Electron Paramagnetic Resonance Imaging Research Articles

Spin Lattice Relaxation EPR pO2 Images May Direct the Location of Radiation Tumor Boosts to Enhance Tumor Cure.

Radiation treatment success and high tumor oxygenation and success have been known to be highly correlated. This suggests that radiation therapy guided by images of tumor regions with low oxygenation, oxygen-guided radiation therapy (OGRT) may be a promising enhancement of cancer radiation treatment. Before applying the technique to human subjects, OGRT needs to be tested in animals, most easily in rodents. Electron paramagnetic resonance imaging provides quantitative maps of tissue and tumor oxygen in rodents with 1 mm spatial resolution and 1 torr pO2 resolution at low oxygen levels. The difficulty of using mouse models is their small size and that of their tumors. To overcome this we used XRAD225Cx micro-CT/ therapy system and 3D printed conformal blocks. Radiation is delivered first to a uniform 15% tumor control dose for the whole tumor and then a boost dose to either hypoxic tumor regions or equal volumes of well oxygenated tumor. Delivery of the booster dose used a multiple beam angles to deliver radiation beams whose shape conforms to that of all hypoxic regions or fully avoids those regions. To treat/avoid all hypoxic regions we used individual radiation blocks 3D-printed from acrylonitrile butadiene styrene polymer infused with tungsten particles fabricated immediately after imaging to determine regions with pO2 less than 10 torr. Preliminary results demonstrate the efficacy of the radiation treatment with hypoxic boosts with syngeneic FSa fibrosarcoma tumors in the legs of C3H mice.

Quantitative imaging of pO2 in orthotopic murine gliomas: hypoxia correlates with resistance to radiation

Hypoxia is considered one of the microenvironmental factors associated with the malignant nature of glioblastoma. Thus, evaluating intratumoural distribution of hypoxia would be useful for therapeutic planning as well as assessment of its effectiveness during the therapy. Electron paramagnetic resonance imaging (EPRI) is an imaging technique which can generate quantitative maps of oxygen in vivo using the exogenous paramagnetic compound, triarylmethyl and monitoring its line broadening caused by oxygen. In this study, the feasibility of EPRI for assessment of oxygen distribution in the glioblastoma using orthotopic U87 and U251 xenograft model is examined. Heterogeneous distribution of pO2 between 0 and 50 mmHg was observed throughout the tumours except for the normal brain tissue. U251 glioblastoma was more likely to exhibit hypoxia than U87 for comparable tumour size (median pO2; 29.7 and 18.2 mmHg, p = .028, in U87 and U251, respectively). The area with pO2 under 10 mmHg on the EPR oximetry (HF10) showed a good correlation with pimonidazole staining among tumours with evaluated size. In subcutaneous xenograft model, irradiation was relatively less effective for U251 compared with U87. In conclusion, EPRI is a feasible method to evaluate oxygen distribution in the brain tumour.

