Ca2+ Overload Decreased Cellular Viability in Magnetic Hyperthermia without a Macroscopic Temperature Rise.

Magnetic hyperthermia is a crucial medical engineering technique for treating diseases, which usually uses alternating magnetic fields (AMF) to interplay with magnetic substances to generate heat. Recently, it has been found that in some cases, there is no detectable temperature increment after applying an AMF, which caused corresponding effects surprisingly. The mechanisms involved in this phenomenon are not yet fully understood. In this study, we aimed to explore the role of Ca2+ overload in the magnetic hyperthermia effect without a perceptible temperature rise. A cellular system expressing the fusion proteins TRPV1 and ferritin was prepared. The application of an AMF (518 kHz, 16 kA/m) could induce the fusion protein to release a large amount of iron ions, which then participates in the production of massive reactive oxygen radicals (ROS). Both ROS and its induced lipid oxidation enticed the opening of ion channels, causing intracellular Ca2+ overload, which further led to decreased cellular viability. Taken together, Ca2+ overload triggered by elevated ROS and the induced oxidation of lipids contributes to the magnetic hyperthermia effect without a perceptible temperature rise. These findings would be beneficial for expanding the application of temperature-free magnetic hyperthermia, such as in cellular and neural regulation, design of new cancer treatment methods.

Tumor inhibition via magneto-mechanical oscillation by magnetotactic bacteria under a swing MF.

Since magnetic micro/nano-materials can serve as multifunctional transducers for remote control of cell functions by applying diverse magnetic fields, magnetic cell manipulation provides a highly promising tool in biomedical research encompassing neuromodulation, tissue regeneration engineering and tumor cell destruction. Magnetotactic bacteria (MTB), which contain natural magnetic materials, can sensitively respond to external magnetic fields via their endogenous magnetosome chains. Here, we developed a technique for magnetotactic bacteria-based cell modulation and tumor suppression combined with a swing magnetic field. We enabled MTB cells to recognize and bind to mammalian tumor cells via functional modification with RGD peptides onto the surfaces of MTB cells, and RGD-modified MTB bacteria could interact with the targeted tumor cells effectively. The magnetic torque, which was due to the interaction of the long magnetosome chain inside the MTB bacterial cell and the applied swing magnetic field, could result in obvious swing magnetic behaviors of the modified MTB bacteria bound to tumor cell surfaces and thus subsequently exert a sustained magnetomechanical oscillation on the tumor cell surfaces, which could induce a significant activation of Ca2+ ion influx in vitro and tumor growth inhibition in vivo. These findings suggest that MTB cells mediated magnetomechanical stimulation, which is remotely controlled by dynamic magnetic fields, as an effective way to regulate cell signaling and treat tumor growth, which will shed the light on further biomedical applications utilizing whole magnetotactic bacteria.

In vitro magnetosome remineralization for silver-magnetite hybrid magnetosome biosynthesis and used for healing of the infected wound.

BACKGROUND: Magnetosomes (BMPs) are organelles of magnetotactic bacteria (MTB) that are responsible for mineralizing iron to form magnetite. In addition, BMP is an ideal biomaterial that is widely used in bio- and nano-technological applications, such as drug delivery, tumor detection and therapy, and immunodetection. The use of BMPs to create multifunctional nanocomposites would further expand the range of their applications. RESULTS: In this study, we firstly demonstrate that the extracted BMP can remineralize in vitro when it is exposed to AgNO3 solution, the silver ions (Ag+) were transported into the BMP biomembrane (MM) and mineralized into a silver crystal on one crystal plane of Fe3O4. Resulting in the rapid synthesis of an Ag-Fe3O4 hybrid BMP (BMP-Ag). The synergy between the biomembrane, Fe3O4 crystal, and unmineralized iron enabled the remineralization of BMPs at an Ag+ concentration ≥ 1.0 mg mL-1. The BMP-Ag displayed good biocompatibility and antibacterial activity. At a concentration of 2.0 mg/mL, the BMP-Ag and biomembrane removed Ag-Fe3O4 NPs inhibited the growth of gram-negative and gram-positive bacteria. Thus using BMP-Ag as a wound dressing can effectively enhance the contraction of infected wounds. CONCLUSIONS: This study represents the first successful attempt to remineralize organelles ex vivo, realizing the biosynthesis of hybrid BMP and providing an important advancement in the synthesis technology of multifunctional biological nanocomposites.

Smart Magnetotactic Bacteria Enable the Inhibition of Neuroblastoma under an Alternating Magnetic Field.

