Received: 3 December 2019 | Revised: 17 July 2020 | Accepted: 22 August 2020

DOI: 10.1002/mrd.23420

REVIEW ARTICLE

Selection of competent oocytes by morphological criteria for

assisted reproductive technologies

Saffet Ozturk

Department of Histology and Embryology,

Akdeniz University School of Medicine, Abstract
Antalya, Turkey
Invasive and noninvasive methods are commonly used to select developmentally
Correspondence competent oocytes that can improve the take‐home baby rates in assisted re-
Saffet Ozturk, Department of Histology and productive technology (ART) centers. One of the noninvasive methods con-
Embryology, Akdeniz University School of
Medicine, Campus, 07070 Antalya, Turkey. ventionally utilized to determine competent oocytes is the morphological analysis of
Email: [email protected] cumulus complex, first polar body, zona pellucida, perivitelline space, meiotic spin-
dle, and ooplasm. Successful fertilization, early embryo development, uterine im-
plantation, and healthy pregnancy depend on the quality of oocytes, and
morphological evaluation is one of the options used to predict quality levels. In this
review, the morphological criteria being utilized in certain ART centers are com-
prehensively evaluated with special references to their predictive values and po-
tential contributions to selecting high‐quality oocytes.

KEYWORDS
assisted reproductive technology, embryo quality, oocyte morphology, oocyte selection

1 | INTRODUCTION (Junk & Yeap, 2012). Then, GV oocytes undergo nuclear and cyto-
plasmic maturation to produce MII oocytes during routine IVM
Infertility rate among couples gradually increases worldwide due to procedure (Hatirnaz et al., 2018). After obtaining MII oocytes from
delaying childbearing, which is the result of various social and eco- either conventional ARTs or IVM, the cumulus cells surrounding
nomic factors (Balasch & Gratacos, 2012). Thus, assisted re- these oocytes can be removed by gentle pipetting to readily ma-
productive technologies (ARTs) acquire more and more importance nipulate denuded oocytes during intracytoplasmic sperm injection
each day. For the purpose of obtaining metaphase II (MII) oocytes in (ICSI) application. The most difficult and valuable part before ap-
ART centers, recombinant follicle‐stimulating hormone (rFSH) or its plying these ART procedures such as ICSI and in vitro fertilization
analogs are routinely used to induce gonadotropin‐dependent folli- (IVF) is to select the best oocytes that enable the production of early
cular development (Schoot et al., 1992). Then, human chronic go- embryos with high quality. Thus, the birth of healthy offspring can be
nadotropin (hCG) is used to resume meiotic progression from increased.
prophase I (PI) to MII stages only after observing follicles greater As the success rate of ART applications remains at ∼35%
than 17 or 18 mm in diameter. Eventually, cumulus–oocyte com- (pregnancy rate per embryo transfer based on the ESHRE annual
plexes (COCs) including MII oocytes are collected from antral folli- report 2016), selection criteria for human oocytes as well as sperm
cles 36 h after injection of hCG. As is known, in vitro maturation cells and early embryos are crucially important. Selecting high‐
(IVM; Cha et al., 1991; Pincus & Enzmann, 1935) is an alternative quality oocytes based on morphological analysis may increase suc-
treatment to conventional ARTs to retrieve MII oocytes from poor cess rates of fertilization and subsequent embryo development
responders and patients with polycystic ovary syndrome (PCOS). To (Gilchrist, Lane, & Thompson, 2008). The main challenge related to
this end, immature germinal vesicle (GV; at the PI stage of first selecting better oocytes is that developmentally incompetent oo-
meiosis) oocytes are recovered from growing small follicles less than cytes may exhibit the same morphologies as the good ones. In ad-
14 mm in diameter following stimulated with rFSH or its analogs dition to morphological analysis, there are molecular biological

Mol Reprod Dev. 2020;87:1021–1036. wileyonlinelibrary.com/journal/mrd © 2020 Wiley Periodicals LLC | 1021
 10982795, 2020, 10, Downloaded from https://onlinelibrary.wiley.com/doi/10.1002/mrd.23420 by CochraneArgentina, Wiley Online Library on [25/03/2024]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
1022 | OZTURK

techniques such as proteomic and genomic applications requiring clots from CC could not rescue the defects observed in the early
samples obtainable only by invasive methods that might influence embryos (Ebner et al., 2008a), suggesting that blood clots may exe-
the developmental potential of early embryos and subsequent terms. cute undesirable effect(s) in the antral follicles. The adverse effects
The first and second polar bodies can also be analyzed by these of blood clots seem to originate from the production of reactive
molecular biological techniques; however, they do not precisely re- oxygen species (ROS) at high levels from existing blood components
flect the developmental potential of relevant oocytes (Cimadomo (Attaran et al., 2000). Taken together, the oocytes having blood clots
et al., 2016; Geraedts et al., 2010). In recent years, as single‐embryo in CC might not be used for ART applications at least in the first
transfer has become an extensively accepted way in many ART attempt if there are more oocytes appearing normal.
centers to decrease the rate of multiple pregnancies that may lead to A few studies also examined whether the expansion status of
medical and perinatal complications, selection of competent oocytes cumulus layers influences the oocytes' quality, early embryos de-
as well as early embryos is also vital for preventing undesirable velopment, and pregnancy rates (Table 1). To this end,
outcomes. It is worth noting that since there are legal, social, and/or Rattanachaiyanont, Leader, and Leveille (1999) evaluated the ex-
religious restrictions for producing supernumerary embryos in some pansion grades of the corona–cumulus layers in COCs using an in-
countries, oocytes selection can further increase the general effi- verted microscope. These researchers revealed that no correlation
ciency of ARTs in these countries. Moreover, in women of advancing exists between corona–cumulus expansion grades and the para-
ages, oocytes quality gradually declines due to deteriorating phy- meters such as nuclear maturity, fertilization, embryonic cleavage,
siological microenvironment around oocytes; therefore, good‐quality and pregnancy rates (Rattanachaiyanont et al., 1999). Likewise,
oocytes should be selected to increase the chance of conceiving in Ebner et al. (2008) observed that the expansion of corona radiata
these women having a limited reproductive window. As a result, and cumulus matrix of COCs could not allow for the prediction of
morphological analysis for selecting competent oocytes can be uti- nuclear maturity and quality of the oocytes undergoing ICSI.
lized in ART applications in spite of its limited contributions. In addition to cumulus expansion, the apoptotic index of cumulus
cells was analyzed to predict the nuclear maturity of oocytes. The
rate of apoptosis in the cumulus cells of immature oocytes at GV or
2 | M O R PH O L O G I C AL E VA L U A T I O N O N metaphase I (MI) stage is at higher levels than in the cumulus cells
H U M A N O O CY TE S from mature oocytes (Host, Gabrielsen, Lindenberg, & Smidt‐Jensen,
2002). Moreover, a higher rate of apoptosis in the cumulus cells of
It is generally accepted that morphological evaluation of oocytes can mature oocytes correlates with the lower fertilization rate in com-
be performed without damaging their initial developmental potential parison with the control group having a lower apoptosis rate (Host
and subsequent embryonic development. Also, this evaluation is et al., 2002).
cost‐effective, noninvasive, compatible with the workflow of ART, Consistently, Faramarzi, Khalili, and Omidi (2019) revealed that
and may contribute to reducing the generation of surplus fertilized there is a close relationship between apoptosis gene expression in-
oocytes and early embryos. A great number of morphological char- cluding BAX, Caspase3, and BCL2 in the cumulus cells and embryo
acteristics of cumulus complex (CC), zona pellucida (ZP), perivitelline morphokinetics (Table 1). The BAX and Caspase3 messenger RNAs
space (PS), shape and volume of oocytes, first polar body (PB), oo- expression were at higher levels in the cumulus cells of ovulated
plasm, meiotic spindle, and multiple properties can be assessed to oocytes that produced embryos with uneven blastomeres than in the
select the oocytes with high quality in certain ART centers (Lasiene, cumulus cells of oocytes that produced embryos without uneven
Vitkus, Valanciute, & Lasys, 2009; Figure 1). blastomeres. Also, there were predominantly higher BAX, Caspase3,
and BAX/BCL2 expression, but a lower BCL2 expression, in the cu-
mulus cells of oocytes that generated embryos with either cell fusion
2.1 | Cumulus complex or trichotomous mitosis when compared with the cumulus cells of
oocytes that generated embryos not having cell fusion or trichoto-
As MII oocytes and surrounding cumulus cells communicate with mous mitosis. These studies suggest that oocytes showing better
each other by using gap junctions for exchanging glucose metabolites developmental potential are the ones whose cumulus cells have
and ions, or with paracrine signaling (Tanghe, Van Soom, Nauwynck, apoptosis rates at lower levels, and lower expression of the
Coryn, & de Kruif, 2002), the morphological features of CC may help apoptosis‐related genes (like BAX and Caspase3) independent of cu-
to determine the oocyte quality. It was noted that the presence of mulus expansion.
blood clots in CC may adversely affect the rates of fertilization and In a different work, Wolf (1988) graded COCs based on the
cleavage of early embryos derived from corresponding oocytes number of corona radiata layers and the expansion state of cumulus
(Daya, Kohut, Gunby, & Younglai, 1990), and decreases these oo- cells. Grade 1 is characterized by absent to sparse cumulus cells and
cytes quality and blastocyst formation rate (Ebner et al., 2008a; 1–3 layers of corona cells; Grade 2 exhibits dense cumulus cells and
Table 1). Blood clots also lead to decreasing the number of harvested tightly packed corona cells; Grade 3 has expanded, fluffy cumulus cells
oocytes when compared with the oocytes number whose CC exhibits and expanded corona cells; Grade 4 is defined as expanded, scanty
normal appearance (Ebner et al., 2008a). Intriguingly, removing blood cumulus cells and partially lost corona cells. Ng, Chang, and Wu (1999)
 10982795, 2020, 10, Downloaded from https://onlinelibrary.wiley.com/doi/10.1002/mrd.23420 by CochraneArgentina, Wiley Online Library on [25/03/2024]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
OZTURK | 1023

FIGURE 1 Morphological properties used to assess oocyte competency. SER, smooth endoplasmic reticulum

reported that the oocytes from grade 3 COCs showed significantly expansion affects the quality of oocytes and which molecular me-
higher fertilization rates than the Grades 1, 2, and 4. Although there chanisms are involved should be evaluated in future studies.
were no differences related to cleavage and polyspermy rates, the As there is bidirectional signaling and exchange of small mole-
pregnancy success rates displayed a higher positive correlation with cules between cumulus cells and oocytes, transcriptome analysis of
the Grade 3 COCs when compared with other grades (Ng et al., 1999). cumulus cells may help to predict the competence of oocytes and
This higher success rate was interpreted as resulting from better cy- subsequent early embryos development (Table 1). For this purpose, a
toplasmic maturation of the oocytes producing better fertilization and more detailed molecular biological analysis of cumulus cells was
pregnancy rates (Ng et al., 1999). On the other hand, Dal Canto et al., performed to identify the oocytes with high quality. The expression
(2012) documented that morphological evaluation of COCs follows levels of cyclooxygenase 2 (COX2, also known as PTGS2), hyaluronic
expanded (EC) and compact cumulus (CC) masses both of which in- acid synthase 2 (HAS2), and gremlin 1 (GREM1) genes were sig-
volve immature oocytes and expanded cumulus mass enclosing ma- nificantly higher in the cumulus cells of oocytes that generated the
ture oocytes (EC–MII) at the time of collection. These COCs were early embryos with high quality (Grades 3, 4, and 5) when compared
analyzed to predict the competence of oocytes. Although there was no with the low‐quality embryos (Grades 1 and 2; McKenzie et al.,
difference in the fertilization rates among groups, both implantation 2004). Moreover, HAS2 and GREM1 were expressed at higher levels
and pregnancy rates increased progressively and significantly from EC in the cumulus cells obtained from oocytes that produced high‐
to EC–MII groups (Dal Canto et al., 2012; Table 1). As a result, there is quality embryos on Day 3 than in the cumulus cells from oocytes that
a positive correlation between cumulus expansion and implantation were not fertilized or produced poor‐quality embryos (Cillo et al.,
and pregnancy rates of the relative oocytes. How cumulus cell 2007). However, there was no significant difference detected for the
 10982795, 2020, 10, Downloaded from https://onlinelibrary.wiley.com/doi/10.1002/mrd.23420 by CochraneArgentina, Wiley Online Library on [25/03/2024]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
1024 | OZTURK