Reoxygenation-Based Radiation Therapy Improve the Tumor Control

Reoxygenation has a deep impact on tumor control in fractionated radiotherapy. With the improvements in radiotherapy technology including IMRT and SRT, it has become much more important to understand the reoxygenation phenomenon clearly. Pulsed electron paramagnetic resonance imaging (EPRI) is a novel imaging method to directly monitor the pO2 in a tumor on a quantitative basis. The purpose of this work was to assess the reoxygenation-based radiation therapy detected by EPRI improve the tumor control using murine models of human cancer. HT29 solid tumors were formed by injecting 1 × 106 cells s.c. into the right hind legs of nude mice. The all experiments were started when tumors grew to approximately 800 mm3. In EPR imaging, it took 12 min to obtain a data set for a 3D pO2 image. The spatial resolution was 1.8 mm. First, we analyzed how irradiation dose affected the reoxygenation, and EPR imaging was performed before and at 30 minutes, 60 minutes, 2 hours, 18 hours, and 24 hours after 3Gy, 10 Gy, 20Gy irradiation in HT29 tumors (n = 6 per time point). Second, we analyzed the tumor control by additional 3Gy, 10 Gy, 20Gy irradiation at 30 minutes, 60 minutes, 2 hours, 18 hours, and 24 hours after 3Gy, 10 Gy, 20Gy irradiation in HT29 tumors (n = 6 per time point). Median pO2 of HT29 tumors was 15.9 ± 1.0 before irradiation, After 3 Gy, 10 Gy, and 20 Gy irradiation, it was, respectively, 15.4 ± 1.1, 14.1 ± 0.3, and 12.7 ± 1.5 at 30 minutes, 14.0 ± 1.0, 13.6 ± 1.0, and 11.8 ± 1.3 at 60 minutes, 15.3 ± 0.6, 13.7 ± 0.5, and 11.4 ± 0.8 at 2 hours, 15.2 ± 0.7, 13.6 ± 1.0, and 13.7 ± 0.5 at 18 hours, 15.4 ± 0.6, 13.6 ± 0.8, and 13.9 ± 0.6 at 24 hours. We could clearly show the phenomenon of reoxygenation after irradiation in HT29 tumors by using EPR pO2 imaging. The median pO2 decreased the most in about 60 minutes after 3Gy, 10 Gy, 20Gy irradiation and the median pO2 was recovered in 24 hours, but as the irradiation dose increased, it took more time to recover, and the median pO2 in 24 hours decreased as the irradiation dose increased. The tumor control by additional irradiation at 60 minutes was worst in additional irradiation at the other after initial 3Gy, 10 Gy, and 20Gy irradiation, and also the tumor control by additional irradiation at 24 hours was best in additional irradiation at the other after initial 3Gy, 10 Gy, and 20Gy irradiation. The difference in radiation response in 3Gy, 10 Gy, and 20Gy irradiation was consistent with the median pO2 of the HT-29 tumor. We could clearly show the phenomenon and effect of reoxygenation in HT29 tumors by using EPR pO2 imaging, and also indicated the possibility and importance of a new radiation therapy concept such as reoxygenation-based radiation therapy that improved the tumor control.

Transplantation of rat bone marrow-derived mesenchymal stem cells regulates oxidative stress in Alzheimer’s disease transgenic mouse model

Background: It is widely recognized that increased oxidative damage on neurons is an early event in Alzheimer’s disease (AD). It has been reported that mesenchymal stem cells (MSC) can reduce oxidative stress on many kinds of disease. Our group have shown that the changes in redox status of the AD model mouse brain can be visualized noninvasively using three-dimensional electron paramagnetic resonance (EPR) imaging (Free Radic Biol Med. 2015).

Resonators for In Vivo Imaging: Practical Experience.

Resonators for preclinical electron paramagnetic resonance imaging have been designed primarily for rodents and rabbits and have internal diameters between 16 and 51 mm. Lumped circuit resonators include loop-gap, Alderman-Grant, and saddle coil topologies and surface coils. Bimodal resonators are useful for isolating the detected signal from incident power and reducing dead time in pulse experiments. Resonators for continuous wave, rapid scan, and pulse experiments are described. Experience at the University of Chicago and University of Denver in design of resonators for in vivo imaging is summarized.

Six Decades of Progress in Magnetic Resonance: The Contributions of James S. Hyde

The development of electron paramagnetic resonance (EPR) and magnetic resonance imaging (MRI) over six decades is sketched with an emphasis on the contributions of James S. Hyde. For twenty years starting three years after the first commercial EPR spectrometer was shipped by Varian, he led commercial EPR developments, and then for more than forty years, he led development of instrumentation and biomedical applications of EPR at the Medical College of Wisconsin. It was there that he also made major contributions to MRI, and especially functional MRI.

