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Comparison of fertilization percentage, embryo quality, pregnancy and implantation rate, between two sperm separation procedures (density gradient centrifugation with surface markers- density gradient centrifugation) in infertile men intra-cytoplasmic sperm injection candidate

Protocol summary

Summary
Objectives: Comparison of fertilization, embryo quality, pregnancy and implantation rates between two sperm separation procedures, density gradient centrifugation (DGC) and Magnetic-Activated Cell Sorting (MACS), followed DGC (MACS-DGC) in infertile men candidates for intra-cytoplasmic sperm injection (ICSI). Design: Infertile couples with primary male factor who referred to Isfahan fertility and infertility center infertility were included in this study. These individuals were informed about the study and were grouped into study (MACS-DGC) and control (DGC). Setting and conduct: In the day of ICSI operation, a total of 113 infertile individuals were randomly assigned (in a 1:1 proportion) to MACS-DGC and DGC groups. In study group, the oocytes were inseminated with sperm selected by MACS-DGC processing. In DGC group, oocytes were inseminated with sperm selected by DGC method (based on viability and morphology). Major Inclusion and Exclusion criteria: Inclusion criteria: Primary male factor infertility (according to the WHO criteria, 2010); with mean age of 20-45. Exclusion criteria: Females factor infertility based on WHO criteria (2010); age higher than 35 years; less than 6 oocytes and poor quality oocyte. Intervention: In study group, all oocytes were inseminated with sperm selected with DGC following MACS processing (separation of intact spermatozoa from those which present apoptotic markers), while in the control group, only routine sperm preparation procedure (DGC) were used. Main outcome measures (variables): In this study, fertilization, embryo quality, implantation and pregnancy rates were compared between two groups. The number of embryos transferred to each patient on day 3 was based on female age, number of previous failed cycles and quality of embryos at the time of embryo transfer and final decision made by the gynecologist. Clinical pregnancy was defined by ultrasonography findings showing at least one embryo with a fetal heartbeat, 5 weeks after transfer. Implantation rate was defined by the number of observed gestational sacs per number of transferred embryos.

General information

Acronym
IRCT registration information
IRCT registration number: IRCT201610317223N8
Registration date: 2017-08-22, 1396/05/31
Registration timing: retrospective

Last update:
Update count: 0
Registration date
2017-08-22, 1396/05/31
Registrant information
Name
Mohammad Hossein Nasr-Esfahani
Name of organization / entity
Royan Institute (Royan Institute for Biotechnology)
Country
Iran (Islamic Republic of)
Phone
+98 31 9501 5688
Email address
mh_nasr@med.mui.ac.ir
Recruitment status
Recruitment complete
Funding source
Royan Institute/Isfahan Fertility and Infertility Center
Expected recruitment start date
2015-04-21, 1394/02/01
Expected recruitment end date
2016-03-10, 1394/12/20
Actual recruitment start date
empty
Actual recruitment end date
empty
Trial completion date
empty
Scientific title
Comparison of fertilization percentage, embryo quality, pregnancy and implantation rate, between two sperm separation procedures (density gradient centrifugation with surface markers- density gradient centrifugation) in infertile men intra-cytoplasmic sperm injection candidate
Public title
Comparison of fertilization percentage, embryo quality, implantation and pregnancy rate, between two sperm separation procedures in infertile men intra-cytoplasmic sperm injection candidate
Purpose
Health service research
Inclusion/Exclusion criteria
Inclusion criteria: Primary male factor infertility (according to the WHO-2010); with mean age of 20-45. Exclusion criteria: Females factor infertility based on WHO criteria (2010); age higher than 35 years; less than 6 oocytes and poor quality oocyte
Age
From 20 years old to 45 years old
Gender
Both
Phase
N/A
Groups that have been masked
No information
Sample size
Target sample size: 113
Randomization (investigator's opinion)
Randomized
Randomization description
Blinding (investigator's opinion)
Single blinded
Blinding description
Placebo
Not used
Assignment
Parallel
Other design features

Secondary Ids

empty

Ethics committees

1

Ethics committee
Name of ethics committee
Ethical Committee of Royan Institute
Street address
Royan Institute, Banihashem St, Resalat Ave, Tehran, Iran
City
Tehran
Postal code
Approval date
2017-01-07, 1395/10/18
Ethics committee reference number
IRI.ACECR.ROYAN.REC.1395.156

Health conditions studied

1

Description of health condition studied
Male factor infertility
ICD-10 code
N46
ICD-10 code description
Male infertility

Primary outcomes

1

Description
Fertilization rate
Timepoint
16-18 h post insemination/injection
Method of measurement
View of 2 pronucleuse with invert microscope,the ratio of fertilized oocytes (2PN) by the total number of injected metaphase II oocytes, multiplied by 100

2

Description
Implantation rate
Timepoint
4 weeks after transfer
Method of measurement
The number of observed gestational sacs per number of transferred

3

Description
Chemical pregnancy
Timepoint
18 days post-ICSI
Method of measurement
Measurement of Beta HCG level in serum

4

Description
Clinical pregnancy
Timepoint
5 weeks after embryo transfer
Method of measurement
ultrasonography findings showing at least one embryo with a fetal heartbeat

