The In Vitro Study Of Levonorgestrel Nanoparticle Sustained-released Composites On Effect Of Embryo Adhesion And Sperm Function

Objective and purposeThe aim is to study the effect of levonorgestrel-loaded nanoparticle composites on embryo adhesion in vitro, human sperm morphology and function. We has a preliminary evaluation on safety and effectiveness of the levonorgestrel-loaded nanoparticle polylactic acid-polystyrene composites, which affords a experimental research basis for levonorgestrel-loaded nanoparticle polylactic acid-polystyrene composites may applicated in clinical as a new intrauterine contraceptive system..Methods1、A co-culture model of Ishikawa monolayer (human endometrial carcinoma cells) and Bewo cell(human choriocarcinoma cells)spheroids was setted up.2、The effect of levonorgestrel-loaded nanoparticle composites on Bewo cell spheroids’adhesion rate and spreading function on Ishikawa cells were observed:(1) Bewo cell spheroids’adhesion rate and spreading rate were calculated after the empty biofilm and drug-loaded biofilm slow-release membrane liquid was added to the co-culture model respectively while no slow-release membrane liquid as blank control group;(2) At the same time, we detected the Ishikawa cell proliferation rate by CCK8 in each group;(3) The mRNA expression of integrin alphav by the methods of real time RT-PCR.3、The influence of levonorgestrel-loaded nanoparticle composites on human sperm morphology and function was observed:(1) The normal fertile men semen was co-cultured with empty biofilm and drug-loaded biofilm after density gradient centrifugation,and the sperm motility was detected by computer assisted sperm analysis (CASA);(2) Sperm morphology was observed under a transmission electron microscope.Results1、Compared with control group and empty biofilm group, drug-loaded biofilm group showed obvious difference in Bewo cell spheroids’adhesion rate and spreading rate as well as the expression level of mRNA of Ishikawa cells (P<0.05).But empty biofilm group had not show significant difference compared with control group respectively(P>0.05);2、Compared with control group, empty biofilm group and drug-loaded biofilm group had no marked difference in proliferation of Ishikawa cells (P>0.05);3、Compared with control group, no significant differences were observed about sperm motility as well as viability in empty biofilm group and drug-loaded biofilm group(P>0.05).And empty biofilm group did not show obvious difference in sperm ultrastructure;4、Remarkable differences were observed in drug-loaded biofilm group including sperm acrosome wave variation, deficiency or disappearance of acrosome, dissolution of plasma membrane, vacuolar degeneration of mitochondria and a significant increase about sperm apoptosis and disintegration.Conclusions1、Polylactic acid-polystyrene biodegradable materials (empty biofilm) itself has no effect on embryo adhesion in vitro and proliferation of human endometrial cells.The material itself has a certain biological safety;2、Levonorgestrel-loaded nanoparticle polylactic acid-polystyrene composites (drug-loaded biofilm) affect the embryo adhesion in vitro, which reveal obvious anti embryo implantation function;3、Polylactic acid-polystyrene composites (empty biofilm) itself has no effect on human sperm motility and ultrastructure;4、Compared with empty biofilm group and control group, levonorgestrel-loaded nanoparticle polylactic acid-polystyrene composites (drug-loaded biofilm) can make the sperms acrosome wave variation, deficiency or disappearance of acrosome, dissolution of plasma membrane, vacuolar degeneration of mitochondria and a significant increase about sperm apoptosis and disintegration.The above reseach show that drug-loaded biofilm can kill sperm;5、Research above provides the experimental basis for biological safety evaluation and effectiveness of levonorgestrel-loaded nanoparticle polylactic acid-polystyrene composites,which can afford a research basis for levonorgestrel-loaded nanoparticle polylactic acid-polystyrene composites applicated in clinical as a new intrauterine contraceptive system.