TABLE 1 The potential outcomes of morphological and molecular changes in the cumulus complex of human oocytes

Cumulus complex Outcome

• Presence of blood clot ↓ Harvested oocyte number (Ebner et al., 2008) ↓ oocyte quality (Ebner et al., 2008) ↓
fertilization rate (Daya et al., 1990) ↓ cleavage rate (Daya et al., 1990) ↓ blastocyst
formation rate (Ebner et al., 2008)

• Higher apoptosis rate ↑ Immaturity (Host et al., 2002) ↓ fertilization (Host et al., 2002)

• Expanded, fluffy cumulus cells and expanded corona ↑ Fertilization rate (Ng et al., 1999) ↑ pregnancy rate (Ng et al., 1999)
cells

• Expanded cumulus mass ↑ Implantation rate (Dal Canto et al., 2012) ↑ pregnancy rate (Dal Canto et al., 2012)

• Higher expression of COX2, HAS2, and GREM1 genes ↑ Embryo quality (McKenzie et al., 2004)

• Higher expression HAS2 and GREM1 genes ↑ Day 3 embryo quality (Cillo et al., 2007) ↑ fertilization rate (Cillo et al., 2007)

• Higher expression of BCL2L11 and PCK1 genes, and ↑ Embryo potential (Assou et al., 2008) ↑ successful pregnancy rate (Assou
lower NFIB expression et al., 2008)

• Higher PTGS2 and lower BDNF genes expression ↑ Normal fertilization (Anderson et al., 2009)

• Higher expression of DPP8, HIST1H4C, UBQLN1, CALM1, ↑ Pregnancy rate (Assidi et al., 2011) ↑ implantation rate (Assidi et al., 2011)
NRP1, and PSMD6 genes

• Downregulation of SPSB2 and TP53I3 genes expression ↑ Chromosomally abnormal oocytes (Fragouli et al., 2012)

• Relative ranking levels of HAS2, FSHR, VCAN, and PR ↑ Oocyte quality (Ekart, McNatty, Hutton, & Pitman, 2013) ↑ blastocyst formation
genes rate (Ekart et al., 2013) ↑ live birth rate (Ekart et al., 2013)

• Higher expression of lncRNA and PSMD6 genes ↑ Pregnancy rate (Li et al., 2015)

• Lower expression of AMHR2 and LIF genes ↑ Embryo quality (Devjak et al., 2016)

• Higher expression of BAX and Caspase3 ↑ Embryos with uneven blastomere, cell fusion, and trichotomous mitoses (Faramarzi
et al., 2019)

• Higher expression of CAMK1D ↑ Blastocyst formation (Scarica et al., 2019)

Note: ↑, increased; ↓, decreased.

pentraxin 3 (PTX3) expression in the same groups (Cillo et al., 2007). The expression of a number of genes in the cumulus cells before
The expression levels of BCL2L11 (BCL‐like protein 11, at the higher ICSI was explored by using microarray technology. The six genes
level), PCK1 (phosphoenolpyruvate carboxykinase 1, at the higher whose expression in the cumulus cells was predominantly decreased
level), and NFIB (nuclear factor IB, at the lower level) genes in the in unsuccessful pregnancies may be related to a successful pregnancy
cumulus cells contributed to the prediction of the early embryos with and implantation rates (Assidi, Montag, Van der Ven, & Sirard, 2011).
higher potential and successful pregnancy outcomes (Assou et al., These genes were dipeptidyl peptidase 8 (DPP8; playing a role in
2008). Furthermore, PTGS2 expression in the cumulus cells was at protein folding and/or apoptosis); histone cluster 1, H4c (HIST1H4C;
higher levels in the mature oocytes, and the brain‐derived neuro- implicating in the regulation of gene expression and cell differ-
trophic factor (BDNF) expression was at lower levels in the cumulus entiation); ubiquilin 1 (UBQLN1; functioning in intercellular signaling
cells whose oocytes underwent normal fertilization (Anderson et al., and cell cycle, and also in ubiquitination machinery); calmodulin 1
2009). And, the GREM1 expression level exhibited a weak positive (CALM1; involving in calcium‐dependent pathways); neuropilin 1
correlation with embryo quality, but BDNF expression level was (NRP1; playing roles in cell assembly and interactions); and protea-
found to be the negative predictor of embryo quality (Anderson some 26S subunit, non‐ATPase, 6 (PSMD6; involved in protein folding
et al., 2009). Consequently, the oocytes whose cumulus cells have and/or apoptosis; Assidi et al., 2011). In addition, Fragouli, Wells,
higher GREM1 expression should be transferred or cryopreserved. Iager, Kayisli, and Patrizio (2012) reported that downregulated ex-
Also, the detectable levels of soluble human leukocyte antigen‐G pression of two genes, splA/ryanodine receptor domain and sup-
(sHLA‐G) molecule ranging from 300 to 800 pg/ml in the medium of pressor of cytokine signaling (SOCS) box containing 2 (SPSB2) and
in vitro matured COCs can also be used as a biomarker for predicting tumor protein p53 inducible protein 3 (TP53I3), in the cumulus cells
oocyte maturity (Rizzo et al., 2009). Overall, higher or lower ex- were significantly associated with chromosomal abnormalities in the
pression profiles of the gene(s) mentioned above could be employed oocytes (Fragouli et al., 2012; Table 1). While SPSB2 implicates in
as a biomarker for determining maturity, fertilization success, em- intracellular signaling and homeostasis, TP53I3 is involved in car-
bryo quality, or pregnancy outcome of the oocytes. bohydrate metabolism and apoptosis (Fragouli et al., 2012).
 10982795, 2020, 10, Downloaded from https://onlinelibrary.wiley.com/doi/10.1002/mrd.23420 by CochraneArgentina, Wiley Online Library on [25/03/2024]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
OZTURK | 1025

A similarly designed study by Ekart et al. (2013) indicated that thickness and embryo quality scores (Host et al., 2002). It is worth
the relative ranking levels of candidate genes, including HAS2, noting that for these analyses polarized microscopes (PolScope) were
follicle‐stimulating hormone receptor (FSHR), versican (VCAN), and commonly utilized to visualize oocyte structural components at
progesterone receptor (PR) in the cumulus cells from young women higher magnification by using polarized light. The studies using
(<38 years old), contributed to selecting the oocytes with higher PolScope revealed that the thickness of the inner layer of ZP seems
quality, resulted in higher blastocyst formation and live birth rates to associate with oocytes' developmental potential (Shen, Stalf,
when compared with the cumulus cells of randomly selected oocytes. Mehnert, Eichenlaub‐Ritter, & Tinneberg, 2005). Analogously, the
The expression of both long noncoding RNAs (lncRNAs, also known oocytes with an inner layer of ZP at 10–12 nm thickness measured
as AK124742) and PSMD6 (natural antisense of AK124742) was by using PolScope exhibited better development than the oocytes
significantly higher in the cumulus cells of a pregnancy group in having a thickness of 8–10 nm or less than 8 nm (Rama Raju, Prakash,
comparison with the cumulus cells from a nonpregnancy group (Li Krishna, & Madan, 2007; Table 2). These studies propose that oo-
et al., 2015). The high or low expression levels of anti‐Mullerian cytes whose ZP or its inner layer thickness is thinner than others
hormone receptor type 2 (AMHR2) and leukemia inhibitory factor may be initially used.
(LIF) genes in the cumulus cells may facilitate the prediction of high The ZP birefringence, a refractive index obtained from polar-
or low‐quality embryo (Devjak et al., 2016; Table 1). Although a ization and propagation direction of light, can be analyzed to predict
limited number of cumulus cells from three women were analyzed by the developmental potential of oocytes and early embryos (Table 2).
using microarray technology, numerous differentially expressed High birefringence of the inner layer of ZP in the oocytes resulted in
genes involved in sugar metabolism, ribosome biogenesis, and the higher implantation, pregnancy, and live birth rates than that of ones
JAK‐STAT signaling pathway were proposed as useful markers for with low birefringence of the inner layer of ZP (Madaschi et al., 2009;
choosing competent embryos (Xu, Chen, Zhang, Wei, & Cao, 2015). Montag et al., 2008). The miscarriage rate was also higher in the
On the other hand, it was demonstrated that RNAseq profiles of transferred embryos obtained from oocytes showing low
cumulus cells could not predict subsequent early embryo develop- birefringence when compared with the ones having high ZP
ment potential (Green et al., 2018). Also, there were no significant birefringence (Madaschi et al., 2009). In addition, Madaschi et al.
differences in the transcriptome of cumulus cells of the oocytes (2009) observed a positive correlation between ZP birefringence and
whose early embryos resulted in live births when compared with the meiotic spindle visualization. The fertilization rate of oocytes show-
cumulus cells of sibling oocytes whose early embryos could not be ing high birefringence was at a similar level with the oocytes having
successfully implanted (Green et al., 2018). In a recently published low birefringence (Madaschi et al., 2009). On the other hand, Ebner
research, Scarica et al. (2019) reported that CAMK1D (calcium/ et al. (2010) demonstrated that the birefringence of the inner layer
calmodulin‐dependent protein kinase type 1d) expression levels in of ZP in the oocytes correlated with predicting blastocyst formation,
cumulus cells can be employed for predicting the developmental but there was no relationship observed with embryo quality or
potential of the oocytes in blastocyst formation. In a conclusion, al- pregnancy proportion. In addition, there were significant positive
though a significant relationship between certain genes expression at correlations observed between ZP birefringence and the rates of
a higher or lower levels in cumulus cells and developmental features fertilization, embryo quality, and conception cycles (Gonzalez‐Ortega
of the oocytes and early embryos has been revealed, there is no et al., 2016). By contrast, no correlation was found between the
consensus on which particular genes can be precisely used and easily expression levels of potential biomarker genes (Calm1, Dpp8,
analyzed. Therefore, new studies especially related to transcriptomic Hist1h4c, Nrp1, Psmd6, Tom1, and Ubqln1) in the cumulus cells playing
and proteomic evaluations in cumulus cells are required for the roles in the oocytes development and ZP birefringence scores
purpose of accurately and expediently predicting which oocytes are (Assidi, Montag, & Sirard, 2015). It is noteworthy that ZP bi-
competent. refringence scores can also be compared with other oocyte‐
development‐related genes to understand the molecular biological
background of ZP birefringence. Consistent with the previous find-
2.2 | Zona pellucida ings in this field mentioned above, ZP birefringence in the oocytes
decreased with increasing age over 35 years old (Gonzalez‐Ortega
It is well known that oocytes in primary follicles and onward are et al., 2016). Importantly, there was no significant correlation de-
surrounded by ZP that is generated both by oocytes and enclosing tected between the embryo morphokinetic features (pronuclear
granulosa cells. Bertrand, Van den Bergh, and Englert (1995) fading timing, cleavage, and multinucleation) and high/low ZP bi-
reported that the oocytes having thinner ZP (the mean of refringence in patients with PCOS (Tabibnejad, Soleimani, &
16.6 ± 3.2 µm) exhibited a higher fertilization rate. This was most Aflatoonian, 2018). Taken together, although there are contradicting
likely because sperm penetration was facilitated in these circum- findings related to ZP birefringence and early development para-
stances when compared with the oocytes whose ZP thickness was at meters, more studies suggest that the oocytes whose ZP or its inner
the mean of 18.9 ± 4.0 µm (Bertrand et al., 1995). In the same study, layer at higher birefringence can be selected. Also, the molecular
no correlation was detected between ZP thickness and ooplasm background of ZP birefringence and its relevance with development
diameter. However, there was a positive correlation between ZP potential should be analyzed in surplus oocytes and early embryos in
 10982795, 2020, 10, Downloaded from https://onlinelibrary.wiley.com/doi/10.1002/mrd.23420 by CochraneArgentina, Wiley Online Library on [25/03/2024]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
1026 | OZTURK

