Jinno  Reproductive Biology

Reproductive Biology and Endocrinology (2025) 23:35

[Link] and Endocrinology

REVIEW Open Access

Ovarian stimulation by promoting basal

follicular growth
Masao Jinno1*

Abstract
Background Most methods of ovarian stimulation rely on gonadotropin modulation. However, abnormal anti-Mül-
lerian hormone concentrations are frequent in infertility, suggesting that defects in the gonadotropin-independent
period of folliculogenesis preceding cyclic recruitment (i.e., basal follicular growth) may often occur. We need to better
understand basal follicular growth and determine how to improve it.
Methods Section I summarizes a literature search concerning preantral and early antral folliculogenesis, cyclic
recruitment, and selection. Section II presents current knowledge about interventions involving early antral folliculo-
genesis and cyclic recruitment.
Results While folliculogenesis following cyclic recruitment is gonadotropin-dependent, basal follicular growth is not.
Basal follicular growth is regulated by follicle-stimulating hormone and local communication between the oocyte
and its granulosa and thecal cells involving gap junctions and many autocrine/paracrine factors. This local communi-
cation sustains growth synergistically with follicle-stimulating hormone, but also suppresses this hormone to induce
granulosa cell differentiation. As a follicle develops, its responsiveness to gonadotropin progressively increases. Sec-
tion II describes 4 interventions affecting early antral folliculogenesis, including granulocyte colony-stimulating factor
priming, bromocriptine rebound, carbohydrate metabolism intervention, and danazol priming, which have improved
embryo development and live birth rate in patients with previous failures.
Conclusion Basal follicular growth modulation can increase live birth rates.
Keywords Preantral and early antral folliculogenesis, Cyclic recruitment, Assisted reproductive technology,
Granulocyte colony-stimulating factor, Prolactin, Advanced glycation end-products, Danazol

Introduction and/or aromatase inhibitors. However, human follicu-

During the 45 years since IVF and ET first succeeded lar growth requires more than 6 months, and the early
using a natural cycle, various methods of ovarian stimu- growth period preceding cyclic recruitment is par-
lation have been developed to increase pregnancy rate. ticularly lengthy [1–6]. Moreover, many paracrine and
These interventions, usually performed at about 2 weeks autocrine factors act importantly in controlling folli-
of follicular growth beginning at cyclic recruitment, culogenesis, especially before cyclic recruitment; after
almost exclusively modulate Gn control by administer- recruitment, Gn becomes pivotal [1–6].
ing Gn, GnRH agonist/antagonists, clomiphene citrate, Poor ovarian reserve and PCOS, respectively charac-
terized by low and high serum concentrations of AMH,
are encountered frequently in the clinical practice of
*Correspondence: ART. These conditions can interfere with obtaining high-
Masao Jinno quality oocytes using conventional ovarian stimulation.
mjinno@[Link]
1
Women’s Clinic Jinno, 3‑11‑7 Kokuryou‑Chou, Choufu City,
AMH is secreted mainly by preantral and early antral fol-
Tokyo 182‑0022, Japan licles before cyclic recruitment, so related major causes

© The Author(s) 2025. Open Access This article is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0
International License, which permits any non-commercial use, sharing, distribution and reproduction in any medium or format, as long
as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if
you modified the licensed material. You do not have permission under this licence to share adapted material derived from this article or
parts of it. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated
otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not
permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To
view a copy of this licence, visit [Link]
Jinno Reproductive Biology and Endocrinology (2025) 23:35 Page 2 of 22

of ovarian dysfunction in these conditions probably act be the primordial follicle in mice [1, 3] and the primor-
during folliculogenesis before cyclic recruitment [7]. dial, transitory, and small primary follicles in humans [2].
Consequently, conventional ovarian stimulation has lim- Section II describes treatment methods that promote
ited efficacy in patients with such impaired folliculogen- basal follicular growth, aiming to provide insight and
esis before cyclic recruitment. perspective on earlier ovarian stimulation.
We need to explore new methods of earlier ovarian
stimulation that improve folliculogenesis before cyclic Preantral and early antral folliculogenesis (Section
recruitment. Among these, we serendipitously discov- I)
ered that G-CSF administration enhances early antral Details of preantral and early antral folliculogenesis
follicle growth, improving embryos and live birth rates in have been increasingly clarified, but many early aspects
patients with poor ovarian reserve [8]. In order to further remain controversial or less clearly known than subse-
advance the researches on earlier ovarian stimulation, quent events in antral folliclulogenesis. Extensive reviews
first we need to know current knowledges on the regula- of preantral and early antral folliculogenesis have been
tions of early folliculogenesis, which are not so familiar to provided by several authors [1–6, 11]. Here I attempt to
many physicians including me. summarize quiescent follicle activation, basal follicular
Section I of this review is an overview of early follicu- growth, cyclic recruitment, and selection, especially the
logenesis including quiescent follicle activation, basal fol- mechanisms regulating them.
licular growth, cyclic recruitment and selection, focusing
especially on the mechanisms regulating them. Basal fol- Overview of folliculogenesis (I‑1)
licular growth is defined as the folliculogenesis from the Around the time of birth, oocytes are encapsulated by
activated quiescent follicle until the early antral selecta- a single layer of squamous somatic pre-granulosa cells,
ble follicle just before cyclic recruitment [2] (See more in forming primordial follicles [12]. Primordial follicles
Fig. 1 and section I-1). The quiescent follicle is believed to (including primordial, transitory, and small primary

Fig. 1 Overview of folliculogenesis. This summary is based on Edson et al., 2009 [1], Gougeon et al., 2010 [2], Rimon-Dahari et al., 2016 [3], Hsueh
et al., 2015 [4], Morton et al., 2023 [5], Gougeon A, 1986 [9], and Paulino et al., 2022 [10]
Jinno Reproductive Biology and Endocrinology (2025) 23:35 Page 3 of 22

follicles in humans) [2] constitute the ovarian quiescent 9]. The selected follicle accelerates its growth, manifest-
follicle reserve, from which only a selected subpopulation ing dominance and experiencing multistep preovulatory
is activated and recruited into the pool of growing fol- events after the LH surge, resulting in ovulation.
licles that proceed to ovulation [1, 3]. According to cur- Folliculogenesis includes 5 main steps (Fig. 1): activa-
rent consensus, no primordial follicles are formed much tion of quiescent follicles including primordial, transi-
later than birth, so this population of primordial follicles tory, and small primary follicles in humans [2]; basal
formed before and just after birth constitutes a fixed follicular growth; cyclic recruitment of selectable fol-
pool of ovarian reserve drawn upon until menopause licles in the mid- and late luteal phase [9]; selection of
[3]. However, in recent experiments involving old female follicle(s) destined to ovulate from among the recruited
rats, intraovarian transplantation of G-CSF-mobilized selectable follicles by the end of the early follicular phase
peripheral blood mononuclear cells recreated follicles [9]; and dominant follicle growth in the mid- and late fol-
at all stages including primordial follicles, restoring fer- licular phase [2]. In humans, preantral folliculogenesis
tility [13]. Similarly, intravenous administration of bone is estimated to take more than 90 days; early antral fol-
marrow mesenchymal stem cells contributed to forma- liculogenesis up to selectable follicles, 70 days; and from
tion of primordial follicles in mice after cyclophospha- cyclic recruitment of selectable follicles until ovulation,
mide-induced ovarian failure [14]. Existence of germline 15–20 days [2, 9] (Fig. 1). Advancing from the primordial
stem cells in adult mouse ovaries [15] suggests that while follicle to the preovulatory follicle, oocytes and follicles
oogenesis in adult females is probably extremely limited respectively enlarge from 30 and 40 μm in diameter to
under normal physiologic conditions, it might become 120–140 μm and 20 mm, respectively [10].
possible after manipulations such as introduction of acti- In humans, the early stages in folliculogenesis, from
vated somatic oogenesis-supporting cells. primordial follicles to selectable small antral follicles
When a primordial follicle is activated, it grows into a that represent basal follicular growth, do not depend on
primary follicle with a single layer of cuboidal granulosa Gn, although they respond to it. In contrast, subsequent
cells (Fig. 1) [1–6, 11]. As these cells proliferate and the stages proceeding to ovulation are Gn-dependent [2].
oocyte increases in size, the follicle becomes a small sec- This distinction is evident from observations that selecta-
ondary follicle with 2 or more layers of granulosa cells. ble small antral follicles can be seen despite congenital
A small secondary follicle is supplied by one or two arte- absence of bioactive FSH [18] and also in infancy, preg-
rioles, terminating in an anastomotic network just out- nancy, hypogonadotropic hypogonadism, and the post-
side the basal lamina [2]. At this stage, cells that form hypophysectomy state, where concentrations of Gn are
the theca interna begin to differentiate. Granulosa cells low [19]. In mice, where the preantral stage of large sec-
further proliferate, the zona pellucida is synthesized, and ondary follicles might be comparable to human selectable
the theca interna with its vasculature fully differentiates, antral follicles, knock-out of Fshβ [20] and Fshr [21] genes
now representing a large secondary follicle. Some theca does not stop folliculogenesis up to the secondary follicle
interna cells take on the appearance of steroid-secreting stage but blocks it prior to antral follicle formation.
cells, termed epithelioid cells; at that point the large sec- Basal follicular growth is regulated by interactions
ondary follicle enters the preantral stage [2]. between FSH and local autocrine/paracrine factors pro-
As the follicle continues to grow, fluid-filled pockets duced by the oocyte, GC, and theca interna cells [2, 3, 5].
begin to form within the granulosa cell layers, eventually In addition, the Hippo signaling pathway constrains fol-
merging to form a single cavity called the antrum; the fol- licular growth [4, 11]. As a follicle develops, its respon-
licle has become an early antral follicle (Fig. 1). As fluid siveness to Gn progressively increases [2, 5] and roles of
accumulates in the antrum and granulosa cells and theca Gn shift from selection of follicles for growth vs. atresia
interna cells proliferate, the follicle progresses to become during basal follicular growth to growth, cell differentia-
a selectable follicle with a size between 2 and 5 mm [2]. tion, and steroidogenesis after cyclic recruitment [5]. In
In humans, selectable follicles may be seen at any time humans, FSH receptor mRNA first can be detected in
throughout the menstrual cycle, but their number and the GC of primary follicles [22], and LH receptor mRNA
quality increase during the late luteal phase in response first can be detected in theca interna cells in the preant-
to increased concentrations of FSH, accompanied by ral stage of large secondary follicles [2, 23]. Although pri-
regression of the corpus luteum [2, 16]. Selectable folli- mordial follicles lack FSH receptor expression, possible
cles are more receptive to cyclic hormonal changes, while indirect roles of FSH include formation of primordial fol-
earlier growing follicles are unresponsive [2, 17]. Among licles in the perinatal period and promotion of primordial
the recruited selectable follicles in the late luteal phase, follicle activation [5].
the follicle destined to ovulate during the subsequent Follicular vascularization increases availability of
cycle is selected by the end of the early follicular phase [2, oxygen, nutrients, and trophic factors such as FSH,
Jinno Reproductive Biology and Endocrinology (2025) 23:35 Page 4 of 22

promoting follicular growth. VEGF participates not only STAT, TGF-β, MAPK, and WNT signaling are thought as
in regulation of follicular angiogenesis but also direct reg- potential regulators [11].
ulation of follicular cell function [6]. In humans, VEGF Awakening signal activates mTORC1 in pre-GCs, stim-
expression first is detectable in both theca and granulosa ulating their differentiations to secrete the Kit ligand [3,
cells of secondary follicles, increasing as follicles enlarge; 11] (Fig. 2). The Kit ligand as well as various growth fac-
VEGF receptors can be detected in follicular cells ranging tors bind to the TK receptor and stimulate PI3K activity,
from primordial to preovulatory follicles [6]. Mediated by increasing ­PIP3. Then PDK1 is recruited and activated,
its receptors on endothelial, thecal, and granulosa cells, phosphorylating AKT and S6K1. Although FOXO3 is
VEGF promotes follicular angiogenesis and growth dur- preventing activation of the oocyte, activated AKT sup-
ing primordial follicle activation, basal follicular growth, presses the inhibitory actions of FOXO3, resulting in
cyclic recruitment, selection, and dominant follicle activation of the oocyte [3, 4, 11]. PTEN converts P ­ IP3 to
growth [6]. ­PIP2, thus counteracting the action of PI3K [3, 4, 11].
Another target of AKT is the TSC1/2 complex-
Activation of quiescent follicles (I‑2) mTORC1-S6K1-rpS6 pathway [3, 11] (Fig. 2). AKT phos-
Inhibitory signals from the oocyte itself and signals phorylates TSC2, destabilizing the TSC1/2 complex and
from somatic cells maintain the ovarian quiescent fol- releasing its inhibitory effect on mTORC1 [3, 11]. Acti-
licle reserve, consisting of primordial follicles in mice vated mTORC1 stimulates S6K1-rpS6 signaling that pro-
and primordial, transitory, and small primary follicles in motes protein translation and ribosome biogenesis in
humans [2–4, 11] (Fig. 2). Activation of quiescent follicles oocytes [3, 11]. AKT also phosphorylates p27Kip1, a cell-
is believed to be regulated in a gonadotropin-independ- cycle inhibitor, opposing G1 arrest [3, 11].
ent manner by paracrine signaling within the follicle and The Hippo signaling pathway regulates organ size
across the local environment [4, 11]. The balance of PI3K/ via control of cell proliferation, apoptosis, and stem
AKT and mTOR cascades (activator) and the Hippo cell self-renewal [4, 11]. It is regulated by the physical
pathway (inhibitor) plays a major role in controlling the and mechanical microenvironment of cells in response
activation of quiescent follicles [11]. In addition, JAK/ to cues such as cell–cell contact, cell polarity, and
energy stress [4, 11]. Hippo signaling consists of the

