MicroRNAs in Apple-Derived Nanoparticles Modulate Intestinal Expression of Organic Anion–Transporting Peptide 2B1/SLCO2B1 in Caco-2 Cells

Abstract

Plant-derived nanoparticles exert cytoprotective effects on intestinal cells by delivering their cargo to intestinal tissues. We previously reported that apple-derived nanoparticles (APNPs) downregulate the mRNA of the human intestinal transporter OATP2B1/SLCO2B1 and that the 3'UTR region is required for the response to APNPs. Here, we investigated the involvement of microRNAs (miRNAs) in APNPs in suppressing OATP2B1 expression to demonstrate that APNP macromolecules directly interact with intestinal tissues. Using in silico analysis, seven apple miRNAs were predicted as candidate miRNAs that interact with the SLCO2B1-3'UTR. The APNP-mediated decrease in luciferase activity of pGL3/SLCO2B1-3'UTR was abrogated by inhibitors of mdm-miR-160a-e, -7121a-c, or -7121d-h. Each miRNA mimic reduced the endogenous expression of SLCO2B1 mRNA in Caco-2 cells. The luciferase activity of the truncated pGL3/SLCO2B1-3'UTR, which contains approximately 200 bp around each miRNA recognition element (MRE), was decreased by the miR-7121d-h mimic but little by the other mimics. APNP also reduced the luciferase activity of truncated pGL3/SLCO2B1-3'UTR containing an MRE for miR-7121d-h. Thus, we demonstrated that mdm-miR-7121d-h contributes to the APNP-mediated downregulation of intestinal OATP2B1. Accordingly, plant macromolecules, such as miRNAs, may directly interact with intestinal tissues via nanoparticles.

Isolation and characterization of ginseng-derived exosome-like nanoparticles with sucrose cushioning followed by ultracentrifugation

“…In comparison with a previous isolation method of plant-derived exosome-like nanoparticles from carrot, apple, lemon, grape, grapefruits, tea leaves, cabbage, ginger, and garlic [32][33][34][35][36][37][38], we established the double fractionation of GDEs by sucrose-based isolation to purify GDEs from vesicle aggregates or contaminated proteins for reproducible high-quality exosome-like nanoparticles. To collect homogeneously sized GDEs with high yield, we first isolated non-contaminated intact EVs from ginseng juice through sucrose cushion of different densities, which can prevent the degradation of EVs during repeated highforce centrifugations.…”

Evidence of Cross-Kingdom Gene Regulation by Plant MicroRNAs and Possible Reasons for Inconsistencies

“…For example, which principle does a plant miRNA functioning in animals obey? The current evidence indicates that plant miRNAs that exert cross-kingdom gene regulation obey the principles of miRNA–target recognition in animals. ,,, …”

“…16,17 These EV range from 30 to 1000 nm and have been isolated and biologically characterized in several species from the perspective of miRNAs, such as ginger, 58 lemon, 59 strawberry, 60 sunflower seeds, 61 A. thaliana, 62 and so on. Furthermore, EV can be artificially generated from plant lipids, 16,31,63 and plant miRNAs can be artificially packaged into EV 16,31 for delivery into body cells. EV-borne plant miRNAs can be specifically internalized by intestinal cells via caveolin-mediated endocytosis and micropinocytosis.…”

Apple juice relieves loperamide-induced constipation in rats by downregulating the intestinal apical sodium-dependent bile acid transporter ASBT

“…Besides small molecules, large molecules such as apple-derived microRNAs may also contribute to a decreased expression of ASBT, since small extracellular vesicles from apples were previously reported to downregulate several intestinal transporters including ASBT in Caco-2 cells 19 and one of the intestinal transporters, OATP2B1, is downregulated by specific apple microRNAs contained in apple-derived small extracellular vesicles. 44 We are now investigating the effect of apple-derived small extracellular vesicles on ASBT expression.…”

“…Besides small molecules, large molecules such as apple-derived microRNAs may also contribute to a decreased expression of ASBT, since small extracellular vesicles from apples were previously reported to downregulate several intestinal transporters including ASBT in Caco-2 cells 19 and one of the intestinal transporters, OATP2B1, is downregulated by specific apple microRNAs contained in apple-derived small extracellular vesicles. 44 We are now investigating the effect of apple-derived small extracellular vesicles on ASBT expression. About the alteration of intestinal luminal BAs, in addition to these gene-regulating effects in apples, the direct effect of AJ components on the ASBT transport activity may also be involved because AJ inhibited the uptake of taurocholic acid by Caco-2 cells in an AJ-concentration-dependent manner with an IC 50 value of 61.9% ± 16.9% AJ (data not shown).…”