title: Human spermatozoa vitrified in the absence of permeable cryoprotectants: birth of two healthy babies.
abstract: Herein, we report the birth of two healthy babies to a woman following intracytoplasmic sperm injection (ICSI) using motile spermatozoa vitrified without permeable cryoprotectants. Spermatozoa (in a case of oligoasthenoteratozoospermia) were cooled in cut standard straws in human tubal fluid supplemented with 0.5% human serum albumin and 0.25 M sucrose. Sperm motility, capacitation-like changes, acrosome reaction and mitochondrial membrane potential (MMP) were compared in fresh and vitrified spermatozoa. Eight mature (MII) oocytes were microinjected with the vitrified-warmed motile spermatozoa. Although the motility of vitrified-warmed spermatozoa was markedly lower than that of fresh spermatozoa (60% v. 90%, respectively), there were no immediate visible differences in the percentages of capacitated and acrosome-reacted vitrified and fresh spermatozoa (10% v. 8% and 5% v. 8%, respectively). However, the MMP in vitrified spermatozoa was apparently adversely affected in the ejaculate used for ICSI compared with fresh spermatozoa (63% v. 96% spermatozoa with high MMP). Eighteen hours later, six oocytes showed signs of normal fertilisation. Two-pronuclear oocytes were cultured in vitro for 24h and two four-blastomere embryos were transferred. Two healthy girls were born at term. Our findings suggest that permeable cryoprotectant-free vitrification can be applied successfully for some procedures in assisted reproduction, in particular in ICSI with motile vitrified spermatozoa, to achieve normal pregnancy and birth.
authors.affiliation: Department of Obstetrics and Gynaecology, University Women Hospital, Kerpener Str. 34, 50931 Cologne, Germany. anna.petrunkina@gmx.de
authors.forename: Vladimir
authors.initials: V
authors.lastname: Isachenko
authors.forename: Evgenia
authors.initials: E
authors.lastname: Isachenko
authors.forename: Anna M
authors.initials: AM
authors.lastname: Petrunkina
authors.forename: Raul
authors.initials: R
authors.lastname: Sanchez
mesh_terms: D000328:Adult; D002463:Cell Membrane Permeability / Q000187:drug effects; D003451:Cryoprotective Agents / Q000494:pharmacology; D005260:Female; D006262:Health; D006801:Humans; D007231:Infant, Newborn; D050498:Live Birth; D008297:Male; D011247:Pregnancy; D059285:Pregnancy, Twin / Q000502:physiology; D012662:Semen Preservation / Q000379:methods*; D020554:Sperm Injections, Intracytoplasmic / Q000379:methods; D013094:Spermatozoa / Q000502:physiology*; D014427:Twins; D058989:Vitrification
chemical_list: D003451:Cryoprotective Agents