A coil system for real-time magnetic fluid hyperthermia microscopy studies.

PURPOSE: We describe the design and application of a new apparatus for applying Radiofrequency (RF) electromagnetic fields to cells in culture on a microscope stage. This new design enables real-time studies of the actuation of magnetic nanoparticles bound to membrane receptors or internalised within cells together with the study of magnetic fluid hyperthermia (MFH)-associated effects. MATERIALS AND METHODS: RF coils were fabricated and electromagnetic simulations were performed along with compatibility evaluations and calorimetric experiments using this apparatus at discreet frequencies between 100 kHz and 1 MHz. Cell killing via MFH was investigated in a neuroblastoma tumour cell line. RESULTS: Simulations and evaluations showed that the field intensity and homogeneity experienced by the cells within the chamber is best with a planar coil configuration. The incubation chamber was suitable for cell culture and the design was compatible with mountings on different makes of microscopes as it mimics a standard 96/24/6 tissue-culture well plate. Successful calorimetric and MFH cytotoxicity proof-of-principle experiments were performed and are presented. CONCLUSIONS: We conclude from these experiments that alternating magnetic field (AMF)-mediated activation and magnetic fluid hyperthermia (MFH) research will benefit from this RF coil that fits inside an incubation chamber, mounted onto a microscope. This new design could be used to assist real-time MFH studies in vitro.

Toxicity testing of indocyanine green and fluorodeoxyglucose conjugated iron oxide nanoparticles with and without exposure to a magnetic field.

Iron nanoparticles (MNPs) are known to induce membrane damage and apoptosis of cancer cells. In our study we determined whether FDG coupled with iron oxide magnetic nanoparticles can exert the same destructive effect on cancer cells. This research study presents data involving NIC-H727 human lung, bronchus epithelial cells exposed to conjugated fluorodeoxyglucose conjugated with iron-oxide magnetic nanoparticles and indocyanine green (ICG) dye (FDG-MNP-ICG), with and without the application of a magnetic field. Cell viability inferred from MTT assay revealed that FDG-MNPs had no significant toxicity towards noncancerous NIC-H727 human lung, bronchus epithelial cells. However, percentage cell death was much higher using a magnetic field, for the concentration of FDG-MNP-ICC used in our experiments. Magnetic field was able to destroy cells containing MNPs, while MNPs alone had significantly lower effects. Additionally, MNPs alone in these low concentrations had less adverse effects on healthy (non-target) cells.

Effects of fluorodeoxyglucose magnetic nanoparticles on NCI-H727 and SH-SY5Y cancer cells.

We present a report regarding the cytotoxic effects of iron-based magnetic nanoparticles conjugated with fluorodeoxyglucose (FDG-mNPs) on the viability of NCI-H727 and SH-SY5Y cancer cells. MTT assays were performed to determine cell viability in treated cancer cells grown under standard 2D culture conditions. FDG-mNP concentrations of 0.075 mg/mL, 0.15 mg/mL, and 0.3 mg/mL decreased mean cell viability of NCI-H727 cells to 92.5%, 82.9%, and 75% respectively. FDG-mNPs was also shown to have a detrimental effect on the viability of SY5Y cells: a decrease of 5.7%, 18.6%, and 36.4% was found for SY5Y cells treated with 0.075 mg/mL, 0.15 mg/mL, and 0.3 mg/mL concentrations of FDG-mNPs, respectively. When NCI-H727 and SH-SY5Y cancer cells were grown as 3D spheroids, morphology was visibly changed and the number of viable cells was decerased in spheroids treated with FDG-mNPs compared with untreated spheroids. The results of our study demonstrated that FDG-mNP has toxic effects on NCI-H7272 and SY5Y cancer cells, and we conclude that conjugated FDG-mNPs are promising in the development of clinical applications for the destruction of cancer cells.

In vitro testing of a new aspiration thrombus device.

