Scanning the Meiotic Spindle in Assisted Reproductive Techniques to Assess Oocyte Quality

NCT ID: NCT06539585

Last Updated: 2024-08-06

Basic Information

• Recruitment Status: ACTIVE_NOT_RECRUITING
• Clinical Phase: NA
• Total Enrollment: 164 participants
• Study Classification: INTERVENTIONAL
• Study Start Date: 2024-07-18
• Study Completion Date: 2028-03-31

Brief Summary

The assisted reproduction success rate is affected by several factors including the age of the women, oocyte quality and maturation state, as well as sperm quality. Imaging of the meiotic spindle may be crucial for determining the oocyte maturation and the optimal time of oocyte fertilization by intracytoplasmic sperm injection (ICSI). A new accurate and non-invasive method for selecting quality maturated oocytes based on meiotic spindle imaging is for women over 35 years of age will be introduced. The evaluation of efficiency using meiotic spindle visibility in polarized light and its relative position to the polar body as indicator of oocyte maturity will be monitored and the optimal time for ICSI will be defined.

Detailed Description

One of the main strategies of infertility treatment is in vitro fertilization (IVF) . The IVF success rate is affected by several key factors including the age of the women, oocyte quality and maturation state, as well as sperm quality. Several authors have suggested that the presence, position and retardance of the optically birefringent meiotic spindle (MS) are related to oocyte developmental competence, affecting the quality of fertilization and embryo development. Oocytes with normal spindle morphology are significantly more likely to produce euploid embryos compared with oocytes with meiotic spindles that are not visible or abnormal. Adjusting the timing of intracytoplasmic sperm injection (ICSI) based on MS morphology has also been proposed. It was shown that adjusting the time of ICSI based on the position of the MS with respect to the polar body (PB) increases the probability of successful fertilization for women over 35 years of age. For the oocytes of patients over 35 years of age, the longer incubation time enabled a greater number of oocytes to become fully mature (MII phase) and prepared for ICSI. In this situation, the MS status was an important indicator of oocyte immaturity.

Patient will be randomized into three groups using Research Electronic Data Capture (REDCap): group 1, standard ICSI without MS imaging; group 2, MS imaging followed by standard ICSI, group 3, MS imaging and ICSI fertilization according MS status. Oocytes from group 3 patients with MS evaluation will be fertilized according to MS status either 5-6 hours after ovum pick-up (OPU) or 7-8 hours after OPU. Oocytes without MS evaluation will all fertilized 5-6 hours after OPU. MS evaluation will take place in pre-prepared glass-bottomed dishes with about 5μl medium with the HEPES buffer covered with paraffin oil, put to heat one hour before use. The oocyte will be rotated using a needle so that Polar Body (PB) and MS are both well visible, and photographs will be taken using optical microscope. The MS status and the angle (α) between MS and PB will be obtained 3-4 hours after oocyte pick-up (OPU). At the same time, the polarized-light microscopy image will be acquired (polarized light microscopy at ×100 magnification. Specially, for oocytes from group 3 patients with PB/MS in close proximity (angle between PB and MS < 5◦) or MS not visible will be predicted as immature oocytes. For these oocytes, ICSI will be performed 4-5 h after the polarization microscopy evaluation, i.e., 7-8 h after OPU. For oocytes with MS clearly visible and PB/MS not in close proximity (angle between PB and MS > 5◦) ICSI will be performed typically 2-3 h after the polarization microscopy evaluation, i.e., 5-6 h after OPU, as for patients from 1 and 2 groups. ICSI will be performed according to standard protocol using ICSI/holding micropipettes (Microtech IVF, Czech Republic), polyvinylpyrrolidone (ICSI™, Vitrolife, Sweden), and Eppendorf (Hamburg, Germany) micromanipulation system equipped with thermoplate (Tokaohit, Japan). The oocytes will be cultivated individually, and their order preserved, so that the other outcomes (data from timelapse, clinical results) can be associated with individual oocytes. We will also monitor the time from the human chorionic gonadotropin (hCG) administration to the completion of the actual fertilization. The oocytes will be denuded (HYASE-10X™, Vitrolife, Sweden) after OPU, and the maturation stage will be examined. Germinal Vesicle (GV) stage oocyte will not be included. Oocytes and embryos in time-lapse incubator from the Japanese manufacturer Astec will be cultivated in a timelapse dish Origio, Denmark Sage 1-Step™, Origio, Denmark under paraffin oil (OVOIL™, VITROLIFE, Sweden) at 37.0 °C, 6% CO2 and 5%O2. Fertilization after ICSI will be defined as the presence of two pronuclei and 2 polar body 16-20 h post ICSI. Embryos will be cultivated for 122-144 h.

