Is the acrosome reaction a prerequisite for sperm incorporation after intra-cytoplasmic sperm injection (ICSI)?

1997 ◽  
Vol 9 (7) ◽  
pp. 703 ◽  
Author(s):  
A. H. Sathananthan ◽  
A. Szell ◽  
S. C. Ng ◽  
A. Kausche ◽  
O. Lacham-Kaplan ◽  
...  
Keyword(s):  
Acrosome Reaction ◽  
Germinal Vesicle ◽  
Sperm Chromatin ◽  
Perivitelline Space ◽  
Chromatin Decondensation ◽  
Immature Oocytes ◽  
Human Oocytes ◽  
Ultrastructural Evidence ◽  
Intra Cytoplasmic Sperm Injection

There is debate as to whether the acrosome reaction is necessary for sperm incorporation after intra-cytoplasmic sperm injection (ICSI). Ultrastructural evidence is presented to show that the acrosome reaction could occur in the ooplasm before sperm incorporation in mature human oocytes or the acrosome could be discarded intact before sperm incorporation in immature oocytes, matured in vitro. Both germinal vesicle and growing follicular oocytes showed sperm chromatin decondensation, with discarded acrosomes close to the sites of incorporation, and were able to form male pronuclei. This is probably the first report of microfertilization of a growing oocyte with a reticulate nucleolus by ICSI. The acrosome reaction, when it occurs, is preceded by acrosome swelling and is followed by vesiculation of surface membranes exposing the inner acrosome membrane, as observed on the surface of the zona during IVF or in the perivitelline space after subzonal sperm injection. These sperm were probably capacitated at the time of ICSI. There was subtle evidence of leaching of the acrosomal matrix from intact discarded acrosomes and from partially depleted acrosomes attached to decondensing spermheads. These sperm were probably not fully capacitated at the time of ICSI. It is concluded that both the acrosome reaction and acrosome deletion are possible prerequisites to sperm incorporation after ICSI.

Developmental potential of in vitro or in vivo matured oocytes

Zygote ◽  
2013 ◽  
Vol 23 (1) ◽  
pp. 93-98 ◽  
Author(s):  
Diego D. Alcoba ◽  
Anita M. Pimentel ◽  
Ilma S. Brum ◽  
Helena E. Corleta
Keyword(s):  
Germinal Vesicle ◽  
Fertilization Rate ◽  
Cleavage Rate ◽  
Suitable Alternative ◽  
Developmental Potential ◽  
Immature Oocytes ◽  
Prophase I ◽  
Intra Cytoplasmic Sperm Injection

SummaryThis study compared the embryological features of mature and immature oocytes (different stages) collected from stimulated cycles of in vitro fertilization (IVF). Immature oocytes were identified, classified as PI (prophase I – germinal vesicle, GV) or MI (metaphase I), were matured in vitro and fertilized using the intra-cytoplasmic sperm injection (ICSI) technique. Fertilization potential, cleavage, and subsequent transfer/cryopreservation of the embryos derived from these in vitro matured oocytes were compared with those of in vivo matured oocytes (collected at the MII stage). The characteristics of embryos derived from gametes recovered in the MI and MII stages were similar. The fertilization rate of immature oocytes recovered in PI was significantly lower than that of MII oocytes (P = 0.031), and the cleavage rate of the PI group was also lower than that of the MI (P = 0.004) and MII (P < 0.001) groups. In vitro maturation of MI oocytes is a suitable alternative when immature oocytes are recovered, as their characteristics and development are similar to those of in vivo matured oocytes. Optimization of outcomes for PI oocytes will require development of techniques that can distinguish which of these gametes will mature and fertilize.