Fig. 2 Summary of activation of quiescent follicles based on Gougeon et al., 2010 [2], Rimon-Dahari et al., 2016 [3], Hsueh et al., 2015 [4], and Telfer
et al., 2023 [11]
Jinno Reproductive Biology and Endocrinology (2025) 23:35 Page 5 of 22

MST1/2-Sav1 complex and the LATS1/2-Mob1. Under communication between the oocyte and its granulosa
basal conditions, the MST1/2-Sav1 phosphorylates the cells and theca internal cells [2–5]. This bidirectional
LATS1/2-Mob1, which in turn phosphorylates and inac- communication sustains follicular growth synergistically
tivates downstream effectors, YAP/TAZ via their seques- with FSH but also suppresses FSH to induce GC differen-
tration and proteolytic degradation in the cytoplasm [4, tiation, according to developmental stage [2]. In addition,
11]. Upon the disruption of the Hippo signaling, unphos- the Hippo signaling pathway, as described in the section
phorylated YAP/TAZ translocates into the nucleus and of I-2, also constrains basal follicular growth according
binds TEAD transcription factors, promoting the expres- to various cues such as inter-follicular communications,
sion of CCN growth factor and BIRC apoptosis inhibi- follicular location within an ovary, and large structural
tors. Thus, disruption of the Hippo signaling pathway changes associated with ovulation [4, 11].
stimulates activation of quiescent follicles, GC prolifera- Communication between the oocyte and its GC and
tion, and even later follicular growth [4, 11] (Fig. 2). TIC involves both direct cell-to-cell communication via
Furthermore, LIF, activin, BMP4, BMP7, androgen, and gap junctions and autocrine/paracrine control (Fig. 3).
insulin stimulate activation of oocytes while AMH and In the former, the core protein that constitutes gap junc-
somatostatin suppress it [2, 3]. Notch signaling in pre- tions, connexins 43 and 37, play pivotal roles [2, 3]. Auto-
granulosa cells is involved in prevention of oocyte activa- crine/paracrine control involves many factors, including
tion [3]. the TGF-β superfamily (TGF-β, BMP, GDF, activin/
inhibin, and AMH), growth factors acting through TKR
Regulation of basal follicular growth (I‑3) (kit ligand, EGF, KGF, HGF, BFGF, IGF-I, and various
Basal follicular growth is regulated by interactions neurotrophic factors), androgen, R-spondin2, and CNP
between FSH and local intra-follicular communica- [2–5]. Oocytes express GDF9 [24], BMP15 [25], and
tion [2, 3, 5] but mostly is directed by local bidirectional R-spondin2 [26], all of which play central roles in cellular

Fig. 3 Regulation of basal follicular growth, according to Gougeon et al., 2010 [2], Rimon-Dahari et al., 2016 [3], Hsueh et al., 2015 [4], Morton et al.,
2023 [5], Shimasaki et al., 2004 [28], and Oktem and Urman, 2010 [31]. For additional references, see text
Jinno Reproductive Biology and Endocrinology (2025) 23:35 Page 6 of 22

communication. GDF9-deficient mice exhibit a block in primordial-to-primary follicle transition and increases
follicular development beyond the primary follicle stage, FSH receptor mRNA in the ovaries [42]. BMP4 and
leading to complete infertility [24]. GDF9 administration BMP7 modulate FSH signaling to promote estradiol pro-
in vivo enhances progression of primordial and primary duction while inhibiting progesterone synthesis [43], act-
follicles to small secondary follicles [27]. GDF9 stimu- ing as luteinization inhibitors such as BMP15 and GDF9
lates inhibin production and basal estradiol synthesis in [28]. Androgen sustains early follicular development in
GC but inhibits FSH actions such as FSH-induced LH monkeys [44] and simulates granulosa cell proliferation
receptor expression and FSH-induced estradiol and pro- at all stages of development [45]. HGF and KGF stimulate
gesterone synthesis [28]. GDF9 is required for granulosa granulosa cells to induce kit ligand, which in turn pro-
cells to produce the paracrine signals of Desert hedgehog motes the expression of HGF and KGF on theca interna
and Indian hedgehog, which in turn induce TIC differen- cells [31].
tiation [29]. BMP15 stimulates proliferation of granulosa EGF, BFGF, and HGF all stimulate granulosa cell pro-
cells in an FSH-independent manner [25], while naturally liferation in the absence of FSH, while EGF and BFGF
occurring point mutations of the BMP15 gene in Inverd- inhibit FSH receptor mRNA expression and subsequent
ale sheep (homozygous) and Hanna sheep cause infertil- GC differentiation during early follicular growth [2].
ity due to a block at the primary stage in folliculogenesis NTF5 and BDNF are involved in progression from pri-
[30]. BMP15 inhibits FSH receptor expression in GC, mary to secondary follicles [3].
resulting in suppression in FSH-induced LH receptor
expression, progesterone synthesis, and activin/inhibin Cyclic recruitment and selection (I‑4)
production [28]. R-spondin2 promotes development of When human follicles grow to become selectable small
primary follicles to the secondary stage in vitro and to the antral follicles (2–5 mm in diameter), they become
antral stage in vivo [26]. responsive to cyclic changes of FSH and are recruited
GCs produce kit ligand, activin, TGF-β, CNP, and into gonadotropin-dependent further growth in response
AMH [2–4, 31]. Kit ligand promotes oocyte growth up to to increases in FSH, or alternatively undergo atresia [2,
the preantral stage on GC monolayers [32]. BMP15 stim- 46, 47]. After recruitment and especially after selection,
ulates kit ligand expression in granulosa cells while this growing follicles are believed to produce factors that
ligand inhibits BMP15 expression in oocytes, constitut- suppress not only growth of less developed antral fol-
ing a negative feedback loop that GC depends upon for licles [48] but also recruitment of following cohorts of
mitotic regulation [33]. Additionally, kit ligand mRNA selectable follicles [17]. These suppressive effects exerted
expression in granulosa cells is suppressed by GDF9 [34]. by growing follicles appear to be much stronger than
Activin and TGF-β promote preantral follicle growth the FSH-stimulatory effect on recruitment because the
in vitro [35]. FSH-induced growth of small secondary fol- number of small antral follicles 2 to 10 mm in diameter
licles, however, is inhibited by activin produced by large decreased even during maximal daily hMG stimulation in
secondary/small antral follicles [36]. TGF-β suppresses ART therapy, while the recruited cohort of follicles grew
androgen production in theca interna cells in most spe- normally; the median follicle number at the beginning of
cies [37]. CNP, secreted by GC of secondary and antral hMG administration, 6 days later, and on the day of hCG
follicles, stimulates preantral and antral follicle growth administration respectively were 8 (interquartile range,
[4]. In addition, in antral and preovulatory follicles, CNP 5–11), 1 (0–6), and 0 (0–2) [49]. Therefore, once a cohort
increases cGMP in cumulus oophorus cells through its of selectable follicles is recruited into further growth by
receptor, NPR2, as well as in oocytes by cGMP transfer an FSH rise, subsequent recruitment is blocked by the
through gap junctions [38]. Elevated cGMP concentra- growing antral follicles despite the FSH elevation until
tions in oocytes inhibit phosphodiesterase 3A, which the growing follicles ovulate or start to regress. Conse-
increases intra-oocytic cAMP concentrations to maintain quently, antral follicle development in the menstrual/
meiotic arrest [4, 38]. AMH inhibits FSH-stimulated pre- estrous cycle occurs in waves over time [9].
antral follicle growth [39]. In humans, AMH expression Conventionally, a cohort of selectable follicles was
in GC is first detected in primary follicles, reaching its thought to be recruited in the late luteal or early follicular
highest level in secondary follicles and small antral folli- phase once in each human menstrual cycle; subsequently,
cles 4 mm or less in diameter, and then gradually disap- one of these ovulated after selection and further growth
pearing in larger (4–8 mm) antral follicles [40]. [9, 46]. However, multiple waves of antral follicle devel-
Theca interna cells produce BMP4, BMP7, TGF-β, opment have been observed during each estrous cycle
androgen, HGF, and KGF [31]. BMP4 promotes survival in cattle (2–3 waves) [50, 51], sheep (3–4 waves) [52,
of primordial follicles and stimulates primordial-to- 53], and other animals including horses, goats, llamas,
primary follicle transition [41], while BMP7 promotes buffalo, deer, and sub-human primates [47]. Each wave
Jinno Reproductive Biology and Endocrinology (2025) 23:35 Page 7 of 22

is preceded a few days earlier by a transient increase in terms of GC proliferation [17] and aromatase activity
circulating FSH [50–52]. In an interovulatory interval [16] greatly increases in the mid- to late-luteal phase even
(IOI), only the final follicular wave is ovulatory, while all though the antral follicle count (2–5 mm) does not differ
preceding waves are anovulatory [51, 52]. In humans, 2 significantly between cycle phases [58].
waves of antral follicle development were demonstrated Selection is the final adjustment of the cohort of
by daily ultrasonographic observations in 68% of women recruited growing follicles to the ovulatory quota, which
and 3 waves in 32% [54]; a nadir in serum FSH occurred is the species-characteristic number of follicles that ovu-
3 days before emergence of any follicular waves [55]. In late in each cycle [2, 59]. Selected follicles are likely to be
women with 2 follicular waves, a first anovulatory wave more sensitive to FSH [60], possibly because of enhanced
emerges at the time of ovulation, followed by emergence FSH receptor expression or increases in local growth
of a second ovulatory wave during the early follicular factors that augment FSH responsiveness [46]. Selecta-
phase; in women showing 3 waves, a first anovulatory ble follicles contain high concentrations of TGF-α, EGF,
wave similarly emerges at the time of ovulation, with a IGFBP, and activin [2, 19]. TGFα and EGF inhibit FSH-
second anovulatory wave emerging during the mid- to induced synthesis of estradiol and stimulate GC pro-
late-luteal phase (earlier than in women with 2 waves), liferation, IGFBP inhibits IGF-II to induce aromatase
and a third ovulatory wave emerges in the early to mid- activity in GC, and activin stimulates GC proliferation
follicular phase [47, 54]. The interwave interval (IWI) and expression of FSH and LH receptors [19]. Conse-
between second and third waves in women with 3 waves quently, GC of selectable follicles respond to FSH by pro-
is significantly shorter than all the other IWIs in women liferating, but not estrogen production, maintaining high
with 2 or 3 waves [55]. Notably, the third wave emerges androgen and low estradiol concentrations in follicular
before the second wave has regressed [56]. Consider- fluid [2]. In selected follicles, however, both proliferation
ing that recruitment occurs despite strong suppressive and differentiation stimulators are influencing their GC,
effects from preceding growing follicles, a recruitment resulting in substantially increased follicular fluid estra-
stimulatory factor in addition to FSH may be at work diol concentration [2].
in the late luteal to early follicular phase, possibly at the During selection, the IGF system plays a key role [2].
time of luteal regression. Disappearance of recruitment- IGF-II increases in GCs of dominant follicles at that time
suppressive effects of the corpus luteum does not fully [19]. Simultaneously, FSH stimulates an IGFBP-4 pro-
account for this phenomenon, considering that in sheep tease, PAPP-A, which cleaves IGFBP-4, subsequently
small FSH surges and corresponding anovulatory folli- increasing bioavailability of IGF-II. The increased free
cular waves occur during the active luteal phase; neither IGF-II stimulates both proliferation and steroidogen-
FSH surges nor waves are suppressible even by adminis- esis in GC and TIC, increasing estradiol and androgen
tration of progesterone in the active luteal phase, indicat- [19], as well as differentiation of LH receptors on GC [2].
ing that the suppressive effect of the corpus luteum is less These effects are strongly enhanced by FSH and LH [19].
potent than the stimulatory effect of FSH [52]. As I sug- An orderly transition in the follicular environment
gest further on in section II-2, the unknown stimulatory from inhibin B and activin to inhibin A and follistatin
factor might be an increase in prolactin during the late appears important for dominant follicle development
luteal phase. [47]. Follicular fluid inhibin B peaks at a follicular diam-
For more than a decade, random start ovarian stimu- eter of 9 to 10 mm, a time of divergence between domi-
lation (RSOS), in which stimulation is initiated regard- nant and subordinate follicles [47]. This is followed by an
less of the day or phase of the menstrual cycle, has been increase in inhibin A, which strongly enhances IGF- and
successfully used in oncofertility and general infertility LH-induced androgen production in TIC [2, 19, 47]. The
practice [56, 57]. Compared with conventional stimula- androgen is then converted to estradiol by aromatase
tion initiated at the early follicular phase, RSOS yields activity in GC [1], which can be detected in follicles of
a similar number of oocytes with similar reproductive 10 mm or larger [2], enhanced by the increases in IGF-
competence, resulting in similar rates of pregnancy and II. Activin inhibits LH-induced progesterone production
live birth after cryopreserved embryo transfers [56, 57]. by GC in preovulatory follicles [19], while follistatin has
Recruitment of selectable follicles thus appears funda- much higher binding affinity for activin than inhibin, so
mentally possible on any day of a menstrual cycle. How- follistatin’s net effect is facilitation of LH-induced proges-
ever, RSOS tends to require longer stimulation and more terone production by GC in preovulatory follicles by sup-
Gn [56, 57], suggesting some differences from physiologic pression of activin effect [19].
recruitment in the late luteal to early follicular phase. Thus, estradiol and inhibin produced by the dominant
Consistently, the GC mitotic index of selectable follicles follicle(s) suppress pituitary FSH release during the mid-
[9] and follicular responsiveness to exogeneous FSH in follicular phase. As a result, other less developed antral
Jinno Reproductive Biology and Endocrinology (2025) 23:35 Page 8 of 22

