy Pubertal Augmentation in Juvenile Rhesus Monkey Testosterone Production Induced by Invariant Gonadotropin Stimulation Is Inhibited by Estrogen J. Clin. Endocrinol. Metab., October 1, 2005; 90(10): 5866 - 5875. [Abstract] [Full Text] [PDF]
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K. L. Matthiesson, P. G. Stanton, L. O'Donnell, S. J. Meachem, J. K. Amory, R. Berger, W. J. Bremner, and R. I. McLachlan Effects of Testosterone and Levonorgestrel Combined with a 5{alpha}-Reductase Inhibitor or Gonadotropin-Releasing Hormone Antagonist on Spermatogenesis and Intratesticular Steroid Levels in Normal Men J. Clin. Endocrinol. Metab., October 1, 2005; 90(10): 5647 - 5655. [Abstract] [Full Text] [PDF]
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A.L. Filby and C.R. Tyler Molecular Characterization of Estrogen Receptors 1, 2a, and 2b and Their Tissue and Ontogenic Expression Profiles in Fathead Minnow (Pimephales promelas) Biol Reprod, October 1, 2005; 73(4): 648 - 662. [Abstract] [Full Text] [PDF]
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R. Yoshida, M. Fukami, I. Sasagawa, T. Hasegawa, N. Kamatani, and T. Ogata Association of Cryptorchidism with a Specific Haplotype of the Estrogen Receptor {alpha} Gene: Implication for the Susceptibility to Estrogenic Environmental Endocrine Disruptors J. Clin. Endocrinol. Metab., August 1, 2005; 90(8): 4716 - 4721. [Abstract] [Full Text] [PDF]
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J. M. Naciff, K. A. Hess, G. J. Overmann, S. M. Torontali, G. J. Carr, J. P. Tiesman, L. M. Foertsch, B. D. Richardson, J. E. Martinez, and G. P. Daston Gene Expression Changes Induced in the Testis by Transplacental Exposure to High and Low Doses of 17{alpha}-Ethynyl Estradiol, Genistein, or Bisphenol A Toxicol. Sci., August 1, 2005; 86(2): 396 - 416. [Abstract] [Full Text] [PDF]
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S. J Meachem, D. M Robertson, N. G Wreford, R. I McLachlan, and P. G Stanton Oestrogen does not affect the restoration of spermatogenesis in the gonadotrophin-releasing hormone-immunised adult rat J. Endocrinol., June 1, 2005; 185(3): 529 - 538. [Abstract] [Full Text] [PDF]
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N. Sofikitis, E. Pappas, A. Kawatani, D. Baltogiannis, D. Loutradis, N. Kanakas, D. Giannakis, F. Dimitriadis, K. Tsoukanelis, I. Georgiou, et al. Efforts to create an artificial testis: culture systems of male germ cells under biochemical conditions resembling the seminiferous tubular biochemical environment Hum. Reprod. Update, May 1, 2005; 11(3): 229 - 259. [Abstract] [Full Text] [PDF]
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 M. E. Juan, E. Gonzalez-Pons, T. Munuera, J. Ballester, J. E. Rodriguez-Gil, and J. M. Planas trans-Resveratrol, a Natural Antioxidant from Grapes, Increases Sperm Output in Healthy Rats J. Nutr., April 1, 2005; 135(4): 757 - 760. [Abstract] [Full Text] [PDF]
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D. P. Mishra and C. Shaha Estrogen-induced Spermatogenic Cell Apoptosis Occurs via the Mitochondrial Pathway: ROLE OF SUPEROXIDE AND NITRIC OXIDE J. Biol. Chem., February 18, 2005; 280(7): 6181 - 6196. [Abstract] [Full Text] [PDF]
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W. de Ronde, A. Hofman, H. A P Pols, and F. H de Jong A direct approach to the estimation of the origin of oestrogens and androgens in elderly men by comparison with hormone levels in postmenopausal women Eur. J. Endocrinol., February 1, 2005; 152(2): 261 - 268. [Abstract] [Full Text] [PDF]
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T. Pakarainen, F.-P. Zhang, S. Makela, M. Poutanen, and I. Huhtaniemi Testosterone Replacement Therapy Induces Spermatogenesis and Partially Restores Fertility in Luteinizing Hormone Receptor Knockout Mice Endocrinology, February 1, 2005; 146(2): 596 - 606. [Abstract] [Full Text] [PDF]