Chromosome and spindle configurations of human oocytes matured in vitro after cryopreservation at the germinal vesicle stage

1997 ◽  
Vol 68 (5) ◽  
pp. 920-926 ◽  
Author(s):  
Sung-Eun Park ◽  
Weon-Young Son ◽  
Sook-Hwan Lee ◽  
Kyung-Ah Lee ◽  
Jung-Jae Ko ◽  
...  
Keyword(s):  
Germinal Vesicle ◽  
Human Oocytes

scholarly journals Progesterone treatment of boar spermatozoa improves male pronuclear formation after intracytoplasmic sperm injection into porcine oocytes

Zygote ◽  
2002 ◽  
Vol 10 (2) ◽  
pp. 95-104 ◽  
Author(s):  
Mike Katayama ◽  
Takashi Miyano ◽  
Masashi Miyake ◽  
Seishiro Kato
Keyword(s):  
Plasma Membrane ◽  
Intracytoplasmic Sperm Injection ◽  
Acrosome Reaction ◽  
Sperm Chromatin ◽  
Oocyte Activation ◽  
Freezing And Thawing ◽  
Chromatin Decondensation ◽  
Progesterone Treatment ◽  
Pronuclear Formation ◽  
Boar Spermatozoa

Boar spermatozoa were prepared for intracytoplasmic sperm injection (ICSI) by two different treatments to facilitate sperm chromatin decondensation and improve fertilisation rates after ICSI in pigs: spermatozoa were either frozen and thawed without cryoprotectants, or treated with progesterone. Morphological changes of the sperm heads after the treatments were examined and then the activation of oocytes and the transformation of the sperm nucleus following ICSI were assessed. After freezing and thawing, the plasma membrane and acrosomal contents over the apical region of sperm head were lost in all the spermatozoa. Following treatment with 1 mg/ml progesterone, the acrosome reaction was induced in 61% of spermatozoa. After injection of three types of spermatozoa, non-treated spermatozoa and progesterone-treated (i.e. acrosome-reacted) spermatozoa induced oocyte activation, but frozen-thawed spermatozoa induced oocyte activation at a significantly lower rate. Sixty-two per cent of sperm heads remained orcein-negative for 6 h, however, resulting in delayed sperm chromatin decondensation and low male pronuclear formation in the oocytes injected with a non-treated spermatazoon. Since the treatments of freezing and thawing and progesterone for spermatozoa accelerated the initial change in sperm chromatin and the latter treatment induced oocyte activation earlier, it is considered that the delay in oocyte activation and decondensation of sperm chromatin after injection of non-treated spermatozoa is caused by the existence of the sperm plasma membrane. These results show that progesterone treatment efficiently induces the acrosome reaction in boar spermatozoa without destroying their potency for oocyte activation, and the induction of the acrosome reaction results in the promotion of male pronuclear formation after ICSI.

scholarly journals Human Sperm Chromatin Undergoes Physiological Remodeling During In Vitro Capacitation and Acrosome Reaction

2012 ◽  
Vol 33 (5) ◽  
pp. 1025-1035 ◽  
Author(s):  
E. de Lamirande ◽  
M. C. San Gabriel ◽  
A. Zini
Keyword(s):  
Acrosome Reaction ◽  
Human Sperm ◽  
Sperm Chromatin

Overheating is detrimental to meiotic spindles within in vitro matured human oocytes

Zygote ◽  
2004 ◽  
Vol 12 (1) ◽  
pp. 65-70 ◽  
Author(s):  
Xiao-Fang Sun ◽  
Wei-Hua Wang ◽  
David L. Keefe
Keyword(s):  
High Temperature ◽  
Light Microscope ◽  
Polarized Light ◽  
Germinal Vesicle ◽  
Meiotic Spindle ◽  
Human Oocytes ◽  
Living State ◽  
Meiotic Spindles ◽  
Metaphase I

