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PURPOSE: The potential for malicious use of radiation, or radiation accidents could potentially lead to acute, high radiation doses to the public. Following acute accidental exposure to high doses of radiation, medical intervention is pivotal to the survivability of the patient, and the sooner the appropriate measures are taken the better the odds for survival. Early estimates of acute accidental radiation doses can be determined via biomarkers such as dicentric chromosome analysis or scenario reconstruction using computer software. However, both take valuable time and can be expensive. Increased frequencies of abnormal neutrophils in peripheral blood, referred to as pseudo Pelger-Huët anomalies (PPHAs), have been shown to be potential biomarkers of radiation exposure in several scenarios, including the 1958 Y-12 criticality accident and the radium dial painters. PPHAs are potentially a faster and cheaper quantitative biomarker for radiation exposure, and here they were evaluated in acutely exposed rhesus macaques. METHODS AND MATERIALS: Peripheral blood smears from acutely exposed rhesus macaques were evaluated for the percentage of neutrophils that displayed the PPHA morphology using light microscopy. Irradiated animals received 0 to 8.5 Gy total body radiation using one of two strategies: (1) linear accelerator-produced 6 MV photons delivered at 80 cGy/minute; or (2) Cobalt 60-produced gamma irradiation delivered at 60 cGy/min. Zero dose animals were used to determine a baseline percentage of PPHAs, and blood smears taken periodically throughout the lifetime of exposed animals post-irradiation were used to determine the persistence and biokinetics of PPHAs. RESULTS: The baseline prevalence of the PPHA in rhesus macaques was determined to be 0.58 ± 0.46%. The dose-response curve with doses ranging from 0 Gy to 8.5 Gy (LD90/30) displayed a strong positive correlation between PPHA percentage and acute radiation dose (R(2) of 0.88 p = 3.62 × 10(−22)). Statistically significant differences were found when animals were separated into dose cohorts of 0, 4, 6.4–6.5, and 8–8.5 Gy. The biokinetics model utilized only 4 Gy exposures and blood smears taken periodically over 3.1 years post-irradiation. PPHA morphology increases quickly following irradiation and appears stable over 3.1 years post-irradiation. CONCLUSION: PPHA morphology was confirmed to be present in rhesus macaques, a dose-response relationship was constructed, and it is stable over 3 years post-irradiation. This study demonstrates that PPHA analysis can be a fast and cheap method of biodosimetry. Future studies will work to determine the accuracy of dose determination and lower limits of detection.

Naturally occurring colorectal cancers (CRC) in rhesus macaques share many features with their human counterparts and emerge at similar life stages as in human patients, but the carcinogenesis and molecular background remain elusive, limiting the value of the model for human disease and cancer therapy. In order to better understand this naturally occurring model, we established a diagnostic and staging pipeline including imaging (18F-FDG PET, plain and contrast-enhanced CT), histopathology, and clinical assessments. We observed cancer symptoms and co-morbidities in our cohort (n=16, all Indian-origin, 11 females, mean age at arrival 20.2y) such as hypoalbuminemia, fecal occult blood, and microcytic anemia, as frequently described in human CRC patients. Pathologically, all cancers were right-sided, involving the proximal colon and/or ileocecocolic junction, and most were densely fibrotic, restricting the colonic lumen. Most cancers appeared with glandular morphology and some (18.8%) had mucinous components. Immunohistochemistry revealed loss of MLH1 and PMS2 in 100% of investigated CRCs, indicating mismatch repair deficiency which furthermore resulted in microsatellite instability (PCR & fragment analysis). Whole exome sequencing revealed the close genetic relatedness to human CRCs, particularly exemplified by mutations affecting KRAS (37.5%, e.g., p.G12D), APC (31.3%), TP53 (18.8%, e.g. p.R175H), ARID1A (56.3%), and ALK (43.8%), as similarly annotated in the human COSMIC database. Nonetheless, somatic mutations do not explain the loss of MLH1 in the entire CRC cohort. Transcriptomics on the other hand revealed the transcriptional suppression of MLH1 but not MSH2, MSH6, or PMS2 in rhesus CRC in comparison to adjacent healthy colon. Moreover, comparison of differentially expressed gene sets of rhesus CRC with a human annotated database (IPA) confirmed the disease similarities observed clinically, genetically, and histopathologically. Subsequently, we investigated DNA methylation of the promoter region of MLH1 and retranslated markers for the CpG island methylator phenotype (CIMP) as described in human CRC. While only 56.3% of CRCs were considered CIMP positive (≥3/5 markers hypermethylated), 100% of investigated CRCs exhibited MLH1 promoter hypermethylation. As a result, epigenetic silencing is suggested to suppress MLH1 transcription, cause the loss of MLH1 protein, and drive mismatch repair deficiency and genomic instability in naturally occurring CRC in rhesus macaques. We therefore consider spontaneous, uninduced CRC in rhesus macaques, their treatment-naïve nature, and their unaltered immune competence an outstanding model for human disease and in particular for human cancer immunotherapy. Citation Format: Simon Deycmar, Brendan Johnson, Karina Ray, David Caudell Caudell, John Olson, Greg Dugan, W. Shane Sills, Declan Ryan, Christopher Whitlow, Kiran K. Solingapuram Sai, Betsy Ferguson, Benjamin Bimber, Cassandra Cullin, Brandy Dozier, Emily Romero, Francois Villinger, Armando Burgos, Jeff Chou, Bruno Gomes, Michael Hettich, Maurizio Ceppi, Jehad Charo, J. Mark Cline. Clinical and molecular characterization of naturally-occurring colorectal cancer in rhesus macaques reveals mismatch repair deficiency driven by epigenetic silencing of MLH1 [abstract]. In: Proceedings of the AACR Special Conference on Colorectal Cancer; 2022 Oct 1-4; Portland, OR. Philadelphia (PA): AACR; Cancer Res 2022;82(23 Suppl_1):Abstract nr A021.