follicles are deprived of sufficient FSH stimulation for none of them achieved pregnancy by these ET, 3 of them
survival [46]. Dominant follicles, however, can maintain conceived spontaneously 2 cycles later; one of these preg-
development despite decreasing FSH given their greater nancies involved twins in a 45-year-old woman without
GC numbers, higher expression of FSH and LH receptors ovulation induction. We therefore suspected that G-CSF
on GC, greater free IGF-II, and richer vascularization [2, administration stimulated preantral or early antral fol-
3, 19, 46, 47]. licle growth to improve ovulation 2 cycles later, result-
Further, concentrations of TGF-α and EGF in follicular ing in pregnancies. We therefore initiated a prospective
fluid decrease during selection [19]. AMH expression in randomized controlled trial in patients with poor ovar-
GC is greatest in secondary follicles and small antral fol- ian reserve to examine whether G-CSF priming preced-
licles 4 mm or less in diameter, followed by gradual dis- ing ART increased serum AMH and improved embryo
appearance in larger antral follicles measuring 4–8 mm development and pregnancy rate [8].
[40]. This AMH disappearance coincides with time of G-CSF (100 μg of lenograstim; Neutrogin, Chyugai
selection, but precise roles of AMH in dominant follicle Pharmaceuticals, Tokyo, Japan) was administered sub-
selection are not known [47]. cutaneously in the early luteal phase (2–5 days after
the ovulation) of 2 consecutive cycles preceding ART
Interventions affecting basal follicular growth (Fig. 4A). Ovulation was determined, based upon basal
(Section II) body temperature records, vaginal ultrasonographic find-
Although various interventions affecting basal follicu- ings (collapse of the dominant follicle), and, if necessary,
lar growth have been proposed to benefit patients with serum progesterone determinations (1.5–3 ng/mL of
poor ovarian response to conventional ovarian stimula- serum progesterone was considered as the value on the
tion, their effectiveness is unclear or doubtful, especially ovulation day). We referred to this treatment as G-CSF
in terms of live birth [61]. Recently I serendipitously dis- priming. Ovarian stimulation using the long protocol and
covered that administration of G-CSF preceding conven- ART followed priming.
tional ovarian stimulation (G-CSF priming) increases One hundred patients with poor ovarian reserve
AMH and improves embryo quality and live birth rate in (AMH < 2 ng/mL, 20 to 42 years old) were enrolled and
patients with poor ovarian reserve [8]. In this section, I prospectively randomized into G-CSF or control groups
first describe G-CSF priming, and then 3 other strategies of 50 patients each. Baseline patient characteristics
favoring basal follicular growth that I have been using were similar between G-CSF and control groups: ages,
successfully in my practice of ART for over 30 years. 36.6 ± 3.8 and 37.5 ± 3.5; BMI, 20.9 ± 2.3 and 21.1 ± 2.8;
Finally, I discuss other previously reported methods. infertility duration, 2.3 ± 2.1 years and 2.4 ± 3.1; gravidity,
In our studies, IBM SPSS Statistics (IBM, Tokyo, Japan) 0.8 ± 0.8 and 1.0 ± 1.1; parity, 0.5 ± 0.6 and 0.4 ± 0.6; AMH,
was used for statistical analyses. Data were tested for 0.98 ± 0.54 ng/mL and 0.91 ± 0.49; and similar prevalence
normality by the Shapiro–Wilk test. If data were not of infertility causes (Refer more details to the reference
normally distributed, analysis was performed using the [8]).
Mann–Whitney U test, the Wilcoxon matched-pairs Both groups initially underwent conventional infer-
signed rank test, or Spearman correlation analyses as tility treatment for 2 consecutive cycles, in which the
appropriate. If data were normally distributed, unpaired t G-CSF group additionally received G-CSF priming. Then
tests, paired t tests, Pearson correlation analyses, analysis each group underwent one cycle of IVF/ICSI and fresh
of variance (ANOVA), or Fisher’ s protected least signifi- ET. If fresh ET failed, cryopreserved ET was performed
cant difference (PLSD) test were performed as appropri- until live birth or embryo depletion. Serum AMH was
ate. Data also were analyzed using the chi-squared test, measured before and after G-CSF priming in the G-CSF
Fisher’s exact test, multiple logistic regression analysis, group, with similar measurements in the control group at
receiver-operating characteristics (ROC) curve analysis, similar time points.
or discriminant analysis as was suitable. P values less than Fertilization rate, embryonic development, implanta-
0.05 were considered to indicate significance. Results are tion rate by fresh ET, and oocyte developmental com-
presented as the mean ± standard deviation (SD), except petence were significantly improved by G-CSF priming
for results of the bromocriptine-rebound method which (Fig. 4B and C). Clinical and ongoing pregnancy rates
are presented as the mean ± standard error measurement using IVF/ICSI-fresh ET were significantly higher with
(SEM). G-CSF priming (30% and 26% of 47 ART patients; 3
patients delivered with conventional treatment) com-
G‑CSF priming (II‑1) pared with controls (12% and 10% of 49 ART patients;
About 10 years ago, we administered G-CSF to overcome 1 dropped out). Significantly more G-CSF patients
RIF in 10 patients with diminished ovarian reserve. While achieved cryopreservation of redundant blastocysts than
Jinno Reproductive Biology and Endocrinology (2025) 23:35 Page 9 of 22

Fig. 4 G-CSF study design (A). Oocyte retrieval, fertilization and embryonic development (B), and oocyte developmental competence (C) were
compared between G-CSF and control groups. Significantly more fertilized oocytes and day-2 embryos, a higher rate of blastocyst acquisition,
and higher embryo quality were obtained in the G-CSF group. Implantation rate per transferred embryo was defined as (number of gestational
sacs / number of transferred embryos) × 100%. Serum AMH significantly increased after G-CSF priming, while in controls AMH decreased, resulting
in higher final concentrations of AMH in the G-CSF group (D). G-CSF, granulocyte colony-stimulating factor; AMH, anti-Müllerian hormone; BBT,
basal body temperature; ART, Assisted reproductive technology. Revised from Jinno et al., 2023 [8]

controls (53% vs. 24%) The cumulative live birth rate was as degeneration and fibrosis of ovarian stroma, while
32% in 50 patients with priming, significantly higher than increasing serum AMH [63]. G-CSF administration also
14% in 49 controls (relative risk, 2.8; 95% confidence significantly increased ovarian preantral follicles and
interval, 1.04–7.7). Infants born after priming had no serum AMH in rats with DOR induced by cisplatin [64].
congenital anomalies, while infant weight, birth week, Improvement of early antral folliculogenesis by G-CSF
and Apgar score were similar between groups. Priming could involve various mechanisms. G-CSF has been
significantly increased serum AMH (Fig. 4D), suggesting reported to act directly against apoptosis [65–67], inflam-
enhancement of early antral folliculogenesis. No adverse mation [63, 67, 68], and oxidation [63, 67] while favoring
effects of priming were observed. No significant differ- angiogenesis [67, 69] and growth promotion [67, 70].
ences in killer-cell immunoglobulin-like receptor (KIR) Autocrine or paracrine influences on folliculogenesis by
genotype were observed between patients in the G-CSF G-CSF [71] might be involved, considering that embryos
group with and without achievement of clinical preg- derived from follicles with higher G-CSF were reported
nancy. G-CSF administration together with ET has been to implant more readily [72].
shown to overcome RIF most effectively in patients lack- G-CSF also promotes egress of bone marrow stem cells
ing 2DS1, 2DS5, and 3DS1 [62]. Accordingly, the mech- (BMSC) into peripheral blood [73], which could aid tis-
anism by which G-CSF priming improves early antral sue regeneration considering that ovarian transplanta-
folliculogenesis is likely to be unrelated to alleviation of tion of autologous BMSC collected by apheresis after
RIF. G-CSF administration for 5 days was found to improve
In diabetic rats, G-CSF administration consist- follicle and oocyte quantity, making pregnancy possible
ently decreased ovarian follicular degeneration as well in poor ART responders [74]. On the other hand, human
Jinno Reproductive Biology and Endocrinology (2025) 23:35 Page 10 of 22

plasma derived from apheresis after daily administration PRL, which evolved from the same ancestor hormone as
of G-CSF for 5 days, enriching it in BMSC-secreted fac- growth hormone, is expressed in all vertebrate species
tors, also improved follicular development and fertility in studied, and has more than 300 diverse effects affecting
a mouse model of chemotherapy-induced ovarian dam- reproduction and lactation, water and electrolyte bal-
age [75]. Further, human umbilical cord mesenchymal ance, growth and development, metabolism and endo-
stem cell-derived conditioned medium (hUCMSC-CM) crinology, brain and behavior, immunologic control, and
opposed granulosa cell apoptosis and depletion of pri- ectodermal integrity [86, 87]. In addition to the pituitary,
mordial follicles in cisplatin-treated mice [71]. Thus, an PRL is also produced and secreted at many other sites
indirect mechanism involving BMSC-secreted factors (extrapituitary PRL), including follicles and endometrium
might be responsible rather than transdifferentiation of [88]. PRL receptor (PRLr) is also expressed nearly eve-
BMSC. rywhere in the body [87]. Following hypophysectomy in
Many previous studies have shown that G-CSF rats, PRL bioactivity initially decreased to 15% of normal
improves endometrial receptivity in the same luteal phase but then rose to 50% because of a compensatory increase
as it is administered. G-CSF Administration together in extrapituitary PRL production; while simultaneous
with ET was found to increase implantation rates and administration of anti-PRL antibody kept PRL at 0%, cul-
clinical pregnancy in ART patients with repeated implan- minating in death of all treated rats [89]. This establishes
tation failure or endometrial thinning [76–81]. G-CSF PRL as a growth factor essential to growth and function
also reduced miscarriage rate and increased live birth of cells, acting in endocrine, paracrine, and autocrine
rate in women with unexplained recurrent miscarriages manners. Provision of PRL, so highly important for tis-
[82]. In the absence of 3 activating KIR genes detected sues, appears to be carried out by both central (pituitary)
particularly frequently in women with unexplained and local (extrapituitary) production.
recurrent miscarriage (i.e., lack of 2DS1, 2DS5, and A midcycle PRL surge in serum and PRLr in GCs were
3DS1) [83], G-CSF has shown high effectiveness in over- demonstrated in humans [90], and abnormal follicular
coming repeated implantation failure [62]. Intrauterine development was observed in a woman with isolated
administration of G-CSF was found to increase endome- PRL deficiency [91]. These observations suggest that PRL
trial thickness in women with endometrial thinning [84]. plays a physiological role in human folliculogenesis. In
Considering such observations, G-CSF administration in human IVF, high concentrations of PRL in follicular fluid
the early- and mid-luteal phase may improve endometrial were associated with maturation of the oocyte-cumulus
receptivity by immunologic interactions and endometrial complex, successful fertilization, and pregnancy [92].
growth promotion. Hypoprolactinemia induced with continuous administra-
In our present study, however, G-CSF was adminis- tion of bromocriptine was associated with lower rates of
tered in the cycles preceding to ART and never in the fertilization [93, 94] and embryo cleavage [94]. Therefore,
ET cycles. Thus, the G-CSF direct effects on endometrial PRL appears to play a stimulatory role in the growth and
receptivity, as described above, is not likely involved in maturation of oocytes. Pathological hyperprolactine-
underlying mechanisms improving clinical pregnancy mia is well known to have detrimental effects on various
rate in our study. Consistently, we observed no effects of ovarian functions [95]. Thus, appropriate levels of PRL
G-CSF priming on miscarriage rates or any association of are probably essential to the normal growth and matura-
G-CSF efficacy with KIR genotype. Furthermore, the sig- tion of oocytes.
nificant increase in serum AMH and improvement of fol- About 33 years ago, malfunction of a PRL analyzer in
licular and embryonic developments in our G-CSF group our hospital made us misdiagnose normoprolactine-
strongly suggest early antral follicle growth enhancement mia as hyerprolactinemia; we erroneously administered
as an underlying mechanism. bromocriptine to affected patients for 1 month, until
Recently, a confirmation of our experience with G-CSF discovery of the malfunction. After bromocriptine was
priming was reported in the annual meeting of Japan discontinued, some of those patients conceived despite
Society for Reproductive Medicine on November 14, a long prior interval of treatment failure. In that man-
2024, showing that 3 of 8 women with AMH < 2 ng/mL ner we discovered that ovulation can improve after ces-
and previous ART failures conceived with G-CSF prim- sation of PRL suppression by bromocriptine [96]. We
ing [85]. confirmed effectiveness of this novel method for ovarian
stimulation, bromocriptine-rebound (BR), in a prospec-
Bromocriptine‑rebound method (II‑2) tive randomized controlled trial [97]. The BR method
While gynecologists recognize prolactin (PRL) as a hor- is the same as the long protocol using a GnRH agonist
mone important in lactation and hyperprolactinemic and hMG, except that bromocriptine (Parlodel, Sandoz,
anovulation, its biologic actions are far more varied. Tokyo, Japan) is administered orally at a dose of 2.5 mg/
Jinno Reproductive Biology and Endocrinology (2025) 23:35 Page 11 of 22

day from the early follicular phase (day 3–5) of the pre- BR method is the same when the interval between bro-
ceding cycle until 7 days before initiation of hMG admin- mocriptine cessation and hMG initiation is 7 to 14 days,
istration (Fig. 5A). In practice, the day of bromocriptine while efficacy appears to decrease when the interval is
cessation was scheduled as follows: when GnRH agonist less than 3 days.
in the long protocol was started on the luteal day 4–6, the Endocrinologically normal ovulatory women with
following onset of menstruation was estimated according at least one unsuccessful ART attempt using the long
to the BBT chart by assuming the luteal phase to con- protocol were assigned at random to receive either the
tinue for a total of 14 days. Then, the day of hMG initia- BR method or a typical long protocol. BR significantly
tion was scheduled on the estimated menstrual day 3 and improved fertilization and embryonic development, sur-
the last administration of bromocriptine was scheduled passing rates of pregnancy and delivery attained with
7 days before the day of hMG initiation. Even if actual repeated long-protocol treatment (Fig. 5E). In initial ART
menstruation started earlier than estimated, hMG was attempts using the long protocol, serum PRL concen-
started on the scheduled date, adhering to 7 days after trations during hMG administration (Fig. 5B) and PRLr
bromocriptine cessation so that the actual cycle day mRNA expression in granulosa cells (Fig. 5D) were sig-
of hMG initiation became a few days later than day 3. nificantly higher in non-pregnant than pregnant patients.
When actual menstruation started later than estimated In the BR method, serum PRL concentrations fell sig-
in fewer patients, however, hMG initiation was delayed nificantly with bromocriptine treatment but increased
until the actual cycle day 3 so that the intervals between significantly beyond the pretreatment value after discon-
the bromocriptine cessation and hMG initiation became tinuation of bromocriptine, showing a rebound phenom-
a few days longer than 7 days. Empirically, the efficacy of enon (Fig. 5C). Subsequently, when ART was repeated

