CAPA-IVM Culture With Low Oxygen Tension

Capacitation in-vitro maturation (CAPA-IVM) has recently been advanced in culturing oocytes from the germinal vesicle (GV) stage following mild or no controlled ovarian stimulation. Recent research suggested that O2 concentration may significantly regulate oocyte maturation and early embryo development through hypoxia-inducible factor (HIF). Nonetheless, it has been challenging to create the environmental culture conditions for addressing the optimal number of oocytes and the highest possibility of embryo development since consensus on the oxygen (O2) concentration index in the IVM culture environment has not been reached. Based on the outcomes of atmospheric O2 concentration (20%) and low O2 concentration (5%) during CAPA-IVM culture in mice, it has been hypothesized that a 5% O2 was the optimal culture condition for the pre-IVM step. A 20% O2 was more suitable for the IVM culture step. Therefore, this study is designed to enhance the CAPA-IVM culture system, improving treatment efficiency and providing various benefits for patients undergoing assisted reproductive technology.

Study Overview

• Status: Completed
• Conditions: In Vitro Fertilization
• Intervention / Treatment: Other: Air Oxygen Concentration CAPA-IVM culture; Other: Low Oxygen Concentration CAPA-IVM culture

Detailed Description

Capacitation in-vitro maturation (CAPA-IVM) has recently been advanced in culturing oocytes from the germinal vesicle (GV) stage. This approach is a modified version of conventional in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI), following mild stimulation or no controlled ovarian stimulation occurred. Specifically, IVM can be indicated for patients diagnosed with polycystic ovary syndrome (PCOS), a higher number of secondary follicles (constituting nearly 15% of total patients), and treat a range of patients with the risks of ovarian hyperstimulation, venous thromboembolism or ovarian torsion. Additionally, CAPA-IVM helps shorten treatment time, is less expensive, and upgrades patient convenience without multiple follow-up examinations. The live birth rate after the first embryo transfer in the CAPA-IVM group was 35.2%, which was not statistically significantly different from the IVF group at 43.2% (risk difference -8.1%; 95% confidence interval from -16.6% to 0.5%). However, the number of high-quality embryos in each cycle and the cumulative clinical pregnancy rate in CAPA-IVM were still lower than in cIVF.

Moreover, further investigation should be considered due to the lack of high-quality evidence of concurrent reports. Therefore, improving the oocyte maturation conditions in CAPA-IVM to harvest the optimal number of oocytes and the highest possibility of embryo development is essential. Many studies conducted on both animal and human models have demonstrated that the effectiveness of CAPA-IVM depends on various factors. Among these, the environmental culture conditions such as oxygen (O2) concentration play a crucial role in producing healthy mature oocytes. O2 is a vital physical and chemical component of the fallopian tube, uterus and ovarian follicle, it is closely related to metabolic activity, oocyte maturation, and early embryo development. Recent research suggested that O2 concentration may significantly regulate oocyte maturation and early embryo development through hypoxia-inducible factor (HIF). A consensus on the O2 concentration index in the IVM culture environment has not been reached. Oocyte-embedded culture systems have been commonly used in two O2 concentrations, 5% and 20% worldwide. In the human body, cumulus-oocyte complexes (COCs) mature in conditions with low O2 concentrations ranging from 2% to 9%.

Conversely, COCs are exposed to an atmospheric O2 concentration of 20% during IVM manipulation and culture. Although the concentration of 5% mimics the most proper environment in the fallopian tube and uterus, the 20% O2 is widely applied in IVM techniques. The use of high concentrations facilitates a better progression of differentiation processes and increases the maturation rate of oocytes. However, some referential frames indicated that a 20% O2 may pose a risk of reactive oxidative stress (ROS), leading to an imbalance in the ratio of pro-oxidants to antioxidants, resulting in cellular damage. Furthermore, real-time respiration analysis of oocytes cultured at 5% O2 is similar to in vivo-developed oocytes but induced cellular activity and oxygen consumption at 20% O2. The impact of atmospheric O2 concentration (20%) and low O2 concentration (5%) during CAPA-IVM culture in mice shown in the study of Vrije Universiteit Brussel (VUB) - Belgium that the respiratory capability of COCs cultured at 5% O2 was relatively similar to COCs developing and maturing in vivo.

