Benjamin B. Williams

Quantitative tumor oxymetric images from 4D electron paramagnetic resonance imaging (EPRI): methodology and comparison with blood oxygen level-dependent (BOLD) MRI.

This work presents a methodology for obtaining quantitative oxygen concentration images in the tumor‐bearing legs of living C3H mice. The method uses high‐resolution electron paramagnetic resonance imaging (EPRI). Enabling aspects of the methodology include the use of injectable, narrow, single‐line triaryl methyl spin probes and an accurate model of overmodulated spectra. Both of these increase the signal‐to‐noise ratio (SNR), resulting in high resolution in space (1 mm)3 and oxygen concentrations (∼3 torr). Thresholding at 15% the maximum spectral amplitude gives leg/tumor shapes that reproduce those in photographs. The EPRI appears to give reasonable oxygen partial pressures, showing hypoxia (∼0–6 torr, 0–103 Pa) in many of the tumor voxels. EPRI was able to detect statistically significant changes in oxygen concentrations in the tumor with administration of carbogen, although the changes were not increased uniformly. As a demonstration of the method, EPRI was compared with nearly concurrent (same anesthesia) T  2* /blood oxygen level‐dependent (BOLD) MRI. There was a good spatial correlation between EPRI and MRI. Homogeneous and heterogeneous T  2* /BOLD MRI correlated well with the quantitative EPRI. This work demonstrates the potential for EPRI to display, at high spatial resolution, quantitative oxygen tension changes in the physiologic response to environmental changes. Magn Reson Med 49:682–691, 2003.

Imaging spin probe distribution in the tumor of a living mouse with 250 MHz EPR : Correlation with BOLD MRI

Electron paramagnetic resonance imaging (EPRI) promises to provide new insights into the physiology of tissues in health and disease. Understanding the in vivo imaging capability of this new modality requires comparison with other physiologically responsive techniques. Here, an initial comparison between 2D EPR spatial imaging of a narrow single line injectable paramagnetic trityl spin probe and 2D slice‐selected carbogen subtraction BOLD MRI is presented. The images were obtained from the same FSa fibrosarcoma grown in the leg of a C3H mouse. This tumor was unusual in comparison with others imaged with subtraction BOLD MRI because of its peripheral distribution of intensity. The spatial distribution of the EPR spin probe showed the same peripheral distribution. The pixel resolutions of these images are comparable. These images provide an early in vivo comparison of EPRI with a well‐established imaging modality. The comparison validates the in vivo distribution of spin probe as imaged with EPRI, and provides a proof of principle for the comparison of BOLD and EPRI. Magn Reson Med 47:634–638, 2002.

The role of oxygen during fracture healing

Oxygen affects the activity of multiple skeletogenic cells and is involved in many processes that are important for fracture healing. However, the role of oxygen in fracture healing has not been fully studied. Here we systematically examine the effects of oxygen tension on fracture healing and test the ability of hyperoxia to rescue healing defects in a mouse model of ischemic fracture healing. Mice with tibia fracture were housed in custom-built gas chambers and groups breathed a constant atmosphere of 13% oxygen (hypoxia), 21% oxygen (normoxia), or 50% oxygen (hyperoxia). The influx of inflammatory cells to the fracture site, stem cell differentiation, tissue vascularization, and fracture healing were analyzed. In addition, the efficacy of hyperoxia (50% oxygen) as a treatment regimen for fracture nonunion was tested. Hypoxic animals had decreased tissue vascularity, decreased bone formation, and delayed callus remodeling. Hyperoxia increased tissue vascularization, altered fracture healing in un-complicated fractures, and improved bone repair in ischemia-induced delayed fracture union. However, neither hypoxia nor hyperoxia significantly altered chondrogenesis or osteogenesis during early stages of fracture healing, and infiltration of macrophages and neutrophils was not affected by environmental oxygen after bone injury. In conclusion, our results indicate that environmental oxygen levels affect tissue vascularization and fracture healing, and that providing oxygen when fractures are accompanied by ischemia may be beneficial.

Spectral fitting: The extraction of crucial information from a spectrum and a spectral image

A highly accurate line‐width simulation computer program is used that can account for both high amplitude and frequency of the Zeeman modulation in an electron paramagnetic resonance (EPR) experiment. This allows for the overmodulation of EPR lines to increase signal‐to‐noise ratio (SNR) in EPR spectra and spectroscopic images, without any sacrifice in the determination of the intrinsic line width (1/γ · T2e). The technique was applied to continuous‐wave EPR spectroscopic images of a narrow, single‐line trityl spin probe wherein a full EPR spectrum was extracted from each 3D spatial voxel. Typical improvements are a three‐ to fivefold increase in SNR in the high‐gradient projections in the image and a reduction in the standard deviation (SD), by a factor of 3, of the line widths in the low‐gradient domain. This method is a general one that is also applicable to the analysis of conventional 14N or 15N nitroxide spin probes. Magn Reson Med 49:1175–1180, 2003.