BACKGROUND: Mechanical thrombectomy can restore blood flow to the brain after acute ischemic stroke, but may be associated with risks, such as breakage of moving parts and clot fragmentation. The aim of this study was to evaluate a new aspiration thrombus device (ATD), the GP ATD, which has no moving parts and extracts clots by suction in a vortex flow pattern. METHODS: The GP ATD is used to extract porcine blood clots inserted into the middle cerebral artery (MCA) of a model of the circle of Willis, and from porcine aorta. RESULTS: The GP ATD is navigable around the acute angles of the circle of Willis model and successfully extracts clots that cause complete occlusion of the MCA. There is a strong correlation between the pressure required for clot extraction (mean 31.8, range 30-34 kPa) and its mass (mean 0.08, range 0.03-0.13 g). Complete clot extraction can be demonstrated by computed tomography scanning. Lysis of a 0.15-g thrombus using alteplase at a concentration of 3.4 microg/mL was more effective when delivered and extracted via the GP ATD than via a catheter without the GP ATD or delivered systemically in our circle of Willis model and extracted without suction (clot mass after extraction 0.07, 0.09, and 0.11 g, respectively). Histologic examination does not show evidence of damage of the arterial wall caused by clot extraction at suction pressures of up to 30 kPa via the GP ATD. CONCLUSION: The GP ATD appears to effectively extract blood clots from models of the MCA without significant clot fragmentation and damage to the arterial wall. Further experiments using arteries in situ are required to confirm these findings.

An investigation into the design of a device to treat haemorrhagic stroke.

In this study, we present the design considerations of a device to assist in the potential treatment of hemorrhagic stroke with the aim of stopping blood from flowing out into brain tissue. We present and model three designs for the clinical scenarios when saccular aneurysms rupture in the middle cerebral artery in the brain. We evaluate and model these three designs using computer aided design software, SolidWorks, which allows the devices to be tested using finite element analysis and also enables us to justify that the materials chosen were suitable for potential use. Computational fluid dynamics modelling were used to demonstrate and analyse the flow of blood through the artery under conditions of normal and ruptured states. We conclude that our device could potentially be useful in the treatment of hemorrhagic stroke, and the modelling process is useful in assisting in determining the performance of our devices.

The "GP" mechanical thrombectomy device.

We present data concerning the extraction of clots using the newly invented "GP" mechanical thrombectomy device (MTD). Artificial and porcine clots of various lengths were used in plastic tube models of an artery. We investigate the pressures and times taken for clot extraction together with the volumes of fluid extracted. We also investigate the impact of using a funnel structure mounted on the end of the device, on clot removal times and fluid removed. Finally, we present results involving clot extraction from the posterior popliteal artery of a cadaver. Our data indicated that: The embedded GP MTD is the most effective device regarding artificial and porcine blood clot removal. This result is consistent with previous published data on this device. The GP MTD was effective in removing clots positioned in the posterior popliteal artery of a cadaver. The embedded GP device removes less fluid compared with the end-mounted GP device. This confirms previous studies. There appears to be a relationship between funnel angle and pressure. Lower extraction pressures are required for larger funnel angles mounted on the GP device. Shorter times of clot extraction are required for larger funnel angles.

Numerical modelling of blood clot extraction by aspiration thrombectomy. Evaluation of aspiration catheter geometry.

Aspiration thrombectomy is one of the most effective systems for blood clot removal and vessel recanalization. We present the results of a study involving the modelling and extraction of blood clots in the arteries of the human body using the following computer tools: Bond-Graph methodology for the fluid domain and Multi-Body Simulation for the mechanical domain. The modelling for the mechanical domain focuses on the clot and the distal end section of an aspiration device. Our final model considers an elastic characterization of the blood clot with progressive detachment from the vessel wall. We conclude that the results of such modelling could potentially improve the effectiveness of blood clot removal by reducing the risk of clot fragmentation. Such modelling could also potentially provide an adjunct technique in improving recanalization of arteries over a range of given parameters (mechanical properties of the vessel, mechanical properties of the blood clot, blood clot length, suction pressure, catheter - clot distance, catheter shape, catheter diameter and vessel occlusion).

Developing a technique to measure intra-limb coordination in gait: applicable to children with cerebral palsy.