Nonetheless, COCs cultured at 20% O2 increased respiratory activity and oxygen consumption remarkably. The study observed that pre-IVM culture of COCs at 20% O2 caused developmental disruptions. Also, the result was unfavorable if mouse COCs were cultured at the IVM step with 5% O2. Based on these analyses, the researchers hypothesized that a 5% O2 was the optimal culture condition for the pre-IVM step, while a 20% O2 was more relevant to the IVM culture step. Combining these findings with results from VUB and characteristics of the differentiation process in CAPA-IVM oocytes, this study is divided into two main groups, including 5% pre-IVM and 20% IVM versus 20% pre-IVM and IVM) and demonstrates whether this hypothesis should be applied CAPA-IVM in human. The enhancement of the CAPA-IVM culture system leads to improved treatment efficiency of this technique and provides various benefits for patients undergoing assisted reproductive technology.

Study procedure:

Screening for eligibility

• This trial will be conducted at My Duc Hospital, Ho Chi Minh City, Viet Nam.
• Women who are potentially eligible will be provided information about the trial at the time of IVM treatment indication.
• Screening for eligibility will be performed on the day of the first visit when the IVM treatment is indicated.
• Patients will be provided information related to the study together with the informed consent documents. Signed informed consent forms will be obtained by the investigators from all women before the enrolment.

Oocytes will be divided into 2 groups:

Group 1 (includes 2 subgroups: 1A and 1B): Air Oxygen Concentration CAPA-IVM culture T = Total number of oocytes after OR and there are two subgroups.

The number of oocytes is divided below:

If T is an even number:

• Number of oocytes in Group 1A: One oocyte.
• Number of oocytes in Group 1B: T1B = (T-2)/2.

If T is an odd number:

• Number of oocytes in Group 1A: One oocyte.
• Number of oocytes in Group 1B: T1B = ([T-1]-2)/2. One oocyte remainder of the first patient will be assigned to group 1B, and the remainder of the next patient will be assigned to group 2B. Continuing to do so sequentially for the next remainder.

Group 2 (includes 2 subgroups: 2A and 2B): Low Oxygen Concentration CAPA- IVM culture T = Total number of oocytes after OR and there are two subgroups.

The number of oocytes is divided below:

If T is an even number:

• Number of oocytes in Group 2A: One oocyte.
• Number of oocytes in Group 2B: T2B = (T-2)/2.

If T is an odd number:

• Number of oocytes in Group 2A: One oocyte.
• Number of oocytes in Group 2B: T2B = ([T-1]-2)/2. One oocyte remainder of the first patient will be assigned to group 1B, and the remainder of the next patient will be assigned to group 2B. Continuing to do so sequentially for the next remainder.

Group 1A, 2A: Collecting after capacitation: oocyte and cumulus cell.

Group 1B, 2B: Collecting after capacitation: spent media, blank well. Collecting after maturation: spent media, cumulus cell, blank well.

• Study Type: Interventional
• Enrollment (Actual): 20
• Phase: Not Applicable

Contacts and Locations

Study Locations

• Vietnam
  • Ho Chi Minh City, Vietnam
    • My Duc Hospital

Participation Criteria

Eligibility Criteria

• Ages Eligible for Study: Adult
• Accepts Healthy Volunteers: No

Description

Inclusion Criteria:

• 18-42 years of age
• Diagnosed with polycystic ovary syndrome according to the Rotterdam criteria (2003)
• Indicating CAPA-IVM treatment
• Having at least 40 antral follicles in two ovaries by transvaginal ultrasound at the time of CAPA-IVM indication
• Agreeing to have frozen embryo transfer
• Agreeing to participate in the trial

Exclusion Criteria:

• Cycles with oocyte donation, preimplantation Genetic Testing (PGT)
• Couples with severe male factor (sperm concentration <5 million/ml, motility < 10%), surgical sperm retrieval
• Previous history of unexplained immature oocytes after IVF treatment
• Uterine abnormalities

Study Plan

How is the study designed?

Design Details

• Primary Purpose: Treatment
• Allocation: Non-Randomized
• Interventional Model: Parallel Assignment
• Masking: None (Open Label)

Number of Arms

2

Arms and Interventions

Participant Group / Arm	Intervention / Treatment
Experimental: Air Oxygen Concentration CAPA-IVM culture; Half of the COCs will be cultured in the CAPA step and IVM step at 37 degrees Celsius in air oxygen concentration (20%) and 6% carbon dioxide.	Other: Air Oxygen Concentration CAPA-IVM culture; Group 1A: COC will be cultured in the CAPA step at an air oxygen concentration (20%) for 24 hours and 6% carbon dioxide at 37 degrees Celsius.; Group 1B: COCs will be cultured in the CAPA step at an air oxygen concentration (20%) for 24 hours and in the IVM step at an air oxygen concentration (20%) for 30 hours; all two steps combine 6% carbon dioxide at 37 degrees Celsius.
Active Comparator: Low Oxygen Concentration CAPA-IVM culture; Half of the COCs will be cultured in the CAPA step at a low oxygen concentration (5%) and IVM step at an air oxygen concentration (20%); all two steps combine 6% carbon dioxide at 37 degrees Celsius.	Other: Low Oxygen Concentration CAPA-IVM culture; Group 2A: COC will be cultured in the CAPA step at a low oxygen concentration (5%) for 24 hours and 6% carbon dioxide at 37 degrees Celsius.; Group 2B: COCs will be cultured in the CAPA step at a low oxygen concentration (5%) for 24 hours and the IVM step at an air oxygen concentration (20%) for 30 hours; all two steps combine 6% carbon dioxide at 37 degrees Celsius.