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E. V. Magnusdottir, T. Thorsteinsson, S. Thorsteinsdottir, M. Heimisdottir, and K. Olafsdottir Persistent organochlorines, sedentary occupation, obesity and human male subfertility Hum. Reprod., January 1, 2005; 20(1): 208 - 215. [Abstract] [Full Text] [PDF]
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A. H. Payne and D. B. Hales Overview of Steroidogenic Enzymes in the Pathway from Cholesterol to Active Steroid Hormones Endocr. Rev., December 1, 2004; 25(6): 947 - 970. [Abstract] [Full Text] [PDF]
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D. M. Selva, O. M. Tirado, N. Toran, C. A. Suarez-Quian, J. Reventos, and F. Munell Estrogen Receptor {beta} Expression and Apoptosis of Spermatocytes of Mice Overexpressing a Rat Androgen-Binding Protein Transgene Biol Reprod, November 1, 2004; 71(5): 1461 - 1468. [Abstract] [Full Text] [PDF]
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Y. Wang, R. Thuillier, and M. Culty Prenatal Estrogen Exposure Differentially Affects Estrogen Receptor-Associated Proteins in Rat Testis Gonocytes Biol Reprod, November 1, 2004; 71(5): 1652 - 1664. [Abstract] [Full Text] [PDF]
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Z.M. Lei, S. Mishra, P. Ponnuru, X. Li, Z.W. Yang, and Ch.V. Rao Testicular Phenotype in Luteinizing Hormone Receptor Knockout Animals and the Effect of Testosterone Replacement Therapy Biol Reprod, November 1, 2004; 71(5): 1605 - 1613. [Abstract] [Full Text] [PDF]
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G. Delbes, C. Levacher, C. Pairault, C. Racine, C. Duquenne, A. Krust, and R. Habert Estrogen Receptor {beta}-Mediated Inhibition of Male Germ Cell Line Development in Mice by Endogenous Estrogens during Perinatal Life Endocrinology, July 1, 2004; 145(7): 3395 - 3403. [Abstract] [Full Text] [PDF]
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V. Pezzi, R. Sirianni, A. Chimento, M. Maggiolini, S. Bourguiba, C. Delalande, S. Carreau, S. Ando, E. R. Simpson, and C. D. Clyne Differential Expression of Steroidogenic Factor-1/Adrenal 4 Binding Protein and Liver Receptor Homolog-1 (LRH-1)/Fetoprotein Transcription Factor in the Rat Testis: LRH-1 as a Potential Regulator of Testicular Aromatase Expression Endocrinology, May 1, 2004; 145(5): 2186 - 2196. [Abstract] [Full Text] [PDF]
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K. Toda, Y. Okada, M. Zubair, K.-i. Morohashi, T. Saibara, and T. Okada Aromatase-Knockout Mouse Carrying an Estrogen-Inducible Enhanced Green Fluorescent Protein Gene Facilitates Detection of Estrogen Actions in Vivo Endocrinology, April 1, 2004; 145(4): 1880 - 1888. [Abstract] [Full Text] [PDF]
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E. D. Albrecht, R. B. Billiar, G. W. Aberdeen, J. S. Babischkin, and G. J. Pepe Expression of Estrogen Receptors {alpha} and {beta} in the Fetal Baboon Testisand Epididymis Biol Reprod, April 1, 2004; 70(4): 1106 - 1113. [Abstract] [Full Text] [PDF]
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S. Aquila, D. Sisci, M. Gentile, E. Middea, S. Catalano, A. Carpino, V. Rago, and S. Ando Estrogen Receptor (ER){alpha} and ER{beta} Are Both Expressed in Human Ejaculated Spermatozoa: Evidence of Their Direct Interaction with Phosphatidylinositol-3-OH Kinase/Akt Pathway J. Clin. Endocrinol. Metab., March 1, 2004; 89(3): 1443 - 1451. [Abstract] [Full Text] [PDF]
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 X. Li, L. Strauss, S. Makela, T. Streng, I. Huhtaniemi, R. Santti, and M. Poutanen Multiple Structural and Functional Abnormalities in the P450 Aromatase Expressing Transgenic Male Mice Are Ameliorated by a P450 Aromatase Inhibitor Am. J. Pathol., March 1, 2004; 164(3): 1039 - 1048. [Abstract] [Full Text] [PDF]