Resilience to stressors is a major component of biological aging and may mediate the onset of multimorbidity in older adults. The Wake Forest Non-Human Primate Radiation Survivor Cohort (RSC) provides a novel opportunity to study aging and resilience in 250 rhesus macaques (Macaca mulatta) with single-dose radiation exposures 0–15 years prior and 50 controls with semi-annual clinical, imaging, and biomarker measurements taken over their lifespan. Multimorbidity is extremely common among irradiated animals. Only 38% of animals have none of 20 common chronic diseases, falling to 16% of animals over age 8 and 11% over age 10 (middle-aged animals). 70% of animals have 5 or more diagnoses in this oldest cohort. The presence of any one disease increases the likelihood of having a second, co-morbid condition. Nevertheless, some animals continue disease-free until late in life, highlighting substantial variability in resilience. To identify patterns of multimorbidity, survival curves for each diagnosis were generated for age and time since radiation and k-median clustered resulting in four groupings of aging-associated morbidities. Bone, brain, and gastrointestinal disorders arise 3.5 years after radiation on average, followed by skin, heart, and cataracts. At 4.65 years, animals are at increased risk of being underweight and overweight and developing diabetes, hypertension, and hepatic dysfunction. Tumor, lung, and kidney disorders arise approximately 6 years after exposure. In all cases, these age-related disorders occur significantly earlier in irradiated animals than controls. These findings highlight the clustering of multimorbidities in aging, radiation-challenged primates and the potential of the RSC in studying resilience.

This paper presents a comprehensive study on degradation monitoring of silicon carbide mosfet s and proposes an early warning method to detect aging. The proposed plug-in tool can be integrated to smart gate drivers or directly to power converters. During the accelerated aging tests (power cycling within safe operating area), several electrical parameters are monitored to find out critical signatures and precursors of failure. Among those, gate leakage current is identified as the most practical precursor, which exhibits consistent changes in all aged devices and is relatively easy to monitor. Due to its simple scheme and low cost, it can potentially be embedded into commercial gate drivers featuring improved reliability options.

In the brain, apolipoprotein E (apoE) plays an important role in lipid transport and response to environmental and age-related challenges, including neuronal repair following injury. While much has been learned from radiation studies in rodents, a gap in our knowledge is how radiation might affect the brain in primates. This is important for assessing risk to the brain following radiotherapy as part of cancer treatment or environmental radiation exposure as part of a nuclear accident, bioterrorism, or a nuclear attack. In this study, we investigated the effects of ionizing radiation on brain volumes and apoE levels in the prefrontal cortex, amygdala, and hippocampus of Rhesus macaques that were part of the Nonhuman Primate Radiation Survivor Cohort at the Wake Forest University. This unique cohort is composed of Rhesus macaques that had previously received single total body doses of 6.5–8.05 Gy of ionizing radiation. Regional apoE levels predicted regional volume in the amygdala and the prefrontal cortex. In addition, apoE levels in the amygdala, but not the hippocampus, strongly predicted relative hippocampal volume. Finally, radiation dose negatively affected relative hippocampal volume when apoE levels in the amygdala were controlled for, suggesting a protective compensatory role of regional apoE levels following radiation exposure. In a supplementary analysis, there also was a robust positive relationship between the neuroprotective protein α-klotho and apoE levels in the amygdala, further supporting the potentially protective role of apoE. Increased understanding of the effects of IR in the primate brain and the role of apoE in the irradiated brain could inform future therapies to mitigate the adverse effects of IR on the CNS.

Purpose Radiation-induced lung injury (RILI) is a progressive condition with an early phase (radiation pneumonitis) and a late phase (lung fibrosis). RILI may occur after partial-body ionizing radiation exposures or internal radioisotope exposure, with wide individual variability in timing and extent of lung injury. This study aimed to provide new insights into the pathogenesis and progression of RILI in the nonhuman primate (NHP) rhesus macaque model. Methods and Materials We used an integrative approach to understand RILI and its evolution at clinical and molecular levels in 17 NHPs exposed to 10 Gy of whole-thorax irradiation in comparison with 3 sham-irradiated control NHPs. Clinically, we monitored respiratory rates, computed tomography (CT) scans, plasma cytokine levels, and bronchoalveolar lavage (BAL) over 8 months and lung samples collected at necropsy for molecular and histopathologic analyses using RNA sequencing and immunohistochemistry. Results Elevated respiratory rates, greater CT density, and more severe pneumonitis with increased macrophage content were associated with early mortality. Radiation-induced lung fibrosis included polarization of macrophages toward the M2-like phenotype, TGF-β signaling, expression of CDKN1A/p21 in epithelial cells, and expression of α-SMA in lung stroma. RNA sequencing analysis of lung tissue revealed SERPINA3, ATP12A, GJB2, CLDN10, TOX3, and LPA as top dysregulated transcripts in irradiated animals. In addition to transcriptomic data, we observed increased protein expression of SERPINA3, TGF-β1, CCL2, and CCL11 in BAL and plasma samples. Conclusions Our combined clinical, imaging, histologic, and transcriptomic analysis provides new insights into the early and late phases of RILI and highlights possible biomarkers and potential therapeutic targets of RILI. Activation of TGF-β and macrophage polarization appear to be key mechanisms involved in RILI.

