Study of Uptake Mechanisms of Halloysite Nanotubes in Different Cell Lines

Abstract

Halloysite nanotubes (HNTs) are a natural aluminosilicate clay with a chemical formula of Al 2 Si 2 O 5 (OH) 4 ×nH 2 O and a hollow tubular structure. Due to their peculiar structure, HNTs can play an important role as a drug carrier system. Currently, the mechanism by which HNTs are internalized into living cells, and what is the transport pathway, is still unclear. Therefore, this study aimed at establishing the in vitro mechanism by which halloysite nanotubes could be internalized, using phagocytic and non-phagocytic cell lines as models. Methods: The HNT/CURBO hybrid system, where a fluorescent probe (CURBO) is confined in the HNT lumen, has been used as a model to study the transport pathway mechanisms of HNTs. The cytocompatibility of HNT/CURBO on cell lines model was investigated by MTS assay. In order to identify the internalization pathway involved in the cellular uptake, we performed various endocytosis-inhibiting studies, and we used fluorescence microscopy to verify the nanomaterial internalization by cells. We evaluated the haemolytic effect of HNT/CURBO placed in contact with human red blood cells (HRBCs), by reading the absorbance value of the supernatant at 570 nm. Results: The HNT/CURBO is highly biocompatible and does not have an appreciable haemolytic effect. The results of the inhibition tests have shown that the internalization process of nanotubes occurs in an energy-dependent manner in both the investigated cell lines, although they have different characteristics. In particular, in non-phagocytic cells, clathrin-dependent and independent endocytosis are involved. In phagocytic cells, in addition to phagocytosis and clathrin-dependent endocytosis, microtubules also participate in the halloysite cellular trafficking. Upon internalization by cells, HNT/CURBO is localized in the cytoplasmic area, particularly in the perinuclear region. Conclusion: Understanding the cellular transport pathways of HNTs can help in the rational design of novel drug delivery systems and can be of great value for their applications in biotechnology.

Novel brain-targeted nanomicelles for anti-glioma therapy mediated by the ApoE-enriched protein corona in vivo

“…In addition, cells treated with chlorpromazine (an inhibitor of clathrin-mediated endocytosis), colchicine (an inhibitor of microtubule formation and macropinocytosis), and brefeldin A (an inhibitor of protein transport from the endoplasmic reticulum to the Golgi apparatus) showed a 36%, 39%, and 31% decrease in cellular uptake, respectively as reflected in the decrease in fluorescence signals. The cellular uptake of rhApoE/Cou-6/Aβ-CN-PMs was partially dependent on clathrin-mediated endocytosis, macropinocytosis, and the Golgi apparatus [ 41 – 43 ]. Sodium azide (NaN 3 ) is used to inhibit energy-dependent endocytosis by disrupting ATP production.…”

Nanomaterials: A Review about Halloysite Nanotubes, Properties, and Application in the Biological Field

“…This result suggests that HNTs do not exhibit a significant haemolytic effect [ 88 ]. It has also been seen that HUVECs (human umbilical vein endothelial cells) and MCF-7 (human breast cancer) cells show high cell viability after being treated with different concentrations of HNTs for 24 h. For both cell lines, vitality remains above 85% even when the concentration of HNTs reaches up to 200 mg/mL.…”

“… In vitro hemocompatibility assay of ( A ) raw HNT and ( B ) HNT/CURBO after incubation at 37 °C for 12 h. The positive control was in ultrapure water (100% lysis), and negative control was in PBS 1× (0% lysis). International Journal of NanoMedicine 2021:16 4755–4768 Originally published by and used with permission from Dove Medical Press Ltd [ 88 ]. …”

Temperature-responsive and biocompatible nanocarriers based on clay nanotubes for controlled anti-cancer drug release

“…Regarding biocompatibility, recent studies have clearly demonstrated that pristine HNTs can easily penetrate the cell membrane and accumulate in the perinuclear areas without causing significant toxic effects. [24][25][26][27] All these appealing features of HNTs encouraged us to explore the potential of these nanotubes for designing a new smart nanocarrier with temperature-mediated release behavior. For this purpose, PNIPAM brushes were grown on the surface of HNTs using a surface-initiated activator regenerated by electron transfer in atom transfer radical polymerization (ARGET-ATRP).…”