RESUMO
The intestinal epithelium constitutes a major barrier for orally delivered nanoparticles (NPs). Although surface ligand modification can increase cellular uptake of NPs, the transepithelial transport of active targeting NPs is relatively limited. The phenomenon is described as "easy entry, hard transcytosis". However, underlying mechanisms and potential solutions of this phenomenon are unclear. Here, butyrate modified polyethylene glycol coated NPs (Bu-PEG NPs) were chosen as the model active targeting NPs. Transport mechanism studies were performed to get a better understanding of intracellular trafficking and exocytosis fate. Results showed that after active binding to monocarboxylate transporter-1 (MCT-1), Bu-PEG NPs went through endolysosomal pathways, endoplasmic reticulum/Golgi recycling routes, and microtubule-dependent shuttling within Caco-2 cells. Then a larger proportion of Bu-PEG NPs was exocytosed from apical side. Notably, increasing the basal expression of MCT-1 by leptin facilitated basal exocytosis and transcytosis of Bu-PEG NPs, which confirmed that enhanced receptor recognition could promote "basal exit". In addition to the effect of receptor recognition, surface properties also influenced the bidirectional exocytosis of Bu-PEG NPs. When surface hydrophobicity increased, Bu-PEG NPs were dominantly exocytosed from basal membrane. Hence, two strategies may help to overcome "hard transcytosis" of active targeting NPs. One is to enhance their affinity with basal membrane by reinforcing the receptor-ligand interaction; the other is to weaken apical exocytosis by optimizing surface hydrophobicity. Thereby, this study might provide important implications for the rational design of NPs to further increase transepithelial transport efficiency.
Assuntos
Administração Oral , Butiratos/química , Nanopartículas/química , Transcitose/fisiologia , Células CACO-2 , Portadores de Fármacos/química , Sistemas de Liberação de Medicamentos/métodos , Exocitose/fisiologia , Humanos , Microscopia de FluorescênciaRESUMO
As endogenous courier vesicles, exosomes play crucial roles in macromolecule transmission and intercellular communication. Therefore, exosomes have drawn increasing attention as biomimetic drug-delivery vehicles over the past few years. However, few studies have investigated the encapsulation of peptide/protein drugs into exosomes for oral administration. Additionally, the mechanisms underlying their biomimetic properties as oral delivery vehicles remain unknown. Herein, insulin-loaded milk-derived exosomes (EXO@INS) were fabricated and the in vivo hypoglycemic effect was investigated on type I diabetic rats. Surprisingly, EXO@INS (50 and 30 IU/kg) elicited a more superior and more sustained hypoglycemic effect compared with that obtained with subcutaneously injected insulin. Further mechanism studies indicated that the origin of excellent oral-performance of milk-derived exosomes combined active multi-targeting uptake, pH adaptation during gastrointestinal transit, nutrient assimilation related ERK1/2 and p38 MAPK signal pathway activation and intestinal mucus penetration. This study provides the first demonstration that multifunctional milk-derived exosomes offer solutions to many of the challenges arising from oral drug delivery and thus provide new insights into developing naturally-equipped nanovehicles for oral drug administration.
RESUMO
The epithelium is a formidable barrier to the absorption of orally delivered nano-vehicles. Here, by exploring a nutrient-absorption pathway, a self-amplified nanoplatform was developed to promote apical-to-basolateral transcytosis across the epithelium. The nanoplatform consisted of fructose-modified polyethylene glycol coated nanoparticles (Fru-PEG NPs) and a sweetener, acesulfame potassium (AceK) in combination. Compared with regular PEGylated nanoparticles, the combination exhibited a 3.9-fold increase of absorption following oral gavage in mice and an 8.8-fold increase of transepithelial transport in vitro. When encapsulated with insulin, the combination regimen elicited a stronger hypoglycemic effect, with a pharmacological bioavailability of 18.56%, which was 3.2-fold higher than that of PEG NPs. We demonstrated that a large proportion of Fru-PEG NPs underwent internalization and basolateral exocytosis via a glucose transporter type 2 (GLUT2)-dependent process, which is an important fructose assimilation pathway. Notably, co-administered AceK could prime the epithelial cells with increased apical distribution of GLUT2, thus amplifying this unidirectional transcytosis of nanoparticles. This work is the first proof-of-concept study of manipulating and amplifying a nutrient-absorption pathway to facilitate the unidirectional trans-epithelial transport of orally administered nano-delivery vehicles.
Assuntos
Nanopartículas , Transcitose , Animais , Células CACO-2 , Humanos , Insulina , Camundongos , NutrientesRESUMO
Surface features are key to the transcellular transport of nanoparticles (NPs) across intestinal epithelium cells. Endowing the NPs with specific surface features adapted to the physiological conditions of the gastrointestinal (GI) tract holds great potential for the oral delivery of peptide/protein drugs. Therefore, in this work, a glutamic acid conjugated amphiphilic dendrimer (Glu-APD) was synthesized to replace the widely used 1,2-distearoyl-sn-glycero-3-phosphatidyl-ethanolamine-polyethylene glycol (DSPE-PEG) in the preparation of poly(lactic-co-glycolic acid) (PLGA)-based NPs. Glu-APD could provide the formed NPs (Glu-APD NPs) with specific surface features of dendritic oligopeptides. With such surface features, Glu-APD NPs exhibited a 7.78-fold increase in cellular uptake and a 2.17-fold increase in the transepithelial transport amount compared with those of the DSPE-PEG2000 modified counterparts (P NPs). Instead of a dominant clathrin-mediated endocytosis as shown by P NPs, Glu-APD can provide the NPs with optional endocytosis pathways (i.e. clathrin-mediated, caveolae-mediated and micropinocytosis pathways), with the involvement of oligopeptide transporters and amino acid transporters, subsequently leading to a broadened intracellular trafficking route via the endoplasmic reticulum (ER) and Golgi apparatus. Furthermore, l-glutamic acid (l-Glu), a natural nutrient, could specifically facilitate the exocytosis of Glu-APD NPs, indicating an amino-acid-associated intracellular trafficking. Oral administration of insulin-loaded Glu-APD NPs could also achieve a good hypoglycemic effect with a relative bioavailability of 10.04%, which is 1.89-fold higher than that of P NPs and 5.20-fold higher than insulin solution. Safety evaluations further verified the biocompatibility of Glu-APD NPs and the related materials. The results confirmed the feasibility of introducing Glu-APD to NPs for improving the oral delivery of insulin. With the surface features of dendritic peptide, Glu-APD could facilitate oligopeptide/amino-acid-associated transport of the related NPs, which might be considered as an advantage under physiological conditions. This work might also be considered as a valid reference for the construction of highly efficient oral delivery systems.