TY - JOUR
T1 - Isolation and Characterization of Functionally Active Extracellular Vesicles from Culture Medium Conditioned by Bovine Embryos In Vitro
AU - Pavani, Krishna Chaitanya
AU - Hendrix, An
AU - Van Den Broeck, Wim
AU - Couck, Liesbeth
AU - Szymanska, Katarzyna
AU - Lin, Xiaoyuan
AU - De Koster, Jenne
AU - Van Soom, Ann
AU - Leemans, Bart
PY - 2019/1/1
Y1 - 2019/1/1
N2 - Extracellular vesicles (EVs) play a possible role in cell-cell communication and are found in various body fluids and cell conditioned culture media. The aim of this study was to isolate and characterize EVs in culture medium conditioned by bovine embryos in group and to verify if these EVs are functionally active. Initially, ultracentrifuged bovine serum albumin (BSA) containing medium was selected as suitable EV-free embryo culture medium. Next, EVs were isolated from embryo conditioned culture medium by OptiPrep(TM) density gradient ultracentrifugation. Isolated EVs were characterized by nanoparticle tracking analysis, western blotting, transmission, and immunoelectron microscopy. Bovine embryo-derived EVs were sizing between 25-230 nm with an average concentration of 236.5 +/- 1.27 x 10(8) particles/mL. Moreover, PKH67 EV pre-labeling showed that embryo-secreted EVs were uptaken by zona-intact bovine embryos. Since BSA did not appear to be a contaminating EV source in culture medium, EV functionality was tested in BSA containing medium. Individual embryo culture in BSA medium enriched with EVs derived from conditioned embryo culture medium showed significantly higher blastocyst rates at day 7 and 8 together with a significantly lower apoptotic cell ratio. In conclusion, our study shows that EVs play an important role in inter embryo communication during bovine embryo culture in group.
AB - Extracellular vesicles (EVs) play a possible role in cell-cell communication and are found in various body fluids and cell conditioned culture media. The aim of this study was to isolate and characterize EVs in culture medium conditioned by bovine embryos in group and to verify if these EVs are functionally active. Initially, ultracentrifuged bovine serum albumin (BSA) containing medium was selected as suitable EV-free embryo culture medium. Next, EVs were isolated from embryo conditioned culture medium by OptiPrep(TM) density gradient ultracentrifugation. Isolated EVs were characterized by nanoparticle tracking analysis, western blotting, transmission, and immunoelectron microscopy. Bovine embryo-derived EVs were sizing between 25-230 nm with an average concentration of 236.5 +/- 1.27 x 10(8) particles/mL. Moreover, PKH67 EV pre-labeling showed that embryo-secreted EVs were uptaken by zona-intact bovine embryos. Since BSA did not appear to be a contaminating EV source in culture medium, EV functionality was tested in BSA containing medium. Individual embryo culture in BSA medium enriched with EVs derived from conditioned embryo culture medium showed significantly higher blastocyst rates at day 7 and 8 together with a significantly lower apoptotic cell ratio. In conclusion, our study shows that EVs play an important role in inter embryo communication during bovine embryo culture in group.
KW - Optiprep (TM) density gradient ultracentrifugation
KW - Bovine
KW - Embryo culture
KW - Embryo-embryo communication
KW - Extracellular vesicles
KW - uptake of embryo-secreted EVs
UR - https://www.webofscience.com/api/gateway?GWVersion=2&SrcApp=d7dz6a2i7wiom976oc9ff2iqvdhv8k5x&SrcAuth=WosAPI&KeyUT=WOS:000459747700038&DestLinkType=FullRecord&DestApp=WOS_CPL
U2 - 10.3390/ijms20010038
DO - 10.3390/ijms20010038
M3 - Article
C2 - 30577682
SN - 1661-6596
VL - 20
JO - International Journal of Molecular Sciences
JF - International Journal of Molecular Sciences
IS - 1
M1 - 38
ER -