5

Description
Quality of embryo
Timepoint
On day 3 after oocyte retrieval
Method of measurement
Using a three-point scoring system including fragmentation rate, number of blastomere, shape and size

Secondary outcomes

1

Description
Perinatal mortality ( fetal or neonatal death)
Timepoint
At 20 completed weeks of gestational age and later, during childbirth, or up to 7 completed days after birth
Method of measurement
Through ultrasound

2

Description
Miscarriage (The spontaneous loss of a clinical pregnancy )
Timepoint
Before 20 completed weeks of gestational age (18 weeks after fertilization)
Method of measurement
Through ultrasound

Intervention groups

1

Description
In study group, total of oocytes were inseminated with sperm that selected with DGC following MACS processing (Separate intact spermatozoa from those that present apoptotic markers by magnetic particles conjugated with Annexin V, which has a high affinity for Phosphatidylserine)(MACS-DGC group). For MACS procedure, semen samples were washed twice with 1X buffer and diluted to ten million sperm per ml. Then, each semen sample was mixed with one hundred microliter of microbeads solution and then, this mixture was incubated for 15 min at room before being loaded to MACS column. Annexin positive sperm was retained within the column while annexin negative sperm were allowed to pass the column.
Category
Other

2

Description
In control group (DGC group), oocytes were inseminated with sperm selected by Density gradient centrifugation method (based on viability and morphology). Density gradient centrifugation was prepared by laying 2 ml of 80% PureSperm into a sterile 15ml conical centrifuge tube. Then, 2 ml 40% PureSperm suspension was slowly layered over lower layer. Subsequently, semen sample were washed (5 min, 3000 rpm) with VitaSperm medium were overlayed on the 40% or upper layer. The layer was centrifuged for 20 min at 1000rpm. The supernatant was aspirated, pellet was resuspended, and washed twice with VitaSperm. After injection of sperm into the oocyte to embryo formation at day 3, events such as fertilization, cleavage and embryo formation occur that these phenomena are in vitro. After 3 days, embryos were transferred into the uterus and then assessed implantation and pregnancy in vivo condition.
Category
Other

Recruitment centers

1

Recruitment center
Name of recruitment center
Isfahan Fertility and Infertility Center
Full name of responsible person
Mohammad Hossein Nasr- Esfahani
Street address
Fertility and Infertility Center, Salman Farsi St, Isfahan, Iran
City
Isfahan

Sponsors / Funding sources

1

Sponsor
Name of organization / entity
Royan Institute
Full name of responsible person
Abdolhossein Shahverdi
Street address
Royan Institute, Banihashem St, Resalat Ave, Tehran, Iran
City
Tehran
Grant name
Grant code / Reference number
Is the source of funding the same sponsor organization/entity?
Yes
Title of funding source
Royan Institute
Proportion provided by this source
Public or private sector
empty
Domestic or foreign origin
empty
Category of foreign source of funding
empty
Country of origin
Type of organization providing the funding
empty

2

Sponsor
Name of organization / entity
Isfahan Fertility and Infertility
Full name of responsible person
Mohammad Hossein Nasr- Esfahani
Street address
Fertility and Infertility Center, Salman Farsi St, Isfahan, Iran
City
Isfahan
Grant name
Grant code / Reference number
Is the source of funding the same sponsor organization/entity?
Yes
Title of funding source
Isfahan Fertility and Infertility
Proportion provided by this source
Public or private sector
empty
Domestic or foreign origin
empty
Category of foreign source of funding
empty
Country of origin
Type of organization providing the funding
empty

Person responsible for general inquiries

Contact
Name of organization / entity
Royan Institute (Royan Institute for Biotechnology)
Full name of responsible person
Mohammad Hossein Nasr-Esfahani
Position
Professor
Other areas of specialty/work
Street address
Royan Institute for Biotechnology, Royan St, Salman St, KHorasgan, Isfahan, Iran
City
Isfahan
Postal code
Phone
+98 31 9501 5688
Fax
+98 31 9501 5687
Email
mh_nasr@med.mui.ac.ir
Web page address

Person responsible for scientific inquiries

Contact
Name of organization / entity
Royan Institute (Royan Institute for Biotechnology)
Full name of responsible person
Mohammad Hossein Nasr-Esfahani
Position
Professor
Other areas of specialty/work
Street address
Royan Institute for Biotechnology, Royan St, Salman St, KHorasgan, Isfahan, Iran
City
Isfahan
Postal code
Phone
+98 31 9501 5688
Fax
Email
mh_nasr@med.mui.ac.ir
Web page address

Person responsible for updating data

Contact
Name of organization / entity
Royan Institute (Royan Institute for Biotechnology)
Full name of responsible person
Mohammad Hossein Nasr- Esfahani
Position
Professor
Other areas of specialty/work
Street address
Royan Institute for Biotechnology, Royan St, Salman St, KHorasgan, Isfahan, Iran
City
Isfahan
Postal code
Phone
+98 31 9501 5688
Fax
Email
mh.nasr-esfahani@royaninstitute.org
Web page address

Sharing plan

Deidentified Individual Participant Data Set (IPD)
empty
Study Protocol
empty
Statistical Analysis Plan
empty
Informed Consent Form
empty
Clinical Study Report
empty
Analytic Code
empty
Data Dictionary
empty
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