TABLE 2 Extracytoplasmic parameters of human oocytes related to zona pellucida, perivitelline space, shape, and giantess

Extracytoplasmic
factor Property Outcome

Zona pellucida • Thin ↑ Fertilization (Bertrand et al., 1995)

• Thick ↑ Oocyte development (Rama Raju et al., 2007; Shen et al., 2005) ↑ embryo quality (Host
et al., 2002)
• High birefringence ↑ Meiotic spindle visualization (Madaschi et al., 2009) ↑ fertilization rate
(Gonzalez‐Ortega et al., 2016) ↑ embryo quality (Gonzalez‐Ortega et al., 2016) ↑
blastocysts formation (Ebner et al., 2010) ↑ implantation (Madaschi et al., 2009; Montag
et al., 2008) ↑ pregnancy (Madaschi et al., 2009; Montag et al., 2008) ↑ live birth
(Madaschi et al., 2009; Montag et al., 2008)
• Low birefringence ↑ Miscarriage rate (Madaschi et al., 2009)

Perivitelline space • Large ↓ Fertilization rate (Xia, 1997) ↓ embryo quality (Xia, 1997); ↑ embryo quality (Ten
et al., 2007)
• Presence of granules ↑ Oocyte maturity (Hassan‐Ali et al., 1998) ↓ implantation rate (Farhi et al., 2002) ↓
pregnancy rate (Farhi et al., 2002)

Shape • Ovoid shape ↓ Cleavage (Ebner et al., 2008) ↑ delayed compaction (Ebner et al., 2008) ↑ delayed
blastocyst formation (Ebner et al., 2008)
• Peripheral defect ↑ Degeneration (Ebner et al., 2001)

Giantess • Presence of giant oocyte ↑ Estradiol level (Balakier et al., 2002; Lehner et al., 2015) ↑ obtained oocyte number
(Balakier et al., 2002; Lehner et al., 2015) ↑ abnormal embryonic cleavage (Machtinger
et al., 2011)

Note: ↑, increased; ↓, decreased.

large groups after providing a consensus on the technical applica- of the embryos derived from these oocytes (Konc, Kanyo, & Cseh,
tions and sampling homogeneity. 2004; Madaschi et al., 2008) when compared to the embryos pro-
In addition, the darkness and hatching of ZP in oocytes was duced from oocytes not having detectable spindles. Petersen et al.
evaluated in a limited number of studies. The darkness of ZP did not (2009) revealed that the oocytes having visible spindles exhibited
exhibit any association with fertilization and implantation rates, or significantly higher developmental parameters including normal
embryo quality (De Sutter, Dozortsev, Qian, & Dhont, 1996; Ten, pronuclei appearance, Day 3 embryo quality, the rates of fertiliza-
Mendiola, Vioque, de Juan, & Bernabeu, 2007). And, Balaban, Ata, tion, cleavage, and blastocyst formation in comparison with non-
Isiklar, Yakin, and Urman (2008) reported that ZP hatching did not visible ones. There was, however, no correlation in the implantation
affect either cryosurvival of embryos or blastocyst formation. To and pregnancy rates when compared with the oocytes whose spin-
determine the potential relationship of darkness and ZP hatching dles were at undetectable levels (Petersen et al., 2009). In addition to
with oocytes quality and subsequent embryo development features, a positive relation with the rates of fertilization and embryo quality,
more studies are required. the conception cycle was also at a significantly higher proportion in
the oocytes with visible spindles than that of nonvisible ones
(Gonzalez‐Ortega et al., 2016). In contrast, Chamayou et al. (2006)
2.3 | Meiotic spindles reported that there was no observed correlation regarding the pre-
sence or absence of spindles and the embryo quality and clinical
Visualization and measuring the length of meiotic spindles were pregnancy rates. In the patients with PCOS, although there was no
examined to determine whether they can be used to predict the significant relationship between the mean timing of embryo mor-
early development‐related features (Table 3). For this purpose, the phokinetics and meiotic spindle visualization, the rates of implanta-
microscope images of oocytes were captured before ICSI by using tion, live birth, chemical, and clinical pregnancy were found to be
PolScope. Then, the length and size of spindles were measured with higher in the transferred embryos derived from oocytes having
software such as the Oosight imaging system (Tomari et al., 2018). visible spindles than those not having visible spindles (Tabibnejad
The visualization of spindles in the oocytes was found to be asso- et al., 2018). It is important to note that the visualization of spindles
ciated with higher ZP birefringence (Madaschi et al., 2009), fertili- also decreased with increasing age (Gonzalez‐Ortega et al., 2016). It
zation rate (Braga et al., 2008; Garcia‐Oro et al., 2017; Rama Raju seems reasonable to conclude that analyzing spindles visualization in
et al., 2007), and progression to the blastocyst stage (Braga et al., oocytes will increase the fertilization, implantation, and pregnancy
2008; Rama Raju et al., 2007). Furthermore, the presence of visible rates, though there are some studies that found no correlation. The
spindles in the oocytes correlated with increased implantation contradicting findings may be the result of using microscopes having
(Garcia‐Oro et al., 2017; Madaschi et al., 2008) and pregnancy rates different technical attachments/software programs and varying
 10982795, 2020, 10, Downloaded from https://onlinelibrary.wiley.com/doi/10.1002/mrd.23420 by CochraneArgentina, Wiley Online Library on [25/03/2024]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
OZTURK | 1027

TABLE 3 The potential outcomes of morphological and structural changes in the meiotic spindles of human oocytes

Meiotic spindle Outcome

• Visualization ↑ ZP birefringence (Madaschi et al., 2009) ↑ fertilization rate (Braga et al., 2008; Garcia‐Oro et al.,
2017; Gonzalez‐Ortega et al., 2016; Petersen et al., 2009; Rama Raju et al., 2007) ↑ normal pronuclei
appearance (Petersen et al., 2009) ↑ cleavage rate (Petersen et al., 2009) ↑ Day 3 embryo quality
(Petersen et al., 2009) ↑ embryo quality (Gonzalez‐Ortega et al., 2016) ↑ blastocyst development
(Braga et al., 2008; Garcia‐Oro et al., 2017; Petersen et al., 2009; Rama Raju et al., 2007) ↑
implantation rate (Garcia‐Oro et al., 2017; Madaschi et al., 2008) ↑ conception cycle (Gonzalez‐Ortega
et al., 2016) ↑ pregnancy rate (Konc et al., 2004; Madaschi et al., 2008)

• Longer spindle length ↑ Fertilization success (Korkmaz et al., 2015) ↑ blastocyst formation (Rama Raju et al., 2007)

• Misaligned meiotic spindle position ↑ Fertilization defect (Rienzi et al., 2003)

• Localization to beneath or adjacent ↑ Fertilization rate (Fang et al., 2007)

to PB

• Higher retardance ↑ Better pronuclear score (Shen et al., 2006) ↑ blastocyst formation (Rama Raju et al., 2007) ↑
pregnancy rate (Shen et al., 2006)

• Higher density ↑ Pregnancy rate (Kilani et al., 2009)

• Normally appearing spindles ↑ Fertilization rate (Kilani et al., 2009; Tilia et al., 2020) ↑ Day 3 embryo development (Tilia et al.,
2020) ↑ blastocyst quality (Tilia et al., 2020) ↑ euploid blastocyst number (Tilia et al., 2020) ↑
implantation rate (Kilani et al., 2009) ↑ pregnancy rate (Kilani et al., 2009) ↑ live birth rate
(Kilani et al., 2009)

• Fine aligned ↑ Embryo cleavage (Asa et al., 2017) ↑ embryo quality (Asa et al., 2017) ↓ embryo fragmentation
(Asa et al., 2017) ↑ implantation rate (Asa et al., 2017)

• Birefringent ↑ Pre‐embryo number (Shen et al., 2006) ↑ fertilization rate (Madaschi et al., 2008) ↑ good quality
pronuclei (Shen et al., 2006) ↑ early embryo cleavage (Madaschi et al., 2008)

Note: ↑, increased; ↓, decreased.

levels of embryologists’ experience in evaluating meiotic spindles as conclude that localization, size, and length of meiotic spindles may be
well as developmental heterogeneity of the oocytes at the time of used in oocytes selection for the purpose of increasing fertilization
analyzing. success rates and early development potential.
The oocytes having meiotic spindle lengths of more than 12 µm The relationship between spindle retardance (order and density
possessed higher chances of producing blastocysts when compared of spindles) and embryo development parameters was also eval-
to the oocytes with meiotic spindle lengths of 10–12 µm or less than uated. Although it was not statistically significant, there was a weak
10 µm (Rama Raju et al., 2007). Similarly, Korkmaz, Tekin, Sakinci, correlation found between low retardance and poor embryo quality
and Ercan (2015) found a correlation between the length of spindles (De Santis et al., 2005). On the other hand, Shen et al. (2006) re-
and fertilization success of women older than 35 years old (Korkmaz ported that high retardance of spindles in oocytes was associated
et al., 2015). In a recent study by Tomari et al. (2018) reported that with better pronuclear scores as well as higher pregnancy (Shen
the oocytes whose meiotic spindle sizes were between 90 and et al., 2006) and blastocyst formation (Rama Raju et al., 2007) rates.
120 μm2 had higher fertilization, blastocyst formation, and clinical Moreover, the oocytes resulting in pregnancy possessed notably
pregnancy rates when compared with the oocytes having larger or higher meiotic spindle densities in comparison with the ones not
smaller spindle sizes. The spindle location with regard to PB also resulting in pregnancy (Kilani, Cooke, Kan, & Chapman, 2009). In the
indicated the first cleavage plane and early embryo development same study, the oocytes having a significantly higher number of
parameters (Cooke, Tyler, & Driscoll, 2003). Indeed, the misalign- normally appearing spindles at 39–40 h following hCG treatment
ment of spindles according to PB resulted in fertilization defects, but resulted in pregnancy when compared with those that did not result
there was no correlation with the cleavage potential of the embryos in successful pregnancies (Kilani et al., 2009). The fertilization rate
derived from these oocytes (Rienzi et al., 2003). The oocytes whose was also significantly higher in the oocytes with normal spindles in
spindles localized beneath or adjacent to the PB had significantly comparison with the oocytes having abnormal spindles (Kilani,
higher fertilization rates when compared with the differently loca- Cooke, Tilia, & Chapman, 2011). The implantation and live birth rates
lized ones (Fang et al., 2007; Table 3). In contrast, Moon et al. (2003) of the embryos derived from oocytes with normal spindles were
demonstrated that no relationship existed between spindle locali- likewise higher than the embryos produced from the oocytes having
zation in the oocytes and fertilization rate as well as normal early abnormal spindles (Kilani et al., 2011). In addition, the fine spindle‐
embryo development (Moon et al., 2003). Based on these studies, we aligned subgroup exhibited positive correlations with embryo
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1028 | OZTURK

