Linh Nguyen, M.S.: No relevant financial relationship(s) with ineligible companies to disclose.
Objectives: Oral administration of garlic chive-derived vesicles-like nanoparticles (GC-VLNs) alleviated chronic inflammation in white adipose tissue of obese mice, suggesting that GC-VLNs were absorbed in the intestinal tract and distributed to target tissues. The objective of this study is to elucidate the absorption mechanisms of GC-VLNs in mice and human intestinal epithelial Caco-2 cells.
Methods: GC-VLNs were isolated from garlic chive by differential centrifugations. These VLNs were labeled with PKH26 red fluorescent dye before being given to the fully differentiated Caco-2 cells and C57BL/6J mice. Biomolecules in GC-VLNs were manipulated to understand how different biomolecules of GC-VLNs mediated their uptake. Small chemical compounds and genetic manipulations were used to investigate the contribution of different endocytic pathways to GC-VLN uptake in Caco-2 cells. The internalization of GC-VLNs was assessed by confocal images, flow cytometry, microRNA enrichment, and fluorescence microplate assay.
Results: GCs-VLNs were absorbed in small intestines of C57BL/6J mice. In the invitro cell culture model, GC-VLNs were taken up and transported across Caco-2 monolayers in a time-, dose-, and energy-dependent manner. Both proteins and lipids of GC-VLNs, but not RNAs, mediated GC-VLN uptake. Chemical inhibition of macropinocytosis by 5-(N-ethyl-N-isopropyl) amiloride led to diminished uptake of GC-VLNs. Moreover, inhibition of fast endophilin-mediated endocytosis (FEME) by dynasore dramatically blocked GC-VLN internalization. Conversely, activation of macropinocytosis or FEME boosted GC-VLN uptake. Other endocytic pathways had no significant contribution to GC-VLN uptake. Genetic manipulation of key molecules in macropinocytosis (Rac1) and FEME (Dynamin and RhoA) confirmed the important contribution of these two specific endocytic pathways toward the cellular uptake of GC-VLNs.
Conclusions: This study demonstrated the intestinal uptake of GC-VLNs in vivo. The detailed mechanistic studies of GC-VLN uptake would facilitate further manipulation and engineering these nanoparticles to enhance their delivery efficacy and maximize their translational potential as therapeutic modalities or drug carriers.
Funding Sources: This work was supported by USDA NIFA and NIH NIDDK.