Fig. 5 The bromocriptine-rebound (BR) method is the same as in the long protocol, except that bromocriptine is administered from the early
follicular phase of the preceding cycle until 7 days before hMG administration (A). In the initial ART with the long protocol, serum PRL during hMG
administration (B) and PRLr mRNA expression in granulosa cells (D) were higher in patients not achieving pregnancy. When ART was repeated
with the BR method in the non-pregnant patients using the initial long protocol, serum PRL during hMG increased further (C), and PRLr mRNA
expression in granulosa cells decreased (D). The BR method improved fertilization, embryo development, and the rates of pregnancy and delivery
(E). Hypothesized mechanisms for the BR method in improving oocyte maturation (F). OPU, oocyte pick-up. Revised from Jinno et al., 1997 [97]
Jinno Reproductive Biology and Endocrinology (2025) 23:35 Page 12 of 22

using the BR method in non-pregnant patients using the coinciding with a physiologic PRL increase in the late
long protocol, serum PRL concentrations during hMG luteal phase, could enhance responsiveness of selecta-
stimulations increased further (Fig. 5C) while PRLr ble follicles to Gn, promoting cyclic recruitment. This
mRNA expression in granulosa cells decreased (Fig. 5D). might be a third mechanism by which the BR method to
As an explanation of these results, we propose that improve oocyte quality.
patients with poor ART outcomes using the long proto- Our observations that the BR method improves embry-
col may have decreased follicular PRL production and/ onic quality and pregnancy rates in patients with previ-
or postreceptor responsiveness of GC to PRL, causing ous ART failures were reconfirmed by 2 clinical trials in
compensatory increases in serum PRL concentrations Tokushima University Hospital [103, 104] and Kitazato
and GC PRLr mRNA and PRLr (Fig. 5F) [97]. Most likely, University Hospital [105].
however, this compensation is inadequate, so that oocyte
maturation is impaired due to inadequate growth-stimu- Interventions affecting carbohydrate metabolism (II‑3)
latory effect of PRL. Bromocriptine suppresses pituitary Glucose metabolism is essential to all human cells as a
PRL production but has no effects on extrapituitary PRL requirement for viability and biologic function, especially
production [88]. Therefore, when the BR method is used in terms of energy production; it is also necessary to sus-
in such patients with poor ART outcomes using the long tain folliculogenesis. Insulin is essential to control of glu-
protocol, bromocriptine administration suppresses pitui- cose metabolism, while insulin resistance (IR) increases
tary, but not follicular, PRL production during the cycle oxidative stress, promoting formation of advanced gly-
preceding to ART cycle. Small antral follicles growing cation end-products (AGE) that induce inflammation,
there are compelled to adapt themselves to hypoprol- oxidative stress, and IR, representing vicious cycles pro-
actinemic circumstances by increasing their follicular moting disease processes [106, 107]. Toxicity of AGE
PRL production and/or postreceptor responsiveness of results directly from macromolecular trapping and
GC to PRL to compensate decreased PRL supply from cross-linking and indirectly from binding to AGE recep-
pituitary. After discontinuation of bromocriptine, the tors (RAGE) [106, 108]. Vicious cycles of IR, oxidative
serum PRL concentrations increase beyond the pre- stress, and AGE toxicity contribute to the pathogenesis
treatment value as a rebound phenomenon by cessation of diseases such as metabolic syndrome, type 2 diabetes,
of suppression. Consequently, recruited antral follicles, hypertension, atherosclerosis, dyslipidemia, nonalcoholic
which are improved of follicular PRL production and/or fatty liver disease, central obesity, polycystic ovary syn-
responsiveness to PRL, receive increased pituitary PRL drome (PCOS), and even conditions such as aging [106,
supply during hMG stimulation, resulting in an improve- 109–114].
ment of oocyte maturation by adequate PRL effects. A IR is strongly associated with aging [115], stress [116],
decrease in PRLr mRNA expression appears to reflect the depression [117], obesity [118], and sedentary lifestyle
recovery of follicular PRL production and/or postrecep- [119]. These conditions are frequent among infertile
tor responsiveness to PRL. women, suggesting a pathogenetic role of IR even in
PRL production by human endometrium has been infertile women without PCOS. Metformin, an insu-
demonstrated, typically following day 23 of the menstrual lin sensitizer, consistently improves pregnancy rates in
cycle [98, 99] and occasionally as early as day 18 [99]. not only patients with PCOS [120] but also non-PCOS
Endometrial PRL production increases as decidualiza- patients with repeated past failures of ART [121]. More-
tion progresses [98, 100], showing more rapid increases over, AGE are linked to causes of infertility including
as implantation progresses [101]. Similarly, PRL is detect- PCOS, ovarian dysfunction, diminished ovarian reserve,
able in endometrial fluid from monkeys on day 24, fol- ovarian aging, endometriosis, and male infertility [122–
lowed by a rapid increase in secretion [102]. Increased 125]. Accordingly, the pathophysiology of infertility
endometrial PRL could reach the ovary by diffusion or via resembles that of other diseases involving IR.
blood vessels and lymphatics, increasing PRL concentra- Given the links between insulin resistance and infer-
tions in mid- and late luteal phase follicles, which might tility described above, we examined whether adminis-
stimulate cyclic recruitment as mentioned in section I-5. tration of low-dose metformin (500 mg/day) improved
I recently attempted to increase PRL concentrations in ART outcomes in non-PCOS patients with repeated
the late luteal phase by direct pharmacologic interven- ART failures in a prospective randomized controlled trial
tion, resulting in beneficial effects on oocyte develop- [121]. The study included infertile women who had failed
mental competence similar to those with the BR method to conceive in 2 or more previous ART attempts and
that were followed by live births (unpublished data). A excluded PCOS women diagnosed according to the 2003
steep increase in PRL concentrations just after discon- Rotterdam criteria. Patients were allocated at random
tinuation of bromocriptine in the BR method (Fig. 5C), to metformin or non-metformin groups, irrespective of
Jinno Reproductive Biology and Endocrinology (2025) 23:35 Page 13 of 22

assessment of insulin resistance. No patients had history (Fig. 6B). When serum TAGE exceeded 7.24 U/mL, num-
of diabetes. In the same study [121], we also considered bers of retrieved oocytes were significantly decreased
whether effectiveness of metformin can be predicted by early on at ages 35–40, with further decreases at ages
a discriminant score (DS) calculated from multiple IR- above 40 (Fig. 6C, left panel). Attainment of ongoing
related parameters assessed before metformin admin- pregnancy was compromised even at ages below 35 years,
istration [121]. We administered metformin from 2 to with greater worsening above 35 years (Fig. 6C, right
3 months before ART until delivery or failure, consider- panel). Even when day-3 FSH was less than 10 IU/L, like-
ing that glucose metabolism makes important contribu- lihood of ongoing pregnancy was significantly lower in
tions to folliculogenesis, implantation, and pregnancy. patients with TAGE exceeding 7.24 U/ml than in patients
Rates of implantation and ongoing pregnancy were sig- with less (6.1% vs. 25%).
nificantly increased by metformin in cases where the DS Finally, we have improved ART outcomes by reducing
predicted its effectiveness (Fig. 6A). AGE [126]. The water chestnut (Trapa bispinosa Roxb.),
Next, we examined correlations between AGE concen- a common aquatic plant in Asia, Europe, and Africa that
trations in blood and follicular fluid (FF) and outcomes in has been consumed safely as food and herbal medicine
157 consecutive cycles of ART [123]. Toxic AGE (TAGE) for over 2000 years, has antidiabetic, analgesic, antibi-
in serum, as well as pentosidine, N-carboxymethyl lysine otic, and immunomodulatory activities [127]. An extract
(CML), and TAGE in FF, showed a significant negative of its dried peel (Hishi extract) used as a cosmetic sup-
correlation with follicular and embryonic development plement was found to inhibit AGE formation and disrupt

Fig. 6 Low-dose metformin increased rates of implantation and ongoing pregnancy when a discriminant score (DS), calculated from IR-related
multiple parameters, predicted it as effective (A). Toxic AGE (TAGE) in serum and pentosidine, N-carboxymethyl lysine (CML), and TAGE in FF
correlated negatively with follicular and embryonic development (B). Number of retrieved oocytes and rate of ongoing pregnancy decreased more
sharply and from an earlier age in patients with higher TAGE in serum (C). Developmental potentials of oocytes and embryos (D), Cumulative live
delivery rate per patient, live delivery rate per ET, and implantation rate (E) were improved by administration of Hishi extract. Hishi extract decreased
­Nδ-(5-hydro-5-methyl-4-imidazolone-2-yl)-ornithine (MG-H1) and N.ω-(carboxymethyl) arginine (CMA) in serum (F) and CMA in follicular fluid (G).
Revised from Jinno et al., 2010 [121], Jinno et al., 2011 [123], and Jinno et al., 2021 [126]
Jinno Reproductive Biology and Endocrinology (2025) 23:35 Page 14 of 22

α-dicarbonyl compounds and AGE crosslinks in vitro mice [130]. Our study [126], however, showed little
[128]. effect of Hishi extract on such glucose metabolism met-
We conducted a prospective randomized controlled rics as fasting plasma glucose, serum insulin, HOMA-
trial to determine whether Hishi extract improved ART R, insulinogenic index, hemoglobin A1c, glycoalbumin,
outcome by reducing AGE [126]. The 64 study patients, C-peptide, and OGTT. Therefore, Hishi extract appeared
with ages ranging from 38 to 42 years, were likely to have unlikely to decrease AGE indirectly by reductions in
age-related AGE accumulation. In Japan, 40.7% of ART blood glucose or enhanced insulin sensitivity or secre-
treatments involve this age group, which has a delivery tion. More likely, Hishi extract directly inhibited produc-
rate of only 9.3% [129]; such patients are on the verge of tion of AGE and their intermediates or disrupted their
becoming too old to overcome infertility. bridging structures, as has been demonstrated in vitro
The patients were randomized to groups receiving ART [128]. MG-H1 and CMA, which are readily generated
with or without administration of Hishi extract (100 mg/ in vitro from the α-dicarbonyl compounds, methylgly-
day; Pregnasupport; Hayashikane Sangyo, Shimonoseki, oxal [131] and glyoxal, respectively [132], were particu-
Japan), continuing from 2 cycles before ART until late larly sensitive to the in vivo effects of Hishi in our study,
pregnancy or failure. Both groups underwent 1 cycle of which inhibits production of α-dicarbonyl compounds
conventional infertility treatment, then 1 cycle of ART and degrades them in vitro [128].
using fresh ET with a long protocol, and, if needed, cryo- Physicians should encourage patients to adopt as
preserved ET until live birth or embryo depletion. Serum healthy lifestyle as possible before any interventions are
AGE were measured before and during ART, as were attempted. Insulin sensitivity is determined physiologi-
AGE in FF. cally by heredity, age, and lifestyle, among which only
Developmental potentials of oocytes and embryos the last factor can be modified. Several points of lifestyle
were significantly greater in the Hishi group than con- modification are emphasized in our clinic: (a) Be happy.
trols (Fig. 6D). Cumulative live birth rate among 32 Hishi Even pretending to be happy is effective. Don’t worry
patients was 47%, signifcantly higher than 16% among about infertility! Worrying about anything worsens IR
31 controls (P < 0.01; RR, 4.6; 95% CI, 1.4–15.0; Fig. 6E); and various disorders, including infertility. (b) Walk con-
1 control patient dropped out. In ART including both tinuously for more than 45 min every day, even on holi-
fresh and cryopreserved ET, the live delivery rate per days, except when ill. Mild to moderate muscle training
ET and implantation rate per embryo both were signifi- exercise also is recommended. (c) Go to sleep before
cantly higher in the Hishi group than in controls (28% vs. 11 pm, wake up around 6 am, having slept for 7–8 h.
10%, P < 0.05 and 23% vs. 8.5%, P < 0.01; Fig. 6E). Among (d) Adjust BMI to between 19 and 23. (e) Absolutely no
4 major fertility-related variables (age, day-3 FSH, AMH, smoking. (f ) If you take an alcoholic drink, abstain for the
and Hishi), only Hishi significantly correlated with cumu- following 3 days. Daily drinking of even small amounts
lative live delivery (P < 0.05; odds ratio, 5.1; 95% CI., increases IR. (g) Partake of healthy food and beverages.
1.4–18.3; logistic regression). Fifteen Hishi and five con- Ordinary meals should be freshly cooked using healthful
trol patients respectively delivered 17 and 5 normal live ingredients. Avoid sugar, especially sweet drinks. Pre-
infants, including 2 and 0 sets of twins. No significant dif- served food can contain AGE, which accumulate in the
ference was evident in body weight or gestational age at body if frequently consumed. Eat at a relaxed pace and
delivery. No adverse effects of Hishi were observed. chew your food.
Serum concentrations of Lifestyle modification is essential for optimum treat-
­N δ-(5-hydro-5-methyl-4-imidazolone-2-yl)-ornithine ment even though it is often difficult. Medical interven-
(MG-H1) and N ­ ω-(carboxymethyl) arginine (CMA) tions cannot achieve desired effects without adequate
fell significantly in the Hishi group but not in controls attention to lifestyle.
(Fig. 6F). Hishi extract also significantly decreased
CMA in FF (Fig. 6G). In addition, reduction of AGE by Danazol priming (II‑4)
Hishi administration correlated with improved follicu- Danazol, a weak androgenic agent used to treat endome-
logenesis, fertilization, embryonic development, and triosis, has been reported to improve pregnancy rate in
implantation [126]. It is suggested, therefore, that AGE patients with ART failures despite more than one embryo
decreases as a therapeutic mechanism underlying fer- having optimal morphology [133, 134]. The original pro-
tility enhancement by Hishi. tocol involved administration of danazol at 400 mg/day
How Hishi decreases AGE in vivo is much less clear. for 12 weeks. ART was repeated within 3 months of the
Polyphenols from water chestnut (Trapa japonica) husk first ovulation after danazol discontinuation. We have
were found to inhibit α-amylase and α-glucosidase, shortened the danazol priming protocol as follows. Dan-
reducing blood glucose and insulin concentrations in azol (Bonzol; Tanabe Co., Ltd., Tokyo, Japan), 200 mg/
Jinno Reproductive Biology and Endocrinology (2025) 23:35 Page 15 of 22