cleavage and implantation rates when compared with the average there was no correlation observed between fertilization/embryo
and poorly aligned subgroups which showed lower embryonic quality quality rates and large PS. Similarly, no relationship was found be-
and more fragmentation (Asa, Tabatabaee, Farrokhi, & Nejatbakhsh, tween PS size and the following morphokinetic values: second PB
2017; Table 3). In conclusion, the retardance, appearance, and extrusion timing, pronuclear morphology, pronuclei fading, 2–8 cell
alignment of spindles can be used as biomarkers to predict the de- embryo formation, and irregular cleavage pattern (Faramarzi et al.,
velopmental potential of the oocytes for subsequent developmental 2019) as well as oocyte yield (Weghofer et al., 2019). Interestingly,
processes such as fertilization, implantation, and pregnancy. De- Ten et al. (2007) documented that large PS caused 1.8 times higher
termining the molecular biological relationship between meiotic embryo quality when compared with the oocytes having normal PS.
spindles and early developmental processes is ideal subjects for fu- Most studies explored this subject suggest that PS size does not
ture investigations. seem to affect the subsequent developmental potential of oocytes.
For the spindle configuration, there was no significant difference When the impact of granules in PS was analyzed, the coarse
detected between fresh and frozen‐ and thawed‐mature oocytes granules decreased the implantation and pregnancy rates of the
when analyzed by using a confocal microscopy (Coticchio et al., oocytes in comparison to control ones (Farhi et al., 2002). Further-
2010). The oocytes with high birefringent spindles produced a sig- more, Hassan‐Ali et al. (1998) revealed that the incidence of granules
nificantly higher number of pre‐embryos with good pronuclei than in PS enhanced due to high dosages of human menopausal gonado-
the oocytes with low birefringent spindles generated pre‐embryos tropin (HMG) treatment, and it was positively associated with the
having low quality (Shen et al., 2006). The rates of fertilization and oocytes maturity. However, there was no correlation observed be-
early embryo cleavage derived from oocytes with birefringent spin- tween granulation in PS and the parameters such as fertilization,
dles were at higher levels when compared with the spindle‐non‐ cleavage, and pregnancy rates (Hassan‐Ali et al., 1998). Based on the
detected oocytes (Madaschi et al., 2008). The birefringent spindles in studies aimed to determine the possible association between PS size/
oocytes could be used in predicting their quality, fertilization, and granulation and oocytes developmental potential: although there is
developmental abilities (Rienzi et al., 2005; Table 3). It should be kept no consensus on these evaluated parameters, it seems reasonable to
in mind that various types of defects in spindles may lead to aneu- prefer the oocytes having small PS size and no granulation. It is
ploidies or maturation arrests in the oocytes (Eichenlaub‐Ritter, important to emphasize that this does not mean that oocytes having
Shen, & Tinneberg, 2002). Therefore, as reported in a recent study, large and granular PS cannot be used for producing embryos.
spindle morphologies of oocytes may be utilized in the prediction of
euploid blastocysts (Tilia, Chapman, Kilani, Cooke, & Venetis, 2020).
In the same study, it was displayed that, based on the intention‐to‐ 2.5 | Shape
treat (ITT) analysis, oocytes having normal spindles exhibited higher
rates of fertilization, Day 3 embryo development and blastocyst A study by De Sutter et al. (1996) documented that there was no
quality when compared with the oocytes with nonvisible spindles or correlation between fertilization rate/embryo quality score and irre-
having a translucent or telophase appearance (Tilia et al., 2020; gular oocyte shape. Similarly, the shapes of oocytes were not found to
Table 3). be associated with cryosurvival (Rienzi et al., 2008), aneuploidy (Yakin,
As a result, the analysis of spindles by using PolScope or other Balaban, Isiklar, & Urman, 2007), and implantation or clinical pregnancy
microscope types which do not seem to damage the developmental rate (Chamayou et al., 2006). Although the oocytes with ovoid shape
potential of the oocytes may be an easy way to predict early de- did not lose their fertilizability, they exhibited abnormalities in clea-
velopment features from fertilization to pregnancy. However, the vage as well as delayed compaction and blastocyst formation (Ebner,
molecular mechanism(s) underlying the correlations between spindle Shebl, Moser, Sommergruber, & Tews, 2008). There was no effect,
morphologies and oocyte quality should be identified in new studies however, on the blastocyst quality. The morphological defects in the
to reach precise, simple, and practical prediction method(s). periphery of oocytes involving fragile oolemma and irregular shape
were related to degeneration rather than cytoplasmic abnormalities,
and did not correlate with the development of the early embryo (Ebner
2.4 | Perivitelline space et al., 2001; Table 2). Overall, the shapes of oocytes except for ovoid
ones do not seem to influence the fertilization, early embryo devel-
The two morphologies involving size (enlarged or not; Xia, 1997) and opment, and the rates of implantation and clinical pregnancy.
content (presence of granulation; Hassan‐Ali et al., 1998) in PS might
be used in assessing oocyte quality (Table 2). Generally, it is accepted
that normal MII oocytes have a small PS including PB and no gran- 2.6 | Giant oocytes
ulation (Ubaldi & Rienzi, 2008). Xia (1997) revealed that large PS in
the oocytes without PB fragmentation and inclusions led to sig- A giant oocyte is characterized by having twice the volume of a
nificantly diminished fertilization and 3–8 cell embryo rates when normal oocyte (∼200 µm in diameter; Balakier, Bouman, Sojecki,
compared with the oocytes in normal appearance. On the other Librach, & Squire, 2002) and by having tetraploidy that is possibly
hand, De Sutter et al. (1996) and Balaban et al. (1998) reported that derived either from the absence of cytogenesis following a nuclear
 10982795, 2020, 10, Downloaded from https://onlinelibrary.wiley.com/doi/10.1002/mrd.23420 by CochraneArgentina, Wiley Online Library on [25/03/2024]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
OZTURK | 1029

division or fusion of two oogonia during early development (Austin, 2.7 | First polar body
1960). It is worth noting that women producing giant oocytes pos-
sessed higher levels of estradiol and an increased number of ob- The extrusion of PB indicates successful meiotic maturation of the
tained oocytes, suggesting that this issue may also originate from oocytes (Eichenlaub‐Ritter, Schmiady, Kentenich, & Soewarto, 1995).
ovarian stimulation protocols (Balakier et al., 2002). The incidence of Therefore, PB can be used as a biomarker of quality and even aging
giant oocytes is normally at a frequency of 0.26–0.31% among re- status of the oocytes because in vivo aged MII oocytes had a de-
covered oocytes (Balakier et al., 2002; Rosenbusch, Schneider, generated PB due to overmaturity, which resulted in developmental
Glaser, & Brucker, 2002). If binucleated giant MII oocytes were disabilities (Eichenlaub‐Ritter et al., 1995). Analogously, Xia (1997)
fertilized by normal spermatozoa, tripronuclear digynic zygotes revealed that intact PB significantly correlated with the fertilization
emerged (Rosenbusch & Schneider, 1998). Interestingly, these fer- rate and embryo quality. The embryos obtained from oocytes with
tilized giant oocytes are capable of undergoing normal cleavage and PB having well‐shaped morphology had higher implantation and
blastocyst formation; however, they should be discarded even if they pregnancy rates, and a decreased multiple pregnancy, when com-
have normally appeared two pronuclei (Ebner, Moser, & Tews, 2006), pared with the control embryos, included embryo fragmentation
due to the possibility of subsequent production of abnormal embryos (Ebner et al., 1999). Similarly, Ebner et al. (2000) showed that PB
(Balakier et al., 2002; Table 2). either in intact or well‐shaped morphology correlated with higher
A study by Machtinger, Politch, Hornstein, Ginsburg, and fertilization and embryo quality rates. Moreover, PB with intact in-
Racowsky (2011) documented that the presence of giant oocytes in a tegrity and having a smooth surface was found to be positively
cohort of retrieved oocytes led to abnormal embryonic cleavage correlated with the fertilization rate and embryo quality as well as
(increased percentage of 1‐ and 10‐cell embryos and reduced per- with the implantation and pregnancy rates (Ebner et al., 2002;
centage of 6‐ and 7‐cell embryos; Table 2), but there were no dif- Table 4).
ferences in the rates of embryos with perfect symmetry, An interesting study by Fancsovits et al. (2006) reported that the
fragmentation and implantation, as well as clinical or ongoing/deliv- oocytes having fragmented PB exhibited higher developmental pro-
ered pregnancy rates when compared with the cohort of oocytes gress than those with intact PB. On the other hand, the oocytes with
without a giant oocyte. In a different study, researchers created two large PB showed lower viability than the normal ones (Fancsovits
groups: a giant group involved cycles with at least one giant oocyte in et al., 2006). Although degeneration or largeness in PB was found to
the cohort of obtained oocytes, and a normal group included cycles be correlated with reduced fertilization rates, no relationship was
with no giant oocytes (Lehner et al., 2015). The giant group had detected with the pronuclear morphology or embryo quality (Rienzi
significantly higher estradiol levels and a higher number of retrieved et al., 2008). And, Navarro et al. (2009) documented a significant
oocytes compared with the normal group. By contrast, there were no association between large PB in the oocytes and the rates of de-
dramatic changes in the embryo quality and clinical pregnancy rate creased fertilization, cleavage, and embryo quality following ICSI
between the two groups (Lehner et al., 2015). It was reported in application. In the same study, no correlation was observed for PB
the same study that ovarian stimulation cannot influence the fragmentation. Also, it was reported that the oocytes with intact PB
incidence of giant oocytes. As a result, the presence of giant oocytes produced significantly higher rates of Day 3 good‐quality embryos
in the cohort of recovered oocytes does not appear to influence the and blastocysts, and more embryos in comparison to the fragmented
developmental features of the sibling oocytes. ones (Zhou, Fu, Sha, Chu, & Li, 2016). However, there were no

TABLE 4 The potential outcomes of morphological changes in the first polar body of human oocytes

First polar body Outcome

• Intact ↑ Fertilization rate (Xia, 1997) ↑ embryo quality (Xia, 1997) ↑ Day 3 good quality embryo (Zhou et al.,
2016) ↑ blastocyst quality (Zhou et al., 2016) ↑ available embryo number (Zhou et al., 2016)

• Well‐shaped morphology ↑ Implantation rate (Ebner et al., 1999) ↑ pregnancy rate (Ebner et al., 1999) ↓ multiple pregnancy rate
(Ebner et al., 1999)

• Intact and well‐shaped morphology ↑ Fertilization rate (Ebner et al., 2000) ↑ embryo quality (Ebner et al., 2000)

• Intact and smooth surface ↑ Fertilization rate (Ebner et al., 2002) ↑ embryo quality (Ebner et al., 2002) ↑ implantation rate
(Ebner et al., 2002) ↑ pregnancy rate (Ebner et al., 2002)

• Large ↓ Viability (Fancsovits et al., 2006) ↓ fertilization rate (Rienzi et al., 2006; Navarro et al., 2009) ↓ cleavage
(Navarro et al., 2009) ↓ embryo quality (Navarro et al., 2009)

• Degenerated ↓ Fertilization rate (Rienzi et al., 2006)

• Fragmented ↑ Developmental progress (Fancsovits et al., 2006

Note: ↑, increased; ↓, decreased.

