Preliminary observations in in vitro development of equine embryo after ICSI.

This research explores the effect of intracytoplasmic sperm injection (ICSI) on in vitro matured equine oocytes and the potential for improved embryonic development with the activation of these oocytes using a calcium ionophore. The findings suggest a high survival rate of injected oocytes and successful fertilization, also resulting in early embryonic development.

Introduction and Methodology

• This study aimed to explore the development potential of equine embryos in vitro after undergoing intracytoplasmic sperm injection (ICSI). ICSI, a procedure wherein a single sperm cell is injected directly into the egg, is often used to overcome fertility issues in various animals.
• Here, matured equine oocytes (eggs) were subjected to ICSI and activated using a calcium ionophore, a compound used to stimulate cellular processes allowing for improved embryonic development.
• The cumulus-oocyte complexes, which represent mature eggs surrounded by cells that aid in their development, were used for this process. These were matured, denuded (removed of surrounding cells), and centrifuged. The oocytes showing the first polar body, indicating maturity, were then targeted for ICSI.
• The ICSI was carried out using fresh semen obtained from three fertile stallions.
• Post-ICSI, the oocytes were activated with a certain concentration of calcium ionophore A23187 and then individually cultured for seven days in a specific environment that encourages growth.

Results

• Out of 353 matured cumulus-oocyte complexes, 103 oocytes were microinjected and eight oocytes were sham microinjected (a control process where the oocytes were not truly injected). Among the microinjected, 85 oocytes (or 82.5%) survived the injection process, suggesting a high success rate of the ICSI procedure.
• Of the 76 successfully microinjected oocytes, a large majority (around 68%) were fertilized. An indication of successful fertilization included the presence of two pronuclei, syngamy stage (a stage where male and female pronuclei have fused), and cleaved ova (fertilized eggs that have started dividing).
• The sham microinjected oocytes were not activated, serving as a control group in the experiment’s methodology.
• Later, an approximately 46% of the ova showed signs of division or cleaving two days after ICSI. Also, three embryos showed early signs of development with a visible increase in cell count five to seven days post-ICSI.