Objectives High-dose glucocorticoids anchor standard care in GCA but are associated with significant toxicity. We aimed to evaluate the safety and effectiveness of a stratified approach to glucocorticoid tapering. The strategy aggressively reduced glucocorticoid doses in those manifesting an adequate early response to treatment, with a view to minimizing glucocorticoid complications. Methods A retrospective, population-based study of GCA was performed. All cases were confirmed by temporal artery biopsy between November 2010 and November 2015. Baseline and outcome data were extracted from secondary and primary care records at diagnosis and 1 year follow-up. The primary outcome was loss of vision. Secondary outcomes included remission and relapse rates and CS-related complications. Results The cohort consisted of 73 patients (76% female; mean age 73.5 years, s.d. 7.6 years). At presentation, a reduction in visual acuity was recorded in 17 patients (22.3%). The median CRP at diagnosis was 69.5 mg/l [interquartile range (IQR) 40.5–101 mg/l], with a median ESR of 80 mm/h (IQR 60–91 mm/h). At 1 year, remission was achieved in 64 patients (87.7%), whereas 10 patients (13.7%) relapsed. A single patient sustained visual loss after initiation of therapy. The median CRP at 1 year was 4 mg/l (IQR 4–9.5 mg/l) and the mean prednisolone dose was 5.4 mg (0–15 mg). CS-related complications were observed in 10 patients (13.7%). Conclusion A stratified approach to CS tapering appeared safe and effective in GCA. It was associated with a high rate of remission and promisingly low rates of relapse at 1 year follow-up. These real-world data indicate that glucocorticoid exposure can be minimized safely in some patients with GCA.

The obligate intracellular parasite Toxoplasma gondii is auxotrophic for several key metabolites and must scavenge these from the host. It is unclear how T. gondii manipulates host metabolism to support its overall growth rate and non-essential metabolites. To investigate this question, we measured changes in the joint host-parasite metabolome over a time course of infection. Host and parasite transcriptomes were simultaneously generated to determine potential changes in expression of metabolic enzymes. T. gondii infection changed metabolite abundance in multiple metabolic pathways, including the tricarboxylic acid cycle, the pentose phosphate pathway, glycolysis, amino acid synthesis, and nucleotide metabolism. Our analysis indicated that changes in some pathways, such as the tricarboxylic acid cycle, were mirrored by changes in parasite transcription, while changes in others, like the pentose phosphate pathway, were paired with changes in both the host and parasite transcriptomes. Further experiments led to the discovery of a T. gondii enzyme, sedoheptulose bisphosphatase, which funnels carbon from glycolysis into the pentose phosphate pathway through an energetically driven dephosphorylation reaction. This additional route for ribose synthesis appears to resolve the conflict between the T. gondii tricarboxylic acid cycle and pentose phosphate pathway, which are both NADP+ dependent. Sedoheptulose bisphosphatase represents a novel step in T. gondii central carbon metabolism that allows T. gondii to energetically-drive ribose synthesis without using NADP+., Author summary The obligate intracellular parasite T. gondii is commonly found among human populations worldwide and poses severe health risks to fetuses and individuals with AIDS. While some treatments are available they are limited in scope. A possible target for new therapies is T. gondii’s incomplete metabolism, which makes it heavily reliant on its host. In this study, we generated a joint host/parasite metabolome to better understand host manipulation by the parasite and to discover unique aspects of T. gondii metabolism that could serve as the next generation of drug targets. Metabolomic analysis of T. gondii infection over time found broad alterations to host metabolism by the parasite in both energetic and biosynthetic pathways. We discovered a new T. gondii enzyme, sedoheptulose bisphosphatase, which redirects carbon from glycolysis into the pentose phosphate pathway. The wholesale remodeling of host metabolism for optimal parasite growth is also of interest, although the mechanisms behind this host manipulation must be further studied before therapeutic targets can be identified.

ObjectivesReliable biomarkers for renal fibrosis are needed for clinical care and for research. Existing non-invasive biomarkers are imprecise, which has limited their utility.MethodsWe developed a method to quantify fibrosis by subject size-adjusted CT Hounsfield units. This was accomplished using CT measurements of renal cortex in previously irradiated non-human primates.ResultsRenal cortex mean CT Hounsfield units that were adjusted for body size had a very good direct correlation with renal parenchymal fibrosis, with an area under the curve of 0.93.ConclusionsThis metric is a promising and simple non-invasive biomarker for renal fibrosis.