day, is administered for 6 weeks beginning on cycle day the differences might be improvement of insulin sensi-
2–3, followed directly by one Kaufmann cycle and then tivity related to increased muscle mass. Alternatively,
ART. For about 25 years, we have been using our danazol androgenic effects of danazol have enhanced preantral/
priming protocol successfully to overcome repeated ART small antral follicle growth, improving oocyte quality.
failures, irrespective of embryo quality in previous ET. In Danazol enhances telomerase activity in vitro [135] and
this opportunity reviewing interventions affecting basal has been used to treat bone marrow failure syndromes
follicular growth, we decided to analyze retrospectively with genetic defects in telomere maintenance and repair,
our ART data for the last 15 years to determine whether increasing both telomere length and peripheral-blood
danazol priming improved embryo development and cell counts [136]. Recently, a randomized clinical pilot
live birth rate as well as the likely mechanism of action. trial attempted to improve telomeric and fertility param-
Unfortunately, we have been using this protocol simply as eters in patients with diminished ovarian reserve by
a routine clinical treatment without interest in research- 3 months of danazol administration, directly followed
ing it further, so we have not measured hormones before by ovarian stimulation for ART without a connecting
and after danazol administration. Kaufmann cycle– differently from our danazol prim-
One hundred thirteen relatively old patients (age: ing protocol [137]. All 5 patients in their danazol group
40.8 ± 3.7 SD) with repeated ART failure (6.1 ± 4.8) under- canceled their post-treatment ART due to poor follicular
went danazol priming and ART using a long/antagonist growth, although their blood examinations suggested an
protocol (danazol group; 139 cycles of ART). The same influence of danazol on telomere maintenance [137]. A
113 patients also underwent 616 cycles of ART with simi- few months after danazol cessation, however, the dana-
lar ovarian stimulation but no danazol priming (control zol patients showed a tendency for higher frequency of
group). ART outcomes were compared between danazol mature oocytes in repeated ART, compared with their
and control groups and also between subjects with and ART before the danazol trial [137]. Taken together with
without endometriosis in the danazol group. Body weight our results, this suggests that an interval of a few weeks
and % body fat were measured based on electrical resist- between danazol cessation and hMG initiation are neces-
ance (Body fat monitor, HBF-306, Omron Healthcare sary to avoid adverse effects of danazol on folliculogen-
Co., Ltd., Kyoto, Japan) before and after danazol priming. esis. Further, to maximize beneficial effects of danazol
In danazol and control groups, numbers of retrieved priming, ovarian stimulation probably should be added at
oocytes (8.4 ± 7.1 vs. 7.6 ± 7.1) and fertilized oocytes an appropriate time–meaning not too late. Consistently,
(5.3 ± 4.5 vs. 4.6 ± 4.1) were not significantly different, danazol treatment alone without ovarian stimulation
but numbers of day-2 embryos (4.8 ± 4.1 vs. 4.1 ± 3.8) did not accomplish any improvement of live birth rate in
and day-3 embryos (5.5 ± 4.0 vs. 4.3 ± 3.4) were signifi- women with unexplained subfertility, even in long term
cantly higher in the danazol group. The rate of blasto- follow-up [138].
cyst acquisition per cycle was significantly higher in the Androgen production begins with the large second-
danazol group than in controls (41% vs. 29%). Rates of ary follicle [2], increasing gradually during small antral
clinical pregnancy (19% vs. 6.5%) and live birth (10.4% folliculogenesis (10 weeks). Aromatase activity quickly
vs. 2.3%) per cycle both were significantly higher in the increases from early- to mid-luteal phase [16], during
danazol group than in controls (P < 0.001; odds ratio, the last 1–2 weeks of small antral folliculogenesis before
3.3 and 4.9, respectively). Comparing danazol cycles cyclic recruitment (prerecruitment period) (sections I-3
in patients with (20 cycles) and without endometriosis and I-4, Fig. 1). Thus, the period of prerecruitment plus
(119 cycles), no significant differences were apparent in recruitment is the transitional period from androgenic
fertilization, embryonic development, or rates of clinical to estrogenic milieu. GS-specific-androgen-receptor-
pregnancy and live birth. Danazol priming significantly knockout mice exhibited a block in folliculogenesis at the
increased body weight (55.1 ± 7.5 kg and 57.1 ± 7.6 before preantral secondary follicle [139], suggesting a critical
and after danazol, respectively; P < 0.001, n = 107, paired role of androgen in small antral folliculogenesis. Conceiv-
t-test) and decreased percent body fat (28.0 ± 4.4% and ably, the first two-thirds of small antral folliculogenesis
27.5 ± 4.4 before and after danazol, respectively; P < 0.05, (about 6.7 weeks) simply in an increasingly androgenic
n = 101, paired t-test), suggesting an increase in amount milieu may safely benefit from androgen augmentation,
of muscle. but the last third of small antral folliculogenesis before
Thus, danazol priming improved embryo development, cyclic recruitment (3.3 weeks) in the transitional period
pregnancy rate, and live birth rate in older women with from androgenic to estrogenic milieu may be too sensi-
repeated ART failure. Efficacy did not vary with presence tive or vulnerable to steroid manipulation. Thus, our
or absence of endometriosis. The mechanism underlying danazol priming protocol, consisting of danazol priming
Jinno Reproductive Biology and Endocrinology (2025) 23:35 Page 16 of 22

for 6 weeks, Kaufmann cycle for 3–4 weeks, and then therapies require a surgical approach. Further, under-
ovarian stimulation, probably suits the changing support- standing of their efficacy, safety, and mechanism remains
ive demands of small antral folliculogenesis according to limited. Ongoing fundamental and clinical research is
developmental stage. needed before wider application of these therapies.

Other strategies (II‑5)

Various pretreatments using androgens such as testos- Conclusion
terone or dehydroepiandrosterone (DHEA), androgen- About 33 years ago I repeatedly used the same long
modulating agents including letrozole, recombinant LH, protocol in 304 patients. The pregnancy rate sharply
and hCG, pretreatment pituitary suppression using a dropped after the second attempt, and the cumulative
GnRH antagonist, an estrogen, or oral contraceptive pills, pregnancy rate plateaued at 35% (Fig. 7A). Since then, I
GH, and coenzyme Q10, have been used to overcome have developed several new methods of ovarian stimula-
poor ovarian response. Recently, Orvieto reviewed those tion, as described in this review, so that pregnancy rate
pretreatments, concluding that none of them showed was improved even in patients with repeated ART fail-
adequately convincing benefit [61]. ures. Naturally, different patients require different ways
A meta-analysis [140] including 3 randomized con- to enhance folliculogenesis. Although various conven-
trolled trials (RCT) in 2006, 2009, and 2011 and one tional methods for ovarian stimulation are in use, the
RCT in 2021 [141] showed significant increases in rates mechanism appears to be similar, involving an increase of
of clinical pregnancy and live birth with transdermal Gn levels during folliculogenesis after cyclic recruitment.
testosterone pretreatment in poor ovarian responders, On the other hand, the high or low AMH values often
while 3 recent RCTs in 2016, 2021, and 2023 showed no encountered in infertility reflect pathogenesis involv-
significant increases in numbers of retrieved and mature ing Gn-responsive but Gn-independent folliculogenesis
oocytes as well as clinical pregnancy rates [142–144]. before cyclic recruitment (Fig. 7B). Therefore, we need
Pretreatment with dehydroepiandrosterone (DHEA) in to accomplish ovarian stimulation for folliculogenesis
poor responders also significantly increased rates of clini- before cyclic recruitment, using means other than Gn
cal pregnancy and live birth in a meta-analysis [145] con- enhancement.
sisting of 21 studies in 2006 to 2015 (including 8 RCTs) Serendipitous discovery of G-CSF priming taught
and a retrospective cohort study in 2018 [146], whereas a me that modulation of basal follicular growth is fea-
recent RCT including a large number of participants (821 sible, encouraging me to seek new ways for the BR
women) showed no beneficial effects [147]. Thus, despite method to improve folliculogenesis since BR has helped
the vast amount of literature on the use of testosterone so many patients in my ART practice over 3 decades. I
and DHEA in poor responders, the bulk of evidence is believe that the steep increase in PRL that immedi-
still too limited to draw definite conclusions [148]. ately follows discontinuation of bromocriptine is likely
As I described in section II-4, small antral folliculogen- to enhance responsiveness of selectable follicles to Gn
esis within 1–2 weeks before cyclic recruitment may be for cyclic recruitment. Consistently in our recent pilot
vulnerable to steroid manipulation since this is a period study, increased PRL in the late luteal phase mimicked
of transition from an androgenic to an estrogenic follicu- beneficial effects of the BR method on oocyte qual-
lar milieu. In all studies for testosterone and DHEA pre- ity. Also, interventions affecting glucose metabolism,
treatment up to now, androgen pretreatment was always such as increasing insulin sensitivity with metformin
followed directly by ovarian stimulation with no inter- and decreasing AGE with Hishi extract, work best when
vening interval, making follicles recruited by Gn spend started 1 to 2 cycles before the ART cycle, i.e., during
the vulnerable period of small antral folliculogenesis early antral folliculogenesis. Danazol, a weak androgen,
under androgen manipulation. This adverse effect during administered during the first two-thirds of that time
prerecruitment period could counteract possible benefi- improves oocyte developmental competence and live
cial effects on earlier small antral folliculogenesis, leading birth rate.
to limited ART outcomes. Without question, the 4 methods just discussed need
Recently, new therapeutic options such as intraovar- to be validated by further investigations. I have published
ian injection of platelet-rich plasma [149], in vitro activa- and presented the BR method in journals and scientific
tion (IVA), and drug-free IVA therapies [150] have been meetings, getting limited attention because of a stereo-
developed for patients with premature ovarian insuffi- type that “increasing PRL is bad” as well as unaware-
ciency, diminished ovarian reserve, and resistant ovary ness of the importance of extrapituitary PRL and PRL
syndrome, resulting in successful pregnancies and births. in general. For 3 decades, however, I still have success-
While these clinical trials report promising results, such fully used these methods in my clinic because they work
Jinno Reproductive Biology and Endocrinology (2025) 23:35 Page 17 of 22

Fig. 7 Pregnancy rate steeply decreased while repeating the same protocol of ovarian stimulation (A). A novel strategy to treat ovarian dysfunction
by improving preantral and early antral folliculogenesis (B)

dependably in my patients, enabling thousands of births. CCN  Cysteine-rich angiogenic protein 61, connective tissue
growth factor, and nephroblastoma overexpressed
I believe that time will bear me out. CI Confidence interval
The methods explained in this review act indepen- CMA Nω-(carboxymethyl) arginine
dently of Gn, improving early antral folliculogenesis CML N-carboxymethyl lysine
CNP C-type natriuretic peptide
before cyclic recruitment, although the BR method, DHEA-S Dehydroepiandrosterone sulfate
metformin, and Hishi extract additionally enhance later DOR Diminished ovarian reserve
antral folliculogenesis. I hope that these treatments will EGF Epidermal growth factor
ET Embryo transfer
help many more patients who are refractory to conven- E2 17β-Estradiol
tional ovarian stimulation, and that basic researchers and FOXO3 Forkhead box O3
physicians will further explore stimulation of preantral FSH Follicle-stimulating hormone
GC Granulosa cell(s)
and early antral folliculogenesis. G-CSF Granulocyte colony-stimulating factor
GDF Growth differentiation factor
Abbreviations
Gn Gonadotropin
AGE Advanced glycation end-products
GnRH Gonadotropin-releasing hormone
AKT Protein kinase B
hCG Human chorionic gonadotropin
AMH Anti-Müllerian hormone
HGF Hepatocyte growth factor
ART​ Assisted reproductive technology
hMG Human menopausal gonadotropin
BDNF Brain-derived neurotrophic factor
ICSI Intracytoplasmic sperm injection
BFGF Basic fibroblast growth factor
IGF Insulin-like growth factor
BIRC  Baculoviral inhibitors of apoptosis repeats-containing
IGFBP Insulin-like growth factor binding protein
proteins
IR Insulin resistance
BMI Body mass index
IVA in vitro activation
BMP Bone morphogenetic protein
IVF in vitro fertilization
BMSC Bone marrow stem cells
IWI Interwave interval
BR method Bromocriptine-rebound method
JAK/STAT​ Janus kinase /signal transducer and activator of transcription
Jinno Reproductive Biology and Endocrinology (2025) 23:35 Page 18 of 22