Design and testing of a 750 MHz CW-EPR digital console for small animal imaging

This paper describes the development of a digital console for three-dimensional (3D) continuous wave electron paramagnetic resonance (CW-EPR) imaging of a small animal to improve the signal-to-noise ratio and lower the cost of the EPR imaging system. A RF generation board, an RF acquisition board and a digital signal processing (DSP) & control board were built for the digital EPR detection. Direct sampling of the reflected RF signal from a resonator (approximately 750MHz), which contains the EPR signal, was carried out using a band-pass subsampling method. A direct automatic control system to reduce the reflection from the resonator was proposed and implemented in the digital EPR detection scheme. All DSP tasks were carried out in field programmable gate array ICs. In vivo 3D imaging of nitroxyl radicals in a mouse’s head was successfully performed.

Time-, spectral- and spatially resolved EPR spectroscopy enables simultaneous monitoring of diffusion of different guest molecules in nano-pores

Diffusion in porous materials is under ongoing active investigation due to its major role in practical applications such as catalysis and chromatography. The complexity of these systems limits the use of the Einstein-Stokes diffusion theory, and it must be distinguished between the microscopic scale of diffusion at a molecular level, which is sensitive to the local surroundings of a diffusing molecule, and the macroscopic scale which takes into account diffusion spanning multiple pores, grain boundaries and inhomogeneity within the material. Here, we employ an in situ approach for quantitative measurements of the diffusion on a macroscopic length scale. For the first time, full time-resolved spectral spatial EPR imaging in combination with the simultaneous iterative reconstruction technique (SIRT) allows the simultaneous observation of the diffusion of two different molecular species inside of an aerogel in a single experiment.

O-94 二次元ESR画像計測による断層摩擦熱の検出(12.岩石・鉱物の破壊と変形,口頭およびポスター発表,一般講演)

O-94 二次元ESR画像計測による断層摩擦熱の検出(12.岩石・鉱物の破壊と変形,口頭およびポスター発表,一般講演)

Abstract 1874: First demonstration that hypoxia targeting with radiation improves tumor control in a mammal tumor model

Abstract For over a century, resistance of radiation to living tissues has been associated with hypoxia, a local lack of molecular oxygen, or low pO2. The focus of this research has been to validate the hypothesis that has been assumed, but not proven over the past century, that radiation treatment focused specifically on hypoxic regions of tumors would improve tumor curability. This research used Electron Paramagnetic Resonance (EPR) imaging to provide absolute pO2 images in volume elements or voxels of murine tumors with 1 torr pO2 resolution and 0.7 mm spatial resolution in FSa fibrosarcomas in the legs of C3H mice. We define hypoxic regions of tumors as those whose image subvolumes or voxels have low pO2, voxels less than 10 torr. We have established EPR pO2 imaging as a reliable locator of relevant radiobiologically relevant hypoxia. To determine if pO2 based dose painting improves tumor cure, we implemented the XRAD225Cx system to deliver gantry based x-ray treatments to mouse tumors accurately registered with EPR pO2 images. All tumors were pretreated with a dose of radiation that had been determined in separate experiments to cure 30% of tumors, a TCD30. Conclusive failure to demonstrate a difference between treating crude spherical volumes including ~ 85% of hypoxic voxels with an additional curative dose compared with treatment with shells of radiation avoiding hypoxia (anti-boosts) proved tenets of radiation biology: a few surviving hypoxic clonogens cause radiation treatment failure. In response, we implemented rapid 3D printing Tungsten loaded, highly conformal plastic blocks to compare treating ~100% of hypoxic tumor voxels with hypoxia avoidance, treating similar volumes of well oxygenated tumor. Only then did we observe significant (p<0.01) tumor control differences between hypoxic boosts and anti-boosts. This is the first validation of the curative effectiveness of focusing hypoxia based dose painting in mammalian tumors. Citation Format: Howard J. Halpern, Boris Epe;, Matthew C. Maggio, Martyna Krzykawska-Serda, Gage H. Redler, Richard C. Miller, Eugene D. Barth, Ralph R. Weichselbaum. First demonstration that hypoxia targeting with radiation improves tumor control in a mammal tumor model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1874. doi:10.1158/1538-7445.AM2017-1874