Children with orthostatic intolerance (OI) have exaggerated decreases in heart rate variability (HRV) and suppression of baroreflex sensitivity (BRS) with standing. Accompanying brain transmitter and metabolite profiles are unknown. In this study, we used proton (1H) magnetic resonance spectroscopy (1H-MRS) to quantify markers of neuronal and glial integrity in a pilot study of children with OI compared with asymptomatic controls. Eighteen participants ages 10–18 yr were evaluated for blood pressure, heart rate (HR), and calculated indexes of autonomic function in supine and upright positions and, within an average of 2 wk, underwent1H-MRS scans of dorsal medulla on a clinical 3T magnet while supine. As a result, of the 18 participants, 11 tested positive for OI and 7 did not. OI subjects exhibited higher HR and lower HRV and high-frequency α-index (HFα), an index of parasympathetic vagal tone, during standing compared with non-OI. HRV, sequence all (Seq All), high- and low-frequency (HFα and LFα) estimates of the spontaneous BRS decreased significantly, while BP variabilty increased significantly during standing only in subjects with OI. OI subjects had higher myoinositol (mIns) and total choline (tCho), markers of glial inflammation. Upright HFα and Seq All inversely correlated to supine tCho and mIns, respectively, independent of age and sex. In conclusions, in this pilot study, children with OI exhibit higher mIns and tCho in the dorsal medulla while supine that may reflect the well-established impairment in regulation of the autonomic nervous system upon standing. Neuroinflammation as an underlying cause or consequence of autonomic dysfunction is an intriguing possibility requiring further study.NEW & NOTEWORTHY (1H) magnetic resonance spectroscopy detected elevated markers of neuroinflammation in the dorsal medulla in children with impaired autonomic responses to head upright tilt. This first report of altered brain metabolites in this population provides a basis for future clinical studies using this methodology to aide in understanding complex autonomic disease states.

Magnetic Resonance Imaging (MRI) is a commonly used, non-invasive imaging technique that provides visualization of soft tissues with high spatial resolution. In both a research and clinical setting, the major challenge has been identifying a non-invasive and safe method for longitudinal tracking of delivered cells in vivo. The labeling and tracking of contrast agent labeled cells using MRI has the potential to fulfill this need. Contrast agents are often used to enhance the image contrast between the tissue of interest and surrounding tissues with MRI. The most commonly used MRI contrast agents contain Gd(III) ions. However, Gd(III) ions are highly toxic in their ionic form, as they tend to accumulate in the liver, spleen, kidney and bones and block calcium channels. Endohedral metallofullerenes such as trimetallic nitride endohedral metallofullerenes (Trimetasphere®) are one unique class of fullerene molecules where a Gd3N cluster is encapsulated inside a C80 carbon cage referred to as Gd3N@C80. These endohedral metallofullerenes have several advantages over small chelated Gd(III) complexes such as increased stability of the Gd(III) ion, minimal toxic effects, high solubility in water and high proton relativity. In this study, we describe the evaluation of gadolinium-based Trimetasphere® positive contrast agent for the ​in vitro labeling and in vivo tracking of human amniotic fluid-derived stem cells within lung tissue. In addition, we conducted a 'proof-of-concept' experiment demonstrating that this methodology can be used to track the homing of stem cells to injured lung tissue and provide longitudinal analysis of cell localization over an extended time course.

Nuclear accidents and acts of terrorism have the potential to expose thousands of people to high-dose total-body iradiation (TBI). Those who survive the acute radiation syndrome are at risk of developing chronic, degenerative radiation-induced injuries [delayed effects of acute radiation (DEARE)] that may negatively affect quality of life. A growing body of literature suggests that the brain may be vulnerable to radiation injury at survivable doses, yet the long-term consequences of high-dose TBI on the adult brain are unclear. Herein we report the occurrence of lesions consistent with cerebrovascular injury, detected by susceptibility-weighted magnetic resonance imaging (MRI), in a cohort of non-human primate [(NHP); rhesus macaque, Macaca mulatta] long-term survivors of high-dose TBI (1.1-8.5 Gy). Animals were monitored longitudinally with brain MRI (approximately once every three years). Susceptibility-weighted images (SWI) were reviewed for hypointensities (cerebral microbleeds and/or focal necrosis). SWI hypointensities were noted in 13% of irradiated NHP; lesions were not observed in control animals. A prior history of exposure was correlated with an increased risk of developing a lesion detectable by MRI (P = 0.003). Twelve of 16 animals had at least one brain lesion present at the time of the first MRI evaluation; a subset of animals (n = 7) developed new lesions during the surveillance period (3.7-11.3 years postirradiation). Lesions occurred with a predilection for white matter and the gray-white matter junction. The majority of animals with lesions had one to three SWI hypointensities, but some animals had multifocal disease (n = 2). Histopathologic evaluation of deceased animals within the cohort (n = 3) revealed malformation of the cerebral vasculature and remodeling of the blood vessel walls. There was no association between comorbid diabetes mellitus or hypertension with SWI lesion status. These data suggest that long-term TBI survivors may be at risk of developing cerebrovascular injury years after irradiation.