Magnetotactic bacteria are ubiquitous microorganisms in nature that synthesize intracellular magnetic nanoparticles called magnetosomes in a gene-controlled way and arrange them in chains. From in vitro to in vivo, we demonstrate that the intact body of Magnetospirillum magneticum AMB-1 has potential as a natural magnetic hyperthermia material for cancer therapy. Compared to chains of magnetosomes and individual magnetosomes, the entire AMB-1 cell exhibits superior heating capability under an alternating magnetic field. When incubating with tumor cells, the intact AMB-1 cells disperse better than the other two types of magnetosomes, decreasing cellular viability under the control of an alternating magnetic field. Furthermore, in vivo experiments in nude mice with neuroblastoma found that intact AMB-1 cells had the best antitumor activity with magnetic hyperthermia therapy compared to other treatment groups. These findings suggest that the intact body of magnetotactic bacteria has enormous promise as a natural material for tumor magnetic hyperthermia. In biomedical applications, intact and living magnetotactic bacteria play an increasingly essential function as a targeting robot due to their magnetotaxis.

In Situ Determination of Sialic Acid on Cell Surface with a pH-Regulated Polymer Enzyme Nanoreactor.

Sialic acid (SA) is an important monosaccharide that is involved in incurable cancer immunotherapy. However, it is difficult to detect SA in situ using the existing strategy based on the SA-terminated glycopeptide extraction from the cell lysate. The countermeasures of the bottleneck caused by cell disruption and peptide extraction should be designed based on a "cell-surface attachment and controlled enzymolysis" protocol. Herein, a poly(styrene-co-maleic anhydride-acrylic acid-concanavalin A) (PSM-PAA-ConA) was synthesized and developed as a pH-regulated enzyme nanoreactor after being loaded with sialidase and myoglobin. The nanoreactor showed controllable biocatalysis induced by a cascade enzyme reaction and applied for the in situ detection of SA on a living cell surface. The addition of an acidic solution resulted in a decrease in the size of the nanoreactor and enhancement of its permeability, triggering an "on" state of the SA catalysis. Subsequent pH increase led to increased hydrophilicity of the nanoreactor, increasing its size and resulting in the catalytic "off" state. ConA assisted the cell-surface attachment of the enzyme reactor. Furthermore, SA on the surface of living cancer cells was successfully monitored by the pH-regulated enzyme nanoreactor, demonstrating the feasibility of high specificity in situ analysis for SA. This pH-induced catalytic efficiency control by the enzyme nanoreactor provides a potential platform for functional stimuli-responsive catalytic systems as well as a strategy for in situ analysis of biomolecules on the cell surface.

Simultaneous Monitoring of Mitochondrial Temperature and ATP Fluctuation Using Fluorescent Probes in Living Cells.

Real-time monitoring of the distribution of energy released during oxidative phosphorylation (OXPHOS) in living cells would advance the understanding of metabolic pathways and cell biology. However, the relationship between intracellular temperature and ATP fluctuation during the OXPHOS process is rarely studied due to the limitation of the sensing approach. Novel fluorescent polymer probes were developed for accurate simultaneous measurements of intracellular temperature and ATP. Utilizing the fluorescence imaging techniques, it was demonstrated for the first time that the temperature in mitochondria increased 2.4 °C and the ATP fluctuation level simultaneously decreased 75% within 2 min during the OXPHOS process. Moreover, the resultant fluorescent polymer probes had good performance and properties for mitochondrial targeting, providing an effective way for investigating mechanisms by which energy is released during the OXPHOS process.

Magnetically targeted photothemal cancer therapy in vivo with bacterial magnetic nanoparticles.

The biomineralized bacterial magnetic nanoparticles (BMPs) have been widely studied for biomedical applications with their magnetic properties and a layer of biomembrane. Herein, BMPs were firstly used for magnetically targeted photothermal cancer therapy in vivo. A self-build C-shaped bipolar permanent magnet was used for magnetic targeting though the generation of a high gradient magnetic field within a small target area. For in vitro simulated experiment, BMPs had a high retention rate in magnetically targeted region with different flow rates. In H22 tumor bearing mice, the magnetic targeting induced a 40% increase of BMPs retention in tumor tissues. In vivo photothermal therapy with 808 nm laser irradiation could induce a complete tumor elimination with magnetic targeting. These results indicated that the systematically administrated BMPs with magnetic targeting would be promising for photothermal cancer therapy.

Structural effect of Fe3O4 nanoparticles on peroxidase-like activity for cancer therapy.