Rapid-scan EPR imaging

In rapid-scan EPR the magnetic field or frequency is repeatedly scanned through the spectrum at rates that are much faster than in conventional continuous wave EPR. The signal is directly-detected with a mixer at the source frequency. Rapid-scan EPR is particularly advantageous when the scan rate through resonance is fast relative to electron spin relaxation rates. In such scans, there may be oscillations on the trailing edge of the spectrum. These oscillations can be removed by mathematical deconvolution to recover the slow-scan absorption spectrum. In cases of inhomogeneous broadening, the oscillations may interfere destructively to the extent that they are not visible. The deconvolution can be used even when it is not required, so spectra can be obtained in which some portions of the spectrum are in the rapid-scan regime and some are not. The technology developed for rapid-scan EPR can be applied generally so long as spectra are obtained in the linear response region. The detection of the full spectrum in each scan, the ability to use higher microwave power without saturation, and the noise filtering inherent in coherent averaging results in substantial improvement in signal-to-noise relative to conventional continuous wave spectroscopy, which is particularly advantageous for low-frequency EPR imaging. This overview describes the principles of rapid-scan EPR and the hardware used to generate the spectra. Examples are provided of its application to imaging of nitroxide radicals, diradicals, and spin-trapped radicals at a Larmor frequency of ca. 250MHz.

Fast backprojection-based reconstruction of spectral-spatial EPR images from projections with the constant sweep of a magnetic field

In this paper, we introduce a procedure for the reconstruction of spectral-spatial EPR images using projections acquired with the constant sweep of a magnetic field. The application of a constant field-sweep and a predetermined data sampling rate simplifies the requirements for EPR imaging instrumentation and facilitates the backprojection-based reconstruction of spectral-spatial images. The proposed approach was applied to the reconstruction of a four-dimensional numerical phantom and to actual spectral-spatial EPR measurements. Image reconstruction using projections with a constant field-sweep was three times faster than the conventional approach with the application of a pseudo-angle and a scan range that depends on the applied field gradient. Spectral-spatial EPR imaging with a constant field-sweep for data acquisition only slightly reduces the signal-to-noise ratio or functional resolution of the resultant images and can be applied together with any common backprojection-based reconstruction algorithm.

In vivo preclinical cancer and tissue engineering applications of absolute oxygen imaging using pulse EPR

The value of any measurement and a fortiori any measurement technology is defined by the reproducibility and the accuracy of the measurements. This implies a relative freedom of the measurement from factors confounding its accuracy. In the past, one of the reasons for the loss of focus on the importance of imaging oxygen in vivo was the difficulty in obtaining reproducible oxygen or pO2 images free from confounding variation. This review will briefly consider principles of electron paramagnetic oxygen imaging and describe how it achieves absolute oxygen measurements. We will provide a summary review of the progress in biomedical EPR imaging, predominantly in cancer biology research, discuss EPR oxygen imaging for cancer treatment and tissue graft assessment for regenerative medicine applications.

250 MHz passive Q-modulator for reflection resonators.

A simple scheme for dynamically switching the quality factor, Q, of a Loop-Gap Resonator (LGR); working at 250 MHz is presented. The addition of this Q-modulator resulted in 30% improvement in Electron Paramagnetic Resonance imager signal-to-noise ratio. During pulse excitation, this scheme lowered the Q, while higher Q was obtained during signal detection. These conditions favored the image acquisition. The Q-modulator is passive; the transition between different states was actuated by the radio frequency power itself.

