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Cryopreservation effects on a viable sperm sterlet (Acipenser ruthenus) subpopulation obtained by a Percoll density gradient method.

ABSTRACT: In many fish species, sperm cryopreservation has deleterious effects and leads to a significant decrease in spermatozoa viability. However, the effect of cryopreservation on sperm cells that survive this process and are still viable is not fully understood. The objective of this study was to compare the viability and proteomes of fresh and cryopreserved sterlet (Acipenser ruthenus) sperm samples before and after live-dead cell separation using Percoll density gradient centrifugation. Both fresh and cryopreserved sperm samples were divided into two groups (with or without application of Percoll separation). At each step of the experiment, sperm quality was evaluated by video microscopy combined with integrated computer-assisted sperm analysis software and flow cytometry for live-dead sperm viability analysis. Sperm motility and the percentage of live cells were reduced in the cryopreserved group compared to the fresh group from 89% to 33% for percentage of motility and from 96% to 70% for live cells. Straight line velocity and linearity of track were significantly lower in cryopreserved samples than in those separated by Percoll before and after cryopreservation. However, the percentages of motile and live spermatozoa were higher than 90% in samples subjected to Percoll separation. Proteomic analysis of spermatozoa by two-dimensional differences in-gel electrophoresis coupled with matrix-assisted laser-desorption/ionization time-of-flight/time-of-flight mass spectrometry revealed that 20 protein spot abundances underwent significant changes in cryopreserved samples compared to fresh ones. However, only one protein spot was significantly altered when samples before and after cryopreservation followed by Percoll separation were compared. Thus, the results of this study show that cryopreservation leads to minimal proteomic changes in the spermatozoa population, retaining high motility and viability parameters. The results also suggest that global differences in protein profiles between unselected fresh and cryopreserved samples are mainly due to protein loss or changes in the lethal and sublethal damaged cell subpopulations.

SUBMITTER: Horokhovatskyi Y

PROVIDER: S-EPMC6095596 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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Cryopreservation effects on a viable sperm sterlet (Acipenser ruthenus) subpopulation obtained by a Percoll density gradient method.

Horokhovatskyi Yevhen Y Dietrich Mariola A MA Lebeda Ievgen I Fedorov Pavlo P Rodina Marek M Dzyuba Borys B

PloS one 20180816 8

In many fish species, sperm cryopreservation has deleterious effects and leads to a significant decrease in spermatozoa viability. However, the effect of cryopreservation on sperm cells that survive this process and are still viable is not fully understood. The objective of this study was to compare the viability and proteomes of fresh and cryopreserved sterlet (Acipenser ruthenus) sperm samples before and after live-dead cell separation using Percoll density gradient centrifugation. Both fresh ...[more]

PMID: 30114243

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Project description:Sturgeons, producers of black caviar, are one of the most valuable wildlife commodities on earth (Pikitch et al., 2005). They appeared approximately 200 million years ago, and are known as living fossils (Bemis et al., 1997). Sturgeon populations dramatically declined as a result of overfishing, poaching and habitat destruction (Birstein, 1993; Billard and Lecointre, 2001; Vidotto et al., 2013). International Union for Conservation of Nature and Natural Resources (IUCN) classified over 85% of sturgeon species as at risk of extinction, more than any other group of species. Therefore, artificial reproduction techniques have been developed for sturgeons to meet the demands of aquaculture and restocking programs. Basic knowledge regarding biology of reproduction such as egg activation and fertilization can help improving the efficiency of artificial reproduction. Fertilization happens when gametes, spermatozoon and egg, are fused together. Egg activation refers to a series of morphological and molecular changes that occur in the egg during and after fertilization. Egg activation is believed to prevent polyspermy, and may also establish a micro-environment to support embryo development (Wong and Wessel, 2006; Niksirat et al., 2015a). Although, earlier researchers have illustrated egg structure and its morphological changes during egg fertilization and activation in sturgeons (Cherr and Clark, 1982, 1985a; Linhart and Kudo, 1997; Debus et al., 2008; Zelazowska, 2010), there is still a lack of molecular knowledge to improve our understanding of this stage of reproduction in these animals. Eggs of sturgeon are released to aqueous environment, where they are fertilized, activated and develop an adhesive layer after contact with freshwater (Cherr and Clark, 1984; Vorobeva and Markov, 1999). Although, egg stickiness is necessary for eggs in nature to help them find a stable position by attaching to substrates such as gravel and stone and increase chance of survival until hatching (Riehl and Patzner, 1998), but can cause high rate of mortality during artificial incubation as a result of attachment of eggs together and subsequent suffocation in limited space of incubators. Using clay suspension in water as an activation medium has been proven to be effective to block egg stickiness in sturgeon eggs (Dettlaff et al., 1993). In recent years, label-free protein quantification techniques have been developed based on the presumption of linear proportionality between peptide mass peak signal intensity and concentration of a given peptide. This approach avoids the limitations and multiple preparation steps of methods such as those based on two-dimensional gel electrophoresis (Niksirat et al., 2015b). The aim of present study was to identify and quantify proteome changes of sterlet eggs during activation in different activation media, water and clay suspension, using label-free protein quantification technique.

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Dataset Information

Cryopreservation effects on a viable sperm sterlet (Acipenser ruthenus) subpopulation obtained by a Percoll density gradient method.

Publications

Cryopreservation effects on a viable sperm sterlet (Acipenser ruthenus) subpopulation obtained by a Percoll density gradient method.

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