Pilot findings will be confirmed in a randomized control trial, i.e. to the efficiency of using MS visibility and relative position to the polar body as indicators of oocyte maturation in order to optimize ICSI timing will be evaluated.

Conditions

Infertility; Infertility, Male; Infertility, Female

Study Design

• Allocation Method: RANDOMIZED
• Intervention Model: PARALLEL
• Primary Study Purpose: DIAGNOSTIC
• Blinding Strategy: SINGLE

Study Groups

Standard ICSI without meiotic spindle imaging.

A control group of patients who underwent a standard IVF cycle without microscopy of the meiotic spindle. In this control group of patients, ICSI fertilization will be performed 5-6 hours after oocyte collection.

ICSI will be performed according to standard protocol [6]. The oocytes will be cultivated individually, and their order preserved, so that the other outcomes (data from timelapse, clinical results) can be associated with individual oocytes. Fertilization after ICSI will be defined as the presence of two pronuclei and 2 polar body. Embryos will be cultivated for 122-144 h.

Group Type: ACTIVE_COMPARATOR

ICSI

Intervention Type: PROCEDURE

ICSI will be performed according to standard protocol using ICSI/holding micropipettes (#002-5-30/#001-120-30, Microtech IVF, Czech Republic), polyvinylpyrrolidone (ICSI™, Vitrolife, Sweden), and Eppendorf (Hamburg, Germany) micromanipulation system equipped with thermoplate (Tokaohit, Japan). The oocytes will be cultivated individually, and their order preserved, so that the other outcomes (data from timelapse, clinical results) can be associated with individual oocytes. Oocytes will be denuded (HYASE-10X™, Vitrolife, Sweden) after OPU, and the maturation stage will be examined.

Meiotic spindle imaging followed by standard ICSI.

MS evaluation will take place in pre-prepared glass-bottomed dishes with about 5μl medium with the HEPES buffer covered with paraffin oil, put to heat one hour before use. The oocyte will be rotated using a needle so that PB and MS are both well visible, and photographs will be taken using optical microscope. The MS status and the angle (α) between MS and PB will be obtained 3-4 hours after oocyte pick-up (OPU). At the same time, the polarized-light microscopy image will be acquired (polarized light microscopy at ×100 magnification. ICSI will be performed typically 2-3 h after the polarization microscopy evaluation, i.e., 5-6 h after OPU.

ICSI will be performed according to standard protocol [6]. Fertilization after ICSI will be defined as the presence of two pronuclei and 2 polar body. Embryos will be cultivated for 122-144 h.

Group Type: ACTIVE_COMPARATOR

Meiotic spindle status observation in polarized light

Intervention Type: PROCEDURE

In patients older than 35 years and younger than 40 years, we will use a microscope with a polarizing filter to evaluate the position of meiotic spindles and polar bodies in oocytes collected from patients who were indicated for IVF and ICSI. Using an optical microscope with a Nikon CEE GmbH polarizing filter, the angle between PB and MS together with MS visibility will be determined.

