Design of Experiment, Preparation, and in vitro Biological Assessment of Human Amniotic Membrane Extract Loaded Nanoparticles

Human amniotic membrane grafting could be potentially useful in ocular surface complications due to tissue similarity and the presence of factors that reduce inflammation, vascularization, and scarring. However, considerations like donor-derived infectious risk and the requirement of an invasive surgery limit the clinical application of such treatments. Moreover, the quick depletion of bioactive factors after grafting reduces the efficacy of treatments. Therefore, in the current study, the possibility of nano delivery of the bioactive factors extracted from the human amniotic membrane to the ocular surface was investigated.Nanoparticles were prepared using polyelectrolyte complexation from chitosan and dextran sulfate. The effect of polymer ratio, pH, and the amount of extract on particle size and encapsulation efficacy were studied using Box-Behnken response surface methodology.The optimum condition was obtained as follows: 4.9:1 ratio of dextran sulfate to chitosan, 600 µL amniotic membrane extract, and pH of 6. The prepared nanoparticles had an average size of 213 nm with 77% encapsulation efficacy. In the release test, after 10 days, approximately 50% of entrapped bioactive proteins were released from the nanocarriers in a controlled manner. Biological activity assessment on endothelial cells revealed amniotic membrane extract loaded nanoparticles had a longer and significant increase in anti-angiogenic effect when compared to the control.Our data elucidate the ability of nanotechnology in ocular targeted nano delivery of bioactive compounds.