The obligate intracellular parasite Toxoplasma gondii is auxotrophic for several key metabolites and must scavenge these from the host. It is unclear how Toxoplasma manipulates host metabolism for its overall growth rate and non-essential metabolites. To address this question, we measured changes in the joint host-parasite metabolome over a time course of infection. Host and parasite transcriptomes were simultaneously generated to determine potential changes in metabolic enzyme levels. Toxoplasma infection increased activity in multiple metabolic pathways, including the tricarboxylic acid cycle, the pentose phosphate pathway, glycolysis, amino acid synthesis, and nucleotide metabolism. Our analysis indicated that changes in some pathways, such as the tricarboxylic acid cycle, derive from the parasite, while changes in others, like the pentose phosphate pathway, were host and parasite driven. Further experiments led to the discovery of a Toxoplasma enzyme, sedoheptulose bisphosphatase, which funnels carbon from glycolysis into ribose synthesis through a energetically driven dephosphorylation reaction. This second route for ribose synthesis resolves a conflict between the Toxoplasma tricarboxylic acid cycle and pentose phosphate pathway, which are both NADP+ dependent. During periods of high energetic and ribose need, the competition for NADP+ could result in lethal redox imbalances. Sedoheptulose bisphosphatase represents a novel step in Toxoplasma central carbon metabolism that allows Toxoplasma to satisfy its ribose demand without using NADP+. Sedoheptulose bisphosphatase is not present in humans, highlighting its potential as a drug target.Author SummaryThe obligate intracellular parasite Toxoplasma is commonly found among human populations worldwide and poses severe health risks to fetuses and individuals with AIDS. While some treatments are available they are limited in scope. A possible target for new therapies is Toxoplasma’s limited metabolism, which makes it heavily reliant in its host. In this study, we generated a joint host/parasite metabolome to better understand host manipulation by the parasite and to discover unique aspects of Toxoplasma metabolism that could serve as the next generation of drug targets. Metabolomic analysis of Toxoplasma during an infection time course found broad activation of host metabolism by the parasite in both energetic and biosynthetic pathways. We discovered a new Toxoplasma enzyme, sedoheptulose bisphosphatase, which redirects carbon from glycolysis into ribose synthesis. Humans lack sedoheptulose bisphosphatase, making it a potential drug target. The wholesale remodeling of host metabolism for optimal parasite growth is also of interest, although the mechanisms behind this host manipulation must be further studied before therapeutic targets can be identified.

Objective To determine whether super-activation of PPARγ can reprogram human myoblasts into brownlike adipocytes and to establish a new cell model for browning research.

Purpose The recent publication of the ACOSOG Z1031 trial results demonstrated that Ki-67 proliferation marker-based neoadjuvant endocrine therapy response monitoring could be used for tailoring the use of adjuvant chemotherapy in ER+HER2-negative breast cancer patients. In this paper, we describe the development of the Ki-67 clinical trial assay used for this study. Methods Ki-67 assay assessment focused on reproducing a 2.7% Ki-67 cut-point (CP) required for calculating the Preoperative Endocrine Prognostic Index and a 10% CP for poor endocrine therapy response identification within the first month of neoadjuvant endocrine treatment. Image analysis was assessed to increase the efficiency of the scoring process. Clinical outcome concordance for two independent Ki-67 scores was the primary performance metric. Results Discordant scores led to a triage approach where cases with complex histological features that software algorithms could not resolve were flagged for visual point counting (17%). The final Ki-67 scoring approach was run on T1/2 N0 cases from the P024 and POL trials (N = 58). The percent positive agreement for the 2.7% CP was 87.5% (95% CI 61.7–98.5%); percent negative agreement 88.9% (95% CI: 65.3–98.6%). Minor discordance did not affect the ability to predict similar relapse-free outcomes (Log-Rank P = 0.044 and P = 0.055). The data for the 10% early triage CP in the POL trial were similar (N = 66), the percentage positive agreement was 100%, and percent negative agreement 93.55% (95% CI: 78.58–99.21%). The independent survival predictions were concordant (Log-rank P = 0.0001 and P = 0.01). Conclusions We have developed an efficient and reproducible Ki-67 scoring system that was approved by the Clinical Trials Evaluation Program for NCI-supported neoadjuvant endocrine therapy trials. Using the methodology described here, investigators are able to identify a subgroup of patients with ER+HER2-negative breast cancer that can be safely managed without the need of adjuvant chemotherapy. Electronic supplementary material The online version of this article (doi:10.1007/s10549-017-4329-y) contains supplementary material, which is available to authorized users.

The blood-brain-barrier (BBB), a network of tight junctions that impedes large molecule transport, limits the usefulness of systemic chemotherapeutic delivery for the treatment of malignant gliomas and other neurological diseases. Here, we present a tool for BBB disruption that uses bursts of sub-microsecond bipolar pulses to enhance the transfer of large molecules to the brain. Blunt needle electrodes were advanced into the motor cortex of anesthetized adult rats, and a series of 90–900 bursts were delivered with voltage-to-distance ratios of 250 or 2000 V/cm, a total programmed energized time of 100 μs, and a repetition rate of 1 Hz. BBB disruption was assessed via a gadolinium-Evans blue albumin tracer, and all experimental conditions were found to cause BBB disruption immediately following treatment without inducing local or distal muscle contractions. The lowest energy condition, 300 bursts consisting of 850 ns bipolar pulses, resulted in significant BBB disruption (0.51 cm3), without displaying necrotic or apoptotic damage to neurological tissue.

