Design and in vitro/in vivo Evaluation of Polyelectrolyte Complex Nanoparticles Filled in Enteric-Coated Capsules for Oral Delivery of Insulin.

Insulin is one of the most important drugs in the treatment of diabetes. There is an increasing interest in the oral administration of insulin as it mimics the physiological pathway and potentially reduces the side effects associated with subcutaneous injection. Therefore, insulin-loaded polyelectrolyte complex (PEC) nanoparticles were prepared by the ionic cross-linking method using protamine sulfate as the polycationic and sodium alginate as the anionic polymer. Taguchi experimental design was used for the optimization of nanoparticles by varying the concentration of sodium alginate, the mass ratio of sodium alginate to protamine, and the amount of insulin. The optimized nanoparticle formulation was used for further in vitro characterization. Then, insulin-loaded PEC nanoparticles were placed in hard gelatin capsules and the capsules were enteric-coated by Eudragit L100-55 (PEC-eCAPs). Hypoglycemic effects PEC-eCAPs were determined in vivo by oral administration to diabetic rats. Furthermore, in vivo distribution of PEC nanoparticles was evaluated by fluorescein isothiocyanate (FITC) labelled nanoparticles. The experimental design led to nanoparticles with a size of 194.4 nm and a polydispersity index (PDI) of 0.31. The encapsulation efficiency (EE) was calculated as 95.96%. In vivo studies showed that PEC-eCAPs significantly reduced the blood glucose level of rats at the 8th hour compared to oral insulin solution. It was concluded that PEC nanoparticles loaded into enteric-coated hard gelatin capsules provide a promising delivery system for the oral administration of insulin.

Contributions of Rho-kinase and AMP-related kinase signaling pathways to responses mediated by endothelium-derived contracting factors in diabetic rat aorta.

Diabetes-induced endothelial damage leads to vascular dysfunction. The current study investigated the effects of short-term (4-week) streptozotocin (STZ)-induced diabetes on responses mediated by endothelium-derived contracting factors (EDCFs) as well as possible contributions of Rho-kinase and AMP-activated kinase (AMPK) signaling pathways. The effects of STZ-diabetes on vascular function were examined in isolated thoracic aorta preparations of 30-week-old rats (n = 27). The diabetes-associated changes in vascular function were studied with calcium ionophore A23187, acetylcholine, Rho-kinase inhibitor Y27632 ((R)-(+)-trans-4-(1-aminoethyl)-N-(4-pyridyl)cyclohexanecarboxamide dihydrochloride), and AMPK activator AICAR (5-aminoimidazole-4-carboxamide-1-ß-d-ribofuranoside). The phosphorylation of acetyl-CoA carboxylase, AMPK, and phospholamban and the protein levels of sarcoplasmic/endoplasmic Ca2+-ATPase 2 (SERCA2) and Rho-associated protein kinase (ROCKII) were measured in aortic preparations. Although the acetylcholine-mediated relaxation responses were preserved in 4-week STZ-induced diabetes, the increased activation of the Rho-kinase pathway was demonstrated via twofold enhancement in A23187-mediated contractile responses and significantly augmented protein levels of ROCKII. The AICAR-activated AMPK-mediated relaxation response was also augmented ∼4-fold in diabetic rats, without any alteration in phospholamban phosphorylation; further, this relaxation response suppressed A23187-mediated contraction in both groups. Diabetic rats showed an increase in AICAR-induced AMPK-mediated vasorelaxation and a 2.5-fold elevation of phosphorylated AMPK levels. These results indicate a possible compensation between hyperglycemia-induced endothelium-dependent hypercontractility and AMPK-mediated vasorelaxation in diabetes.