The present study was designed to examine the effects of overheating on meiotic spindle morphology within in vitro matured human oocytes using a polarized light microscope (Polscope). Immature human oocytes at either germinal vesicle or metaphase I stage were cultured in vitro for 24–36 h until they reached metaphase II (M-II) stage. After maturation, oocytes at M-II stage were imaged in the living state with the Polscope at 37, 38, 39 and 40 °C for up to 20 min. After heating, oocytes were returned to 37 °C and then imaged for another 20 min at 37 °C. The microtubules in the spindles were quantified by their maximum retardance, which represents the amount of microtubules. Spindles were intact at 37 °C during 40 min of examination and their maximum retardance (1.72–1.79) did not change significantly during imaging. More microtubules were formed in the spindles heated to 38 °C and the maximum retardance was increased from 1.77 before heating to 1.95 at 20 min after heating. By contrast, spindles started to disassemble when the temperature was increased to 39 °C for 10 min (maximum retardance was reduced from 1.76 to 1.65) or 40 °C for 1 min (maximum retardance was reduced from 1.75 to 1.5). At the end of heating (20 min), fewer microtubules were present in the spindles and the maximum retardance was reduced to 0.8 and 0.78 in the oocytes heated to 39 °C and 40 °C, respectively. Heating to 40 °C also induced spindles to relocate in the cytoplasm in some oocytes. After the temperature was returned to 37 °C, microtubules were repolymerized to form spindles, but the spindles were not reconstituted completely compared with the spindles imaged before heating. These results indicate that spindles in human eggs are sensitive to high temperature. Moreover, maintenance of an in vitro manipulation temperature of 37 °C is crucial for normal spindle morphology.

Mouse sperm fertilising capacity following subzonal microinjection is dependent on sperm washing and response to solubilised zonae pellucidae

Zygote ◽  
1995 ◽  
Vol 3 (1) ◽  
pp. 9-16 ◽  
Author(s):  
Orly Lacham Kaplan ◽  
Alan Trounson
Keyword(s):  
High Speed ◽  
Acrosome Reaction ◽  
Reaction Rate ◽  
Culture Medium ◽  
Perivitelline Space ◽  
Hybrid Male ◽  
Fertilisation Rate ◽  
Fresh Culture Medium ◽  
Fresh Culture

SummarySpermatozoa obtained from hybrid male mice were treated with solubilised zonae pellucidae after a period of capacitation in vitro to induce the acrosome reaction. Single spermatozoa were selected and microinjected into the perivitelline space of mature oocytes. A high proportion of the spermatozoa acrosome-reacted affter treatment with solubilised zonae (63%). However, the fertilisation rate (37%) after subzonal microinjection of oocytes by the treated spermatozoa was not different to the fertilisation rate (40%) of oocytes microinjected with untreated spermatozoa which had a lower rate of acrosome reaction (39%). When spermatozoa were washed by high-speed centrifugation before treatment with solubilised zonae pellucidae and subzonal microinjection, the fertilisation rate (68%) was significantly higher (p < 0.001) than that (29%) for occytes microinjected with untreated spermatozoa and was found to be correlated with a high acrosome reaction rate (74%) (r = 0.8). The washing of spermatozoa by centrifugation itself did not increase the acrosome reaction rate or fertilisation rate of oocytes after microinjection. The results of this study suggests that some modifications other than the acrosome reaction are needed to enable capacitated mouse spermatozoa to fuse with the oocyte plasma membrane. These modifications were achieved by washing spermatozoa by high-speed centrifugation and the replacement of the supernatant with fresh culture medium used for capacitation. Induction of the acrosome reaction by solubilised zonae pellucidae following this treatment leads to a high fertilisation rate of oocytes by subzonal sperm microinjection.

scholarly journals In vitro development of human oocytes reconstructed by sequential transfer of germinal vesicle and MII spindle

2017 ◽  
Vol 108 (3) ◽  
pp. e57-e58 ◽  
Author(s):  
H. Liu ◽  
Z. Lu ◽  
M. Yang ◽  
Z. Liu ◽  
Z. Merhi ◽  
...  
Keyword(s):  
Germinal Vesicle ◽  
In Vitro Development ◽  
Human Oocytes ◽  
Vitro Development