Lower limb intra-limb coordination was investigated using sagittal plane kinematic data extracted from gait data recorded using a Vicon system (Vicon Motion Systems Ltd., Oxford, UK) of 20 normal (N) and 20 children with cerebral palsy (CP). Walking speed, maximum and minimum flexion and range of motion (ROM) were calculated. The repeatability of the data was checked by calculating the coefficient of multiple correlation. Data were also processed to determine angular velocity of hip and knee joints. A logical spreadsheet was devised to determine when both joints moved in the same direction (in-phase), in different directions (antiphase, AP) or if either joint was immobile (JS). In-phase joint motion was further subdivided into in-phase flexion (IPF) and in-phase extension (IPE), which comprises in-phase during stance phase (IPEst) and in-phase during swing phase (IPEsw). Data were processed using two threshold values for angular velocity below which the joint was considered to be immobile. The threshold values used were 0.05 degrees /% of gait cycle and 0.025 degrees /% of gait cycle. Children with cerebral palsy had reduced ROM and walked more slowly than normal children. There are significant differences between N and cerebral palsy coordination phases with marginally greater significance at the 0.05 degrees /% threshold for most component parameters; the exception being in-phase flexion. It is therefore suggested that this threshold value (0.05 degrees /%) is used for future work.

Tissue Morphology and Gene Expression Characterisation of Transplantable Adenocarcinoma Bearing Mice Exposed to Fluorodeoxyglucose-Conjugated Magnetic Nanoparticles.

Fluorodeoxyglucose-conjugated magnetic nanoparticles, designed to target cancer cells with high specificity when heated by an alternating magnetic field, could provide a low-cost, non-toxic treatment for cancer. However, it is essential that the in vivo impacts of such technologies on both tumour and healthy tissues are characterised fully. Profiling tissue gene expression by semi-quantitative reverse transcriptase real-time PCR can provide a sensitive measurement of tissue response to treatment. However, the accuracy of such analyses is dependent on the selection of stable reference genes. In this study, we determined the impact of fluorodeoxyglucose-conjugated magnetic nanoparticles on tumour and non-tumour tissue gene expression and morphology in MAC16 adenocarcinoma established male NMRI mice. Mice received an injection of 8 mg/kg body weight fluorodeoxyglucose-conjugated magnetic nanoparticles either intravenously in to the tail vein, directly into the tumour or subcutaneously directly overlying the tumour. Tissues from mice were sampled between 70 minutes and 12 hours post injection. Using the bioinformatic geNorm tool, we established the stability of six candidate reference genes (Hprt, Pgk1, Ppib, Sdha, Tbp and Tuba); we observed Pgk1 and Ppib to be the most stable. We then characterised the expression profiles of several apoptosis genes of interest in our adenocarcinoma samples, observing differential expression in response to mode of administration and exposure duration. Using histological assessment and fluorescent TUNNEL staining, we observed no detrimental impact on either tumour or non-tumour tissue morphology or levels of apoptosis. These observations define the underlying efficacy of fluorodeoxyglucose-conjugated magnetic nanoparticles on tumour and non-tumour tissue morphology and gene expression, setting the basis for future studies.

An in-vivo pilot study into the effects of FDG-mNP in cancer in mice.

PURPOSE: Previously, fluorodeoxy glucose conjugated magnetite nanoparticles (FDG-mNPs) injected into cancer cells in conjunction with the application of magnetic hyperthermia have shown promise in new FDG-mNPs applications. The aim of this study was to determine potential toxic or unwanted effects involving both tumour cells and normal tissue in other organs when FDG-mNPs are administered intravenously or intratumourally in mice. MATERIALS AND METHODS: FDG-mNPs were synthesized. A group of six prostate-tumour bearing mice were injected with 23.42 mg/ml FDG-mNPs (intravenous injection, n = 3; intratumoural injection into the prostate tumour, n = 3). Mice were euthanized and histological sampling of tissue was conducted for the prostate tumour, as well as for lungs, lymph nodes, liver, kidneys, spleen, and brain, at 1 hour (n = 2) and 7 days (n = 4) post-injection. A second group of two normal (non-cancerous) mice received the same injection intravenously into the tail vein and were euthanised at 3 and 6 months post-injection, respectively, to investigate if FDG-mNPs remained in organs at those time points. RESULTS: In prostate-tumour bearing mice, FDG-mNPs concentrated in the prostate tumour, while relatively small amounts were found in the organs of other tissues, particularly the spleen and the liver; FDG-mNP concentrations decreased over time in all tissues. In normal mice, no detrimental effects were found in either mouse at 3 or 6 months. CONCLUSION: Intravenous or intratumoural FDG-mNPs can be safely administered for effective cancer cell destruction. Further research on the clinical utility of FDG-mNPs will be conducted by applying hyperthermia in conjunction with FDG-mNPs in mice.

