Bronchopulmonary dysplasia (BPD) is the most common respiratory disease in preterm infants born at less than 28 weeks gestation. Most existing clinical prediction models for BPD show limited accuracy in predicting BPD development when validated using external data, stressing the need for novel biomarkers to identify at-risk infants for early and effective interventions. We leveraged existing frozen umbilical cord blood samples from the Northwestern University Cord Blood Biobank (NUCord) to perform parallel transcriptional and DNA methylation profiling. BPD-associated differentially expressed genes (DEGs) in our cohort included markers previously established in clinical and animal BPD studies, such as genes related to NF-κB signaling and immune responses. We also identified that BPD development is associated with disrupted methylation signatures in microRNA genes and genes associated with glucose metabolism. Our results suggest that BPD development is associated with distinct transcriptomic and epigenetic signatures when compared with healthy term and preterm infants. These signatures may represent biomarkers measurable at birth that predict BPD development during a time window when preventative or therapeutic interventions could be applied.

Fractionated whole-brain irradiation (fWBI), used to treat brain metastases, often leads to neurologic injury and cognitive impairment. The cognitive effects of irradiation in nonhuman primates (NHP) have been previously published; this report focuses on corresponding neuropathologic changes that could have served as the basis for those effects in the same study. Four rhesus monkeys were exposed to 40 Gy of fWBI [5 Gy × 8 fraction (fx), 2 fx/week for four weeks] and received anatomical MRI prior to, and 14 months after fWBI. Neurologic and histologic sequelae were studied posthumously. Three of the NHPs underwent cognitive assessments, and each exhibited radiation-induced impairment associated with various degrees of vascular and inflammatory neuropathology. Two NHPs had severe multifocal necrosis of the forebrain, midbrain and brainstem. Histologic and MRI findings were in agreement, and the severity of cognitive decrement previously reported corresponded to the degree of observed pathology in two of the animals. In response to fWBI, the NHPs showed pathology similar to humans exposed to radiation and show comparable cognitive decline. These results provide a basis for implementing NHPs to examine and treat adverse cognitive and neurophysiologic sequelae of radiation exposure in humans.

There are several ways to score normal tissue injury. Imaging is an attractive tool since it allows for an objective assessment of normal tissue changes in vivo. Regional changes in tissue structure/function, detected by imaging, can be related to the different radiation doses delivered to different regions. This approach has been used for several organs, and for some, there is a clear dose-response relationship for changes in normal tissue (e.g., lung density or perfusion). Further, degree/extent of changes in regional imaging may be associated with changes in global organ function. Future improvements in imaging (e.g., more functional versus structural assessments) will afford additional opportunities for studies to better understand RT-induced normal tissue injury.

PURPOSE: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.MATERIALS AND METHODS: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.RESULTS:Irradiation reduced brain size (p < 0.001) and body weight (p < 0.001) proportionate to dose. At 1 month post-fWBI and throughout follow-up, diffusion was reduced in IR-30 and IR-39 relative to shams (p < 0.001). IR-30 but not IR-39 rats were impaired relative to Shams on the reversal trial of the Morris Water Maze (p < 0.05), and IR-30 rats preferred a striatum- mediated strategy (p < 0.06).CONCLUSIONS: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.

Future space missions are expected to include increased extravehicular activities (EVAs) during which astronauts are exposed to high-energy space radiation while breathing 100% oxygen. Given that brain irradiation can lead to cognitive impairment, and that oxygen is a potent radiosensitizer, there is a concern that astronauts may be at greater risk of developing cognitive impairment when exposed to space radiation while breathing 100% O(2) during an EVA. To address this concern, unanesthetized, unrestrained, young adult male Fischer 344 × Brown Norway rats were allowed to breathe 100% O(2) for 30 min prior to, during and 2 h after whole-body irradiation with 0, 1, 3, 5 or 7 Gy doses of 18 MV X rays delivered from a medical linear accelerator at a dose rate of ~425 mGy/min. Irradiated and unirradiated rats breathing air (~21% O(2)) served as controls. Cognitive function was assessed 9 months postirradiation using the perirhinal cortex-dependent novel object recognition task. Cognitive function was not impaired until the rats breathing either air or 100% O(2) received a whole-body dose of 7 Gy. However, at all doses, cognitive function of the irradiated rats breathing 100% O(2) was improved over that of the irradiated rats breathing air. These data suggest that astronauts are not at greater risk of developing cognitive impairment when exposed to space radiation while breathing 100% O(2) during an EVA.

