Biocompatible nanoparticle PLGA is a noble safe delivery system for embryo development and next generations

Nano-sized particles (NPs) of various materials have been extensively utilized as therapeutic and diagnostic agents, drug delivery systems and biomedical devices. Previous studies showed that biodegradable poly (Poly lactic-co-glycolic acid) (PLGA) NPs complexed with cationic polymer of poly (ethyleneimine) (PEI) (PEI-PLGA NPs) are non-toxic and biocompatible, and are suitable for in vivo gene delivery applications. However, it has not been investigated whether NPs are practically usable to deliver nucleotides or proteins to mammalian oocytes and embryos. Furthermore, toxicity of these nanoparticles to embryo development and further reproduction has not been paid attention. We evaluated reproductive toxicity of PEI-PLGA NPs as a new delivery system to mammalian embryos. A superovulation regime was given to immature female mice to induce ovulation. The embryos at various stages were flushed from oviducts or uteri and cultured in KSOM. Embryos were treated with TRITC-labeled PEI-PLGA NPs (110±10 nm size) in serum-free media. The images were acquired using a confocal laser scanning microscope. TUNEL assay and OCT4 immunostaining were performed to investigate apoptosis and the number of inner cell mass (ICM) at blastocyst stage after 2-cell embryos were treated with PEI-PLGA NPs. Two-cell embryos exposed to TRITC PEI-PLGA NPs were transferred into pseudopregnant mice to evaluate post-implantation development and live birth of embryos exposed to NPs. Genetic and morphologic characteristics of live pups were evaluated and long-term toxicity of NPs in reproduction was also examined. Time-lapse confocal microscopic images showed that TRITC fluorescence quickly accumulated and the maximum levels were detected within 60 min after NPs treatment. Treatment of TRITC PEI-PLGA NPs for 4 h in serum free condition was enough to visualize successful delivery of NPs into oocytes and embryos and did not compromise embryo development to the blastocyst stage. Furthermore, treatment of TRITC PEI-PLGA NPs did not have any significant effect on apoptosis and the number of ICM. Two-cell embryos transferred into recipients after NP exposure were born healthy without any distinct morphologic and genetic abnormalities. They showed normal growth rates and regular reproductive cycles, and produced similar number of pups. PEI-PLGA NPs can be used to develop a new safe carrier to introduce exogenous materials into mouse embryos without apparent toxicity.