Ferromagnetic nanoparticles (Fe3O4 NPs) have been proven to have the intrinsic peroxidase-like activity. This property has been used for analyte detection, tumor tissue visualization, and cancer therapy, etc. However, the effect of particle structure and morphology on its peroxidase-like activity has been rarely reported. In this work, we fabricated Fe3O4 nanoparticles with different structures (nanoclusters, nanoflowers, and nanodiamonds) by facilely tuning the pH values in the hydrothermal reaction. Their in vitro peroxidase-like activity was evaluated via chromogenic reaction of 3,3',5,5'-tetramethylbenzidine (TMB) by the reduction of H2O2 to H2O. It was found the nanostructures had a great influence on their peroxidase-like activity, following the order of nanoclusters>nanoflowers>nanodiamonds. With this activity, the peroxidase-like activity of Fe3O4 NPs was used for cancer therapy with the addition of low-concentration H2O2. The cancer cell-killing activity was due to the intracellular generated reactive oxygen species (ROS) after endocytosis of Fe3O4 NPs into the Hela cells. It was interesting that the cell killing ability of these three kinds of Fe3O4 NPs was not consistent with the in vitro enzyme-like activity. It was deduced that the cell endocytosis of the nanoparticles along with their enzyme-like activity co-determined their cancer cell-killing performance.

Bacterial magnetic nanoparticles for photothermal therapy of cancer under the guidance of MRI.

The bacterial magnetic nanoparticles (BMPs) are biomineralized by the magnetotactic bacteria and naturally covered with a layer of biomembrane. Herein, BMPs were isolated and firstly used for the photothermal therapy (PTT) of cancer under the guidance of magnetic resonance imaging (MRI) in vitro and in vivo. The results showed that BMPs could rapidly convert the energy of 808 nm near-infrared (NIR) light into heat. After internalization by the HepG2 tumor cells, BMPs with good biocompatibility could induce an efficient killing effect after NIR light irradiation, along with a change of mitochondrial membrane potential (ΔΨm) and level of intracellular reactive oxygen species (ROS). The in vivo therapy also confirms that PTT with BMPs could effectively and completely ablate the tumor in mice without inducing observable toxicity. T2-weighted MRI showed a clear tumor boundary and a 25% enhancement of negative contrast enhancement at the tumor site, suggesting that BMPs can act as an effective MRI contrast agent for guiding the PTT. Our results indicate that BMPs could be a potential theranostic agent for simultaneous MRI and PTT of cancer.

Applications of Bacterial Magnetic Nanoparticles in Nanobiotechnology.

The bacterial magnetic nanoparticle (BMP) has been well researched in nanobiotechnology as a new magnetic crystal. The BMPs are extracted from magnetotactic bacteria and under precise biological control. Compared with engineered magnetic nanoparticles synthesized by chemical approaches, BMPs have the properties of large production, monodispersity, high crystallinity, and close-to-bulk magnetization, which enable BMPs to be the highly promising magnetic nanoparticles for nanobiotechnology. In this paper, we review the biomedical applications of BMPs in magnetic hyperthermia, drug treatment with tumour and bioseparation. In addition, the biodistribution and toxicity are also reviewed.

Microenvironment-Driven Bioelimination of Magnetoplasmonic Nanoassemblies and Their Multimodal Imaging-Guided Tumor Photothermal Therapy.

Biocompatibility and bioelimination are basic requirements for systematically administered nanomaterials for biomedical purposes. Gold-based plasmonic nanomaterials have shown potential applications in photothermal cancer therapy. However, their inability to biodegrade has impeded practical biomedical application. In this study, a kind of bioeliminable magnetoplasmonic nanoassembly (MPNA), assembled from an Fe3O4 nanocluster and gold nanoshell, was elaborately designed for computed tomography, photoacoustic tomography, and magnetic resonance trimodal imaging-guided tumor photothermal therapy. A single dose of photothermal therapy under near-infrared light induced a complete tumor regression in mice. Importantly, MPNAs could respond to the local microenvironment with acidic pH and enzymes where they accumulated including tumors, liver, spleen, etc., collapse into small molecules and discrete nanoparticles, and finally be cleared from the body. With the bioelimination ability from the body, a high dose of 400 mg kg(-1) MPNAs had good biocompatibility. The MPNAs for cancer theranostics pave a way toward biodegradable bio-nanomaterials for biomedical applications.

Ratiometric Fluorescent Polymeric Thermometer for Thermogenesis Investigation in Living Cells.

Intracellular temperature has a fundamental effect on cellular events. Herein, a novel fluorescent polymer ratiometric nanothermometer has been developed based on transferrin protein-stabilized gold nanoclusters as the targeting and fluorescent ratiometric unit and the thermosensitve polymer as the temperature sensing unit. The resultant nanothermometer could feature a high and spontaneous uptake into the HeLa cells and the ratiometric temperature sensing over the physiological temperature range. Moreover, the precise temperature sensing for intracellular heat generation in HeLa cells following calcium ions stress has been achieved. This practical intracellular thermometry could eliminate the interference of the intracellular surrounding environment in cancer cells without a microinjection procedure, which is user-friendly. The prepared new nanothermometer can provide tools for unveiling the intrinsic relationship between the intracellular temperature and ion channel function.