Non-invasive imaging of the levels and effects of glutathione on the redox status of mouse brain using electron paramagnetic resonance imaging

Glutathione (GSH) is the most abundant non-protein thiol that buffers reactive oxygen species in the brain. GSH does not reduce nitroxides directly, but in the presence of ascorbates, addition of GSH increases ascorbate-induced reduction of nitroxides. In this study, we used electron paramagnetic resonance (EPR) imaging and the nitroxide imaging probe, 3-methoxycarbonyl-2,2,5,5-tetramethyl-piperidine-1-oxyl (MCP), to non-invasively obtain spatially resolved redox data from mouse brains depleted of GSH with diethyl maleate compared to control. Based on the pharmacokinetics of the reduction reaction of MCP in the mouse heads, the pixel-based rate constant of its reduction reaction was calculated as an index of the redox status in vivo and mapped as a “redox map”. The obtained redox maps from control and GSH-depleted mouse brains showed a clear change in the brain redox status, which was due to the decreased levels of GSH in brains as measured by a biochemical assay. We observed a linear relationship between the reduction rate constant of MCP and the level of GSH for both control and GSH-depleted mouse brains. Using this relationship, the GSH level in the brain can be estimated from the redox map obtained with EPR imaging.

Pro-oxidant activity of α-tocopherol during photooxidative degradation of polyolefins. ESRI and IR microspectroscopy studies

Photooxidation processes inside high-density polyethylene (HDPE) and cyclic olefin copolymer (COC) poly(ethylene-co-norbornene), both stabilized with biological antioxidant α-tocopherol (α-T-OH), has been investigated. Plaques made of the polymers stabilized with α-T-OH were subjected to long-term accelerated weathering in a Weather-Ometer (WOM). Concentration profiles of radicals generated in the plaques in dependence on WOM exposure time were determined by electron spin resonance imaging (ESRI). Concentration profiles of oxidation products and stabilizer, together with local crystallinity profiles inside the polymer plaques were determined by infrared (IR) microspectroscopy. The ESRI and IR profiles, supplemented by light microscopy (LM) and microindentation hardness measurements (MH), revealed the heterogeneity of the oxidation process and, surprisingly, the pro-oxidative activity of α-T-OH. Increasing concentration of products of polymer oxidation was found in the surface layers of the plaques stabilized with α-T-OH since the beginning of WOM treatment unlike the same control experiments with polymer plaques stabilized with HAS Tinuvin® 770, where no oxidation products were observed after the same treatment before HAS depletion. We have confirmed that α-T-OH exhibits antioxidant activity during thermooxidation but it acts as a pro-oxidant agent during photooxidation.

EPR imaging and HPLC characterization of the pigment-based organic free radical in black soybean seeds

We investigated the location and distribution of paramagnetic species in dry black, brown, and yellow (normal) soybean seeds using electron paramagnetic resonance (EPR), X-band (9 GHz) EPR imaging (EPRI), and HPLC. EPR primarily detected two paramagnetic species in black soybean. These two different radical species were assigned as stable organic radical and Mn2+ species based on the g values and hyperfine structures. The signal from the stable radical was noted at g ≈ 2.00 and was relatively strong and stable. Subsequent noninvasive two-dimensional (2D) EPRI of the radical present in black soybean revealed that the stable radical was primarily located in the pigmented region of the soybean coat, with very few radicals observed in the soybean cotyledon (interior). Pigments extracted from black soybean were analyzed using HPLC. The major compound was found to be cyanidin-3-glucoside. Multi-EPR and HPLC results indicate that the stable radical was only found within the pigmented region of the soybean coat, and it could be cyanidin-3-glucoside or an oxidative decomposition product.

Electron spin resonance studies on deuterated nitroxyl spin probes used in Overhauser-enhanced magnetic resonance imaging.