KGF Keratinocyte growth factor Consent for publication

KIR Killer-cell immunoglobulin-like receptor Not applicable.
LATS1/2-Mob1 Large tumor suppressor homolog 1/2-MOB kinase activator
1 Competing interests
LH Luteinizing hormone The authors declare no competing interests.
LIF Leukemia inhibitory factor
MAPK Mitogen-activated protein kinase
MG-H1 Nδ-(5-hydro-5-methyl-4-imidazolone-2-yl)-ornithine Received: 22 November 2024 Accepted: 4 February 2025
MST1/2-Sav1 Mammalian Ste-20 like kinase 1/2-Salvador 1
mTORC1 Mammalian target of rapamycin complex 1
NPR2 Natriuretic peptide receptor 2
NTF5 Neurotrophin 5
OPU Oocyte pick-up References
OS Ovarian stimulation 1. Edson MA, Nagaraja AK, Matzuk MM. The mammalian ovary from
PAPP-A Pregnancy-associated plasma protein-A genesis to revelation. Endocr Rev. 2009;30:624–712. [Link]
PCOS Polycystic ovary syndrome 1210/​er.​2009-​0012.
PDK1 Phosphatidylinositol-dependent kinase 1 2. Gougeon A. Human ovarian follicular development: from activa-
PETEN Phosphatase and tensin homolog tion of resting follicles to preovulatory maturation. Ann Endocrinol.
PI3K Phosphatidylinositol 3-kinase 2010;71:132–43. [Link]
PIP2 Phosphatidylinositol-4,5-biphosphate 3. Rimon-Dahari N, Yerushalmi-Heinemann L, Alyagor L, Dekel N. Ovarian
PIP3 Phosphatidylinositol-3,4,5-triphosphate folliculogenesis. Results Probl Cell Differ. 2016;58:167–90. [Link]
PRL Prolactin org/​10.​1007/​978-3-​319-​31973-5_7.
PRP Platelet-rich plasma 4. Hsueh AJW, Kawamura K, Cheng Y, Fauser BCJM. Intraovarian control
RAGE AGE receptor of early folliculogenesis. Endocr Rev. 2015;36:1–24. [Link]
RCT​ Randomized controlled trial 1210/​er.​2014-​1020.
RIF Repeated implantation failure 5. Morton AJ, Candelaria JI, McDonnell SP, Zgodzay DP, Denicol AC.
rpS6 Ribosomal protein S6 Review: Roles of follicle-stimulating hormone in preantral folliculogen-
RR Relative risk esis of domestic animal: what can we learn from model species and
RSOS Random start ovarian stimulation where do we go from here? Animal. 2023;17:100743. [Link]
SD Standard deviation 1016/j.​animal.​2023.​100743.
S6K1 P70 S6 kinase 1 6. Guzmán A, Hernández-Coronado CG, Gutiérrez CG, Rosales-Torres
TAGE Toxic AGE AM. The vascular endothelial growth factor (VEGF) systems as a key
TEAD  Transcription factors containing the TEA (transcriptional regulator of ovarian follicle angiogenesis and growth. Mol Reprod Dev.
enhancer activator) DNA binding domain 2023;90:201–17. [Link]
TIC Theca interna cell(s) 7. Dewailly D, Andersen CY, Balen A, Broekmans F, Dilaver N, Fanchin R,
TGF Transforming growth factor Griesinger G, Kelsey TW, Marca AL, Lambalk C, Mason H, Nelson SM, Vis-
TKR Tyrosine kinase receptor ser JA, Wallace WH, Anderson RA. The physiology and clinical utility of
TSC1/2 Tuberous sclerosis complex 1/2 anti-Mullerian hormone in women. Hum Reprod Update. 2014;20:370–
VEGF Vascular endothelial growth factor 85. [Link]
WNT Wingless 8. Jinno M, Tamaoka Y, Teruya K, Watanabe A, Hatakeyama N, Goda T,
YAP/TAZ Yes-associated protein/transcriptional coactivator with PDZ- Kimata H, Jinno Y. Granulocyte colony-stimulating factor priming
binding motif improves embryos and pregnancy rate in patients with poor ovar-
ian reserve: a randomized controlled trial. Reprod Biol Endocrinol.
Acknowledgements 2023;21:29. [Link]
I thank Mr. Naohisa Hatakeyama for his excellent graphic work, Ms. Aiko 9. Gougeon A. Dynamics of follicular growth in the human: a model from
Watanabe, Mr. Tomoya Gouda, and Mr. Hayato Kimata for their diligent clinical preliminary results. Hum Reprod. 1986;1:81–7.
work, Ms. Sayuri Hoshi for her diligent management of patient charts, and Dr. 10. Paulino LRFM, de Assis EIT, Azevedo VAN, Silva BR, da Cunha EV, Silva
Jeffrey Smith for checking English usage in my manuscript. JRV. Why is it so difficult to have competent oocytes from in vitro cul-
tured preantral follicles? Reprod Sci. 2022;29:3321–34. [Link]
Author’s contributions 10.​1007/​s43032-​021-​00840-8.
Masao Jinno, MD, PhD made almost all contributions to study design and to 11. Telfer EE, Grosbois J, Odey YL, Rosario R, Anderson RA. Making a good
acquisition, analysis, and interpretation of data. He wrote all part of this article. egg: human oocyte health, aging, and in vitro development. Physiol
He gave final approval for this version of the manuscript to be published. Rev. 2023;103:2623–77. [Link]
12. Pepling ME, Spradling AC. Mouse ovarian germ cell cysts undergo
Funding programmed breakdown to form primordial follicles. Dev Biol.
I received no grant or financial support for this study. 2001;234:339–51. [Link]
13. Yuan L, Huang W, Bi Y, Chen S, Wang X, Li T, et al. G-CSF-mobilized
Data availability peripheral blood mononuclear cells combined with platelet-rich
No datasets were generated or analysed in the part of review in this study. The plasma restored the ovarian function of aged rats. J Reprod Immunol.
datasets used and/or analyzed in the section of II-4. danazol priming in this 2023;158:103953. [Link]
study are available from the corresponding author on reasonable request. 14. Badawy A, Sobh MA, Ahdy M, Abdelhafez MS. Bone marrow mesenchy-
mal stem cell repair of cyclophosphamide-induced ovarian insuffi-
ciency in a mouse model. Int J Women’s Health. 2017;9:441–7.
Declarations 15. Tilly JL, Telfer EE. Purification of germline stem cells from adult mam-
malian ovaries: a step closer towards control of the female biological
Ethics approval and consent to participate
clock? Mol Hum Reprod. 2009;15:393–8. [Link]
Informed consent was obtained from all patients. The study was approved by
molehr/​gap036.
the Ethics Committees of Women’s Clinic Jinno. Clinical trials referenced in this
16. McNatty KP, Hillier SG, van den Boogaard AMJ, Trimbos-Kemper TCM,
review were registered with the UMIN in Japan and each registration number
Reichert LE, van Hall EV. Follicular development during the luteal phase
was stated in each published paper. Our research was conducted in accord-
of the human menstrual cycle. J Clin Endocrinol Metab. 1983;56:1022–
ance with the Declaration of Helsinki.
31. [Link]
Jinno Reproductive Biology and Endocrinology (2025) 23:35 Page 19 of 22

17. Gougeon A, Testart J. Influence of human menopausal gonado- 38. Zhang M, Su YQ, Sugiura K, Xia G, Eppig JJ. Granulosa cell ligand NPPC
tropin on the recruitment of human ovarian follicles. Fertil Steril. and its receptor NPR2 maintain meiotic arrest in mouse oocytes. Sci-
1990;54:848–52. ence. 2010;330:366–9. [Link]
18. Matthews CH, Borgato S, Beck-Peccoz P, Adams M, Tone Y, Gambino 39. Durlinger ALL, Gruijters MJG, Kramer P, Karels B, Kumar TR, Matzuk MM,
G, Casagrande S, Tedeschini G, Benedetti A, Chatterjee VKK. Primary Rose UM, de Jong FH, Uilenbroek JTJ, Grootegoed JA, Themmen APN.
amenorrhoea and infertility due to a mutation in the β-subunit of Anti-Müllerian hormone attenuates the effects of FSH on follicle devel-
follicle-stimulating hormone. Nat Genet. 1993;5:83–6. opment in the mouse ovary. Endocrinology. 2001;142:4891–9.
19. Gougeon A. Regulation of ovarian follicular development in primates: 40. Weenen C, Laven JSE, Von Bergh ARM, Cranfield M, Groome NP, Visser
facts and hypotheses. Endocr Rev. 1996;17:121–55. JA, Kramer P, Fauser BCJM, Themmen APN. Anti-Müllerian hormone
20. Kumar TR, Wang Y, Lu N, Matzuk MM. Follicle stimulating hormone is expression pattern in the human ovary: potential implications for initial
required for ovarian follicle maturation but not male fertility. Nat Genet. and cyclic follicle recruitment. Mol Hum Reprod. 2004;10:77–83. [Link]
1997;15:201–4. doi.​org/​10.​1093/​molehr/​gah015.
21. Dierich A, Sairam MR, Monaco L, Fimia GM, Gansmuller A, LeMeur M, 41. Nilsson EE, Skinner MK. Bone morphogenetic protein-4 acts as an
Sassone-Corsi P. Impairing follicle-stimulating hormone (FSH) signal- ovarian follicle survival factor and promotes primordial follicle develop-
ing in vitro: targeted disruption of the FSH receptor leads to aberrant ment. Biol Reprod. 2003;69:1265–72. [Link]
gametogenesis and hormonal imbalance. Proc Natl Acad Sci USA. eprod.​103.​018671.
1998;95:13612–7. 42. Lee WS, Yoon SJ, Yoon TK, Cha KY, Lee SH, Shimasaki S, Lee S, Lee KA.
22. Oktay K, Briggs D, Gosden RG. Ontogeny of follicle-stimulating hor- Effects of bone morphogenetic protein-7 (BMP-7) on primordial fol-
mone receptor gene expression in isolated human ovarian follicles. J licular growth in the mouse ovary. Mol Reprod Dev. 2004;69:159–63.
Clin Endocrinol Metab. 1997;82:3748–51. [Link]
23. Shima K, Kitayama S, Nakano R. Gonadotropin binding sites in human 43. Shimasaki S, Zachow RJ, Li D, Kim H, Iemura S, Ueno N, Sampath K,
ovarian follicles and corpora lutea during the menstrual cycle. Obstet Chang RJ, Erickson GF. A functional bone morphogenetic protein
Gynecol. 1987;69:800–6. system in the ovary. Proc Natl Acad Sci USA. 1999;96:7282–7.
24. Dong J, Albertini DF, Nishimori K, Kumar TR, Lu N, Matzuk MM. Growth 44. Vendola KA, Zhou J, Adesanya OO, Weil SJ, Bondy CA. Androgens stimu-
differentiation factor-9 is required during early ovarian folliculogenesis. late early stages of follicular growth in the primate ovary. J Clin Invest.
Nature. 1996;383:531–5. 1998;101:2622–9.
25. Otsuka F, Yao Z, Lee T, Yamamoto S, Erickson GF, Shimasaki S. Bone 45. Weil S, Vendola K, Zhou J, Bondy CA. Androgen and follicle-stimulating
morphogenetic protein-15. Identification of target cells and biological hormone interactions in primate ovarian follicle development. J Clin
functions. J Biol Chem. 2000;50:39523–8. Endocrinol Metab. 1999;84:2951–6.
26. Cheng Y, Kawamura K, Takae S, Deguchi M, Yang Q, Kuo C, Hsueh AJ. 46. McGee EA, Hsueh AJW. Initial and cyclic recruitment of ovarian follicles.
Oocyte-derived R-spondin2 promotes ovarian follicle development. Endocr Rev. 2000;21:200–14.
FASEB J. 2013;27:2175–84. 47. Baerwald AR, Adams GP, Pierson RA. Ovarian antral folliculogenesis
27. Vitt UA, McGee EA, Hayashi M, Hsueh AJ. In vivo treatment with during the human menstrual cycle: a review. Hum Reprod Update.
GDF-9 stimulates primordial and primary follicle progression and 2012;18:73–91. [Link]
theca cell marker CYP17 in ovaries of immature rats. Endocrinology. 48. diZerega GS, Hodgen GD. The primate ovarian cycle: suppression
2000;141:3814–20. of human menopausal gonadotropin-induced follicular growth
28. Shimasaki S, Moore RK, Otsuka F, Erickson GF. The bone morphogenetic in the presence of the dominant follicle. J Clin Endocrinol Metab.
protein system in mammalian reproduction. Endocr Rev. 2004;25:72– 1980;50:819–25. [Link]
101. [Link] 49. Grynnerup AG, Lǿssl K, Pilsgaard F, Bogstad JW, Prætorius L, Zedeler
29. Liu C, Peng J, Matzuk MM, Yao HH-C. Lineage specification of ovarian A, Lunding SA, Bungum L, Andersen AN, Pinborg A. Contribution of
theca cells requires multicellular interactions via oocyte and granulosa recruitable follicles to circulating anti-Müllerian hormone levels fol-
cells. Nat Commun. 2015;6:6934. [Link] lowing maximal gonadotrophin stimulation in patients with limited
30. Galloway SM, McNatty KP, Cambridge LM, et al. Mutations in an oocyte- ovarian reserve. Gynecol Endocrinol. 2020;36:273–6. [Link]
derived growth factor gene (BMP15) cause increased ovulation rate 1080/​09513​590.​2019.​16484​14.
and infertility in a dosage-sensitive manner. Nat Genet. 2000;25:279–83. 50. Adams GP, Matteri RL, Kastelic JP, Ko JCH, Ginther OJ. Association
31. Oktem O, Urman B. Understanding follicle growth in vivo. Hum Reprod. between surges of follicle-stimulating hormone and the emergence of
2010;25:2944–54. [Link] follicular waves in heifers. J Reprod Fert. 1992;94:177–88.
32. Klinger FG, De Felici M. In vitro development of growing oocytes from 51. Adams GP, Pierson RA. Bovine model for study of ovarian follicular
fetal mouse oocytes: stage-specific regulation by stem cell factor and dynamics in humans. Theriogenology. 1995;43:113–20.
granulosa cells. Dev Biol. 2002;244:85–95. [Link] 52. Seekallu SV, Toosi BM, Rawlings NC. LH pulse frequency and the emer-
2002.​0592. gence and growth of ovarian antral follicular waves in the ewe during
33. Otsuka F, Shimasaki S. A negative feedback system between oocyte the luteal phase of the estrous cycle. Reprod Biol Endocrinol. 2009;7:78.
bone morphogenetic protein 15 and granulosa cell kit ligand: its role in [Link]
regulating granulosa cell mitosis. Proc Natl Acad Sci USA. 2002;99:8060– 53. Baby TE, Bartlewski PM. Progesterone as the driving regulatory force
5. [Link] behind serum FSH concentrations and antral follicular development in
34. Joyce IM, Clark AT, Pendola FL, Eppig JJ. Comparison of recombinant cycling ewes. Reprod Fertil Development. 2011;23:303–10.
growth differentiation factor-9 and oocyte regulation of kit ligand 54. Baerwald AR, Adams GP, Pierson RA. A new model for ovarian fol-
messenger ribonucleic acid expression in mouse ovarian follicles. Biol licular development during the human menstrual cycle. Fertil Steril.
Reprod. 2000;63:1669–75. 2003;80:116–22. [Link]
35. Liu X, Andoh K, Abe Y, Kobayashi J, Yamada K, Mizunuma H, Ibuki Y. A 55. Baerwald AR, Adams GP, Pierson RA. Characterization of ovarian fol-
comparative study on transforming growth factor-β and activin A for licular wave dynamics in women. Biol Reprod. 2003;69:1023–31. [Link]
preantral follicles from adult, immature, and diethylstilbestrol-primed doi.​org/​10.​1095/​biolr​eprod.​103.​017772.
immature mice. Endocrinology. 1999;140:2480–5. 56. Ata B, La Marca A, Polyzos NP. Free your patients and yourself from
36. Mizunuma H, Liu X, Andoh K, Abe Y, Kobayashi J, Yamada K, Yokota H, day 2–3: start ovarian stimulation any time in freeze-all cycles. Reprod
Ibuki Y, Hasegawa Y. Activin from secondary follicles causes small pre- Biomed Online. 2023;47:103305. [Link]
antral follicles to remain dormant at the resting stage. Endocrinology. 103305.
1999;140:37–42. 57. Vaiarelli A, Climadomo D, Petriglia C, Conforti A, Alviggi C, Ubaldi N,
37. Juengel JL, McNatty KP. The role of proteins of the transforming growth Ledda S, Ferrero S, Rienzi L, Ubaldi FM. DuoSti - a reproducible strategy
factor-β superfamily in the intraovarian regulation of follicular develop- to obtain more oocytes and competent embryos in a short time-frame
ment. Hum Reprod Update. 2005;11:144–61. [Link] aimed at fertility preservation and IVF purposes. A systematic review.
humupd/​dmh061.
Jinno Reproductive Biology and Endocrinology (2025) 23:35 Page 20 of 22