A new device for the treatment of thromboembolic strokes.

We present data concerning the design and testing of a new clot-sucking device for possible use in the extraction of blood clots that arise during thromboembolic strokes. Our data indicate that a jelly-like mass of 0.02 g (of an artificial clot) can be removed from plastic tubing using this vortex-creating device, which has an optimal diameter of 1 mm, in conjunction with a 6F arterial catheter that is 110-cm long and using applied suction pressures of about 50 cm Hg.

Effects of Fluorodeoxyglucose Conjugated and Unconjugated Iron Oxide Magnetic Nanoparticles on Macrophages: a Pilot Study

Abstract The effects of fluorodeoxyglucose conjugated iron oxide magnetic nanoparticles (FDGMNP) on macrophages are presented using a yeast substrate. Iron oxide magnetic nanoparticles (MNP) were synthesized by partially reducing FeCl3, then conjugated with (3-aminopropyl) triethoxysilane (APTES) after silication with tetraethyl orthosilicate. Silanated MMP nanoparticles were combined with fluorodeoxyglucose (FDG). Fluorodeoxyglucose iron oxide magnetic nanoparticles (FDGMNP) and its unconjugated control (MNP) were added (1mL) to the cells from the murine macrophage-like, Abelson murine leukemia virus transformed cell line RAW 264.7 (American Type Culture Collection number TIB-71) cell culture wells at different concentrations from 90-3.6 μg/mL. Cells were placed on the magnet plate for 30 min before incubating at 37°C, 5% CO2 overnight. MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide tetrazolium) assay was performed to measure cell viability. Our results demonstrate that iron based nanoparticles can be linked to macrophages (elements of the immune system that attack bacteria) without the function of the macrophages being affected, ie no detrimental effects to the macrophages were evident in these experiments. We conclude that neither FDGMNP nor MNP had a detrimental effect on macrophage function.

A Pilot Study Into the Use of FDG-mNP as an Alternative Approach in Neuroblastoma Cell Hyperthermia.

Herein, we present a pilot study concerning the use of fluorodeoxy glucose conjugated magnetite nanoparticles (FDG-mNP) as a potential agent in magnetic nanoparticle mediated neuroblastoma cancer cell hyperthermia. This approach makes use of the 'Warburg effect', utilizing the fact that cancer cells have a higher metabolic rate than normal cells. FDG-mNP were synthesized, then applied to the SH-SY5Y neuroblastoma cancer cell line and exposed to an ac magnetic field. 3D Calorimetry was performed on the FDG-mNP compound. Simulations were performed using SEMCAD X software using Thelonious, (an anatomically correct male child model) in order to understand more about the end requirements with respect to cancer cell destruction. We investigated FDG-mNP mediated neuroblastoma cytotoxicity in conjunction with ac magnetic field exposure. Results are presented for 3D FDG-mNP SAR mnp (10.86 ± 0.99 W/g of particles) using a therapeutic dose of 0.83 mg/ mL. Human model simulations suggest that 43 W/kg SAR Theo would be required to obtain 42 °C within the centre of a liver tumor (Tumor size, bounding box x = 64, y = 61, z = 65 [mm]), and that the temperature distribution is inhomogeneous within the tumor. Our study suggests that this approach could potentially be used to increase the temperature within cells that would result in cancer cell death due to hyperthermia. Further development of this research will also involve using whole tumors removed from living organisms in conjunction with magnetic resonance imaging and positron emission tomography.

Antiviral efficacy of abacavir in antiretroviral therapy-experienced adults harbouring HIV-1 with specific patterns of resistance to nucleoside reverse transcriptase inhibitors.

