Alternating magnetic field-induced hyperthermia increases iron oxide nanoparticle cell association/uptake and flux in blood-brain barrier models

Abstract

Purpose: Superparamagnetic iron oxide nanoparticles (IONPs) are being investigated for brain cancer therapy because alternating magnetic field (AMF) activates them to produce hyperthermia. For central nervous system applications, brain entry of diagnostic and therapeutic agents is usually essential. We hypothesized that AMF-induced hyperthermia significantly increases IONP blood-brain barrier (BBB) association/uptake and flux.

Methods: Cross-linked nanoassemblies loaded with IONPs (CNA-IONPs) and conventional citrate-coated IONPs (citrate-IONPs) were synthesized and characterized in house. CNA-IONP and citrate-IONP BBB cell association/uptake and flux were studied using two BBB Transwell(®) models (bEnd.3 and MDCKII cells) after conventional and AMF-induced hyperthermia exposure.

Results: AMF-induced hyperthermia for 0.5 h did not alter CNA-IONP size but accelerated citrate-IONP agglomeration. AMF-induced hyperthermia for 0.5 h enhanced CNA-IONP and citrate-IONP BBB cell association/uptake. It also enhanced the flux of CNA-IONPs across the two in vitro BBB models compared to conventional hyperthermia and normothermia, in the absence of cell death. Citrate-IONP flux was not observed under these conditions. AMF-induced hyperthermia also significantly enhanced paracellular pathway flux. The mechanism appears to involve more than the increased temperature surrounding the CNA-IONPs.

Conclusions: Hyperthermia induced by AMF activation of CNA-IONPs has potential to increase the BBB permeability of therapeutics for the diagnosis and therapy of various brain diseases.

Fig. 1

Schema of the Transwell® system and AMF used for cell association/uptake and flux studies.

Fig. 2

AMF did not change CNA-IONP nanoparticle size but produced hyperthermia. Particle stability characterization results determined by DLS after exposure to 37°C for 30 h (n=6), 43°C (0.5 h) followed by 37°C for 30 h (n=6), or AMF (0.5 h), followed by 37°C for 30 h (n=4) (a). Representative heating profile of CNA-IONPs in cell culture medium for fine-tuned remote heating with 0.5 h application of AMF (b). Cell culture medium exposed to AMF that contained no CNA-IONPs (37°C) served as the control.

Fig. 3

Citrate-IONP and CNA-IONP cell association/uptake and flux across bEnd.3 cells. Citrate-IONP bEnd.3 cell association/uptake after 37°C for 6 h (n=3), conventional hyperthermia at 43°C (0.5 h) followed by 37°C for 6 h (n=3), or AMF-induced hyperthermia (0.5 h) followed by 37°C for 6 h (n=3) (a). CNA-IONP bEnd.3 cell association/uptake after 37°C for 6 h (n=3), conventional hyperthermia at 43°C (0.5 h) followed by 37°C for 6 h (n=3), or AMF-induced hyperthermia (0.5 h) followed by 37°C for 6 h (n=4) (b). Citrate-IONP flux across bEnd.3 cells (c). CNA-IONP flux across bEnd.3 cells (d). The paracellular flux across bEnd.3 cells measured by LY for the first 6 h in the Citrate-IONP treated group (e) and CNA-IONP treated group (f). Error bars not seen in panels C, D, E, and F are less than the symbol height. * Significantly different compared to 37°C in panel A or compared to 37°C and conventional hyperthermia in panel D and panel F.

Fig. 4

Citrate IONP and CNA-IONP cell localization. Transmission electron microscopy results of the cellular localization of the Citrate-IONPs (a and b) and CNA-IONPs (c) in bEnd.3 cells at 2 h. The arrows indicate the location of IONPs inside the cells.

Fig. 5

Paracellular permeability and CNA-IONP flux across the MDCKII in vitro BBB model. LY (a) and CNA-IONP (b) flux at 2 concentrations and 2 temperatures (n=3).

Fig. 6

CNA-IONP cell association/uptake and CNA-IONP and Lucifer yellow flux across MDCKII cells. CNA-IONP MDCKII cell association/uptake after 37°C for 6 h (n=3), conventional hyperthermia at 43°C (0.5 h) followed by 37°C for 6 h (n=3), or AMF-induced hyperthermia (0.5 h) followed by 37°C for 6 h (n=4) (a). CNA-IONP flux across a MDCKII in vitro BBB model (b). The effect of conventional and AMF-induced hyperthermia on LY flux across MDCKII cells after 43°C incubation (n=3) or AMF for 0.5 h followed by 37°C for 6 h (n=4), compared to 37°C control (n=3) (c). * Significantly different compared to AMF (0.5 h) in panel A or compared to 37°C and conventional hyperthermia in panels B and C.