UPSALA J Med Sci. 2020;125:121–30. [Link] 77. Aleyasin A, Abediasl Z, Nazari A, Sheikh M. Granulocyte colony-stimu-
2020.​17346​94. lating factor in repeated IVF failure, a randomized trial. Reproduction.
58. Van Disseldorp J, Lambalk CB, Kwee J, Looman CWN, Eijkemans MJC, 2016;151:637–42.
Fauser BC, Broekmans FJ. Comparison of inter- and intra-cycle variability 78. Kamath MS, Chittawar PB, Kirubakaran R, Mascarenhas M. Use of granu-
of anti-Müllerian hormone and antral follicle counts. Hum Reprod. locyte-colony stimulating factor in assisted reproductive technology: A
2010;25:221–7. [Link] systematic review and meta-analysis. Eur J Obstet Gynecol Reprod Biol.
59. Goodman AL, Hodgen GS. The ovarian triad of the primate menstrual 2017;214:16–24. [Link]
cycle. Recent Prog Horm Res. 1983;39:1–73. 79. Zhang L, Xu W, Fu X, Huang Q, Guo X, Zhang L, et al. Therapeutic role
60. Fauser BC, Van Heusden AM. Manipulation of human ovarian func- of granulocyte colony-stimulating factor (G-CSF) for infertile women
tion: physiological concepts and clinical consequences. Endocr Rev. under in vitro fertilization and embryo transfer (IVF-ET) treatment: a
1997;18:71–106. meta-analysis. Arch Gynecol Obstet. 2018;298:861–71. [Link]
61. Orvieto R. Pretreatment: Does it improve quantity or quality? Fertil 10.​1007/​s00404-​018-​4892-4.
Steril. 2022;117:657–63. [Link] 80. Kamath MS, Kirubakaran R, Sunkara SK. Granulocyte-colony stimulat-
62. Würfel W, Santjohanser C, Hirv K, Bühl M, Meri O, Laubert I, et al. High ing factor administration for subfertile women undergoing assisted
pregnancy rates with administration of granulocyte colony-stimulating reproduction. Cochrane Database Syst Rev. 2020;1:CD013226. [Link]
factor in ART-patients with repetitive implantation failure and lacking doi.​org/​10.​1002/​14651​858.​CD013​226.​pub2.
killer-cell immunoglobulin-like receptors. Hum Reprod. 2010;25:2151–3. 81. Fu L, Xu Y, Yan J, Zhang X, Li D, Zheng L. Efficacy of granulocyte colony-
63. Pala HG, Pala EE, Ulkumen BA, Aktug H, Yavasoglu A, Korkmaz HA, et al. stimulating factor for infertility undergoing IVF: a systematic review and
The protective effect of granulocyte colony-stimulating factor on endo- meta-analysis. Reprod Biol Endocrinol. 2023;21:34. [Link]
metrium and ovary in a rat model of diabetes mellitus. Gynecol Obstet 1186/​s12958-​023-​01063-z.
Invest. 2014;78:94–100. [Link] 82. Scarpellini F, Sbracia M. Use of granulocyte colony-stimulating factor
64. Akdemir A, Zeybek B, Akman L, Ergenoglu AM, Yeniel AO, Erbas O, et al. for the treatment of unexplained recurrent miscarriage: a randomised
Granulocyte-colony stimulating factor decreases the extent of ovarian controlled trial. Hum Reprod. 2009;24:2703–8.
damage caused by cisplatin in an experimental rat model. J Gynecol 83. Hiby SE, Regan L, Lo W, Farrell L, Carrington M, Moffett A. Association of
Oncol. 2014;25:328–33. [Link] maternal killer-cell immunoglobulin-like receptors and parental HLA-C
65. Kojima H, Otani A, Oishi A, Makiyama Y, Nakagawa S, Yoshimura N. Gran- genotypes with recurrent miscarriage. Hum Reprod. 2008;23:972–6.
ulocyte colony-stimulating factor attenuates oxidative stress-induced [Link]
apoptosis in vascular endothelial cells and exhibits functional and 84. Xie Y, Zhang T, Tian Z, Zhang J, Wang W, Zhang H, et al. Efficacy of
morphologic protective effect in oxygen-induced retinopathy. Blood. intrauterine perfusion of granulocyte colony-stimulating factor (G-CSF)
2011;117:1091–100. [Link] for infertile women with thin endometrium: a systematic review and
66. Harada M, Qin Y, Takano H, Minamino T, Zou Y, Toko H, et al. G-CSF pre- meta-analysis. Am J Reprod Immunol. 2017;78:e12701. [Link]
vents cardiac remodeling after myocardial infarction by activating the 10.​1111/​aji.​12701.
Jak-Stat pathway in cardiomyocytes. Nat Med. 2005;11:305–11. [Link] 85. Kato S, Yoshida S, Morikawa M, Miyamoto A, Habara Y, Oki M, et al. Three
doi.​org/​10.​1038/​nm1199. women with low AMH conceived by conventional infertility treatments
67. Patel AMR, Apaijai N, Chattipakorn N, Chattipakorn SC. The protective following G-CSF priming. J Jpn Soc Reprod Med. 2024;69:416 (in
and reparative role of colony-stimulating factors in the brain with cer- Japanese).
ebral ischemia/reperfusion injury. Neuroendocrinology. 2021;111:1029– 86. Kelly PA, Djiane J, Postel-Vinay M, Edery M. The prolactin/growth hor-
65. [Link] mone receptor family. Endocr Rev. 1991;12:235–51.
68. Michailov Y, AbuMadighem A, Lunenfeld E, Kapelushnik J, Huleihel M. 87. Bole-Feysot C, Goffin V, Edery M, Binart N, Kelly P. Prolactin (PRL) and
Granulocyte colony-stimulating factor restored impaired spermatogen- its receptor: actions, signal transduction pathways and phenotypes
esis and fertility in an AML-chemotherapy mice model. Int J Mol Sci. observed in PRL receptor knockout mice. Endocr Rev. 1998;19:225–68.
2021;22:11157. [Link] 88. Ben-Jonathan N, Mershon JL, Allen DL, Steinmetz RW. Extrapituitary
69. Lee ST, Chu K, Jung KH, Ko SY, Kim EH, Shinn DI, et al. Granulocyte prolactin: distribution, regulation, functions, and clinical aspects. Endocr
colony-stimulating factor enhances angiogenesis after focal cerebral Rev. 1996;17:639–69.
ischemia. Brain Res. 2005;1058:120–8. 89. Nagy E, Berczi I. Hypophysectomized rats depend on residual prolactin
70. Würfel W. Treatment with granulocyte colony-stimulating factor in for survival. Endocrinology. 1991;128:2776–84.
patients with repetitive implantation failures and/or recurrent sponta- 90. McNeilly AS. Prolactin and ovarian function. In: Muller EE, MacLeod
neous abortions. J Reprod Immunol. 2015;108:123–35. [Link] RM, eds. Neuroendocrine perspectives. Amsterdam: Elsevier,
10.​1016/j.​jri.​2015.​01.​010. 1984;3:279–316.
71. Hong L, Yan L, Xin Z, Hao J, Liu W, Wang S, et al. Protective effects of 91. Kauppila A, Chatelain P, Kirkinen P, Kivinen S, Ruokonen A. Isolated
human umbilical cord mesenchymal stem cell-derived conditioned prolactin deficiency in a woman with puerperal alactogenesis. J Clin
medium on ovarian damage. J Mol Cell Biol. 2020;12:372–85. [Link] Endocrinol Metab. 1987;64:309–12.
doi.​org/​10.​1093/​jmcb/​mjz105. 92. Laufer N, Botero-Ruiz W, DeCherney AH, Haseltine F, Polan ML, Behrman
72. Lédée N, Gridelet V, Ravet S, Jouan C, Gaspard O, Wenders F, et al. HR. Gonadotropin and prolactin levels in follicular fluid of human ova
Impact of follicular G-CSF quantification on subsequent embryo trans- successfully fertilized in vitro. J Clin Endocrinol Metab. 1984;58:430–4.
fer decisions: a proof of concept study. Hum Reprod. 2013;28:406–13. 93. Gonen Y, Casper RF. The influence of transient hyperprolactinemia on
[Link] hormonal parameters, oocyte recovery, and fertilization rates in in vitro
73. de Kruijf EFM, Fibbe WE, van Pel M. Cytokine-induced hematopoietic fertilization. J In Vitro Fertil Embryo Transf. 1989;6:155–9.
stem and progenitor cell mobilization: unraveling interactions between 94. Oda T, Yoshimura Y, Takehara Y, Kohriyama S, Sano Y, Tanabe K, et al.
stem cells and their niche. Ann N Y Acad Sci. 2020;1466:24–38. [Link] Effects of prolactin on fertilization and cleavage of human oocytes.
doi.​org/​10.​1111/​nyas.​14059. Horm Res. 1991;35:33–8.
74. Herraiz S, Romeu M, Buigues A, Martinez S, Diaz-Garcia C, Gómez- 95. Evans WS, Cronin MJ, Thorner MO. Hypogonadism in hyperprolactine-
Segui I, et al. Autologous stem cell ovarian transplantation to increase mia and proposed mechanisms. In: Ganong WF, Martini L, eds. Frontiers
reproductive potential in patients who are poor responders. Fertil Steril. in neuroendocrinology. New York; Raven Press, 1982;7:77–122.
2018;110:496–505. [Link] 96. Jinno M, Yoshimura Y, Ubukata Y, Nakamura Y. A novel method of ovar-
75. Buigues A, Marchante M, de Miguel-Gómez L, Martinez J, Cervelló I, Pel- ian stimulation for in vitro fertilization: bromocriptine-rebound method.
licer A, et al. Stem cell-secreted factor therapy regenerates the ovarian Fertil Steril. 1996;66:271–4.
niche and rescues follicles. Am J Obstet Gynecol. 2021;225(65):e1-14. 97. Jinno M, Katsumata Y, Hoshiai T, Nakamura Y, Matsumoto K, Yoshimura Y.
[Link] A therapeutic role of prolactin supplementation in ovarian stimula-
76. Würfel W. Approaches to better implantation. J Ass Reprod Genet. tion for in vitro fertilization: the bromocriptine-rebound method. J Clin
2000;17:473. Endocrinol Metab. 1997;82:3603–11.
Jinno Reproductive Biology and Endocrinology (2025) 23:35 Page 21 of 22