The electron spin resonance studies were carried out for 2mm concentration of 14 N-labeled and 15 N-labeled 3-carbamoyl-2,2,5,5-tetramethyl-pyrrolidine-1-oxyl, 3-carboxy-2,2,5,5-tetramethyl-pyrrolidine-1-oxyl, 3-methoxycarbonyl-2,2,5,5-tetramethyl-pyrrolidine-1-oxyl and their deuterated nitroxyl radicals using X-band electron spin resonance spectrometer. The electron spin resonance line shape analysis was carried out. The electron spin resonance parameters such as linewidth, Lorentzian component, signal intensity ratio, rotational correlation time, hyperfine coupling constant and g-factor were estimated. The deuterated nitroxyl radicals have narrow linewidth and an increase in Lorentzian component, compared with undeuterated nitroxyl radicals. The dynamic nuclear polarization factor was observed for all nitroxyl radicals. Upon 2 H labeling, about 70% and 40% increase in dynamic nuclear polarization factor were observed for 14 N-labeled and 15 N-labeled nitroxyl radicals, respectively. The signal intensity ratio and g-value indicate the isotropic nature of the nitroxyl radicals in pure water. Therefore, the deuterated nitroxyl radicals are suitable spin probes for in vivo/in vitro electron spin resonance and Overhauser-enhanced magnetic resonance imaging modalities. Copyright © 2017 John Wiley & Sons, Ltd.

Feasibility of in vivo three-dimensional T 2* mapping using dicarboxy-PROXYL and CW-EPR-based single-point imaging.

The aim of this study was to demonstrate the feasibility of in vivo three-dimensional (3D) relaxation time T 2* mapping of a dicarboxy-PROXYL radical using continuous-wave electron paramagnetic resonance (CW-EPR) imaging. Isotopically substituted dicarboxy-PROXYL radicals, 3,4-dicarboxy-2,2,5,5-tetra(2H3)methylpyrrolidin-(3,4-2H2)-(1-15N)-1-oxyl (2H,15N-DCP) and 3,4-dicarboxy-2,2,5,5-tetra(2H3)methylpyrrolidin-(3,4-2H2)-1-oxyl (2H-DCP), were used in the study. A clonogenic cell survival assay was performed with the 2H-DCP radical using squamous cell carcinoma (SCC VII) cells. The time course of EPR signal intensities of intravenously injected 2H,15N-DCP and 2H-DCP radicals were determined in tumor-bearing hind legs of mice (C3H/HeJ, male, n=5). CW-EPR-based single-point imaging (SPI) was performed for 3D T 2* mapping. 2H-DCP radical did not exhibit cytotoxicity at concentrations below 10mM. The in vivo half-life of 2H,15N-DCP in tumor tissues was 24.7±2.9min (mean±standard deviation [SD], n=5). The in vivo time course of the EPR signal intensity of the 2H,15N-DCP radical showed a plateau of 10.2±1.2min (mean±SD) where the EPR signal intensity remained at more than 90% of the maximum intensity. During the plateau, in vivo 3D T 2* maps with 2H,15N-DCP were obtained from tumor-bearing hind legs, with a total acquisition time of 7.5min. EPR signals of 2H,15N-DCP persisted long enough after bolus intravenous injection to conduct in vivo 3D T 2* mapping with CW-EPR-based SPI.

Investigating the Distribution of Stable Paramagnetic Species in an Apple Seed Using X-Band EPR and EPR Imaging.

This study investigated the location and distribution of paramagnetic species in apple seeds using electron paramagnetic resonance (EPR) and X-band (9 GHz) EPR imaging (EPRI). EPR primarily detected two paramagnetic species per measured seed. These two different radical species were assigned as stable radicals and Mn2+ species based on the g values and hyperfine components. The signal from the stable radical was noted at g ≈ 2.00 and was strong and relatively stable. The subsequent noninvasive EPRI of the radical present in each seed revealed that the stable radicals were located primarily in the seed coat, with very few radicals observed in the cotyledon of the seed. These results indicate that the stable radical species were only found within the seed coat, and few radical species were found in other seed parts.