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1030 | OZTURK

differences for pregnancy or implantation rates between the two defining PB morphologies; (b) the experience of laboratory techni-
groups. In addition to PB in oocytes, the second PB angle below cian analyzing PB; (c) the numbers of cases enrolled in each study;
24.25° in zygotes contributed to the prediction of good‐quality em- and (d) the timing of PB morphology evaluation; for example, the
bryos when compared with the zygotes whose PB exhibited a greater frequency of PB fragmentation increases during the interval from 2 h
angle between them (Zhang, Guo, & Li, 2016; Table 4). In light of 31 min to 3 h 30 min, that is, the time of passage from oocyte de-
these investigations, the oocytes with intact structure and well‐ nudation to ICSI (Ciotti et al., 2004).
shaped PB seem to have a higher developmental potential than that
of the ones displaying the opposite features. Consequently, these
oocytes may be selected above all others to improve the rate of take‐ 2.8 | Cytoplasmic changes
home babies.
In contrast to the studies mentioned above, Verlinsky et al. Cytoplasmic changes such as vacuolization and various types of in-
(2003) revealed that the abnormal PB morphologies involving frag- corporations including refractile bodies, granulation, inclusion, dis-
mentation and irregular shape did not correlate with fertilization coloration, and clustering of the smooth endoplasmic reticulum (SER)
rate, embryo quality, embryo survival to blastocyst stage, embryo in the oocytes were evaluated for the purpose of predicting their
transfer outcome, and aneuploidies. In parallel with this study, no potential effects on developmental competence (Ebner et al., 2001;
relationship was found between fragmentation in PB and the fol- Table 5). De Sutter et al. (1996) reported that there was no observed
lowing parameters: fertilization and cleavage rates, embryo quality, correlation between the fertilization rate/embryo quality score and
pregnancy and implantation rates (Ciotti et al., 2004). Similarly, De the presence of vacuoles in the ooplasm. In contrast, it was docu-
Santis et al. (2005) evaluated the following parameters such as size mented that the vacuoles derived from tubular‐type SER clusters
(normal and large) and shape (smooth, fragmented and rough) of PB, were associated with an increased biochemical pregnancy rate and
and determined that there were no significant differences between serum estradiol level at the day of hCG treatment, and with the
fertilization rate or embryo quality and the PB morphologies. The reduced clinical pregnancy rate (Otsuki, Okada, Morimoto, Nagai, &
contradicting findings for the PB morphologies may derive from the Kubo, 2004). Also, the oocytes with vacuolar cytoplasm or central
differences in (a) the criteria and technical equipment being used for granulation exhibited declined cryosurvival, and were unable to

TABLE 5 Cytoplasmic changes and their potential outcomes in the human oocytes

Cytoplasmic changes Outcome

• Presence of vacuole ↑ Serum estradiol level (Otsuki et al., 2004) ↓ oocyte cryosurvival (Balaban et al., 2008) ↓ good
quality blastocyst (Balaban et al., 2008) ↓ zona pellucida hatching (Balaban et al., 2008) ↑
biochemical pregnancy (Otsuki et al., 2004) ↓ clinical pregnancy (Otsuki et al., 2004)

• Presence of granulation ↓ Oocyte cryosurvival (Balaban et al., 2008) ↑ fertilization rate (Wilding et al., 2007) ↑
compromised pronuclear morphology (Rienzi et al., 2008) ↓ good quality blastocyst
(Balaban et al., 2008) ↓ zona pellucida hatching (Balaban et al., 2008) ↓ pregnancy rate
(Serhal et al., 1997)

• Presence of inclusion ↓ Fertilization rate (Xia, 1997) ↓ embryo quality (Xia, 1997) ↓ implantation rate (Serhal et al.,
1997) ↓ pregnancy rate (Serhal et al., 1997)

• Presence of SER aggregate ↓ Oocyte maturation (Setti et al. 2016) ↓ fertilization rate (Sa et al., 2011) ↓ embryo quality
(Ebner et al. 2008b; Sa et al. 2011) ↑ embryo transfer cancellation (Restelli, et al., 2015) ↓
implantation rate (Setti et al. 2016) ↓ pregnancy rate (Otsuki et al. 2004) ↑ miscarriage rate
(Braga et al. 2013) ↓ transcriptome level (Stigliani et al., 2018)

• High viscosity ↓ Fertilization rate (Ebner et al., 2003) ↓ embryo quality (Ebner et al., 2003) ↓ blastocyst
formation (Ebner et al., 2003)

• Presence of refractile body ↓ Fertilization rate (De Sutter et al., 1996; Otsuki et al., 2007) ↓ blastocyst formation (De Sutter
et al., 1996; Otsuki et al., 2007)

• Presence of darkness ↓ Embryo quality (Ten et al., 2007)

• Presence of multiple cytoplasmic ↓ Embryo quality (Chamayou et al., 2006) ↓ implantation rate (Serhal et al., 1997) ↓ pregnancy
dysmorphisms rate (Serhal et al., 1997)

• Texture, inclusion, vacuole and central ↓ Embryo quality (Chamayou et al., 2006)
granulation

• Granulation, inclusion, SER cluster and ↓ Implantation rate (Serhal et al., 1997) ↓ pregnancy rate (Serhal et al., 1997)
refractile body

Note: ↑, increased; ↓, decreased.

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OZTURK | 1031

develop into good quality blastocysts able to reach hatching stage organization of cytoskeleton and microtubules, and mitochondrial
(Balaban et al., 2008). Accordingly, although the transferred embryos function (Stigliani et al., 2018). Although these aggregates seem to be
derived from normal oocytes reached 24% pregnancy rate, it was related to the altered genes expression in the oocytes, it remains
only 3% in the embryos produced from oocytes having homogeneous elusive whether this type of oocytes should be used in ART centers.
granular cytoplasm (Serhal et al., 1997) and 12.8% in the oocytes In a limited number of studies, other cytoplasmic changes such
with granular cytoplasm (Kahraman et al., 2000). On the other hand, as cytoplasmic viscosity and refractile bodies were explored. As is
there was no relationship found between oocytes with centrally lo- known, cytoplasmic viscosity is briefly defined as the persistence of
cated granular cytoplasm and fertilization, embryo development, and injection funnel after withdrawing into ICSI pipette (Ebner et al.,
pregnancy rates (Kahraman et al., 2000). 2003). It influences the movement of cellular organelles and micro-
In other similarly designed studies, no correlation was detected tubule organization. Therefore, high cytoplasmic viscosity in the
between cytoplasmic inclusions and the development‐related para- oocytes adversely affected the fertilization, embryo quality, and
meters such as implantation, fertilization, and embryo quality rates blastocyst formation rates (Ebner et al., 2003). Also, the presence of
(Balaban et al., 1998; De Sutter et al., 1996). Interestingly, Wilding refractile bodies derived from lipofuscin inclusions correlated with
et al. (2007) revealed that the oocytes having cytoplasmic granula- the decreased fertilization rate and defective blastocyst formation
tion possessed a higher fertilization rate than the oocytes not having (De Sutter et al., 1996; Otsuki, Nagai, & Chiba, 2007). The darkness
any type of granularity. On the other hand, Rienzi et al. (2008) re- of ooplasm was found to be related to the reduced possibility of
ported that diffuse peripheral granulation in the MII oocytes resulted obtaining good‐quality embryos (Ten et al., 2007; Table 5). Overall,
in compromised pronuclear morphology. It is important to note that although oocytes showing granulation, cytoplasmic inclusions, SER
approximately 90% of the oocytes exhibited circular waves of cy- clusters, and refractile bodies exhibit normal fertilization and early
toplasmic granulation at the time period of 20–53 min after ICSI embryo development, the resultant embryos have lower implanta-
(Payne, Flaherty, Barry, & Matthews, 1997). To establish accurate tion and pregnancy rates when compared with the embryos derived
correlations of presence and localization of the vacuoles and cyto- from oocytes with normally appearing cytoplasm as documented in
plasmic granulation with the oocytes developmental potential, more the study by (Serhal et al., 1997). Also, multiple cytoplasmic dys-
studies in large groups are needed to reach a consensus on how to morphisms in the oocytes such as texture, inclusions, vacuoles, and
predict competent oocytes. central granulation adversely affected the embryo quality (Table 5);
The potential effects of cytoplasmic inclusion on the early de- however, there was no relationship with implantation and pregnancy
velopmental progress were also analyzed. Xia (1997) reported that rates (Chamayou et al., 2006). As a result, the oocytes having no
cytoplasmic inclusion in the oocytes was at higher levels in the pa- inclusions and aggregates in the cytoplasm and lower cytoplasmic
tients older than 35 years old when compared with the patients viscosity should be chosen to achieve higher pregnancy rates.
younger than 35 years old, and is correlated with a decreased fer-
tilization rate and embryo quality. Although there were no pregnancy
and implantation observed upon transfer of the embryos derived 2.9 | Multiple morphological parameters
from the oocytes with cytoplasmic inclusions, the fertilization, and
early embryo development were at similar rates between the two Some studies were examined the probable effects of multiple mor-
groups (Serhal et al., 1997). This study suggests that cytoplasmic phological changes on the developmental potential of oocytes
inclusions might exhibit their adverse effect(s) at the later terms of (Table 6). Xia (1997) demonstrated that oocytes with large PS,
prenatal development. fragmented PB, and cytoplasmic inclusions exhibited significantly
According to the Alpha/EHSRE consensus, the percentage of decreased fertilization rate and embryo quality. Moreover, the oo-
embryo transfer cancellation of the oocytes with aggregates of SER cytes showing dark cytoplasm with many vacuoles or fragments
was significantly higher when compared with the embryos resulting produced poor quality embryos, resulted in lower pregnancy rates
from oocytes without aggregates (Restelli et al., 2015). Indeed, SER when compared with the oocytes with normal morphology (5.5% vs.
aggregates were associated with lower oocyte maturation (Setti 29.4%; Loutradis et al., 1999). A significant relationship was dis-
et al., 2016) and embryo quality (Ebner, Moser, Shebl, Sommerguber, covered between oocyte morphologies (presence of vacuoles, ab-
& Tews, 2008b; Sa et al., 2011), and reduction of fertilization (Sa normal PB, large PS, diffused cytoplasmic granularity, and centrally
et al., 2011), implantation (Setti et al., 2016), and pregnancy rates located granular area) and the clinical pregnancy rate, a number of
(Otsuki et al., 2004), even increased miscarriage rates (Braga et al., retrieved oocytes, female ages, and basal FSH levels on Day 3 (Rienzi
2013) when compared with the oocytes lacking of SER aggregates et al., 2008). Yakin et al. (2007) found similar results and reported
(Ferreux et al., 2019). The potential impact of SER aggregates on the that the morphological changes in oocytes related to the meiotic
transcriptome of MII oocytes was also evaluated by using microarray spindle, ZP, PS, PB as well as the presence of vacuoles or granula-
(Stigliani et al., 2018). The oocytes with these aggregates showed tions in the ooplasm remarkably decreased the blastocyst formation.
significantly decreased expression of the genes being involved in As expected, high‐quality oocytes based on the morphological
certain biological activities such as cellular division, spindle organi- characteristics (granularity, vacuoles, and inclusions in the ooplasm,
zation, chromosome segregation, G2/M mitotic cell cycle transition, and the resistance and fragility of the oocytes plasma membrane at
 10982795, 2020, 10, Downloaded from https://onlinelibrary.wiley.com/doi/10.1002/mrd.23420 by CochraneArgentina, Wiley Online Library on [25/03/2024]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
1032 | OZTURK

TABLE 6 Multiple morphological, structural, and metabolic parameters in the human oocytes

Multiple morphologies Outcome

• Large PS, fragmented PB, and cytoplasmic inclusion ↓ Fertilization rate (Xia, 1997) ↓ embryo quality (Xia, 1997)

• Dark cytoplasm, many vacuoles or fragments in cytoplasm ↓ Embryo quality (Loutradis et al., 1999) ↓ Pregnancy rate (Loutradis
et al., 1999)

• Vacuoles, abnormal PB, large PS, diffused cytoplasmic granularity ↓ Oocyte number (Rienzi et al., 2008) ↓ clinical pregnancy (Rienzi
et al., 2008) ↑ basal FSH (Rienzi et al., 2008)

• Single PB, normally appearing cytoplasm, proper ZP thickness, and PS ↑ Transferable embryo number (Swain et al., 2008)

• Dismorphologies in meiotic spindle, ZP, PS, PB as well as the presence ↓ Blastocyst formation (Yakin et al., 2007)
of vacuoles or granulations in the ooplasm

• Abnormal PB, existing vacuoles, and large PS ↓ Fertilization rate (Rienzi et al., 2008)

• Granularity, vacuoles, and inclusions in the ooplasm, and the resistance ↓ Fertilization rate (Wilding et al., 2007) ↓ embryo quality (Wilding
and fragility of the oocyte plasma membrane et al., 2007) ↓ clinical pregnancy rate (Wilding et al., 2007)

• Large PS, diffused/central cytoplasmic granulation ↓ Pronuclear quality (Rienzi et al., 2008) ↓ Day 2 embryo quality
(Rienzi et al., 2008)

• Lower apoptotic index in cumulus cells and higher oxygen consumption ↑ Fertilization (Ruvolo et al., 2013) ↑ embryo cleavage (Ruvolo
of oocytes et al., 2013)

Note: ↑, increased; ↓, decreased.

Abbreviations: FSH, follicle‐stimulating hormone; PB, polar body; PS, perivitelline space; ZP, zona pellucida.