98. Maslar IA, Riddick DH. Prolactin production by human endome- 122. Diamanti-Kandarakis E, Piperi C, Kalofoutis A, Creatsas G. Increased
trium during the normal menstrual cycle. Am J Obstet Gynecol. levels of serum advanced glycation end-products in women with
1979;135:751–4. polycystic ovary syndrome. Clin Endocrinol. 2005;62:37–43.
99. Heffner LJ, Iddenden DA, Lyttle CR. Electrophoretic analyses of secreted 123. Jinno M, Takeuchi M, Watanabe A, Teruya K, Hirohama J, Eguchi N,
human endometrial proteins: identification and characterization of Miyazaki A. Advanced glycation end-products accumulation compro-
luteal phase prolactin. J Clin Endocrinol Metab. 1986;62:1288–95. mises embryonic development and achievement of pregnancy by
100. Daly DC, Maslar IA, Rosenberg SM, Tohan N, Riddick DH. Prolactin assisted reproductive technology. Hum Reprod. 2011;26:604–10.
production by luteal phase defect endometrium. Am J Obstet Gynecol. 124. Merhi Z. Advanced glycation end products and their relevance in
1981;140:587–91. female reproduction. Hum Reprod. 2014;29:135–45.
101. Maslar IA, Kaplan BM, Luciano AA, Riddick DH. Prolactin production by 125. Zhu J, Cai Y, Long S, Chen Z, Mo Z. The role of advanced glycation end
the endometrium of early human pregnancy. J Clin Endocrinol Metab. products in human infertility. Life Sci. 2020;255:117830. [Link]
1980;51:78–83. 10.​1016/j.​lfs.​2020.​117830.
102. Ying Y, Randolph JF, Maier DB, Kuslis ST, Chapitis J, Riddick DH. Uterine 126. Jinno M, Nagai R, Takeuchi M, Watanabe A, Teruya K, Sugawa H,
fluid and prolactin secretion in the ovulating cynomolgus monkey. Am Hatakeyama N, Jinno Y. Trapa bispinosa Roxb. extract lowers advanced
J Obstet Gynecol. 1986;155:677–80. glycation end-products and increases live births in older patients
103. Moride N, Nakagawa K, Tsutsumi Y, Yamashita M, Matsumoto M, with assisted reproductive technology: a randomized controlled
Komatsu J, Yamano S, Aono T. Does the bromocriptine-rebound trial. Reprod Biol Endocrinol. 2021;19:149. [Link]
method improve embryo quality? Jpn J Fertil Steril. 2000;45:541 (in s12958-​021-​00832-y.
Japanese). 127. Adkar P, Dongare A, Ambavada S, Bhaskar VH. Trapa bispinosa Roxb.: a
104. Moride N, Nakagawa K, Tsutsumi Y, Yamashita M, Matsumoto M, review on nutritional and pharmacological aspects. Adv Pharmacol Sci.
Komatsu J, Yamano S, Aono T. Does the bromocriptine-rebound 2014;2014:959830. [Link]
method improve embryo quality? Acta Obst Gynaec Jpn 2001;53:417(S- 128. Takeshita S, Yagi M, Uemura T, Yamada M, Yonei Y. Peel extract of
343). (in Japanese) water chestnut (Trapa bispinosa Roxb.) inhibits glycation, degrades
105. Nakamura M, Kawauchi H, Fujita K, Takei E, Motohashi E, Ishikawa M, α-dicarbonyl compound, and breaks advanced glycation end product
Nishijima M. Usefulness of the bromocriptine-rebound method in IVF crosslinks. Glycative Stress Res. 2015;2:72–9.
treatment. Annual Meeting of Japan Fertilization Implantation Society; 129. JSOG-ART. 2018 ART data book, 2018. [Link]
2002 Oct 5; Gifu City, Japan. art/​2018d​ata_​20201​001.​pdf.
106. Thomas MC, Baynes JW, Thorpe SR, Cooper ME. The role of AGEs and 130. Yasuda M, Yasutake K, Hino M, Ohwatari H, Ohmagari N, Takedomi
AGE inhibitors in diabetic cardiovascular disease. Curr Drug Targets. K, Tanaka T, Nonaka G. Inhibitory effects of polyphenols from water
2005;6:453–74. chestnut (Trapa japonica) husk on glycolytic enzymes and postprandial
107. Unoki H, Yamagishi S. Advanced glycation end products and insulin blood glucose elevation in mice. Food Chem. 2014;165:42–9.
resistance. Curr Pharm Design. 2008;14:987–9. 131. Yamaguchi H, Nagai M, Sugawa H, Yasuda H, Nagai R. Development of
108. Ulrich P, Cerami A. Protein glycation, diabetes, and aging. Recent Prog a conventional immunochemical detection system for determination
Horm Res. 2001;56:1–21. of ­Nδ-(5-hydro-5-methyl-4-imidazolone-2-yl)-ornithine in methylg-
109. Ferrannini E, Buzzigoli G, Bonadonna R, et al. Insulin resistance in essen- lyoxal-modified proteins. Glycoconj J. 2020. [Link]
tial hypertension. N Engl J Med. 1987;317:350–7. s10719-​020-​09957-5.
110. Reaven GM. Role of insulin resistance in human disease. Diabetes. 132. Kinoshita S, Mera K, Ichikawa H, Shimasaki S, Nagai M, Taga Y, Iijima K,
1988;37:1595–607. Hattori S, Fujiwara Y, Shirakawa J, Nagai R. Nω-(carboxymethyl)arginine
111. Patrick L. Nonalcoholic fatty liver disease: relationship to insulin is one of the dominant advanced glycation end products in glycated
sensitivity and oxidative stress. Treatment approaches using vitamin E, collagens and mouse tissues. Oxid Med Cell Longev. 2019;14:9073451.
magnesium, and betaine. Altern Med Rev 2002;7:276–91. [Link]
112. Terman A, Brunk UT. Aging as a catabolic malfunction. Int J Biochem 133. Miyazaki T, Kuji N, Sueoka K, Natori M, Kobayashi T, Nozawa S. Effect of
Cell Biol. 2004;36:2365–75. administration of danazol on the pregnancy rate of initially unsuc-
113. Takeuchi M. Toxic AGEs (TAGE) theory: a new concept for preventing cessful in vitro fertilization-embryo transfer patients. Jpn J Fertil Sterl.
the development of diseases related to lifestyle. Diabetol Metab Syndr. 1995;40:380–5 (Japanese).
2020;12:105. [Link] 134. Tei C, Miyazaki T, Kuji N, Tanaka M, Sueoka K, Yoshimura Y. Effect of
114. Ruiz HH, Ramasamy R, Schmidt AM. Advanced glycation end products: danazol on the pregnancy rate in patients with unsuccessful in vitro
building on the concept of the “common soil” in metabolic disease. fertilization-embryo transfer. J Reprod Med. 1998;43:541–6.
Endocrinology. 2020;161:bqz006. [Link] 135. Vieri M, Kirschner M, Tometten M, Abels A, Rolles B, Isfort S, et al. Com-
bqz006. parable effects of the androgen derivatives danazol, oxymetholone
115. Paolisso G, Tagliamonte MR, Rizzo MR, Giugliano D. Advancing age and and nandrolone on telomerase activity in human primary hemat-
insulin resistance: new facts about an ancient history. Eur J Clin Invest. opoietic cells from patients with dyskeratosis congenita. Int J Mol Sci.
1999;29:758–69. 2020;21:7196. [Link]
116. Vanltallie TB. Stress: a risk factor for serious illness. Metabolism. 136. Townsley DM, Dumitriu B, Liu D, Biancotto A, Weinstein B, Chen C, et al.
2002;51:40–5. Danazol treatment for telomere diseases. N Engl J Med. 2016;374:1922–
117. Wolkowitz OM, Epel ES, Reus VI. Stress hormone-related psychopa- 31. [Link]
thology: pathophysiological and treatment implications. World J Biol 137. Córdova-Oriz I, Kohls G, Iglesias C, Polonio AM, Chico-Sordo L, Toribio
Psychiatry. 2001;2:115–43. M, et al. A randomized controlled intervention trial with danazol to
118. Bjorntorp P, Rosmond R. The metabolic syndrome - a neuroendocrine improve telomeric and fertility parameters in women with diminished
disorder? Br J Nutr. 2000;83:S49–57. ovarian reserve: a pilot study. Women’s Health Rep. 2023;4(1):305–18.
119. Rosenthal M, Haskell WL, Solomon R, Widstrom A, Reaven GM. Dem- [Link]
onstration of a relationship between levels of physical training and 138. Hughes E, Brown J, Tiffin G. Danazol for unexplained subfertility.
insulin-stimulated glucose utilisation in normal humans. Diabetes. Cochrane Database Sys Rev. 2007;1:CD000069. [Link]
1983;32:408–11. 14651​858.​CD000​069.​pub2.
120. Dunaif A. Insulin resistance and the polycystic ovary syndrome: mecha- 139. Sen A, Hammes SR. Granulosa cell-specific androgen receptors are criti-
nism and implications for pathogenesis. Endocr Rev. 1997;18:774–800. cal regulators of ovarian development and function. Mol Endocrinol.
121. Jinno M, Kondou K, Teruya K. Low-dose metformin improves pregnancy 2010;24:1393–403. [Link]
rate in in vitro fertilization repeaters without polycystic ovary syndrome: 140. González-Comadran M, Durán M, Solá I, Fábregus F, Carreras R, Checa
prediction of effectiveness by multiple parameters related to insulin MA. Effects of transdermal testosterone in poor responders undergo-
resistance. Hormones (Athens). 2010;9:161–70. ing IVF: systematic review and meta-analysis. Reprod Biomed Online.
2012;25:450–9. [Link]
Jinno Reproductive Biology and Endocrinology (2025) 23:35 Page 22 of 22

141. Hoang QH, Ho HS, Do HT, Nguyen TV, Nguyen HP, Le MT. Therapeu-
tic effect of prolonged testosterone pretreatment in women with
poor ovarian response: a randomized control trial. Reprod Med Biol.
2021;20:305–12. [Link]
142. Bosdou JK, Venetis CA, Dafopoulos K, Zepiridis L, Chatzimeletiou K,
Anifandis G, et al. Transdermal testosterone pretreatment in poor
responders undergoing ICSI: a randomized clinical trial. Hum Reprod.
2016;31:977–85. [Link]
143. Subirá J, Algaba A, Vázquez S, Dasí RT, Robles GM, Fabuel SM, et al.
Testosterone does not improve ovarian response in Bologna poor
responders: a randomized controlled trial (TESTOPRIM). Reprod Biomed
Online. 2021;43:466–74. [Link]
144. Polyzos NP, Martinez F, Blockeel C, Gosalvez A, De la Fuente L, Pinborg
A, et al. Transdermal testosterone prior to ovarian stimulation for in vitro
fertilization in women with poor ovarian response. A multicenter
multinational double-blind placebo-controlled randomized trial
(The T-TRANSPORT). Hum Reprod. 2023;38:Suppl(1):i41. June 2023,
dead093.080. [Link]
145. Zhang M, Niu W, Wang Y, Xu J, Bao X, Wang L, et al. Dehydroepiandros-
terone treatment in women with poor ovarian response undergoing
IVF or ICSI: a systematic review and meta-analysis. J Assist Reprod
Genet. 2016;33:981–91. [Link]
146. Chern C, Tsui K, Vitale SG, Chen S, Wang P, Cianci A, et al. Dehydroepi-
androsterone (DHEA) supplementation improves in vitro fertilization
outcomes of poor ovarian responders, especially in women with low
serum concentration of DHEA-S: a retrospective cohort study. Reprod
Biol Endocrinol. 2018;16:90. [Link]
147. Wang Z, Yang A, Bao H, Wang A, Deng X, Xue D, et al. Effect of dehy-
droepiandrosterone administration before in vitro fertilization on the
live birth rate in poor ovarian responders according to the Bologna
criteria: a randomised controlled trial. BJOG. 2021;129:1030–8. [Link]
doi.​org/​10.​1111/​1471-​0528.​17045.
148. Neves AR, Montoya-Botero P, Polyzos NP. The role of androgen sup-
plementation in women with diminished ovarian reserve: time to rand-
omize, not meta-analyze. Front Endocrinol (Lausanne). 2021;12:653857.
[Link]
149. Atkinson L, Martin F, Sturmey RG. Intraovarian injection of platelet-rich
plasma in assisted reproduction: too much too soon? Hum Reprod.
2021;36:1737–50. [Link]
150. Hsueh AJW, Kawamura K. Hippo signaling disruption and ovarian fol-
licle activation in infertile patients. Fertil Steril. 2020;114:458–64. [Link]
doi.​org/​10.​1016/j.​fertn​stert.​2020.​07.​031.

Publisher’s Note
Springer Nature remains neutral with regard to jurisdictional claims in pub-
lished maps and institutional affiliations.