Radio-wave heating of iron oxide nanoparticles can regulate plasma glucose in mice

Abstract

Medical applications of nanotechnology typically focus on drug delivery and biosensors. Here, we combine nanotechnology and bioengineering to demonstrate that nanoparticles can be used to remotely regulate protein production in vivo. We decorated a modified temperature-sensitive channel, TRPV1, with antibody-coated iron oxide nanoparticles that are heated in a low-frequency magnetic field. When local temperature rises, TRPV1 gates calcium to stimulate synthesis and release of bioengineered insulin driven by a Ca(2+)-sensitive promoter. Studying tumor xenografts expressing the bioengineered insulin gene, we show that exposure to radio waves stimulates insulin release from the tumors and lowers blood glucose in mice. We further show that cells can be engineered to synthesize genetically encoded ferritin nanoparticles and inducibly release insulin. These approaches provide a platform for using nanotechnology to activate cells.

Fig. 1

Nanoparticles induced cell excitation to increase insulin expression and release in vitro. (A) Schema of nanoparticle-induced cell activation and gene expression. Antibody-coated ferrous oxide nanoparticles bind to a unique epitope, His × 6, in the first extracellular loop of the temperature-sensitive TRPV1 channel. Exposure to a RF field induces local nanoparticle heating, which opens temperature-sensitive TRPV1 channels. Calcium entry triggers downstream pathways, such as activation of calcineurin, leading to dephosphorylation of NFAT and translocation to the nucleus. Here, NFAT binds to upstream response elements to initiate gene expression of a bioengineered human insulin gene. Additional calcium-dependent signal transduction pathways also stimulate gene expression via binding to SRE and CRE. P indicates a phosphate group. (B) RF treatment increases proinsulin release and insulin gene expression in vitro. Nanoparticle-decorated HEK293T cells transfected with TRPV1^His and calcium-dependent insulin show a significant increase in proinsulin release and insulin gene expression with RF treatment that is blocked by the TRP antagonist ruthenium red. (Columns marked with the same letter indicate significance, P < 0.05. Error bars indicate SEM) (C) Time courses of proinsulin release and insulin gene expression from nanoparticle-decorated HEK293T cells transfected with TRPV1^His and calcium-dependent insulin with RF treatment.

Fig. 2

Nanoparticle regulation of blood glucose in vivo. (A) Effects of RF treatment on blood glucose in PBS and nanoparticle-treated mice with tumors expressing TRPV1^His and calcium-dependent human insulin. RF treatment significantly reduces blood glucose in nanoparticle-treated mice compared with that of PBS-treated mice. (Asterisks indicate P < 0.05. Error bars indicate SEM.) (B) RF treatment of mice with tumors expressing TRPV1^His and calcium-dependent human insulin injected with nanoparticles significantly reduces blood glucose over the course of the study as assessed by the area under the curve. There is no effect in mice with tumors expressing calcium-dependent insulin alone without TRPV1^His. (Same letter indicates P < 0.05.) (C) Plasma insulin is significantly increased by RF treatment in nanoparticle-treated but not PBS-treated mice with tumors expressing TRPV1^His and calcium-dependent human insulin. There is no effect in mice with tumors expressing calcium-dependent insulin alone without TRPV1^His. (Same letter indicates P < 0.05). (D) Insulin gene expression is significantly increased in the tumors expressing TRPV1^His and calcium-dependent human insulin treated with nanoparticles and RF magnetic field but not in tumors expressing calcium-dependent human insulin alone without TRPV1^His.

Fig. 3

Intracellular nanoparticle synthesis and cell activation. (A) Schema of intracellular nanoparticle synthesis and cell activation. A ferritin fusion protein is composed of a ferritin light chain fused to ferritin heavy chain with a flexible linker region. Heating of the iron core by a RF magnetic field opens the TRPV1 channel to trigger calcium entry, as previously described. (B) RF treatment increases proinsulin release in vitro. HEK293T cells transiently transfected with TRPV1, ferritin, and calcium-dependent human insulin show a significant increase in proinsulin release in response to RF treatment. (Same letter indicates significance, P < 0.05.) RF treatment does not increase proinsulin release from cells expressing ferritin in the absence of TRPV1. (C) RF treatment increases insulin gene expression in vitro. Insulin gene expression is significantly increased by RF treatment in cells transfected with TRPV1, ferritin, and calcium-dependent human insulin. (Same letter indicates significance, P < 0.05.) RF treatment does not increase insulin gene expression in cells expressing ferritin in the absence of TRPV1.