Effects of Porcine Immature Oocyte Vitrification on Actin Microfilament Distribution and Chromatin Integrity During Early Embryo Development in vitro

Abstract

Vitrification is mainly used to cryopreserve female gametes. This technique allows maintaining cell viability, functionality, and developmental potential at low temperatures into liquid nitrogen at −196°C. For this, the addition of cryoprotectant agents, which are substances that provide cell protection during cooling and warming, is required. However, they have been reported to be toxic, reducing oocyte viability, maturation, fertilization, and embryo development, possibly by altering cell cytoskeleton structure and chromatin. Previous studies have evaluated the effects of vitrification in the germinal vesicle, metaphase II oocytes, zygotes, and blastocysts, but the knowledge of its impact on their further embryo development is limited. Other studies have evaluated the role of actin microfilaments and chromatin, based on the fertilization and embryo development rates obtained, but not the direct evaluation of these structures in embryos produced from vitrified immature oocytes. Therefore, this study was designed to evaluate how the vitrification of porcine immature oocytes affects early embryo development by the evaluation of actin microfilament distribution and chromatin integrity. Results demonstrate that the damage generated by the vitrification of immature oocytes affects viability, maturation, and the distribution of actin microfilaments and chromatin integrity, observed in early embryos. Therefore, it is suggested that vitrification could affect oocyte repair mechanisms in those structures, being one of the mechanisms that explain the low embryo development rates after vitrification.

Unveiling how vitrification affects the porcine blastocyst: clues from a transcriptomic study

“…To date, vitrification is the most efficient and widely used method in pig embryo cryopreservation, particularly the ultra–rapid vitrification systems [ 11 , 25 – 29 ]. Vitrification has been successfully used on embryos at early stages of development [ 14 , 30 ] and also on oocytes [ 20 , 31 , 32 ]. However, the lower pregnancy rates achieved for vitrified embryos (ranging from 39% to 75%) when compared to fresh ones [ 24 , 33 ], the need of highly qualified personnel for vitrification and surgical embryo transfer to achieve high pregnancy and prolificacy rates, represent major hurdles for pig embryo cryopreservation.…”

Oocyte Cryopreservation in Domestic Animals and Humans: Principles, Techniques and Updated Outcomes

“…Modifications of vitrification devices, the type of cryoprotectant and the procedure used were tested, aiming at reducing cryoinjuries and cellular changes of the vitrified-warmed oocytes. Notably, vitrification induces several changes within porcine oocytes, such as in the cytoskeleton [100], mitochondrial abnormalities [101], epigenetic changes [102] and decreased expression level of the Type 1 inositol 1,4,5 trisphosphate receptor [103]. More recently, vitrification of porcine oocytes has been associated with alterations at transcriptomic and proteomic levels [37,104].…”

Antioxidant procyanidin B2 protects oocytes against cryoinjuries via mitochondria regulated cortical tension

“…Vitrification is an ultrafast cooling method performed with a very high concentration cryoprotectant for dehydration to avoid the formation of ice crystals during oocyte cryopreservation, which ultimately leads to higher cell survival, fertilization, embryo development and pregnancy rates compared with traditional slow-freezing [3][4][5][6]. However, studies reveal that impaired oolemma permeability [5], oxidative stress [7] and the toxicity of cryoprotectants during vitrification [8] can still cause damages to the oocytes [9,10], especially deteriorating cytoskeleton [11] and mitochondrial function [12,13], which eventually obstruct subsequent embryo development and need further study [14][15][16]. It is reported that mechanical properties play a pivotal role in oocyte development [17].…”