What is the study measuring?

Primary Outcome Measures

Outcome Measure	Measure Description	Time Frame
Maturation rate	The oocyte maturation rate was usually defined by MII oocyte number divided by total COCs number	Two day after oocyte retrieval

Secondary Outcome Measures

Outcome Measure	Measure Description	Time Frame
Mode of delivery	Vaginal delivery, C-section (elective, suspected fetal distress, non-progressive labor)	At birth
Total number of oocytes retrieval	Counting the number of oocytes retrieved	On the day of oocyte retrieval
Number of patients with no oocyte retrieved	Counting the number of patients with no oocyte retrieved	On the day of oocyte retrieval
Number of MII oocytes	The oocyte maturation was usually defined by MII oocyte number	Two day after oocyte retrieval
Number of GV oocytes	Counting the number of GV oocytes	Two day after oocyte retrieval
Number of patients with no matured oocyte	Counting the number of patients with no matured oocyte	Two day after oocyte retrieval
Number of 2PN oocytes	Number of zygotes with 2 pronuclei after ICSI	16-18 hours after ICSI
Fertilization rate	Number of fertilized oocytes / number of oocytes inseminated	16-18 hours after ICSI
Abnormal fertilization rate	The percentage of zygotes with 1,3, or 4 pronuclei after ICSI / number of oocytes inseminated	16-18 hours after ICSI
Number of patients with no day-3 embryo	Counting the number of patients with no embryo	Five day after oocyte retrieval
Number of day-3 embryos	Counting the number of day-3 embryos at 64±2h after ICSI	Three days after intra-cytoplasmic sperm injection
Number of good quality Day-3 embryos	Number of grade 1 and grade 2 day-3 embryos	Three days after intra-cytoplasmic sperm injection
Number of frozen day-3 embryos	Counting the numer of frozen day-3 embryos	Three days after ICSI
Number of blastocyst (day 5 or day 6 embryo)	Counting the number of blastocyst at 114±2h/140±2h after ICSI	Five or six days after ICSI
Number of patients with no blastocyst	Counting the number of patients with no blastocyst	Five or six days after ICSI
Number of good quality blastocysts	Number of grade 1 and grade 2 blastocysts	Five days after intra-cytoplasmic sperm injection
Number of frozen blastocysts	Counting the numer of frozen blastocysts	Three days after ICSI
Number of embryos transferred	Total embryos transferred	On the day of embryo transfer
Quality of embryos transferred (Grade 1, Grade 2, Grade 3)	The quality of transferred embryo is classified according to Alpha Scientists in Reproductive Medicine and ESHRE Special Interest Group of Embryology, 2011; D. Gardner, 1999; D. K. Gardner & Schoolcrati, 1999	On the day of embryo transfer
Positive pregnancy test rate	Positive pregnancy test defined as serum human chorionic gonadotropin level greater than 25 mIU/mL	11 days after the day of blastocyst transfer and 13 days after the day of day-3 embryo transfer
Implantation rate	Implantation rate is explained as the number of gestational sacs per number of embryos transferred.	At 3 weeks after embryo transfer after the completion of the embryo transfer
Clinical pregnancy rate	Diagnosed by ultrasonographic visualization of one or more gestational sacs or definitive clinical signs of pregnancy at 6 weeks or more after the onset of last menstrual period. In addition to intra-uterine pregnancy, it includes a clinically documented ectopic pregnancy	5 weeks after embryo transfer
Ectopic pregnancy rate	A pregnancy outside the uterine cavity, diagnosed by ultrasound, surgical visualization or histopathology	3 weeks after embryo transfer
Ongoing pregnancy rate	Ongoing pregnancy is defined as pregnancy with a detectable heart rate at 12 weeks' gestation or beyond.	10 weeks after embryo transfer
Miscarriage <12 weeks rate (Early miscarriage)	Spontaneous loss of pregnancy up to 12 weeks of gestation is referred to as an early	2-10 weeks after embryo transfer
Miscarriage <22 weeks rate (late miscarriage)	Spontaneous loss of pregnancy between 12 to 22 weeks is termed as late miscarriage	At >10 to 20 weeks after the transfer