We hypothesized that dietary administration of the peroxisomal proliferator-activated receptor α agonist, fenofibrate, to young adult male rats would prevent the fractionated whole-brain irradiation (fWBI)-induced reduction in cognitive function and neurogenesis and prevent the fWBI-induced increase in the total number of activated microglia. Eighty 12–14-week-old young adult male Fischer 344 × Brown Norway rats received either: (1) sham irradiation, (2) 40 Gy of fWBI delivered as two 5 Gy fractions/week for 4 weeks, (3) sham irradiation dietary fenofibrate (0.2% w/w) starting 7 days prior to irradiation, or (4) fWBI fenofibrate. Cognitive function was measured 26–29 weeks after irradiation using: (1) the perirhinal cortex (PRh)-dependent novel object recognition task; (2) the hippocampal-dependent standard Morris water maze (MWM) task; (3) the hippocampal-dependent delayed match-to-place version of the MWM task; and (4) a cue strategy preference version of the MWM to distinguish hippocampal from striatal task performance. Neurogenesis was assessed 29 weeks after fWBI in the granular cell layer and subgranular zone of the dentate gyrus using a doublecortin antibody. Microglial activation was assessed using an ED1 antibody in the dentate gyrus and hilus of the hippocampus. A significant impairment in perirhinal cortex-dependent cognitive function was measured after fWBI. In contrast, fWBI failed to alter hippocampal-dependent cognitive function, despite a significant reduction in hippocampal neurogenesis. Continuous administration of fenofibrate prevented the fWBI-induced reduction in perirhinal cortex-dependent cognitive function, but did not prevent the radiation-induced reduction in neurogenesis or the radiation-induced increase in activated microglia. These data suggest that fenofibrate may be a promising therapeutic for the prevention of some modalities of radiation-induced cognitive impairment in brain cancer patients.

PURPOSE:To determine if the brain's response to single doses predicts its response to 'biologically equivalent' fractionated doses. METHODS:Young adult male Fischer 344 rats were whole-brain irradiated with either single 11, 14, or 16.5 Gy doses of (137)Cs γ rays or their 'biologically equivalent' 20, 30, or 40 Gy fractionated doses (fWBI) delivered in 5 Gy fractions, twice/week for 2, 3, or 4 weeks, respectively. At 2 months post-irradiation, cellular markers of inflammation (total, activated, and newborn microglia) and neurogenesis (newborn neurons) were measured in 40 μm sections of the dentate gyrus (DG). RESULTS:Although the total number of microglia in the DG/hilus was not significantly different (p > 0.7) in unirradiated, single dose, and fWBI rats, single doses produced a significant (p < 0.003) increase in the percent-activated microglia; fWBI did not (p > 0.1). Additionally, single doses produced a significant (p < 0.002) dose-dependent increase in surviving newborn microglia; fWBI did not (p < 0.8). Although total proliferation in the DG was reduced equally by single and fWBI doses, single doses produced a significant dose-dependent (p < 0.02) decrease in surviving newborn neurons; fWBI did not (p > 0.6). CONCLUSIONS:These data demonstrate that the rat brain's cellular response to single doses often does not predict its cellular response to 'biologically equivalent' fWBI doses.

About 500,000 new cancer patients will develop brain metastases in 2013. The primary treatment modality for these patients is partial or whole brain irradiation which leads to a progressive, irreversible cognitive impairment. Although the exact mechanisms behind this radiation-induced brain injury are unknown, neuroinflammation in glial populations is hypothesized to play a role. Blockers of the renin-angiotensin system (RAS) prevent radiation-induced cognitive impairment and modulate radiation-induced neuroinflammation. Recent studies suggest that RAS blockers may reduce inflammation by increasing endogenous concentrations of the anti-inflammatory heptapeptide angiotensin-(1-7) [Ang-(1-7)]. Ang-(1-7) binds to the AT(1-7) receptor and inhibits MAP kinase activity to prevent inflammation. This study describes the inflammatory response to radiation in astrocytes characterized by radiation-induced increases in (i) IL-1β and IL-6 gene expression; (ii) COX-2 and GFAP immunoreactivity; (iii) activation of AP-1 and NF-κB transcription factors; and (iv) PKCα, MEK, and ERK (MAP kinase) activation. Treatment with U-0126, a MEK inhibitor, demonstrates that this radiation-induced inflammation in astrocytes is mediated through the MAP kinase pathway. Ang-(1-7) inhibits radiation-induced inflammation, increases in PKCα, and MAP kinase pathway activation (phosphorylation of MEK and ERK). Additionally Ang-(1-7) treatment leads to an increase in dual specificity phosphatase 1 (DUSP1). Furthermore, treatment with sodium vanadate (Na3VO4), a phosphatase inhibitor, blocks Ang-(1-7) inhibition of radiation-induced inflammation and MAP kinase activation, suggesting that Ang-(1-7) alters phosphatase activity to inhibit radiation-induced inflammation. These data suggest that RAS blockers inhibit radiation-induced inflammation and prevent radiation-induced cognitive impairment not only by reducing Ang II but also by increasing Ang-(1-7) levels.