Meiosis progression and donor age affect expression profile of DNA repair genes in bovine oocytes

Zygote ◽  
2013 ◽  
Vol 23 (1) ◽  
pp. 11-18 ◽  
Author(s):  
S. Bilotto ◽  
R. Boni ◽  
G.L. Russo ◽  
M.B. Lioi
Keyword(s):  
Dna Repair ◽  
Dna Sequences ◽  
Calf Serum ◽  
Internal Standard ◽  
Germinal Vesicle ◽  
Cumulus Cells ◽  
Gene Marker ◽  
Donor Age ◽  
Immature Oocytes

SummarySeveral genetic and physiological factors increase the risk of DNA damage in mammalian oocytes. Two critical events are: (i) meiosis progression, from maturation to fertilization, due to extensive chromatin remodelling during genome decondensation; and (ii) aging, which is associated with a progressive oxidative stress. In this work, we studied the transcriptional patterns of three genes, RAD51, APEX-1 and MLH1, involved in DNA repair mechanisms. The analyses were performed by real-time quantitative PCR (RT-qPCR) in immature and in vitro matured oocytes collected from 17 ± 3-month-old heifers and 94 ± 20-month-old cows. Batches of 30–50 oocytes for each group (three replicates) were collected from ovarian follicles of slaughtered animals. The oocytes were freed from cumulus cells at the time of follicle removal, or after in vitro maturation (IVM) carried out in M199 supplemented with 10% fetal calf serum, 10 IU luteinising hormone (LH)/ml, 0.1 IU follicle-stimulating hormone (FSH)/ml and 1 μg 17β-oestradiol/ml. Total RNA was extracted by Trizol method. The expression of bovine GAPDH gene was used as the internal standard, while primers for bovine RAD51, APEX-1 and MLH1 genes were designed from DNA sequences retrieved from GenBank. Results obtained indicate a clear up-regulation of RAD51, APEX-1 and MLH1 genes after IVM, ranging between two- and four-fold compared with germinal vesicle (GV) oocytes. However, only RAD51 showed a significant transcript increase between the immature oocytes collected from young or old individuals. This finding highlights RAD51 as a candidate gene marker for discriminating bovine immature oocytes in relation to the donor age.

scholarly journals Surubim-do-Paraíba oocytes viability after being exposed to different cryoprotectants

2018 ◽  
Vol 48 (6) ◽  
Author(s):  
Taís da Silva Lopes ◽  
Eduardo Antônio Sanches ◽  
Danilo Caneppele ◽  
Elizabeth Romagosa
Keyword(s):  
Dimethyl Sulfoxide ◽  
Propylene Glycol ◽  
Germinal Vesicle ◽  
Blue Staining ◽  
Mature Stage ◽  
Germinal Vesicle Breakdown ◽  
Immature Oocytes ◽  
Stage Of Development ◽  
Oocyte Stage

ABSTRACT: To know the non-toxic cryoprotectants to fish oocytes is of extreme importance for tests that aim to increase oocyte resistance to cold, thus allowing more advanced studies in cryopreservation. Therefore, commonly used cryoprotectants such as methanol, dimethyl sulfoxide, ethylene glycol, propylene glycol, sucrose and fructose were studied. Immature oocytes from the initial to vitelogenic (diameter <1.7 mm) and mature (diameter >1.8 mm) stages of Steindachneridion parahybae were evaluated. Four distinct experiments were performed, three using immature oocytes and one using oocytes at the mature stage. For each oocyte stage, the best maintenance solution to be used: Hank or 50% L15 and; viability after baths for 30min (room temperature) at cryoprotectant concentrations ranging from 0.25 to 4M were evaluated. Different tests were used to evaluate oocyte viability: in vitro maturation followed by observation of germinal vesicle breakdown (only for immature oocytes), Trypan Blue staining (all stages) and fertilization and hatching rates (mature stage only). Results showed that the toxic effect of cryoprotectants on oocytes generally increases with increasing concentrations. Sensitivity of oocytes to cryoprotectants increases according to the stage of development, with mature oocytes being more sensitive. Sucrose, fructose, methanol, propylene glycol and dimethyl sulfoxide can be used as cryoprotectants for S. parahybae oocytes.

