Study On Nano-sized Polymer Vesicles As Carriers For Oral Insulin Delivery

Oral insulin delivery has been the most convenient way for patients due to its painless treatment in recent years. However, as a protein drug used to treat diabetes, insulin is easily damaged its inherent instability in the gastrointestinal (GI) tract. Therefore, as oral insulin carriers, the exploitation of polymers prepared into microspheres or vehicles becomes an important object.Our groups had synthesized amphipathic block copolymer PLA-Pluronic P85-PLA nanoparticles. However, the biocompatibility of vesicles, release of insulin in vitro and in vivo uptake and transport studies had no reports. Therefore, the following aspects would be researched in this study:1. Release of insulin-loaded vesicles in vitro:Dialysis method was applied to prepare insulin loaded PLA-P85-PLA nanoparticles, the mean size of PLA-P85-PLA nanoparticles determined by dynamic light scattering (DLS) technique was240.2nm. Encapsulation capacity of insulin was up to59.6%, and the drug loading capacity was4.6%., release of insulin in vitro was designed at pH1.2,2.0,6.8,7.2, and the cumulative release rate of insulin in vitro at pH7.2was highest (25.2%), free insulin was completely released within4h (98%), and insulin-loaded vesicles was released45%within12h. It clearly concluded that PLA-P85-PLA vesicles had obviously controlled release for insulin.2. Biocompatibility of vesicles in vitro&in vivo:MTT assay, neutral red uptake (NRU) method and lactate dehydrogenase (LDH) release method were used to analyse the cytotoxicity of PLA-P85-PLA vesicles on intestinal epithelial cells (Caco-2cells). Results suggested that PLA-P85-PLA vesicles have good cytocompatibility. Kunming mice were selected as the animals model, the mice were administrated with different concentrations (20,100,500mg/L) of PLA-P85-PLA vesicles during28days. the body weights, food consumptions and water intakes, organ weights, physiological and biochemical parameters, and histopathological findings of mice were were measured in predetermined times. The results demonstrated that PLA-P85-PLA vesicles produced no toxicity and good biocompatibility in vivo.3. Uptake and transport of insulin-loaded vesicles:FITC-labeled insulin was applied in this study, and then was loaded in PLA-P85-PLA vesicles in order to research the absorption of insulin in vitro&in vivo. PLA-P85-PLA vesicles were administrated to Kunming mice (fasting for24h), and then segments of stomach and intestine were quickly dissected out at0.5h,1h,2h,4h. Small intestine was found stronger fluorescence, indicating insulin was mainly uptaked through the small intestine. Therefore, Caco-2cells (similar to characters of intestinal epithelial cells) were also selected to quantificationally analyze the effects of concentrations of insulin, incubation time, pH and incubation temperature on the cellular uptake in vitro of free insulin and insulin loaded vesicles. The results suggested that PLA-P85-PLA vesicles promoted significantly the absorption of insulin. Caco-2cell monolayer model were established to study the transport of insulin and insulin-loaded vesicles across cells. The results were observed that PLA-P85-PLA vesicles could also enhance the transport of insulin.In conclusion, PLA-P85-PLA vesicles have observably control release of insulin, good biocompatibility, and promote uptake and transport of insulin. As a result, this material could be alternative carriers and provide theoretical basis for further research.