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O. M. Tirado, D. M. Selva, N. Toran, C. A. Suarez-Quian, M. Jansen, D. P. McDonnell, J. Reventos, and F. Munell Increased Expression of Estrogen Receptor {beta} in Pachytene Spermatocytes After Short-Term Methoxyacetic Acid Administration J Androl, January 1, 2004; 25(1): 84 - 94. [Abstract] [Full Text] [PDF]
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A. Guais, B. Solhonne, N. Melaine, G. Guellaen, and F. Bulle Goliath, a Ring-H2 Mitochondrial Protein, Regulated by Luteinizing Hormone/Human Chorionic Gonadotropin in Rat Leydig Cells Biol Reprod, January 1, 2004; 70(1): 204 - 213. [Abstract] [Full Text] [PDF]
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S. Bourguiba, S. Chater, C. Delalande, M. Benahmed, and S. Carreau Regulation of Aromatase Gene Expression in Purified Germ Cells of Adult Male Rats: Effects of Transforming Growth Factor {beta}, Tumor Necrosis Factor {alpha}, and Cyclic Adenosine 3',5'-Monosphosphate Biol Reprod, August 1, 2003; 69(2): 592 - 601. [Abstract] [Full Text] [PDF]
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J. Lassurguere, G. Livera, R. Habert, and B. Jegou Time- and Dose-Related Effects of Estradiol and Diethylstilbestrol on the Morphology and Function of the Fetal Rat Testis in Culture Toxicol. Sci., May 1, 2003; 73(1): 160 - 169. [Abstract] [Full Text] [PDF]
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S. Ramaswamy, G. R. Marshall, C. R. Pohl, R. L. Friedman, and T. M. Plant Inhibitory and Stimulatory Regulation of Testicular Inhibin B Secretion by Luteinizing Hormone and Follicle-Stimulating Hormone, Respectively, in the Rhesus Monkey (Macaca mulatta) Endocrinology, April 1, 2003; 144(4): 1175 - 1185. [Abstract] [Full Text] [PDF]
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 H. Sipahutar, P. Sourdaine, S. Moslemi, B. Plainfosse, and G.-E. Seralini Immunolocalization of Aromatase in Stallion Leydig Cells and Seminiferous Tubules J. Histochem. Cytochem., March 1, 2003; 51(3): 311 - 318. [Abstract] [Full Text] [PDF]
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 S. Lambard, I. Galeraud-Denis, H. Bouraima, S. Bourguiba, A. Chocat, and S. Carreau Expression of aromatase in human ejaculated spermatozoa: a putative marker of motility Mol. Hum. Reprod., March 1, 2003; 9(3): 117 - 124. [Abstract] [Full Text] [PDF]
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R. Thuillier, Y. Wang, and M. Culty Prenatal Exposure to Estrogenic Compounds Alters the Expression Pattern of Platelet-Derived Growth Factor Receptors {alpha} and {beta} in Neonatal Rat Testis: Identification of Gonocytes as Targets of Estrogen Exposure Biol Reprod, March 1, 2003; 68(3): 867 - 880. [Abstract] [Full Text] [PDF]
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K. Golovine, M. Schwerin, and J. Vanselow Three Different Promoters Control Expression of the Aromatase Cytochrome P450 Gene (Cyp19) in Mouse Gonads and Brain Biol Reprod, March 1, 2003; 68(3): 978 - 984. [Abstract] [Full Text] [PDF]
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R. Nair and C. Shaha Diethylstilbestrol Induces Rat Spermatogenic Cell Apoptosis in Vivo through Increased Expression of Spermatogenic Cell Fas/FasL System J. Biol. Chem., February 14, 2003; 278(8): 6470 - 6481. [Abstract] [Full Text] [PDF]
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R. A. Anderson and D. T. Baird Male Contraception Endocr. Rev., December 1, 2002; 23(6): 735 - 762. [Abstract] [Full Text] [PDF]
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B. L. Herrmann, B. Saller, O. E. Janssen, P. Gocke, A. Bockisch, H. Sperling, K. Mann, and M. Broecker Impact of Estrogen Replacement Therapy in a Male with Congenital Aromatase Deficiency Caused by a Novel Mutation in the CYP19 Gene J. Clin. Endocrinol. Metab., December 1, 2002; 87(12): 5476 - 5484. [Abstract] [Full Text] [PDF]
