Transport Pathways of Solid Lipid Nanoparticles Across Madin–Darby Canine Kidney Epithelial Cell Monolayer

Abstract

An understanding of drug delivery system transport across epithelial cell monolayer is very important for improving the absorption and bioavailability of the drug payload. The mechanisms of epithelial cell monolayer transport for various nanocarriers may differ significantly due to their variable components, surface properties, or diameter. Solid lipid nanoparticles (SLNs), conventionally formed by lipid materials, have gained increasing attention in recent years due to their excellent biocompatibility and high oral bioavailability. However, there have been few reports about the mechanisms of SLNs transport across epithelial cell monolayer. In this study, the molecular mechanisms utilized by SLNs of approximately 100 nm in diameter crossing intestinal epithelial monolayer were carefully studied using a simulative intestinal epithelial monolayer formed by Madin-Darby canine kidney (MDCK) epithelial cells. The results demonstrated that SLNs transportation did not induce a significant change on tight junction structure. We found that the endocytosis of SLNs into the epithelial cells was energy-dependent and was significantly greater than nanoparticle exocytosis. The endocytosis of SLNs was found to be rarely mediated via macropinocytosis, as confirmed by the addition of 5-(N-ethyl-N-isopropyl)amiloride (EIPA) as an inhibitory agent, and mainly depended on lipid raft/caveolae- and clathrin-mediated pathways. After SLNs was internalized into MDCK cells, lysosome was one of the main destinations for these nanoparticles. The exocytosis study indicated that the endoplasmic reticulum, Golgi complex, and microtubules played important roles in the transport of SLNs out of MDCK cells. The transcytosis study indicated that only approximately 2.5% of the total SLNs was transported from the apical side to the basolateral side. For SLNs transportation in MDCK cell monolayer, greater transport (approximately 4-fold) was observed to the apical side than to the basolateral side. Our findings may present a more comprehensive understanding on the transport of SLNs across epithelial cell monolayer.

The cellular uptake mechanism, intracellular transportation, and exocytosis of polyamidoamine dendrimers in multidrug-resistant breast cancer cells

“…The MCF-7 and MCF-7/ADR cells were preincubated with brefeldin A, monensin, and bafilomycin A1 at a given concentration 18 (Table 1) for 30 minutes in a humidified atmosphere with 5% CO 2 at 37°C, and incubated with PAMAM-NH 2 -FITC (10 μg/mL) for another 3 hours. The cells were then processed and examined using a BD FACS Caliber flow cytometer.…”

“…The cells were then incubated with PAMAM-NH 2 -FITC (10 μg/mL) in a fresh serum-free medium for 3 hours, rinsed three times with cold PBS, and incubated in a fresh medium containing various inhibitors 18 (the intracellular transport inhibitors in Table 1) for another 5 hours. Finally, the cells were washed with PBS and resuspended in PBS, and the samples were analyzed with a BD FACS Caliber flow cytometer.…”

“…18 Brefeldin A, a fungal metabolite, blocks forward transportation between the ER and Golgi complex, 35 whereas monensin, another inhibitor that is often used to disrupt the Golgi complex, has been shown to effectively block the transportation of macromolecules from the Golgi complex to the plasma membrane. 36 As shown in Figure 5, the addition of brefeldin A and monensin did not influence the intracellular amount of PAMAM-NH 2 , indicating that neither the ER nor Golgi complex participates in the endocytosis process of PAMAM-NH 2 .…”

Synergistic and complete reversal of the multidrug resistance of mitoxantrone hydrochloride by three-in-one multifunctional lipid-sodium glycocholate nanocarriers based on simultaneous BCRP and Bcl-2 inhibition

“…47,48 To elucidate the underlying MDR reversal mechanisms, the uptake mechanism of MTOTMLGNs was investigated through temperature and endocytosis inhibition experiments in resistant MCF-7/MX cells. In addition, the different dosage regimens were compared to investigate the synergic MDR reversal effect of CsA and GcNa.…”

“…Clathrin-mediated endocytosis is a classical internalization pathway for macromolecules. [47][48][49][50] Extracellular substances are incorporated into clathrin-coated pits, which pinch off from plasma membrane, and are then internalized by the cell along with these pits.…”