Radiation Dose Prediction Using Data on Time to Emesis in the Case of Nuclear Terrorism

Abstract Demidenko, E., Williams, B. B. and Swartz, H. M. Radiation Dose Prediction Using Data on Time to Emesis in the Case of Nuclear Terrorism. Radiat. Res. 171, 310–319 (2009). A rigorous statistical analysis of the retrospective estimation of radiation dose received using time to emesis and its uncertainty is provided based on 108 observations associated with accidents with significant exposures to ionizing radiation in the period 1956–2001. The standard error, confidence interval, specificity and sensitivity, and Receiver Operating Characteristic (ROC) curve are used to characterize the uncertainty of the dose prediction. The relative error of the dose prediction using time to emesis data is about 200%. Consequently, if D is the dose assessment, the 95% confidence interval is approximately (D/4, 4D). Our assessment of the precision is applied to computation of the probabilities in triage medical management in the case of a nuclear terrorism event. We also note several factors that indicate that there are additional problems in the use of time to emesis for triage, including a lack of consideration of individuals that do not vomit, differences between the conditions under which the data were obtained and the conditions under which they are likely to be used, and the potential for the incidence of vomiting to be altered by factors unrelated to radiation exposure such as psychogenic factors and the use of emetic agents. In summary, while time to emesis is a rapid and inexpensive method for estimating the radiation dose, it should be used with caution because it is imprecise and may lead to a very high false positive rate. More reliable methods for after-the-fact assessment of radiation dose are needed to complement the use of time to emesis.

Seizure‐induced formation of isofurans: novel products of lipid peroxidation whose formation is positively modulated by oxygen tension

We have previously shown that seizures induce the formation of F2‐isoprostanes (F2‐IsoPs), one of the most reliable indices of oxidative stress in vivo. Isofurans (IsoFs) are novel products of lipid peroxidation whose formation is favored by high oxygen tensions. In contrast, high oxygen tensions suppress the formation of F2‐IsoPs. The present study determined seizure‐induced formation of IsoFs and its relationship with cellular oxygen levels (pO2). Status epilepticus (SE) resulted in F2‐IsoP and IsoF formation, with overlapping but distinct time courses in hippocampal subregions. IsoF, but not F2‐IsoP formation coincided with mitochondrial oxidative stress. SE resulted in a transient decrease in hippocampal pO2 measured by in vivo electron paramagnetic resonance oximetry suggesting an early phase of seizure‐induced hypoxia. Seizure‐induced F2‐IsoP formation coincided with the peak hypoxia phase, whereas IsoF formation coincided with the ‘reoxygenation’ phase. These results demonstrate seizure‐induced increase in IsoF formation and its correlation with changes in hippocampal pO2 and mitochondrial dysfunction.

The effect of oxygen therapy on brain damage and cerebral pO(2) in transient focal cerebral ischemia in the rat.

We examined the effect of hyperbaric oxygen (HBO) and normobaric oxygen (NBO) on neurologic damage and brain oxygenation before and after focal cerebral ischemia in rats. A middle cerebral artery occlusion (MCAO)/reperfusion rat model was used. The rats were sacrificed 22 h after reperfusion, and the infarct volume was evaluated. In study A, HBO (2.0 ATA), NBO (100% oxygen) and normobaric air (NBA) were each administered for 60 min in five different rat groups. The sizes of the infarcts after HBO and NBO applied during ischemia were 8.8 +/- 2.8% and 22.8 +/- 3.7% respectively of the ipsilateral non-occluded hemisphere. The infarct size after HBO applied during ischemia was statistically smaller than for NBO and NBA exposure (p < 0.01). In study B, cerebral pO(2) was measured before and after MCAO and HBO exposure (2.0 ATA for 60 min) in six rats using electron paramagnetic resonance (EPR) oximetry. The pO(2) in the ischemic hemisphere fell markedly following ischemia, while the pO(2) in the contralateral hemisphere remained within the normal range. Measurements of the pO(2) performed minutes after HBO exposure did not show an increase in the ischemic or normal hemispheres. The mean relative infarct size was consistent with the changes observed in study A. These data confirm the neuroprotective effects of HBO in cerebral ischemia and indicate that in vivo EPR oximetry can be an effective method to monitor the cerebral oxygenation after oxygen therapy for ischemic stroke. The ability to measure the pO(2) in several sites provides important information that should help to optimize the design of hyperoxic therapies for stroke.