ICSI

Intervention Type: PROCEDURE

ICSI will be performed according to standard protocol using ICSI/holding micropipettes (#002-5-30/#001-120-30, Microtech IVF, Czech Republic), polyvinylpyrrolidone (ICSI™, Vitrolife, Sweden), and Eppendorf (Hamburg, Germany) micromanipulation system equipped with thermoplate (Tokaohit, Japan). The oocytes will be cultivated individually, and their order preserved, so that the other outcomes (data from timelapse, clinical results) can be associated with individual oocytes. Oocytes will be denuded (HYASE-10X™, Vitrolife, Sweden) after OPU, and the maturation stage will be examined.

MS imaging and ICSI fertilization according MS status.

Oocytes with MS evaluation will be fertilized according to MS status either 5-6 hours after ovum pick-up (OPU) or 7-8 hours after OPU. MS evaluation will take place in pre-prepared glass-bottomed dishes. The MS status and the angle (α) between MS and PB will be obtained 3-4 hours after oocyte pick-up (OPU). At the same time, the polarized-light microscopy image will be acquired (polarized light microscopy at ×100 magnification. For oocytes with PB/MS in close proximity (angle between PB and MS < 5◦) or MS not visible ICSI will be performed 4-5 h after the polarization microscopy evaluation, i.e., 7-8 h after OPU (these oocytes are supposed to be not fully mature). For oocytes with MS clearly visible and PB/MS not in close proximity (angle between PB and MS > 5◦) ICSI will be performed typically 2-3 h after the polarization microscopy evaluation, i.e., 5-6 h after OPU.

ICSI will be performed according to standard protocol [6].

Group Type: ACTIVE_COMPARATOR

Meiotic spindle status observation in polarized light

Intervention Type: PROCEDURE

In patients older than 35 years and younger than 40 years, we will use a microscope with a polarizing filter to evaluate the position of meiotic spindles and polar bodies in oocytes collected from patients who were indicated for IVF and ICSI. Using an optical microscope with a Nikon CEE GmbH polarizing filter, the angle between PB and MS together with MS visibility will be determined.

ICSI

Intervention Type: PROCEDURE

ICSI will be performed according to standard protocol using ICSI/holding micropipettes (#002-5-30/#001-120-30, Microtech IVF, Czech Republic), polyvinylpyrrolidone (ICSI™, Vitrolife, Sweden), and Eppendorf (Hamburg, Germany) micromanipulation system equipped with thermoplate (Tokaohit, Japan). The oocytes will be cultivated individually, and their order preserved, so that the other outcomes (data from timelapse, clinical results) can be associated with individual oocytes. Oocytes will be denuded (HYASE-10X™, Vitrolife, Sweden) after OPU, and the maturation stage will be examined.

Interventions

Meiotic spindle status observation in polarized light

In patients older than 35 years and younger than 40 years, we will use a microscope with a polarizing filter to evaluate the position of meiotic spindles and polar bodies in oocytes collected from patients who were indicated for IVF and ICSI. Using an optical microscope with a Nikon CEE GmbH polarizing filter, the angle between PB and MS together with MS visibility will be determined.

Intervention Type: PROCEDURE

ICSI

ICSI will be performed according to standard protocol using ICSI/holding micropipettes (#002-5-30/#001-120-30, Microtech IVF, Czech Republic), polyvinylpyrrolidone (ICSI™, Vitrolife, Sweden), and Eppendorf (Hamburg, Germany) micromanipulation system equipped with thermoplate (Tokaohit, Japan). The oocytes will be cultivated individually, and their order preserved, so that the other outcomes (data from timelapse, clinical results) can be associated with individual oocytes. Oocytes will be denuded (HYASE-10X™, Vitrolife, Sweden) after OPU, and the maturation stage will be examined.

