Pretreating porcine sperm with lipase enhances developmental competence of embryos produced by intracytoplasmic sperm injection

Abstract

Intracytoplasmic sperm injection (ICSI) has been widely applied in humans, mice, and some domestic animals to cure human infertility, or produce genetically superior or genetically engineered animals. However, the production efficiency of ICSI in pigs remains quite low. In this study, we developed a new sperm pretreatment method to improve production efficiency of ICSI in pigs. Experiment 1 revealed that pretreating porcine sperm with 2.5 mg/ml lipase before ICSI operation, not only can reduce the adhesion between sperm and the injection pipette without adding polyvinylpyrrolidone (PVP) in the operating medium, but also significantly improve male pronuclei (MPN) formation rate (55.56% vs. 40.00% (0 mg/ml), 42.59% (5.0 mg/ml), 40.00% (10.0 mg/ml), P < 0.05) and enhance developmental competence of ICSI embryos (26.03% vs. 10.87% (0 mg/ml), 10.00% (5.0 mg/ml), 10.13% (10.0 mg/ml), P < 0.05). Experiment 2 showed that this method has a higher MPN formation rate (50.47% vs. 30.78%, P < 0.05) and blastocyst rate (18.81% vs. 7.41%, P < 0.05) than the PVP method, and was better than the Triton X-100 treatment method (50.47% vs. 46.23%, 18.81% vs. 12.75%). Therefore, pretreating porcine sperm with 2.5 mg/ml lipase before ICSI operation is highly recommended, instead of adding PVP in the operating medium.

An efficiency comparison of different in vitro fertilization methods: IVF, ICSI, and PICSI for embryo development to the blastocyst stage from vitrified porcine immature oocytes

“…Compared to IVF and PICSI, during ICSI, PVP is often used for sperm manipulation decreasing motility and facilitating capture. However, it was reported that PVP could be toxic for the spermatozoa [ 47 ], reducing fertilization [ 27 ], male PN formation and blastocyst development [ 48 ]. These observations agree with our results, where the PVP exposed spermatozoa displayed a higher proportion of D/AR sperm than those exposed to HA and control, suggesting that the HA does not affect sperm viability.…”

“…In the present study, the use of HA increased 2PN formation up to 45%. Recently, it was reported that the use of lipase to select and inject oocytes, resulted in 29% of 2PN formation [ 48 ]. In addition, this study also evaluated the efficiency of PVP, reporting 24% of 2PN formation [ 48 ], similar to those obtained in the present study (35%).…”

“…Recently, it was reported that the use of lipase to select and inject oocytes, resulted in 29% of 2PN formation [ 48 ]. In addition, this study also evaluated the efficiency of PVP, reporting 24% of 2PN formation [ 48 ], similar to those obtained in the present study (35%). Fertilization and ED success depends on the quality of the spermatozoa selected for injection, but this is not possible to achieve by ICSI.…”

Efecto de la vitrificación en la fertilización y desarrollo embrionario in vitro de ovocitos inmaduros porcinos

“…Hasta el momento, no se ha reportado el daño en uniones comunicantes en COCs porcinos Los resultados mostraron que los espermatozoides seleccionados con AH tuvieron mayores tasas de viabilidad y reacción acrosomal (V/RA) en comparación con el grupo control y con aquellos expuestos a PVP. Estos resultados coinciden con los reportados anteriormente porWei et al, (2016). Los espermatozoides que fueron expuestos a PVP presentaron altas tasas de espermatozoides muertos que estaban reaccionados M/RA comparados con los seleccionados con AH y el control, lo que indica que el AH puede ser otro método que reemplace el uso de la PVP.…”

“…Por lo anterior, se evaluaron los porcentajes de viabilidad y estado acrosomal de los espermatozoides expuestos a la PVP o seleccionados con AH (Experimento 3). con lo anterior, se reportó que la PVP es capaz de disminuir la formación de PN, así como el desarrollo de blastocistos(Wei et al, 2016). La PVP en el procedimiento de ICSI, es tóxica para el espermatozoide, lo que reduce su capacidad fertilizante (Kato y Nagao., 2012).…”