Premature musculoskeletal joint failure is a major source of morbidity among childhood cancer survivors. Radiation effects on synovial joint tissues of the skeleton are poorly understood. Our goal was to assess long-term changes in the knee joint from skeletally mature rats that received total-body irradiation while skeletal growth was ongoing.14 week-old rats were irradiated with 1, 3 or 7 Gy total-body doses of 18 MV X-rays. At 53 weeks of age, structural and compositional changes in knee joint tissues (articular cartilage, subchondral bone, and trabecular bone) were characterized using 7T MRI, nanocomputed tomography (nanoCT), microcomputed tomography (microCT), and histology.T2 relaxation times of the articular cartilage were lower after exposure to all doses. Likewise, calcifications were observed in the articular cartilage. Trabecular bone microarchitecture was compromised in the tibial metaphysis at 7 Gy. Mild to moderate cartilage erosion was scored in the 3 and 7 Gy rats.Late degenerative changes in articular cartilage and bone were observed after total-body irradiation in adult rats exposed prior to skeletal maturity. 7T MRI, microCT, nanoCT, and histology identified potential prognostic indicators of late radiation-induced joint damage.

To assess the long-term effects of fractionated whole brain irradiation (fWBI) using diffusion tensor imaging (DTI) and behavior in a pediatric rodent model for the clinical presentation of adult pediatric cancer survivors.Five-week-old, male F344xBN rats were randomized to receive 0, 5, or 6.5 Gy fractions biweekly for 3 weeks, resulting in Sham, Irradiated-30 (IR-30) and IR-39 Gy total dose groups. Magnetic Resonance Imaging occurred at 1, 3, 6 and 9 months with behavioral assessment at 10-11 months post-fWBI.Irradiation reduced brain size (p0.001) and body weight (p0.001) proportionate to dose. At 1 month post-fWBI and throughout follow-up, diffusion was reduced in IR-30 and IR-39 relative to shams (p0.001). IR-30 but not IR-39 rats were impaired relative to Shams on the reversal trial of the Morris Water Maze (p0.05), and IR-30 rats preferred a striatum- mediated strategy (p0.06).Hippocampal performance was impaired in IR-30 but not IR-39 animals. While gross size differences exist, white matter integrity is preserved in rats after fWBI at 5 weeks. This significant departure from childhood cancer survivors and single fraction rodent studies where white matter degradation is a prominent feature are discussed.

Tissue growth and function depend on vascularization, and vascular insufficiency or excess exacerbates many human diseases. Identification of the biological processes involved in angiogenesis will dictate strategies to modulate reduced or excessive vessel formation. We examine the essential role of pericytes. Their heterogeneous morphology, distribution, origins, and physiology have been described. Using double-transgenic Nestin-GFP/NG2-DsRed mice, we identified two pericyte subsets. We found that Nestin-GFP−/NG2-DsRed+(type-1) and Nestin-GFP+/NG2-DsRed+(type-2) pericytes attach to the walls of small and large blood vessels in vivo; in vitro, type-2, but not type-1, pericytes spark endothelial cells to form new vessels. Matrigel assay showed that only type-2 pericytes participate in normal angiogenesis. Moreover, when cancer cells were transplanted into Nestin-GFP/NG2-DsRed mice, type-1 pericytes did not penetrate the tumor, while type-2 pericytes were recruited during its angiogenesis. As inhibition of angiogenesis is a promising strategy in cancer therapy, type-2 pericytes may provide a cellular target susceptible to signaling and pharmacological manipulation in treating malignancy. This work also reports the potential of type-2 pericytes to improve blood perfusion in ischemic hindlimbs, indicating their potential for treating ischemic illnesses.

Background: In the US, the majority of BC is diagnosed by screening XRAY mammography (MM) but up to ∼20% of BC are undetected by MM. The development of reliable, blood-based BC screening test to increase the sensitivity and specificity of currently existing BC screening methods such as MM. Rationale: GP88 is expressed & secreted by BC cells & is not expressed by normal mammary epithelial cells. Two retrospective randomized multi-site trials (one training study & one validating study with about 300 cases each) have demonstrated that elevated GP88 expression in ER+ IDC was statistically correlated with a 4-fold increase in the risk of 5-yr disease recurrence. Multivariate analysis showed that GP88 as a risk predictor was independent from PR expression, tumor size, grade, lymph node status & disease stage. The quantitative GP88 EIA developed to determine the amount of GP88 in biological fluids was developed at A&G. The assay is highly specific for GP88 & both sensitive & linear over a wide dynamic range, i.e. detection of GP88 concentrations from 0.1 to 20ng/ml. A baseline (28.4 ± 5 ng/ml) was established for healthy volunteers (HV). In BC pts a statistically significant increase of serum GP88 was seen for early stage pts (40.7 ± 16 ng/ml; p=0.007). Additionally, a stratification of pts according to their clinical outcomes shows that pts having no evidence of disease (NED) have serum GP88 levels within the range of HV. These data suggest that pts with breast tumors express & secrete high levels of GP88. Objectives: 1. To determine prospectively GP-88 blood levels in HV at average risk of developing BC who are undergoing screening MM & in women with biopsy-confirmed BC. 2. To establish the statistical distribution of GP88 serum levels in these women by baseline BIRAD classification (1-6). 3. To determine if baseline GP88 level is predictive of change in BIRADS classification from baseline to 12-mos follow-up. 4. To determine if baseline GP88 level is predictive of the appearance of BC at 12 mos follow-up in HV who were cancer-free at baseline. Inclusion Criteria: Female, aged >=40 yrs old, presenting to UM Breast Center for screening MM or for diagnostic MM or diagnostic workup and/or biopsy due to BIRADS 0 MM at an outside facility Study procedures: Serum levels of GP88 in subjects with average BC risk factors will be measured prospectively at baseline; 3-6 mos & 6-12 mos. & correlated with BIRADS reading of the screening MM, BIRADS 1 or 2 (benign), BIRADS 3 (short term MM follow-up) & BIRADS 4 or 5 (suspicious and tissue diagnosis needed immediately). GP88 blood level will be correlated with pathologic results of breast biopsies performed on subjects with suspicious BIRADS (4 & 5) MM & final pathologically confirmed diagnosis of BC as BIRADS 6. Statistical Considerations The total number of pts will be 725 & screened up to 1400 subjects. Study is UM IRB approved & just started accruing. Funding is provided by Maryland Industry Partnership Grant (MIPS)& Avon Grant No. 02-2013-018. Contact: Ntait@umm.edu. Citation Format: Katherine HR Tkaczuk, Cristina Campassi, Susan Kesmodel, Emily Bellavance, John Olson, Elizabeth Nichols, Steven J Feigenberg, Binbin Yue, David Hicks, Ginette Serrero. A prospective study of glycoprotein 88 GP-88 blood test in healthy women with Gail model risk