Intervention Type: PROCEDURE

Eligibility Criteria

Inclusion Criteria

• Women who underwent ICSI;
• Spermiogram containing at least 0.5 mil/ml sperms;
• Morphlogy: normal morphology >1%;
• Stimulated cycles

Exclusion Criteria

• Age under 35 years or over 40;
• Native cycles;
• Severe uterine abnormalities (submucosal fibroid, fibroid ≥5cm, uterine septum, endometrial polyps, uterus duplex)

• Minimum Eligible Age: 35 Years
• Maximum Eligible Age: 40 Years
• Eligible Sex: FEMALE
• Accepts Healthy Volunteers: No

Sponsors

Czech Academy of Sciences

OTHER

Sponsor Role: collaborator

Czech Technical University in Prague

OTHER

Sponsor Role: collaborator

General University Hospital, Prague

OTHER

Sponsor Role: collaborator

Charles University, Czech Republic

OTHER

Sponsor Role: lead

Responsible Party

Jaromír Mašata

Leading doctor of the Center of Urogynecology and Pelvic Recontructive Surgery

Responsibility Role: PRINCIPAL_INVESTIGATOR

Locations

General University Hospital in Prague

Prague, , Czechia

Countries

Czechia

References

van Loendersloot LL, van Wely M, Limpens J, Bossuyt PM, Repping S, van der Veen F. Predictive factors in in vitro fertilization (IVF): a systematic review and meta-analysis. Hum Reprod Update. 2010 Nov-Dec;16(6):577-89. doi: 10.1093/humupd/dmq015. Epub 2010 Jun 25.

Reference Type: BACKGROUND

PMID: 20581128 (View on PubMed)

Wu B, Shi J, Zhao W, Lu S, Silva M, Gelety TJ. Understanding reproducibility of human IVF traits to predict next IVF cycle outcome. J Assist Reprod Genet. 2014 Oct;31(10):1323-30. doi: 10.1007/s10815-014-0288-y. Epub 2014 Aug 15.

Reference Type: BACKGROUND

PMID: 25119191 (View on PubMed)

Hanevik HI, Hessen DO. IVF and human evolution. Hum Reprod Update. 2022 Jun 30;28(4):457-479. doi: 10.1093/humupd/dmac014.

Reference Type: BACKGROUND

PMID: 35355060 (View on PubMed)

Rienzi L, Ubaldi F, Martinez F, Iacobelli M, Minasi MG, Ferrero S, Tesarik J, Greco E. Relationship between meiotic spindle location with regard to the polar body position and oocyte developmental potential after ICSI. Hum Reprod. 2003 Jun;18(6):1289-93. doi: 10.1093/humrep/deg274.

Reference Type: BACKGROUND

PMID: 12773461 (View on PubMed)

Innocenti F, Fiorentino G, Cimadomo D, Soscia D, Garagna S, Rienzi L, Ubaldi FM, Zuccotti M; SIERR. Maternal effect factors that contribute to oocytes developmental competence: an update. J Assist Reprod Genet. 2022 Apr;39(4):861-871. doi: 10.1007/s10815-022-02434-y. Epub 2022 Feb 15.

Reference Type: BACKGROUND

PMID: 35165782 (View on PubMed)

Rienzi L, Vajta G, Ubaldi F. Predictive value of oocyte morphology in human IVF: a systematic review of the literature. Hum Reprod Update. 2011 Jan-Feb;17(1):34-45. doi: 10.1093/humupd/dmq029. Epub 2010 Jul 16.

Reference Type: BACKGROUND

PMID: 20639518 (View on PubMed)

Tepla O, Topurko Z, Jirsova S, Moosova M, Fajmonova E, Cabela R, Komrskova K, Kratochvilova I, Masata J. Timing of ICSI with Respect to Meiotic Spindle Status. Int J Mol Sci. 2022 Dec 21;24(1):105. doi: 10.3390/ijms24010105.

Reference Type: RESULT

PMID: 36613547 (View on PubMed)

Other Identifiers

MW24-08-00048

Identifier Type: -

Identifier Source: org_study_id