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Q. Zhou, R. Nie, G. S. Prins, P. T. K. Saunders, B. S. Katzenellenbogen, and R. A. Hess Localization of Androgen and Estrogen Receptors in Adult Male Mouse Reproductive Tract J Androl, November 1, 2002; 23(6): 870 - 881. [Abstract] [Full Text] [PDF]
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M. Kos, S. Denger, G. Reid, and F. Gannon Upstream Open Reading Frames Regulate the Translation of the Multiple mRNA Variants of the Estrogen Receptor alpha J. Biol. Chem., September 27, 2002; 277(40): 37131 - 37138. [Abstract] [Full Text] [PDF]
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 K. L. BRITT, J. KERR, L. O'DONNELL, M. E. E. JONES, A. E. DRUMMOND, S. R. DAVIS, E. R. SIMPSON, and J. K. FINDLAY Estrogen regulates development of the somatic cell phenotype in the eutherian ovary FASEB J, September 1, 2002; 16(11): 1389 - 1397. [Abstract] [Full Text] [PDF]
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K. M. Robertson, L. O'Donnell, E. R. Simpson, and M. E. E. Jones The Phenotype of the Aromatase Knockout Mouse Reveals Dietary Phytoestrogens Impact Significantly on Testis Function Endocrinology, August 1, 2002; 143(8): 2913 - 2921. [Abstract] [Full Text] [PDF]
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M. H. Tong and W.-C. Song Estrogen Sulfotransferase: Discrete and Androgen-Dependent Expression in the Male Reproductive Tract and Demonstration of an in Vivo Function in the Mouse Epididymis Endocrinology, August 1, 2002; 143(8): 3144 - 3151. [Abstract] [Full Text] [PDF]
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R. Luboshitzky, Z. Shen-Orr, R. Nave, S. Lavi, and P. Lavie Melatonin Administration Alters Semen Quality in Healthy Men J Androl, July 1, 2002; 23(4): 572 - 578. [Abstract] [Full Text] [PDF]
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J. C. Achermann, G. Ozisik, J. J. Meeks, and J. L. Jameson Genetic Causes of Human Reproductive Disease J. Clin. Endocrinol. Metab., June 1, 2002; 87(6): 2447 - 2454. [Full Text] [PDF]
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P. T. K. Saunders, M. R. Millar, S. Macpherson, D. S. Irvine, N. P. Groome, L. R. Evans, R. M. Sharpe, and G. A. Scobie ER{beta}1 and the ER{beta}2 Splice Variant (ER{beta}cx/{beta}2) Are Expressed in Distinct Cell Populations in the Adult Human Testis J. Clin. Endocrinol. Metab., June 1, 2002; 87(6): 2706 - 2715. [Abstract] [Full Text] [PDF]
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C. A. Oliveira, Q. Zhou, K. Carnes, R. Nie, D. E. Kuehl, G. L. Jackson, L. R. Franca, M. Nakai, and R. A. Hess ER Function in the Adult Male Rat: Short- and Long-Term Effects of the Antiestrogen ICI 182,780 on the Testis and Efferent Ductules, without Changes in Testosterone Endocrinology, June 1, 2002; 143(6): 2399 - 2409. [Abstract] [Full Text] [PDF]
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G. Stelzer and J. Don Atce1: A Novel Mouse Cyclic Adenosine 3',5'-Monophosphate-Responsive Element-Binding Protein-Like Gene Exclusively Expressed in Postmeiotic Spermatids Endocrinology, May 1, 2002; 143(5): 1578 - 1588. [Abstract] [Full Text] [PDF]
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Z.-x. Zhou, B. He, S. H. Hall, E. M. Wilson, and F. S. French Domain Interactions between Coregulator ARA70 and the Androgen Receptor (AR) Mol. Endocrinol., February 1, 2002; 16(2): 287 - 300. [Abstract] [Full Text] [PDF]
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 R.I. McLachlan, L. O'Donnell, S.J. Meachem, P.G. Stanton, D.M. de Kretser, K. Pratis, and D.M. Robertson Identification of Specific Sites of Hormonal Regulation in Spermatogenesis in Rats, Monkeys, and Man Recent Prog. Horm. Res., January 1, 2002; 57(1): 149 - 179. [Abstract] [Full Text] [PDF]
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