Live birth rate	Live birth is defined as the complete expulsion or extraction from a woman of a product of fertilization, after 22 completed weeks of gestational age; which, after such separation, breathes or shows any other evidence of life, such as heartbeat, umbilical cord pulsation or definite movement of voluntary muscles, irrespective of whether the umbilical cord has been cut or the placenta is attached. A birth weight of 500 grams or more can be used if gestational age is unknown. Twins counted as one live birth.	At 22 weeks of gestation
Multiple pregnancy rate	Defined as the presence of more than one gestational sac at early pregnancy ultrasound (6-9 weeks gestation)	4 weeks after embryo transfer
Multiple delivery rate	Defined as the complete expulsion or extraction from a woman of more than one fetus, after 22 completed weeks of gestational age, irrespective of whether it is a live birth or stillbirth	At 22 weeks' gestation
Gestational age at birth	Calculated by gestational age of all live births	At birth
Birth weight	in grams; of singletons and twins	At the time of delivery
Very low birth weight rate	Birth weight less than 1.500 g	At birth
Low birth weight rate	Birth weight less than 2.500 g	At birth
High birth weight rate	Implies growth beyond an absolute birth weight, historically 4.000 g or 4.500 g, regardless of the gestational age	At birth
Very high birth weight rate	Birth weight over than 4.500 g for women with diabetes, and a threshold of 5000 g for women without diabetes	At birth
Small for gestational age rate	Large for gestational age was defined as a birth weight below the 10th percentile	At birth
Large for gestational age rate	Large for gestational age was defined as a birth weight above the 90th percentile	At birth
Hypertension in pregnancy rate	Comprising pregnancy-induced hypertension (PIH), pre-eclampsia (PET), eclampsia, and HELLP syndrome.	At 20 weeks of gestation or beyond
Gestational diabetes mellitus rate	Diagnosed according to the latest version of ADA guidelines. a 75-g OGTT, with plasma glucose measurement when patient is fasting and at 1 and 2 h, at 24-28 weeks of gestation in women not previously diagnosed with diabetes.; Fasting: 92 mg/dL (5.1 mmol/L); 1 h: 180 mg/dL (10.0 mmol/L); 2 h: 153 mg/dL (8.5 mmol/L)	At 24 to 28 weeks of gestation
Still birth rate	defined as the death of a fetus prior to the complete expulsion or extraction from its mother after 20 completed weeks of gestational age. The death is determined by the fact that, after such separation, the fetus does not breathe or show any other evidence of life, such as heartbeat, umbilical cord pulsation, or definite movement of voluntary muscles. It includes deaths occurring during labor	After 20 completed weeks of gestational age
Premature birth rate	Defined as delivery at <24, <28, <32, <37 completed weeks. A birth that takes place after 22 weeks and before 37 completed weeks of gestational age.	On the day of delivery
Antepartum haemorrhage rate	Defined as bleeding from or into the genital tract, occurring from 24 weeks of pregnancy and prior to the birth of the baby.	At birth
Major congenital abnormalities rate	Structural, functional, and genetic anomalies, that occur during pregnancy, and identified antenatally, at birth, or later in life, and require surgical repair of a defect, or are visually evident, or are life-threatening, or cause death. Any congenital anomaly will be included as followed definition of congenital abnormalities in Surveillance of Congenital Anomalies by Division of Birth Defects and Developmental Disabilities, NCBDDD, Centers for Disease Control and Prevention (2020).	At birth
Neonatal mortality rate	Neonatal mortality defined as the death of a live-born baby within 28 days of birth. This can be divided into early neonatal mortality, if death occurs in the first seven days after birth, and late neonatal if death occurs between eight and 28 days after delivery	between eight and 28 days after delivery
NICU admission rate	Counting number of babies admited to neonatal intensive care unit	At birth
Reason for NICU admission	Respiratory distress, Intraventricular Hemorrhage, Necrotizing enterocolitis, Sepsis	At birth

Collaborators and Investigators

• Sponsor: Mỹ Đức Hospital
• Collaborators: Vrije Universiteit Brussel
• Investigators: Principal Investigator: Lan N Vuong, MD, PhD, University of Medicine and Pharmacy at Ho Chi Minh City