P–269 Effect of well-of-the-well culture as in vitro maturation system for human oocytes

2021 ◽  
Vol 36 (Supplement_1) ◽  
Author(s):  
B Dura. Lopez ◽  
I Moya ◽  
P Torres ◽  
M J Gomez-Torres ◽  
A Monzo ◽  
...  
Keyword(s):  
Culture System ◽  
In Vitro Maturation ◽  
Germinal Vesicle ◽  
Study Groups ◽  
Culture Conditions ◽  
Human Oocytes ◽  
Nuclear Maturation ◽  
Secreted Factors ◽  
Experimental Group

Abstract Study question Can the Well-of-the-Well system (WOW), applied on denuded oocytes, improve germinal vesicle breakdown (GVBD) and maturation rate? Summary answer In vitro maturation (IVM) of denuded germinal vesicle (GV) oocyte using WOW culture system increases nuclear maturation competence when compared with droplet conventional culture What is known already Further research remains necessary to address the mechanism of oocyte maturation in order to refine culture conditions and improve the implantation rate of in vitro matured oocytes. Several studies on bovine oocytes have shown that oocyte-secreted factors (an uncharacterized mix of growth factors secreted by the oocyte) enhance oocyte developmental competence during in vitro maturation. These oocyte-secreted factors may accumulate at the bottom of the micro-well, as suggested for the WOW culture system. Previous reports suggested that diffusible factors secreted by individual oocytes probably accumulated in a micro-well WOW dish, may provide a suitable microenvironment for their in vitro maturation. Study design, size, duration A total of 879 GV collected between 2017 and 2019 were included in this study. They were randomly allocated into two experimental groups: (1) single-cultured oocytes (SC) that were cultured individually in micro-droplets, and (2) group-cultured oocytes (WOW) that were cultured in a microwell culture system using the WOW dish (culture dish for time lapse incubator). The nuclear maturation was assessed after 24 hours and 48 hours of IVM Participants/materials, setting, methods GV oocytes were obtained from 609 patients undergoing controlled ovarian stimulation cycles. Oocytes from the experimental group (1) were placed individually in conventional 25μl micro-droplets in a 35 mm dish. Oocytes from the experimental group (2) were placed in 80 μl droplet individually in each of 9 microwells of WOW dish. All GV oocytes were matured in a single step embryo culture medium, supplemented with human menopausal gonadotropin and synthetic serum substitute. Main results and the role of chance Mature oocyte (MII) was considered when we observed rupture of the GV and the presence of a first polar body in the perivitelline space during the first 24 or 48 hours of culture under inverted optical microscope. GVBD noted significant differences (p-valor = 0.000) between the study groups after culturing of 24 hours [GVBD: SC group; 70% (318/455) vs. WOW group; 83% (352/424)] and 48 hours [GVBD: SC group; 77% (319/416) vs. WOW group; 94% (398/424)]. The maturation rates (MR) showed significant differences (p-valor = 0.000) between the study groups after culturing of 24 hours [MR: SC group; 51% (233/455) vs. WOW group; 80% (338/424)] and 48 hours [MR: SC group; 71% (295/416) vs. WOW group; 91% (387/424)]. Limitations, reasons for caution There is no data on cleavage and blastocyst rates. There are no previous reports comparing the maturation rates in denuded human oocytes single-cultured in individually droplet or group-cultured in WOW dish. Wider implications of the findings: Our results must be taken into account in order to improve the culture conditions for the optimization of the in vitro maturation technique in human oocytes from stimulated cycles. We now provide evidence that group-cultured oocytes in WOW dish increase GVBD and maturation rates. Trial registration number Not applicable

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