ICSI) possessed higher fertilization, embryo quality, and clinical with the previous studies, morphological changes in the oocytes as-
pregnancy rates than the low‐quality group (Wilding et al., 2007). sociated with the meiotic spindle, ZP, PS, PB, vacuolization, or
Also, it was documented that abnormal PB, existing vacuoles, and granulation did not influence the rates of fertilization, cleavage, and
large PS in the oocytes led to lower fertilization rates (Rienzi et al., aneuploidy (Yakin et al., 2007). Likewise, there was no correlation
2008). In the same study, large PS and diffused/central cytoplasmic determined between the morphological changes in COC, ZP, PS, PB,
granulation in the oocytes were found to affect the pronuclear vacuolization or granulation of the oocytes, and the rates of fertili-
morphology and Day 2 embryo quality. In addition to the morpho- zation, implantation, pregnancy, take‐home baby, and multiple
logical analysis, lower apoptotic index (<25%) in the cumulus cells pregnancy (La Sala et al., 2009).
and higher oxygen consumption of individual oocytes seemed to be Based on the studies presented in this section, there are con-
an indicator of better fertilization and embryo cleavage abilities tradictory findings regarding the ability of multiple morphological
(Ruvolo, Fattouh, Bosco, Brucculeri, & Cittadini, 2013). As a result, analyses to predict oocytes with high quality. The opposite findings
the oocytes having single and intact PB, normally appearing cyto- for similar multiple morphologies suggest that distinct molecular
plasm and meiotic spindle, proper ZP thickness, and PS exhibit a biological mechanisms having different effects on the developmental
higher potential for producing transferable embryos, as elegantly potential of oocytes may exhibit a correlation with the same
reviewed by (Swain & Pool, 2008; Table 6). morphologies. Therefore, to reach a consensus on the multiple
On the other hand, the morphologies such as extracytoplasmic morphologies, more studies are required to elicit the signaling
changes (dark ZP and enlarged PS), cytoplasmic abnormalities (dark pathways showing direct or indirect interaction(s) with these
cytoplasm, granular appearance, and refractile bodies), and shape morphologies. Until these investigations are carried out, multiple
defects in the oocytes did not influence fertilization and clinical morphologic analysis can be used to predict competent oocytes by
pregnancy rates as well as implantation potential and embryo quality employing a mathematical formula based on the prediction value of
when compared with the normal ones (Balaban et al., 1998). Cor- each morphology revealed in these studies.
respondingly, Chamayou et al. (2006) observed no correlation be-
tween the clinical pregnancy/implantation rates and the oocyte
morphological parameters (ooplasm texture, PS size/granulation, PB 3 | CO N C L U S I O N
shape, and meiotic spindle visualization). Also, no association was
detected between the oocyte dysmorphisms related to PS content, The morphological criteria being evaluated in some ART centers by
shape, PB, cytoplasmic granularity, cytoplasmic vacuoles as well as using light or polarized microscopy may contribute to successfully
with semen source, donor age, number of days of stimulation, plasma selecting good quality oocytes for the purpose of increasing the rate
estradiol concentration, and the development parameters including of take‐home baby. The most commonly analyzed morphologies in
fertilization rate and embryo quality (Ten et al., 2007). In agreement the oocytes are related to the meiotic spindle, CC, ZP, PS, vacuoles
 10982795, 2020, 10, Downloaded from https://onlinelibrary.wiley.com/doi/10.1002/mrd.23420 by CochraneArgentina, Wiley Online Library on [25/03/2024]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
OZTURK | 1033

or refractile bodies, shape, granulation, and viscosity of ooplasm. assessing embryo potential by gene expression profiles of human
There is, however, still no consensus on which of these morphologies cumulus cells: A proof of concept study. Molecular Human
Reproduction, 14(12), 711–719.
is the best one for oocyte selection. Therefore, multiple morpholo-
Attaran, M., Pasqualotto, E., Falcone, T., Goldberg, J. M., Miller, K. F.,
gical analyses seem to be a good way of predicting the oocytes Agarwal, A., & Sharma, R. K. (2000). The effect of follicular fluid
having a higher potential for producing competent early embryos. reactive oxygen species on the outcome of in vitro fertilization.
The oocytes having a clear or moderate granular cytoplasm, International Journal of Fertility and Women's Medicine, 45(5),
314–320.
narrow PS, PB in an intact appearance, normally appearing meiotic
Austin, C. R. (1960). Anomalies of fertilization leading to triploidy. Journal
spindles and CC, and colorless and birefringent ZP should be se- of Cellular and Comparative Physiology, 56(Suppl. 1), 1–15.
lected for the initial ARTs application if there are no restrictive Balaban, B., Ata, B., Isiklar, A., Yakin, K., & Urman, B. (2008). Severe
factors such as patient's age, harvested oocyte numbers, and pre- cytoplasmic abnormalities of the oocyte decrease cryosurvival and
subsequent embryonic development of cryopreserved embryos.
vious ART failures. It is noteworthy that these morphological criteria
Human Reproduction, 23(8), 1778–1785.
must be evaluated in a large sample of homogenous cases with the
Balaban, B., Urman, B., Sertac, A., Alatas, C., Aksoy, S., & Mercan, R.
participation of multiple ART centers. Together with morphological (1998). Oocyte morphology does not affect fertilization rate,
analysis, more sophisticated technologies such as genomics, tran- embryo quality and implantation rate after intracytoplasmic sperm
scriptomics, proteomics, and metabolomics can be used in more ac- injection. Human Reproduction, 13(12), 3431–3433.
Balakier, H., Bouman, D., Sojecki, A., Librach, C., & Squire, J. A. (2002).
curately predicting the oocytes with high quality.
Morphological and cytogenetic analysis of human giant oocytes and
In addition to choosing oocytes, the selection of competent giant embryos. Human Reproduction, 17(9), 2394–2401.
embryos is also a predominant strategy in infertility treatment for Balasch, J., & Gratacos, E. (2012). Delayed childbearing: Effects on fertility
the purpose of increasing the success rates and decreasing multiple and the outcome of pregnancy. Current Opinion in Obstetrics and
Gynecology, 24(3), 187–193.
pregnancy rates. if there are no limitations of obtaining oocytes
Bertrand, E., Van den Bergh, M., & Englert, Y. (1995). Does zona pellucida
fertilized by IVF or ICSI, the produced embryos can be selected thickness influence the fertilization rate? Human Reproduction, 10(5),
based on their morphological parameters and preimplantation ge- 1189–1193.
netic testing for aneuploidy analysis in some cases. In addition, Braga, D. P., Figueira Rde, C., Rodrigues, D., Madaschi, C.,
Pasqualotto, F. F., Iaconelli, A., & Borges, E. (2008). Prognostic value
morphokinetic, metabolomic, proteomic, epigenetic, and genomic
of meiotic spindle imaging on fertilization rate and embryo
evaluations on the early embryos are being investigated for selecting development in in vitro‐matured human oocytes. Fertility and
the ones having higher quality. In the cases generating a limited Sterility, 90(2), 429–433.
number of oocytes due to low fertility characteristics or legal re- Braga, D. P., Setti, A. S., Figueira Rde, C., Machado, R. B., Iaconelli, A., Jr., &
Borges, E., Jr. (2013). Influence of oocyte dysmorphisms on
strictions, the morphological criteria for determining competent
blastocyst formation and quality. Fertility and Sterility, 100(3),
oocytes can be used despite its clinical value lower than expected. 748–754.
Cha, K. Y., Koo, J. J., Ko, J. J., Choi, D. H., Han, S. Y., & Yoon, T. K. (1991).
C O NF L IC T O F IN T E R ES T Pregnancy after in vitro fertilization of human follicular oocytes
collected from nonstimulated cycles, their culture in vitro and their
The author declares that there is no conflict of interest.
transfer in a donor oocyte program. Fertility and Sterility, 55(1),
109–113.
ORCID Chamayou, S., Ragolia, C., Alecci, C., Storaci, G., Maglia, E., Russo, E., &
Saffet Ozturk http://orcid.org/0000-0002-8089-7078 Guglielmino, A. (2006). Meiotic spindle presence and oocyte
morphology do not predict clinical ICSI outcomes: A study of 967
transferred embryos. Reproductive BioMedicine Online, 13(5),
R EF E RE N C E S
661–667.
Anderson, R. A., Sciorio, R., Kinnell, H., Bayne, R. A., Thong, K. J., Cillo, F., Brevini, T. A., Antonini, S., Paffoni, A., Ragni, G., & Gandolfi, F.
de Sousa, P. A., & Pickering, S. (2009). Cumulus gene expression as a (2007). Association between human oocyte developmental
predictor of human oocyte fertilisation, embryo development and competence and expression levels of some cumulus genes.
competence to establish a pregnancy. Reproduction, 138(4), Reproduction, 134(5), 645–650.
629–637. Cimadomo, D., Capalbo, A., Ubaldi, F. M., Scarica, C., Palagiano, A.,
Asa, E., Tabatabaee, R., Farrokhi, A., & Nejatbakhsh, R. (2017). Canipari, R., & Rienzi, L. (2016). The impact of biopsy on human
Relationship between meiotic spindles visualization and embryo developmental potential during preimplantation genetic
intracytoplasmic sperm injection outcomes in human oocytes. diagnosis. BioMed Research International, 2016, 7193075.
Anatomy & Cell Biology, 50(1), 26–32. Ciotti, P. M., Notarangelo, L., Morselli‐Labate, A. M., Felletti, V., Porcu, E.,
Assidi, M., Montag, M., & Sirard, M. A. (2015). Use of both cumulus cells' & Venturoli, S. (2004). First polar body morphology before ICSI is
transcriptomic markers and zona pellucida birefringence to select not related to embryo quality or pregnancy rate. Human
developmentally competent oocytes in human assisted reproductive Reproduction, 19(10), 2334–2339.
technologies. BMC Genomics, 16(Suppl. 1), S9. Cooke, S., Tyler, J. P., & Driscoll, G. L. (2003). Meiotic spindle location and
Assidi, M., Montag, M., Van der Ven, K., & Sirard, M. A. (2011). Biomarkers identification and its effect on embryonic cleavage plane and early
of human oocyte developmental competence expressed in cumulus development. Human Reproduction, 18(11), 2397–2405.
cells before ICSI: a preliminary study. Journal of Assisted Reproduction Coticchio, G., Sciajno, R., Hutt, K., Bromfield, J., Borini, A., & Albertini, D. F.
and Genetics, 28(2), 173–188. (2010). Comparative analysis of the metaphase II spindle of human
Assou, S., Haouzi, D., Mahmoud, K., Aouacheria, A., Guillemin, Y., oocytes through polarized light and high‐performance confocal
Pantesco, V., … Hamamah, S. (2008). A non‐invasive test for microscopy. Fertility and Sterility, 93(6), 2056–2064.
 10982795, 2020, 10, Downloaded from https://onlinelibrary.wiley.com/doi/10.1002/mrd.23420 by CochraneArgentina, Wiley Online Library on [25/03/2024]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
1034 | OZTURK