Publications and helpful links

General Publications

• Rotterdam ESHRE/ASRM-Sponsored PCOS consensus workshop group. Revised 2003 consensus on diagnostic criteria and long-term health risks related to polycystic ovary syndrome (PCOS). Hum Reprod. 2004 Jan;19(1):41-7. doi: 10.1093/humrep/deh098.
• Fischer B, Bavister BD. Oxygen tension in the oviduct and uterus of rhesus monkeys, hamsters and rabbits. J Reprod Fertil. 1993 Nov;99(2):673-9. doi: 10.1530/jrf.0.0990673.
• Paulson RJ, Fauser BCJM, Vuong LTN, Doody K. Can we modify assisted reproductive technology practice to broaden reproductive care access? Fertil Steril. 2016 May;105(5):1138-1143. doi: 10.1016/j.fertnstert.2016.03.013. Epub 2016 Apr 4.
• Vuong LN, Ho VNA, Ho TM, Dang VQ, Phung TH, Giang NH, Le AH, Pham TD, Wang R, Smitz J, Gilchrist RB, Norman RJ, Mol BW. In-vitro maturation of oocytes versus conventional IVF in women with infertility and a high antral follicle count: a randomized non-inferiority controlled trial. Hum Reprod. 2020 Nov 1;35(11):2537-2547. doi: 10.1093/humrep/deaa240.
• Akin N, Ates G, von Mengden L, Herta AC, Meriggioli C, Billooye K, Stocker WA, Ghesquiere B, Harrison CA, Cools W, Klamt F, Massie A, Smitz J, Anckaert E. Effects of lactate, super-GDF9, and low oxygen tension during bi-phasic in vitro maturation on the bioenergetic profiles of mouse cumulus-oocyte complexdagger. Biol Reprod. 2023 Oct 13;109(4):432-449. doi: 10.1093/biolre/ioad085.
• Practice Committees of the American Society for Reproductive Medicine, the Society of Reproductive Biologists and Technologists, and the Society for Assisted Reproductive Technology. Electronic address: jgoldstein@asrm.org. In vitro maturation: a committee opinion. Fertil Steril. 2021 Feb;115(2):298-304. doi: 10.1016/j.fertnstert.2020.11.018. Epub 2020 Dec 24.
• Gilchrist RB, Ho TM, De Vos M, Sanchez F, Romero S, Ledger WL, Anckaert E, Vuong LN, Smitz J. A fresh start for IVM: capacitating the oocyte for development using pre-IVM. Hum Reprod Update. 2024 Jan 3;30(1):3-25. doi: 10.1093/humupd/dmad023.
• Zhao X, Liu X, Feng Y, Shi D, Lu F. Regulation of hypoxia-inducible factor 1alpha by optimal oxygen concentration enhances oocyte maturation and early embryonic development in buffalo. Theriogenology. 2023 Aug;206:50-59. doi: 10.1016/j.theriogenology.2023.05.006. Epub 2023 May 8.
• Bahrami M, Cottee PA. Culture conditions for in vitro maturation of oocytes - A review. Reproduction and Breeding. 2022 Jun 1;2(2):31-6
• Increased susceptibility to oxidative stress as a proximate cost of reproduction - Alonso-Alvarez - 2004 - Ecology Letters - Wiley Online Library [Internet]. [cited 2023 Sep 19]. Available from: https://onlinelibrary.wiley.com/doi/10.1111/j.1461-0248.2004.00594.x
• Herta AC, von Mengden L, Akin N, Billooye K, Coucke W, van Leersum J, Cava-Cami B, Saucedo-Cuevas L, Klamt F, Smitz J, Anckaert E. Characterization of carbohydrate metabolism in in vivo- and in vitro-grown and matured mouse antral folliclesdagger. Biol Reprod. 2022 Oct 11;107(4):998-1013. doi: 10.1093/biolre/ioac124.

Study record dates

Study Major Dates

• Study Start (Actual): May 13, 2024
• Primary Completion (Actual): July 11, 2024
• Study Completion (Actual): June 1, 2025

Study Registration Dates

• First Submitted: March 26, 2024
• First Submitted That Met QC Criteria: April 10, 2024
• First Posted (Actual): April 16, 2024

Study Record Updates

• Last Update Posted (Actual): June 12, 2025
• Last Update Submitted That Met QC Criteria: June 9, 2025
• Last Verified: June 1, 2025

More Information

Terms related to this study

• Keywords: PCOS; Oxygen; Culture; CAPA-IVM; Low Oxygen Tension
• Other Study ID Numbers: 03/24/DD-BVMD

Drug and device information, study documents

• Studies a U.S. FDA-regulated drug product: No
• Studies a U.S. FDA-regulated device product: No
• product manufactured in and exported from the U.S.: No