Enhancement of the hydrolysis activity of F0F1-ATPases using 60 Hz magnetic fields.

The effects of extremely low frequency (ELF) magnetic fields on membrane F(0)F(1)-ATPase activity have been studied. When the F(0)F(1)-ATPase was exposed to 60 Hz magnetic fields of different magnetic intensities, 0.3 and 0.5 mT magnetic fields enhanced the hydrolysis activity, whereas 0.1 mT exposure caused no significant changes. Even if the F(0)F(1)-ATPase was inhibited by N,N-dicyclohexylcarbodiimide, its hydrolysis activity was enhanced by a 0.5 mT 60 Hz magnetic field. Moreover, when the chromatophores which were labeled with F-DHPE were exposed to a 0.5 mT, 60 Hz magnetic field, it was found that the pH of the outer membrane of the chromatophore was unchanged, which suggested that the magnetic fields used in this work did not affect the activity of F0. Taken together, our results show that the effects of magnetic fields on the hydrolysis activity of the membrane F(0)F(1)-ATPases were dependent on magnetic intensity and the threshold intensity is between 0.1 and 0.3 mT, and suggested that the F1 part of F(0)F(1)-ATPase may be an end-point affected by magnetic fields.

[Effects of extremely low frequency magnetic fields on hydrolysis of F0F1-ATPases and their relationship with turnover rates of F1].

OBJECTIVE: To study the effects of extremely low frequency sinusoidal magnetic fields on hydrolysis of F(0)F(1)-ATPase and its mechanism. METHODS: The F(0)F(1)-ATPases which was localized on the outer surface of chromatophores were prepared from the cells of Rhodospirillum rubrum and were exposed to 0.1 approximately 0.5 mT, 4.7 approximately 96.0 Hz magnetic fields. RESULTS: The hydrolysis activity of F(0)F(1)-ATPase was stimulated by 0.5 mT, 4.7, 12.0, 60.0, 72.0, 84.0 and 96.0 Hz magnetic fields respectively and inhibited by 0.5 mT, 24.0 Hz magnetic field (P < 0.05); 0.3 mT, 4.7, 24.0 and 60.0 Hz magnetic fields also distinctly affected F(0)F(1)-ATPases activity respectively (P < 0.05), whereas 0.1 mT exposure caused no significant changes on that activity. When the hydrolysis activity of the F(0)F(1)-ATPases was inactivated by its inhibitor DCCD, the 0.5 mT, 24.0 Hz magnetic field still inhibited the hydrolysis activity of the F(0)F(1)-ATPase and 0.5 mT, 60.0 Hz magnetic field also had stimulating effects (P < 0.05). CONCLUSION: The effects of magnetic fields on the hydrolysis activity of the F(0)F(1)-ATPases depend on not only magnetic frequency but also magnetic intensity. The threshold of magnetic intensity is between 0.1 mT and 0.3 mT. F(0)F(1)-ATPases, especially F1-portion may be an end-point of magnetic fields.

Preliminary estimation of rotary torque produced by proton-motive force in fully functional F0F1-ATPase.

F(0)F(1)-ATPase is a rotary molecular motor. It is well known that the rotary torque is generated by ATP hydrolysis in F(1) but little is known about how it produces the proton-motive force (PMF) in F(0). Here a cross-linking approach was used to estimate the rotary torque produced by PMF. Three mutant E. coli strains were used in this study: SWM92 (deltaW28L F(0)F(1), as control), MM10 (alphaP280CgammaA285C F(0)F(1)) and PP2 (alphaA334C/gammaL262C F(0)F(1)). The oxidized inner membranes from mutant MM10 having a disulfide bridge in the top of gamma subunit exhibited good ATP synthesis activity, while the oxidized PP2 inner membranes having a disulfide bridge in the middle of gamma subunit synthesized ATP very poorly. We conclude that the rotary torque generated by PMF is sufficient to uncoil the alpha-helix in the top of gamma subunit (MM10) and to overcome the Ramachandran activation barriers (25-30 kJ/mol, i.e. about 40-50pNnm), but cannot cleave the disulfide bond in the middle of the gamma subunit (200 kJ/mol, i.e. 330pNnm) (PP2). Consequently a preliminary estimation is that the rotary torque generated by PMF in the fully functional F(0)F(1) motor is greater than 40-50pNnm but less than 330pNnm.