Dal Canto, M., Brambillasca, F., Mignini Renzini, M., Coticchio, G., Fang, C., Tang, M., Li, T., Peng, W. L., Zhou, C. Q., Zhuang, G. L., &
Merola, M., Lain, M., … Fadini, R. (2012). Cumulus cell‐oocyte Leong, M. (2007). Visualization of meiotic spindle and subsequent
complexes retrieved from antral follicles in IVM cycles: relationship embryonic development in in vitro and in vivo matured human
between COCs morphology, gonadotropin priming and clinical oocytes. Journal Of Assisted Reproduction And Genetics, 24(11),
outcome. Journal Of Assisted Reproduction And Genetics, 29(6), 547–551.
513–519. Faramarzi, A., Khalili, M. A., & Omidi, M. (2019). Morphometric analysis of
Daya, S., Kohut, J., Gunby, J., & Younglai, E. (1990). Influence of blood human oocytes using time lapse: does it predict embryo
clots in the cumulus complex on oocyte fertilization and cleavage. developmental outcomes? Human Fertility, 22(3), 171–176.
Human Reproduction, 5(6), 744–746. Farhi, J., Nahum, H., Weissman, A., Zahalka, N., Glezerman, M., &
Devjak, R., Burnik Papler, T., Verdenik, I., Fon Tacer, K., Vrtacnik, & Levran, D. (2002). Coarse granulation in the perivitelline space and
Bokal, E. (2016). Embryo quality predictive models based on IVF‐ICSI outcome. Journal of Assisted Reproduction And Genetics,
cumulus cells gene expression. Balkan Journal of Medical Genetics: 19(12), 545–549.
BJMG, 19(1), 5–12. Ferreux, L., Sallem, A., Chargui, A., Gille, A. S., Bourdon, M., Maignien, C., …
Ebner, T., Balaban, B., Moser, M., Shebl, O., Urman, B., Ata, B., & Tews, G. Pocate‐Cheriet, K. (2019). Is it time to reconsider how to manage
(2010). Automatic user‐independent zona pellucida imaging at the oocytes affected by smooth endoplasmic reticulum aggregates?
oocyte stage allows for the prediction of preimplantation Human Reproduction, 34(4), 591–600.
development. Fertility and Sterility, 94(3), 913–920. Fragouli, E., Wells, D., Iager, A. E., Kayisli, U. A., & Patrizio, P. (2012).
Ebner, T., Moser, M., Shebl, O., Sommerguber, M., & Tews, G. (2008). Alteration of gene expression in human cumulus cells as a potential
Prognosis of oocytes showing aggregation of smooth endoplasmic indicator of oocyte aneuploidy. Human Reproduction, 27(8), 2559–2568.
reticulum. Reproductive BioMedicine Online, 16(1), 113–118. Garcia‐Oro, S., Rey, M. I., Rodriguez, M., Duran, A., Devesa, R., &
Ebner, T., Moser, M., Shebl, O., Sommergruber, M., Yaman, C., & Tews, G. Valverde, D. (2017). Predictive value of spindle retardance in
(2008). Blood clots in the cumulus‐oocyte complex predict poor embryo implantation rate. Journal of Assisted Reproduction And
oocyte quality and post‐fertilization development. Reproductive Genetics, 34(5), 617–625.
BioMedicine Online, 16(6), 801–807. Geraedts, J., Collins, J., Gianaroli, L., Goossens, V., Handyside, A.,
Ebner, T., Moser, M., Sommergruber, M., Puchner, M., Wiesinger, R., & Harper, J., … Schmutzler, A. (2010). What next for preimplantation
Tews, G. (2003). Developmental competence of oocytes showing genetic screening? A polar body approach! Human Reproduction,
increased cytoplasmic viscosity. Human Reproduction, 18(6), 25(3), 575–577.
1294–1298. Gilchrist, R. B., Lane, M., & Thompson, J. G. (2008). Oocyte‐secreted
Ebner, T., Moser, M., Sommergruber, M., Yaman, C., Pfleger, U., & Tews, G. factors: regulators of cumulus cell function and oocyte quality.
(2002). First polar body morphology and blastocyst formation rate Human Reproduction Update, 14(2), 159–177.
in ICSI patients. Human Reproduction, 17(9), 2415–2418. Gonzalez‐Ortega, C., Cancino‐Villarreal, P., Alonzo‐Torres, V. E., Martinez‐
Ebner, T., Moser, M., & Tews, G. (2006). Is oocyte morphology prognostic Robles, I., Perez‐Pena, E., & Gutierrez‐Gutierrez, A. M. (2016).
of embryo developmental potential after ICSI? Reproductive [Polarized light microscopy for evaluation of oocytes as a
BioMedicine Online, 12(4), 507–512. prognostic factor in the evolution of a cycle in assisted
Ebner, T., Moser, M., Yaman, C., Feichtinger, O., Hartl, J., & Tews, G. reproduction]. Ginecologia Y Obstetricia de Mexico, 84(4), 217–227.
(1999). Elective transfer of embryos selected on the basis of first Green, K. A., Franasiak, J. M., Werner, M. D., Tao, X., Landis, J. N.,
polar body morphology is associated with increased rates of Scott, R. T., Jr., & Treff, N. R. (2018). Cumulus cell transcriptome
implantation and pregnancy. Fertility and Sterility, 72(4), 599–603. profiling is not predictive of live birth after in vitro fertilization: a
Ebner, T., Shebl, O., Moser, M., Sommergruber, M., & Tews, G. (2008). paired analysis of euploid sibling blastocysts. Fertility and Sterility,
Developmental fate of ovoid oocytes. Human Reproduction, 23(1), 109(3), 460–466e462.
62–66. Hassan‐Ali, H., Hisham‐Saleh, A., El‐Gezeiry, D., Baghdady, I., Ismaeil, I., &
Ebner, T., Yaman, C., Moser, M., Sommergruber, M., Feichtinger, O., & Mandelbaum, J. (1998). Perivitelline space granularity: A sign of
Tews, G. (2000). Prognostic value of first polar body morphology on human menopausal gonadotrophin overdose in intracytoplasmic
fertilization rate and embryo quality in intracytoplasmic sperm sperm injection. Human Reproduction, 13(12), 3425–3430.
injection. Human Reproduction, 15(2), 427–430. Hatirnaz, S., Ata, B., Hatirnaz, E. S., Dahan, M. H., Tannus, S., Tan, J., &
Ebner, T., Yaman, C., Moser, M., Sommergruber, M., Jesacher, K., & Tan, S. L. (2018). Oocyte in vitro maturation: A sytematic review.
Tews, G. (2001). A prospective study on oocyte survival rate after Turkish Journal of Obstetrics and Gynecology, 15(2), 112–125.
ICSI: Influence of injection technique and morphological features. Host, E., Gabrielsen, A., Lindenberg, S., & Smidt‐Jensen, S. (2002).
Journal Of Assisted Reproduction And Genetics, 18(12), 623–628. Apoptosis in human cumulus cells in relation to zona pellucida
Eichenlaub‐Ritter, U., Schmiady, H., Kentenich, H., & Soewarto, D. (1995). thickness variation, maturation stage, and cleavage of the
Recurrent failure in polar body formation and premature corresponding oocyte after intracytoplasmic sperm injection.
chromosome condensation in oocytes from a human patient: Fertility and Sterility, 77(3), 511–515.
indicators of asynchrony in nuclear and cytoplasmic maturation. Junk, S. M., & Yeap, D. (2012). Improved implantation and ongoing
Human Reproduction, 10(9), 2343–2349. pregnancy rates after single‐embryo transfer with an optimized
Eichenlaub‐Ritter, U., Shen, Y., & Tinneberg, H. R. (2002). Manipulation of protocol for in vitro oocyte maturation in women with polycystic
the oocyte: possible damage to the spindle apparatus. Reproductive ovaries and polycystic ovary syndrome. Fertility and Sterility, 98(4),
BioMedicine Online, 5(2), 117–124. 888–892.
Ekart, J., McNatty, K., Hutton, J., & Pitman, J. (2013). Ranking and selection Kahraman, S., Yakin, K., Donmez, E., Samli, H., Bahce, M., Cengiz, G., …
of MII oocytes in human ICSI cycles using gene expression levels from Imirzalioglu, N. (2000). Relationship between granular cytoplasm of
associated cumulus cells. Human Reproduction, 28(11), 2930–2942. oocytes and pregnancy outcome following intracytoplasmic sperm
Fancsovits, P., Tothne, Z. G., Murber, A., Takacs, F. Z., Papp, Z., & injection. Human Reproduction, 15(11), 2390–2393.
Urbancsek, J. (2006). Correlation between first polar body Kilani, S., Cooke, S., Kan, A., & Chapman, M. (2009). Are there non‐
morphology and further embryo development. Acta Biologica invasive markers in human oocytes that can predict pregnancy
Hungarica, 57(3), 331–338. outcome? Reproductive BioMedicine Online, 18(5), 674–680.
 10982795, 2020, 10, Downloaded from https://onlinelibrary.wiley.com/doi/10.1002/mrd.23420 by CochraneArgentina, Wiley Online Library on [25/03/2024]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
OZTURK | 1035