The present study represents the first longitudinal, within-subject 1H MRS investigation of the developing rat brain spanning infancy, adolescence, and early adulthood. We obtained neurometabolite profiles from a voxel located in a central location of the forebrain, centered on the striatum, with smaller contributions for cortex, thalamus, and hypothalamus, on postnatal days 7, 35, and 60. Water-scaled metabolite signals were corrected for T1 effects and quantified using the automated processing software LCModel, yielding molal concentrations. Our findings indicate age-related concentration changes in N-acetylaspartate + N-acetylaspartylglutamate, myo-inositol, glutamate + glutamine, taurine, creatine + phosphocreatine, and glycerophosphocholine + phosphocholine. Using a repeated measures design and analysis, we identified significant neurodevelopment change across all three developmental ages and identified adolescence as a distinctive phase in normative neurometabolic brain development. Between postnatal days 35 and 60, changes were observed in concentrations of N-acetylaspartate + N-acetylaspartylglutamate, glutamate + glutamine, and glycerophosphocholine + phosphocholine observed between postnatal days 35 and 60. Our data replicate past studies of early neurometabolite development and, for the first time, link maturational profiles in the same subjects across infancy, adolescence, and adulthood.

There is little known about the effect of both reduced weight bearing and exposure to radiation during spaceflight on the mechanically-sensitive cartilage lining the knee joint. In this study, we characterized cartilage damage in rat knees after periods of reduced weight bearing with/without exposure to solar-flare-relevant radiation, then cartilage recovery after return to weight bearing. Male Sprague Dawley rats (n = 120) were either hindlimb unloaded (HLU) via tail suspension or remained weight bearing in cages (GROUND). On day 5, half of the HLU and GROUND rats were 1 Gy total-body X-ray irradiated during HLU, and half were sham irradiated (SHAM), yielding 4 groups: GROUND-SHAM; GROUND-IR; HLU-SHAM; and HLU-IR. Hindlimbs were collected from half of each group of rats on day 13. The remaining rats were then removed from HLU or remained weight bearing, and hindlimbs from these rats were collected on day 62. On day 13, glycosaminoglycan (GAG) content in cartilage lining the tibial plateau and femoral condyles of HLU rats was lower than that of the GROUND animals. Likewise, on day 13, immunoreactivity of the collagen type II-degrading matrix metalloproteinase-13 (MMP-13) and of a resultant metalloproteinase-generated neoepitope VDIPEN was increased in all groups versus GROUND-SHAM. Clustering of chondrocytes indicating cartilage damage was present in all HLU and IR groups versus GROUND-SHAM on day 13. On day 62, after 49 days of reloading, the loss of GAG content was attenuated in the HLU-SHAM and HLU-IR groups, and the increased VDIPEN staining in all treatment groups was attenuated. However, the increased chondrocyte clustering remained in all treatment groups on day 62. MMP-13 activity also remained elevated in the GROUND-IR and HLU-IR groups. Increased T2 relaxation times, measured on day 62 using 7T MRI, were greater in GROUND-IR and HLU-IR knees, indicating persistent cartilage damage in the irradiated groups. Both HLU and total-body irradiation resulted in acute degenerative and pre-arthritic changes in the knee articular cartilage of rats. A return to normal weight bearing resulted in some recovery from cartilage degradation. However, radiation delivered as both a single challenge and when combined with HLU resulted in chronic cartilage damage. These findings suggest that radiation exposure during spaceflight leads to and/or impairs recovery of cartilage upon return to reloading, generating long-term joint problems for astronauts.

