Effect of culture media on porcine embryos produced by in vitro fertilization or parthenogenetic activation after oocyte maturation with cycloheximide

Abstract

This study evaluated the effects of reversible meiotic inhibition and different culture media (PZM3 or NCSU23) on production of porcine embryos by either in vitro fertilization (IVF) or parthenogenetic activation (PA). Oocytes from abattoir-derived ovaries were allocated into two groups for maturation: CHX (5 μg/ml cycloheximide for 10 h) or Control (no CHX). The percentage of metaphase II (MII) oocytes was determined at 36, 40 or 44 h of in vitro maturation. For IVF and PA, denuded oocytes were fertilized with purified sperm for 6 h or activated by electric stimuli. Zygotes were then subdivided into two culture groups: NCSU23 or PZM3. No effect of treatment with CHX and culture media was observed on cleavage (D3) and blastocyst (D7) rates in IVF and PA groups. There are no differences of quality or development rates between IVF-derived embryos cultured in NCSU23 or PZM3. However, we observed high quality PA embryos in PZM3 compared with NCSU23. Maturation arrest with CHX decreased the average blastocyst cell number in IVF while it was increased in PA embryos. As older oocytes are more effectively activated, CHX- blocked oocytes reached the mature stage faster than the control group. In conclusion, the CHX treatment for 10 h, followed by oocyte maturation for 40 h, is an efficient protocol to produce high quality parthenote embryos, especially when they are cultured in PZM3. However, this protocol is not satisfactory for IVF embryos production. In this case, a shorter maturation period could provide better embryo quality.

Modern achievements of the development of scientific research at obtaining in vitro sheep embryos

“…Однак результати виявили суперечливий вплив цих інгібіторів на зупинку мейотичного клітинного циклу, швидкість дозрівання і пізніше на потенціал розвитку. Крім того, деякі інгібітори, такі як СНХ, позитивно впливають на зупинку мейотичного клітинного циклу впродовж 24 год і на потенціал розвитку ооцитів окремих видів, включаючи свиней та велику рогату худобу (Saeki et al, 1997;Marques et al, 2011). Отже, використання інгібіторів може не тільки підвищити IVP овець, а й забезпечити транспортування ооцитів на велику відстань та гнучкий час початку IVM ооцитів.…”

Advances in in vitro production of sheep embryos

“…Nonetheless, agents that modulate oocyte cAMP during IVM showed greater potential, possibly as these compounds could extend oocyte-cumulus cells gap- junctional communication [36], [66]. Additionally, some inhibitors such as CHX seem to have a positive effect on meiotic cell cycle arrest within 24 h and on developmental potential in some species including porcine and bovine [67], [68]. Therefore, use of the inhibitors might pave not only the way for improving ovine IVP, but also would benefit researchers with a flexible alternative in working practice without compromising developmental competence of oocytes, such as oocyte transportation for a long distance and a flexible starting time for oocyte IVM in 24 h. Nevertheless, the similar reports on ovine IVM are few compared to those in bovine.…”

Meiotic arrest as an alternative to increase the production of bovine embryos by somatic cell nuclear transfer

“…However, it can be very useful for routine SCNT by providing matured oocytes different times, allowing more flexibility in manipulations, and thus, enabling the use of the oocytes from a single batch of ovaries in 2 or more consecutive sets of micromanipulation. To block the spontaneous resumption of meiosis, various pharmacological substances have been used in the bovine oocyte, such as 6-dimethylaminopurine (6-DMAP), which acts on the meiosis promoting factor (MPF) and specific inhibitors of cyclin-dependent kinases (CDKs), such as butyrolactone-I and roscovitine (Dode & Adona, 2001; Adona & Leal, 2004; Adona et al , 2008; Barretto et al , 2011; Marques et al , 2011). More recently, inhibitors of phosphodiesterase type 3 A (PDE-3A), which are specific to the oocytes (Sasseville et al , 2009), such as cilostamide and C-type natriuretic peptide (NPPC), have been extensively used for the pre-maturation of bovine oocytes to be used in IVEP, in an attempt to increase the oocyte and embryonic development competence (Albuz et al , 2010; Dieci et al , 2013; Franciosi et al , 2014; De Cesaro et al , 2015; Farghaly et al , 2015; Guimaraes et al , 2015; Guimaraes et al , 2016).…”