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Selected Publication:

Type of publication: Journal Article
Type of document: Full Paper

Year: 2011

Authors: Kuzmany, A; Havlicek, V; Brem, G; Walter, I; Besenfelder, U

Title: Assessment of actin cytoskeleton and nuclei in bovine blastocysts developed under different culture conditions using a novel computer program.

Source: Reprod Domest Anim. 2011; 46(1):e46-e53

Authors Vetmeduni Vienna:

Besenfelder Urban
Brem Gottfried
Havlicek Vitezslav
Kuzmany Anna
Walter Ingrid

Vetmed Research Units
Institute of Animal Breeding and Genetics
Institute of Morphology
Institute of Animal Breeding and Genetics, Unit of Reproductive Biology

This study was performed to investigate the effects, in terms of nuclear material and actin cytoskeleton quantities (fluorescent pixel counts), of four different bovine blastocyst culturing techniques (in vitro, stepwise in vitro-to-in vivo, or purely in vivo). Cumulus oocyte complexes from abattoir-sourced ovaries were matured in vitro and allocated to four groups: IVP-group embryos developed up to blastocyst stage in vitro. Gamete intra-fallopian transfer (GIFT)-group oocytes were co-incubated with semen for 4 h before transfer to oviducts of heifers. Following in vitro fertilization, cleaved embryos (day 2 of embryo development, day 2-7 group) were transferred into oviducts on day 2. Multiple ovulation embryo transfer (MOET)-group embryos were obtained by superovulating and inseminating heifers; the heifers" genital tracts were flushed at day 7 of blastocyst development. Within each group, ten blastocysts were selected to be differentially dyed (for nuclei and actin cytoskeleton) with fluorescent stains. A novel computer program (ColorAnalyzer) provided differential pixel counts representing organelle quantities. Blastocysts developed only in vivo (MOET group) showed significantly more nuclear material than did blastocysts produced by any other technique. In terms of actin cytoskeleton quantity, blastocysts produced by IVP and by day 2-7 transfer did not differ significantly from each other. Gamete intra-fallopian transfer- and MOET-group embryos showed significantly larger quantities of actin cytoskeleton when compared with any other group and differed significantly from each other. The results of this study indicate that culturing under in vitro conditions, even with part time in vivo techniques, may adversely affect the quantity of blastocyst nuclear material and actin cytoskeleton. The software employed may be useful for culture environment evaluation/developmental competence assessment.

Keywords Pubmed: Actins/analysis*
Cell Nucleus/ultrastructure*
Embryo Culture Techniques/methods
Embryo Culture Techniques/veterinary*
Embryo Transfer/methods
Embryo Transfer/veterinary
Fertilization in Vitro/veterinary
Gamete Intrafallopian Transfer/veterinary
Insemination, Artificial/veterinary
Tissue and Organ Harvesting/methods
Tissue and Organ Harvesting/veterinary

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