Kilani, S., Cooke, S., Tilia, L., & Chapman, M. (2011). Does meiotic spindle Payne, D., Flaherty, S. P., Barry, M. F., & Matthews, C. D. (1997).
normality predict improved blastocyst development, implantation Preliminary observations on polar body extrusion and pronuclear
and live birth rates? Fertility and Sterility, 96(2), 389–393. formation in human oocytes using time‐lapse video cinematography.
Konc, J., Kanyo, K., & Cseh, S. (2004). Visualization and examination of the Human Reproduction, 12(3), 532–541.
meiotic spindle in human oocytes with polscope. Journal Of Assisted Petersen, C. G., Oliveira, J. B., Mauri, A. L., Massaro, F. C., Baruffi, R. L.,
Reproduction And Genetics, 21(10), 349–353. Pontes, A., & Franco, J. G., Jr. (2009). Relationship between
Korkmaz, C., Tekin, Y. B., Sakinci, M., & Ercan, C. M. (2015). Effects of visualization of meiotic spindle in human oocytes and ICSI outcomes:
maternal ageing on ICSI outcomes and embryo development in a meta‐analysis. Reproductive BioMedicine Online, 18(2), 235–243.
relation to oocytes morphological characteristics of birefringent Pincus, G., & Enzmann, E. V. (1935). The comparative behavior of
structures. Zygote, 23(4), 550–555. mammalian eggs in vivo and in vitro: I. The activation of ovarian egg.
Lasiene, K., Vitkus, A., Valanciute, A., & Lasys, V. (2009). Morphological Journal of Experimental Medicine, 6, 66–67.
criteria of oocyte quality. Medicina (Kaunas), 45(7), 509–515. Rama Raju, G. A., Prakash, G. J., Krishna, K. M., & Madan, K. (2007).
Lehner, A., Kaszas, Z., Murber, A., Rigo, J., Jr., Urbancsek, J., & Meiotic spindle and zona pellucida characteristics as predictors of
Fancsovits, P. (2015). Giant oocytes in human in vitro fertilization embryonic development: A preliminary study using PolScope
treatments. Archives of Gynecology and Obstetrics, 292(3), 697–703. imaging. Reproductive BioMedicine Online, 14(2), 166–174.
Li, J., Cao, Y., Xu, X., Xiang, H., Zhang, Z., Chen, B., … Chen, D. (2015). Rattanachaiyanont, M., Leader, A., & Leveille, M. C. (1999). Lack of
Increased New lncRNA‐mRNA gene pair levels in human cumulus correlation between oocyte‐corona‐cumulus complex morphology
cells correlate with oocyte maturation and embryo development. and nuclear maturity of oocytes collected in stimulated cycles for
Reproductive Sciences, 22(8), 1008–1014. intracytoplasmic sperm injection. Fertility and Sterility, 71(5), 937–940.
Loutradis, D., Drakakis, P., Kallianidis, K., Milingos, S., Dendrinos, S., & Restelli, L., Delle Noci, S., Mangiarini, A., Ferrari, S., Somigliana, E., &
Michalas, S. (1999). Oocyte morphology correlates with embryo Paffoni, A. (2015). The impact of Alpha/ESHRE consensus regarding
quality and pregnancy rate after intracytoplasmic sperm injection. oocytes with aggregates of smooth endoplasmic reticulum (SERa) on
Fertility and Sterility, 72(2), 240–244. in vitro fertilization outcome. Journal of Assisted Reproduction And
Machtinger, R., Politch, J. A., Hornstein, M. D., Ginsburg, E. S., & Genetics, 32(11), 1629–1635.
Racowsky, C. (2011). A giant oocyte in a cohort of retrieved oocytes: Rienzi, L., Ubaldi, F., Iacobelli, M., Minasi, M. G., Romano, S., & Greco, E.
does it have any effect on the in vitro fertilization cycle outcome? (2005). Meiotic spindle visualization in living human oocytes.
Fertility and Sterility, 95(2), 573–576. Reproductive BioMedicine Online, 10(2), 192–198.
Madaschi, C., Aoki, T., de Almeida Ferreira Braga, D. P., Rienzi, L., Ubaldi, F. M., Iacobelli, M., Minasi, M. G., Romano, S., Ferrero, S.,
de Cassia Savio Figueira, R., Semiao Francisco, L., Iaconelli, A., Jr., & Sapienza, F., Baroni, E., Litwicka, K., & Greco, E. (2008). Significance
Borges, E., Jr. (2009). Zona pellucida birefringence score and meiotic of metaphase II human oocyte morphology on ICSI outcome. Fertility
spindle visualization in relation to embryo development and ICSI and Sterility, 90(5), 1692–1700.
outcomes. Reproductive BioMedicine Online, 18(5), 681–686. Rienzi, L., Ubaldi, F. M., Martinez, F., Iacobelli, M., Minasi, M. G.,
Madaschi, C., de Souza Bonetti, T. C., de Almeida Ferreira Braga, D. P., Ferrero, S., Tesarik, J., & Greco, E. (2003). Relationship between
Pasqualotto, F. F., Iaconelli, A., Jr., & Borges, E., Jr. (2008). Spindle meiotic spindle location with regard to the polar body position and
imaging: a marker for embryo development and implantation. oocyte developmental potential after ICSI. Human Reproduction,
Fertility and Sterility, 90(1), 194–198. 18(6), 1289–1293.
McKenzie, L. J., Pangas, S. A., Carson, S. A., Kovanci, E., Cisneros, P., Rizzo, R., Dal Canto, M. B., Stignani, M., Fadini, R., Fumagalli, D.,
Buster, J. E., … Matzuk, M. M. (2004). Human cumulus granulosa cell Renzini, M. M., Borgatti, M., Gambari, R., & Baricordi, O. R. (2009).
gene expression: A predictor of fertilization and embryo selection in Production of sHLA‐G molecules by in vitro matured cumulus‐
women undergoing IVF. Human Reproduction, 19(12), 2869–2874. oocyte complex. International Journal of Molecular Medicine, 24(4),
Montag, M., Schimming, T., Koster, M., Zhou, C., Dorn, C., Rosing, B., … 523–530.
der Ven, K. (2008). Oocyte zona birefringence intensity is associated Rosenbusch, B., & Schneider, M. (1998). Maturation of a binuclear oocyte
with embryonic implantation potential in ICSI cycles. Reproductive from the germinal vesicle stage to metaphase II: Formation of two
BioMedicine Online, 16(2), 239–244. polar bodies and two haploid chromosome sets. Human
Moon, J. H., Hyun, C. S., Lee, S. W., Son, W. Y., Yoon, S. H., & Lim, J. H. Reproduction, 13(6), 1653–1655.
(2003). Visualization of the metaphase II meiotic spindle in living Rosenbusch, B., Schneider, M., Glaser, B., & Brucker, C. (2002).
human oocytes using the Polscope enables the prediction of Cytogenetic analysis of giant oocytes and zygotes to assess their
embryonic developmental competence after ICSI. Human relevance for the development of digynic triploidy. Human
Reproduction, 18(4), 817–820. Reproduction, 17(9), 2388–2393.
Navarro, P. A., de Araujo, M. M., de Araujo, C. M., Rocha, M., dos Reis, R., Ruvolo, G., Fattouh, R. R., Bosco, L., Brucculeri, A. M., & Cittadini, E.
& Martins, W. (2009). Relationship between first polar body (2013). New molecular markers for the evaluation of gamete
morphology before intracytoplasmic sperm injection and quality. Journal of Assisted Reproduction and Genetics, 30(2), 207–212.
fertilization rate, cleavage rate, and embryo quality. International Sa, R., Cunha, M., Silva, J., Luis, A., Oliveira, C., Teixeira da Silva, J.,
Journal of Gynecology & Obstetrics, 104(3), 226–229. Barros, A., & Sousa, M. (2011). Ultrastructure of tubular smooth
Ng, S. T., Chang, T. H., & Wu, T. C. (1999). Prediction of the rates of endoplasmic reticulum aggregates in human metaphase II oocytes
fertilization, cleavage, and pregnancy success by cumulus‐coronal and clinical implications. Fertility and Sterility, 96(1), 143–149e147.
morphology in an in vitro fertilization program. Fertility and Sterility, La Sala, G. B., Nicoli, A., Villani, M. T., Di Girolamo, R., Capodanno, F., &
72(3), 412–417. Blickstein, I. (2009). The effect of selecting oocytes for insemination
Otsuki, J., Nagai, Y., & Chiba, K. (2007). Lipofuscin bodies in human and transferring all resultant embryos without selection on
oocytes as an indicator of oocyte quality. Journal of Assisted outcomes of assisted reproduction. Fertility and Sterility, 91(1),
Reproduction And Genetics, 24(7), 263–270. 96–100.
Otsuki, J., Okada, A., Morimoto, K., Nagai, Y., & Kubo, H. (2004). The De Santis, L., Cino, I., Rabellotti, E., Calzi, F., Persico, P., Borini, A., &
relationship between pregnancy outcome and smooth endoplasmic Coticchio, G. (2005). Polar body morphology and spindle imaging as
reticulum clusters in MII human oocytes. Human Reproduction, 19(7), predictors of oocyte quality. Reproductive BioMedicine Online, 11(1),
1591–1597. 36–42.
 10982795, 2020, 10, Downloaded from https://onlinelibrary.wiley.com/doi/10.1002/mrd.23420 by CochraneArgentina, Wiley Online Library on [25/03/2024]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
1036 | OZTURK

Scarica, C., Cimadomo, D., Dovere, L., Giancani, A., Stoppa, M., Capalbo, A., Tilia, L., Chapman, M., Kilani, S., Cooke, S., & Venetis, C. (2020). Oocyte meiotic
& Canipari, R. (2019). An integrated investigation of oocyte spindle morphology is a predictive marker of blastocyst ploidy‐a
developmental competence: expression of key genes in human prospective cohort study. Fertility and Sterility, 113(1), 105–113e101.
cumulus cells, morphokinetics of early divisions, blastulation, and Tomari, H., Honjo, K., Kunitake, K., Aramaki, N., Kuhara, S., Hidaka, N.,
euploidy. Journal of Assisted Reproduction, 36(5), 875–887. Nishimura, K., Nagata, Y., & Horiuchi, T. (2018). Meiotic spindle size
Schoot, D. C., Coelingh Bennink, H. J., Mannaerts, B. M., Lamberts, S. W., is a strong indicator of human oocyte quality. Reproductive Medicine
Bouchard, P., & Fauser, B. C. (1992). Human recombinant follicle‐ and Biology, 17(3), 268–274.
stimulating hormone induces growth of preovulatory follicles Ubaldi, F., & Rienzi, L. (2008). Morphological selection of gametes.
without concomitant increase in androgen and estrogen Placenta, 29(Suppl. B), 115–120.
biosynthesis in a woman with isolated gonadotropin deficiency. Verlinsky, Y., Lerner, S., Illkevitch, N., Kuznetsov, V., Kuznetsov, I.,
Journal of Clinical Endocrinology and Metabolism, 74(6), 1471–1473. Cieslak, J., & Kuliev, A. (2003). Is there any predictive value of first
Serhal, P. F., Ranieri, D. M., Kinis, A., Marchant, S., Davies, M., & Khadum, I. M. polar body morphology for embryo genotype or developmental
(1997). Oocyte morphology predicts outcome of intracytoplasmic potential? Reproductive BioMedicine Online, 7(3), 336–341.
sperm injection. Human Reproduction, 12(6), 1267–1270. Weghofer, A., Kushnir, V. A., Darmon, S. K., Jafri, H., Lazzaroni‐Tealdi, E.,
Setti, A. S., Figueira, R. C., de Almeida Ferreira Braga, D. P., Zhang, L., Albertini, D. F., Barad, D. H., & Gleicher, N. (2019). Age,
Azevedo, M. C., Iaconelli, A., Jr., & Borges, E., Jr. (2016). Oocytes body weight and ovarian function affect oocyte size and morphology
with smooth endoplasmic reticulum clusters originate blastocysts in non‐PCOS patients undergoing intracytoplasmic sperm injection
with impaired implantation potential. Fertility and Sterility, 106(7), (ICSI). PLoS One, 14(10), e0222390.
1718–1724. Wilding, M., Di Matteo, L., D'Andretti, S., Montanaro, N., Capobianco, C., &
Shen, Y., Stalf, T., Mehnert, C., Eichenlaub‐Ritter, U., & Tinneberg, H. R. Dale, B. (2007). An oocyte score for use in assisted reproduction.
(2005). High magnitude of light retardation by the zona pellucida is Journal of Assisted Reproduction and Genetics, 24(8), 350–358.
associated with conception cycles. Human Reproduction, 20(6), Wolf, D. P. (1988). Oocyte quality and fertilization. In D. P. Wolf (Ed.), In
1596–1606. vitro fertilization and embryo transfer (pp. 129–138). New York, NY:
Shen, Y., Stalf, T., Mehnert, C., De Santis, L., Cino, I., Tinneberg, H. R., & Plenum Press.
Eichenlaub‐Ritter, U. (2006). Light retardance by human oocyte Xia, P. (1997). Intracytoplasmic sperm injection: correlation of oocyte
spindle is positively related to pronuclear score after ICSI. grade based on polar body, perivitelline space and cytoplasmic
Reproductive BioMedicine Online, 12(6), 737–751. inclusions with fertilization rate and embryo quality. Human
Stigliani, S., Moretti, S., Casciano, I., Canepa, P., Remorgida, V., Anserini, P., Reproduction, 12(8), 1750–1755.
& Scaruffi, P. (2018). Presence of aggregates of smooth endoplasmic Xu, X., Chen, D., Zhang, Z., Wei, Z., & Cao, Y. (2015). Molecular signature
reticulum in MII oocytes affects oocyte competence: Molecular‐ in human cumulus cells related to embryonic developmental
based evidence. Molecular Human Reproduction, 24(6), 310–317. potential. Reproductive Sciences, 22(2), 173–180.
De Sutter, P., Dozortsev, D., Qian, C., & Dhont, M. (1996). Oocyte Yakin, K., Balaban, B., Isiklar, A., & Urman, B. (2007). Oocyte dysmorphism
morphology does not correlate with fertilization rate and embryo is not associated with aneuploidy in the developing embryo. Fertility
quality after intracytoplasmic sperm injection. Human Reproduction, and Sterility, 88(4), 811–816.
11(3), 595–597. Zhang, Q., Guo, X. M., & Li, Y. (2016). Optimal polar bodies angle for
Swain, J. E., & Pool, T. B. (2008). ART failure: Oocyte contributions to higher subsequent embryo viability: A pilot study. Fertility and
unsuccessful fertilization. Human Reproduction Update, 14(5), 431–446. Sterility, 105(3), 670–675.
Tabibnejad, N., Soleimani, M., & Aflatoonian, A. (2018). Zona pellucida Zhou, W., Fu, L., Sha, W., Chu, D., & Li, Y. (2016). Relationship of polar
birefringence and meiotic spindle visualization are not related to the bodies morphology to embryo quality and pregnancy outcome.
time‐lapse detected embryo morphokinetics in women with Zygote, 24(3), 401–407.
polycystic ovarian syndrome. European Journal of Obstetrics,
Gynecology and Reproductive Biology, 230, 96–102.
Tanghe, S., Van Soom, A., Nauwynck, H., Coryn, M., & de Kruif, A. (2002).
Minireview: Functions of the cumulus oophorus during oocyte How to cite this article: Ozturk S. Selection of competent
maturation, ovulation, and fertilization. Molecular Reproduction and oocytes by morphological criteria for assisted reproductive
Development, 61(3), 414–424.
technologies. Mol Reprod Dev. 2020;87:1021–1036.
Ten, J., Mendiola, J., Vioque, J., de Juan, J., & Bernabeu, R. (2007). Donor
oocyte dysmorphisms and their influence on fertilization and https://doi.org/10.1002/mrd.23420
embryo quality. Reproductive BioMedicine Online, 14(1), 40–48.