Iodine environmental measurements have consistently been validated in the literature using the standard material IAEA-375, soil collected approximately 160 miles northeast of Chernobyl, which is currently the only soil/sediment material with a certified

A gated-7T magnetic resonance imaging (MRI) application is described that can accurately and efficiently measure the size of in vivo mouse lung tumors from ∼0.1 mm(3) to4 mm(3). This MRI approach fills a void in radiation research because the technique can be used to noninvasively measure the growth rate of lung tumors in large numbers of mice that have been irradiated with low doses (50 mGy) without the additional radiation exposure associated with planar X ray, CT or PET imaging. High quality, high resolution, reproducible images of the mouse thorax were obtained in ∼20 min using: (1) a Bruker 7T micro-MRI scanner equipped with a 60 mm inner diameter gradient insert capable of generating a maximum gradient of 1000 mT/m; (2) a 35 mm inner diameter quadrature radiofrequency volume coil; and (3) an electrocardiogram and respiratory gated Fast Low Angle Shot (FLASH) pulse sequence. The images had an in-plane image resolution of 98 μm and a 0.5 mm slice thickness. Tumor diameter measured by MRI was highly correlated (R(2) = 0.97) with the tumor diameter measured by electronic calipers. Data generated with an initiation/promotion mouse model of lung carcinogenesis and this MRI technique demonstrated that mice exposed to 4 weekly fractions of 10, 30 or 50 mGy of CT radiation had the same lung tumor growth rate as that measured in sham-irradiated mice. In summary, this high-field, double-gated MRI approach is an efficient way of quantitatively tracking lung tumor development and progression after exposure to low doses of ionizing radiation.

Glioblastoma multiforme (GBM) accounts for the majority of primary malignant brain tumors and remains virtually incurable despite extensive surgical resection, radiotherapy, and chemotherapy. Treatment difficulty is due to its exceptional infiltrative nature and proclivity to integrate into normal brain tissue. Long-term survivors are rare, and median survival for patients is about 1 year. Use of adult stem cells as cellular delivery vehicles for anticancer agents is a novel attractive therapeutic strategy. We hypothesized that adipose-derived stem cells (ADSCs) possess the ability to home and deliver myxoma virus to glioma cells and experimental gliomas. We infected ADSCs with vMyxgfp and found them to be permissive for myxoma virus replication. ADSCs supported single and multiple rounds of replication leading to productive infection. Further, we observed no significant impact on ADSC viability. We cocultured fluorescently labeled GBM cells with myxoma virus–infected ADSCs in three-dimensional assay and observed successful cross infection and concomitant cell death almost exclusively in GBM cells. In vivo orthotopic studies injected with vMyxgfp-ADSCs intracranially away from the tumor demonstrated that myxoma virus was delivered by ADSCs resulting in significant survival increase. Our data suggest that ADSCs are promising new carriers of oncolytic viruses, specifically myxoma virus, to brain tumors.

We investigated the cytogenetic and gene expression responses of peripheral blood cells of non-human primates (NHP, Macaca mulatta) that were whole-thorax irradiated with a single dose of 10 Gy. In this model, partial irradiation of NHPs in the thoracic region (Whole Thorax Lung Irradiation, WTLI) allows the study of late radiation-induced lung injury, while avoiding acute radiation syndromes related to hematopoietic and gastrointestinal injury. A transient drop in circulating lymphocytes and platelets was seen by 9 days, followed by elevations in respiratory rate, circulating neutrophils, lymphocytes, and monocytes at 60-100 days, corresponding to computed tomography (CT) and histologic evidence of pneumonitis, and elective euthanasia of four animals. To evaluate long-term DNA damage in NHP peripheral blood lymphocytes after 10 Gy WTLI, we used the cytokinesis-block micronucleus (CBMN) assay to measure chromosomal aberrations as post-mitotic micronuclei in blood samples collected up to 8 months after irradiation. Regression analysis showed significant induction of micronuclei in NHP blood cells that persisted with a gradual decline over the 8-month study period, suggesting long-term DNA damage in blood lymphocytes after WTLI. We also report transcriptomic changes in blood up to 30 days after WTLI. We isolated total RNA from peripheral blood at 3 days before and then at 2, 5 and 30 days after irradiation. We identified 1187 transcripts that were significantly changed across the 30-day time course. From changes in gene expression, we identified biological processes related to immune responses, which persisted across the 30-day study. Response to oxygen-containing compounds and bacteria were implicated by gene-expression changes at the earliest day 2 and latest, day 30 time-points. Gene expression changes suggest a persistent altered state of the immune system, specifically response to infection, for at least a month after WTLI.

Gamma Knife stereotactic radiosurgery is capable of providing small, high gradient dose distributions to a target with a high level of precision, which makes it an excellent choice for studies of focal irradiations with small animals. However, the Gamma Knife stereotactic radiosurgery process makes use of a human-sized fiducial marker system that requires a field of view of at least 200 mm(2) to relate computed tomography and magnetic resonance images to the Gamma Knife treatment planning software. Thus the Gamma Knife fiducial marker system is five to six times larger than a typical small animal subject. The required large field of view limits the spatial resolution and structural detail available in the animal treatment planning image set. In response to this challenge we have developed a custom-designed stereotactic jig and miniature fiducial marking system that allow small bore high-resolution micro-imaging techniques, such as 7T MR and micro-CT, to be used for treatment planning of Gamma Knife stereotactic radiosurgery focal irradiation of small animals.