OBJECTIVE: To evaluate HIV-1 reverse transcriptase genotypic and phenotypic indicators of resistance to abacavir (ABC) as predictors of ABC antiviral efficacy. DESIGN: The study was a retrospective, combined analysis of five multicentre trials in which ABC was added as a single agent to background antiretroviral therapy in experienced adults. METHODS: Baseline HIV-1 genotype and phenotypic susceptibility to ABC were determined and the association of genotype and phenotype with virological response after addition of ABC was analysed. RESULTS: Overall, 68% of these therapy-experienced subjects had a virological response (>0.5 log10 or <400 copies/ml; 42% <400 copies/ml) 4 weeks after addition of ABC. Multivariable analyses revealed no significant difference in the response rate between subjects with wild-type virus and those carrying virus with 1-2 nucleoside reverse transcriptase inhibitor (NRTI)-associated mutations. At the 4-week time-point subjects harbouring virus with > or = 3 mutations associated with NRTI resistance were significantly less likely to respond to ABC than were subjects harbouring wild-type virus (P=0.015). However, at the last viral RNA measurement after addition of ABC (12-28 weeks), > or = 4 mutations were required to diminish virological response significantly (P=0.012). Phenotypic resistance was also predictive of antiviral response. Significant breakpoints were identified for virological responses for the PhenoSense HIV assay and the Antivirogram assay. CD4 responses generally paralleled the antiviral responses with a median increase of 55 cells/microl by weeks 12-28. CONCLUSIONS: Virological response to ABC may be diminished significantly by multiple NRTI-associated mutations and/or by reductions in phenotypic susceptibility to ABC. However, many subjects with baseline samples showing evidence of resistance to NRTIs respond to ABC.

Safety and efficacy of the Aperio thrombectomy device when compared to the Solitaire AB/FR and the Revive devices in a pulsatile flow system.

BACKGROUND AND PURPOSE: There are a limited number of studies comparing the Aperio mechanical thrombectomy device to other stent-based devices. In this paper, we compared the Aperio thrombectomy device to the Solitaire AB, FR and Revive devices in a model of the middle cerebral artery (MCA) within a modified pulsatile flow system. METHODS: Thrombi made of lamb's blood were placed into a pulsatile flow system perfused with Hartmann's solution at 80 bpm with a mean pressure of 90 mm Hg. 30 experiments were run with each device. RESULTS: Recanalization rates were similar for all three devices (90% with the Solitaire AB, FR, 80% with the Revive, and 90% with the Aperio). The mean number of attempts to retrieve the thrombus was also similar for all three devices (1.7 with the Solitaire AB, FR, 2.1 with the Revive, 1.6 with the Aperio). Clot fragmentation and embolization rates revealed no statistical significance but there was a trend towards lower embolization rates with the Aperio (23% compared to 40% with the Solitaire AB, FR and 47% with the Revive). The Aperio was the fastest to recanalize the MCA (mean of 66 seconds compared to 186 seconds for the Solitaire AB, FR and 169 seconds for the Revive). CONCLUSIONS: In this in vitro setting, the Aperio device seems to be an efficacious and safe device when compared to other similar clinically used mechanical thrombectomy devices. Larger clinical trials are warranted.

Comparison of the Effectiveness of Three Methods of Recanalization in a Model of the Middle Cerebral Artery: Thrombus Aspiration via a 4F Catheter, Thrombus Aspiration via the GP Thromboaspiration Device, and Mechanical Thrombectomy Using the Solitaire Thrombectomy Device.

Introduction. This paper compares different approaches to recanalization in a model of the middle cerebral artery (MCA). Methods. An occlusive thrombus (lamb's blood) was introduced into the MCA of a model of the cerebral circulation perfused with Hartmann's solution (80 pulsations/min, mean pressure 90 mm Hg). Three methods of clot retrieval were tested: thrombus aspiration via a 4F catheter (n = 26), thrombus aspiration via the GP thrombus aspiration device (GPTAD) (n = 30), and mechanical thrombectomy via the Solitaire Device (n = 30). Results. Recanalization rate was similar for all 3 approaches (62%, 77%, and 85%). Time to recanalization was faster with aspiration devices (41 SD 42 s for 4F and 61 SD 21 s for GPTAD) than with the Solitaire (197 SD 64 s P < .05 Kruksal-Wallis). Clot fragmentation was the same in the Solitaire (23%) and the GPTAD (23%), but higher with the 4F (53%, P < .05). Conclusion. In this model, thrombus aspiration was faster than mechanical thrombectomy, and similarly effective at recanalization. These results should be confirmed in vivo.