Clinical EPR: Unique Opportunities and Some Challenges

Electron paramagnetic resonance (EPR) spectroscopy has been well established as a viable technique for measurement of free radicals and oxygen in biological systems, from in vitro cellular systems to in vivo small animal models of disease. However, the use of EPR in human subjects in the clinical setting, although attractive for a variety of important applications such as oxygen measurement, is challenged with several factors including the need for instrumentation customized for human subjects, probe, and regulatory constraints. This article describes the rationale and development of the first clinical EPR systems for two important clinical applications, namely, measurement of tissue oxygen (oximetry) and radiation dose (dosimetry) in humans. The clinical spectrometers operate at 1.2 GHz frequency and use surface-loop resonators capable of providing topical measurements up to 1 cm depth in tissues. Tissue pO2 measurements can be carried out noninvasively and repeatedly after placement of an oxygen-sensitive paramagnetic material (currently India ink) at the site of interest. Our EPR dosimetry system is capable of measuring radiation-induced free radicals in the tooth of irradiated human subjects to determine the exposure dose. These developments offer potential opportunities for clinical dosimetry and oximetry, which include guiding therapy for individual patients with tumors or vascular disease by monitoring of tissue oxygenation. Further work is in progress to translate this unique technology to routine clinical practice.

Clinical electron paramagnetic resonance (EPR) oximetry using India ink.

Electron paramagnetic resonance (EPR) oximetry can be used to provide direct absolute measurements of pO(2) in living tissue using India ink as an O(2) reporter. In vivo measurements are made using low frequency (1.2 GHz) EPR spectroscopy and surface loop resonators, which enable measurements to be made at superficial sites through a non-invasive (after placing the ink in the tissues) and repeatable measurement procedure. Ongoing EPR oximetry studies in human subjects include measurement of subcutaneous pO(2) in the feet of healthy volunteers to develop procedures that could be used in the treatment of peripheral vascular disease and oximetry in tumors during courses of radiation and chemotherapy, to follow pO(2) so oxygen-dependent therapies can be optimized. In each case, we aim to provide quantitative measurements of tissue pO(2) which will aid physicians in the characterization of disease status and the effects of therapeutic measures, so that treatments can be applied with optimal effectiveness by taking into account the oxygen-dependent aspects of the therapy. The overall goal is to enhance clinical outcomes. Oximetry measurements of subcutaneous tissue on dorsal and plantar foot surfaces have been made in 9 volunteers, with measurements ongoing for each and the longest set of measurements carried out successfully over the last 5 years. Tumor oximetry measurements have been performed in tumor tissues of 10 patients during courses of radiation and chemotherapy. Tumor types include melanoma, basal cell, soft tissue sarcoma, and lymphoma, and measurement sites have ranged from the feet to the scalp. These studies demonstrate the feasibility of EPR oximetry in a clinical setting and the potential for more widespread use in the treatment of these and other oxygen-dependent diseases.

DEVELOPMENT OF IN VIVO TOOTH EPR FOR INDIVIDUAL RADIATION DOSE ESTIMATION AND SCREENING

The development of in vivo EPR has made it feasible to perform tooth dosimetry measurements in situ, greatly expanding the potential for using this approach for immediate screening after radiation exposures. The ability of in vivo tooth dosimetry to provide estimates of absorbed dose has been established through a series of experiments using unirradiated volunteers with specifically irradiated molar teeth placed in situ within gaps in their dentition and in natural canine teeth of patients who have completed courses of radiation therapy for head and neck cancers. Multiple measurements in patients who have received radiation therapy demonstrate the expected heterogeneous dose distributions. Dose-response curves have been generated using both populations and, using the current methodology and instrument, the standard error of prediction based on single 4.5-min measurements is approximately 1.5 Gy for inserted molar teeth and between 2.0 and 2.5 Gy in the more irregularly shaped canine teeth. Averaging of independent measurements can reduce this error significantly to values near 1 Gy. Developments to reduce these errors are underway, focusing on geometric optimization of the resonators, detector positioning techniques, and optimal data averaging approaches. In summary, it seems plausible that the EPR dosimetry techniques will have an important role in retrospective dosimetry for exposures involving large numbers of individuals.