Hypotaurine Supplementation Benefits in Cryopreservation

Completed

• Conditions: Human Spermatozoa Parameters; Hypotaurine Supplementation; Cryopreservation; Spermatozoa DNA Alterations

• Registration Number: NCT04011813
• Lead Sponsor: University Hospital, Clermont-Ferrand

Brief Summary

Although it is widely used, slow freezing can induce strong functional and nuclear spermatic alterations reducing the chances of pregnancy. The study objective is to determinate the effects of the combination of hypotaurine supplementation and spermatozoa selection by Density Gradient Centrifugation (DGC) on human sperm functions and DNA quality during a freezing-thawing cycle.

Detailed Description

This prospective study was performed on surplus semen after a density gradient centrifugation-frozen-thawing cycle. Samples were obtained from men undergoing routine semen analysis at the Center for Reproductive Medicine. Spermatozoa were selected by density gradient centrifugation, washed and frozen using a programmable device. Each step was performed in parallel with (H+ arm) or without (H- arm) 50mM hypotaurine supplementation. After thawing, investigator team compared for both conditions the total and progressive mobility, vitality, integrity of the acrosome, markers of Protein Kinase A (PKA) dependent capacitation intracellular signaling pathway and nuclear quality by measuring chromatin packaging, DNA fragmentation and oxidation and vacuoles presence in the spermatozoa head.

Study Timeline

• Study Start: 2014-05-01
• Primary Completion: 2018-08-01
• Study Completion: 2019-05-01
• Status Verified: 2019-06
• Study First Submitted: 2019-06-21
• Study First Submitted QC: 2019-07-03
• Last Update Submitted: 2019-07-03
• Study First Posted: 2019-07-09
• Last Update Posted: 2019-07-09

Recruitment & Eligibility

• Status: COMPLETED
• Sex: Male
• Target Recruitment: 33

Inclusion Criteria

• None

Exclusion Criteria

• None

Study & Design

• Study Type: OBSERVATIONAL
• Study Design: Not specified

Primary Outcome Measures

Name	Time	Method
DNA fragmentation using TUNEL assay	Day 0	Comparison after a cycle of freezing-thawing for both conditions, with or without hypotaurine supplementation of DNA fragmentation using TUNEL assay
chromatin packaging labelled using aniline blue and chromomycin A3	Day 0	Comparison after a cycle of freezing-thawing for both conditions, with or without hypotaurine supplementation of chromatin packaging labelled using aniline blue and chromomycin A3
DNA oxidation assessed by 8-OHdG immunodetections	Day 0	Comparison after a cycle of freezing-thawing for both conditions, with or without hypotaurine supplementation of DNA oxidation assessed by 8-OHdG immunodetections
vacuoles presence in the spermatozoa head using Motile Sperm Organelle Morphology Examination (MSOME)	Day 0	Comparison after a cycle of freezing-thawing for both conditions, with or without hypotaurine supplementation of vacuoles presence in the spermatozoa head using Motile Sperm Organelle Morphology Examination (MSOME)

Secondary Outcome Measures

Name	Time	Method
vitality using Eosin Nigrosin	Day 0	Comparison after a cycle of freezing-thawing for both conditions, with or without hypotaurine supplementation of vitality using Eosin Nigrosin
markers of PKA-dependent capacitation intracellular signaling pathway assessing western blot	Day 0	Comparison after a cycle of freezing-thawing for both conditions, with or without hypotaurine supplementation of markers of PKA-dependent capacitation intracellular signaling pathway assessing western blot
integrity of the acrosome using Fluorescein IsoThioCyanate-Pisum Sativum Agglutinin (FITC-PSA) labelling	Day 0	Comparison after a cycle of freezing-thawing for both conditions, with or without hypotaurine supplementation of integrity of the acrosome using Fluorescein IsoThioCyanate-Pisum Sativum Agglutinin (FITC-PSA) labelling
motility total and progressive	Day 0	Comparison after a cycle of freezing-thawing for both conditions, with or without hypotaurine supplementation of motility total and progressive