Abstract About 500,000 new cancer patients will develop brain metastases in 2012. The primary treatment modality for ∼250,000 of these patients is partial or whole brain irradiation. While effective for controlling metastatic brain tumors, there are significant late effects (> 6 months) following radiotherapy which include a progressive, irreversible cognitive impairment. Although the exact mechanism(s) behind this radiation-induced brain injury are unknown, preclinical studies suggest that radiation alters neuronal plasticity. Indeed irradiating the rodent brain decreases long term potentiation, alters N-methyl-D-aspartate (NMDA) and gamma-aminobutyric acid (GABA) receptor expression and ablates neurogenesis. In this study, we used microarray analysis to show that Homer1a, a transcriptionally-regulated early response gene, was transiently upregulated in the hippocampus and downregulated in the cortex at 48 hours after fractionated (total dose of 40 Gy delivered as 2 Gy/fraction, 2X/week) whole brain irradiation (fWBI) in young adult male Fischer 344 X Brown Norway rats. Two months after fWBI Homer1a gene expression was downregulated in both the hippocampus and cortex. Homer1a binds the consensus sequence on the C-terminus of the group I metabotropic glutamate receptors (mGluR); thereby, disrupting the cross-linking of constitutively expressed forms of Homer1 and activating mGluR in the absence of glutamate. Radiation did not alter phosphorylation of mGluR1, total GluR2 or phosphorylation of GluR2/3 in the hippocampus or cortex at 2 months after fWBI. However, the radiation-induced changes in Homer1a expression appeared to produce a significant downregulation in mGluR1 in the hippocampus, and a significant upregulation in the cortex. Memory/learning paradigms suggest that trafficking of 2-amino-3-(3-hydroxy-5-methyl-isoxazol-4-yl)propanoic acid (AMPA) receptors containing mGluR1 from intracellular compartments to the plasma membrane is an “early” event during neural plasticity; thus decreased mGluR1 may result in alterations in hippocampal neural plasticity following fWBI. These data suggest a novel mechanism for the development of radiation-induced cognitive impairment. (Supported by CA122318) Citation Format: Elizabeth Moore, Mitra Kooshki, Lance Miller, Mike Robbins. Fractionated whole brain irradiation modulates Homer1a expression in a brain region specific manner. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 78. doi:10.1158/1538-7445.AM2013-78

Each year, approximately 200,000 patients in the United States will receive partial- or whole-brain irradiation for the treatment of primary or metastatic brain cancer. Early and delayed radiation effects are transient and reversible with modern therapeutic standards; yet, late radiation effects (>= 6 months postirradiation) remain a significant risk, resulting in progressive cognitive impairment. These risks include functional deficits in memory, attention, and executive function that severely affect the patient's quality of life. The mechanisms underlying radiation-induced cognitive impairment remain ill defined. Classically, radiation-induced alterations in vascular and neuroinflammatory glial cell clonogenic populations were hypothesized to be responsible for radiation-induced brain injury. Recently, preclinical studies have focused on the hippocampus, one of two sites of adult neurogenesis within the brain, which plays an important role in learning and memory. Radiation ablates hippocampal neurogenesis, alters neuronal function, and induces neuroinflammation. Neuronal stem cells implanted into the hippocampus prevent the decrease in neurogenesis and improve cognition after irradiation. Clinically prescribed drugs, including PPAR alpha and PPAR gamma agonists, as well as RAS blockers, prevent radiation-induced neuroinflammation and cognitive impairment independent of improved neurogenesis. Translating these exciting findings to the clinic offers the promise of improving the quality of life of brain tumor patients who receive radiotherapy. (c) 2013 AACR.

The use of behavioral testing has become an invaluable tool for assessing the efficacy of therapeutics for a variety of disorders of the central nervous system. This chapter will describe in detail several behavioral paradigms to evaluate the efficacy of PPAR agonists to modulate cognitive impairments in rodent models. When used together as a battery these procedures allow for a global assessment of cognition. These tests are explained in detail below, and include: (1) Novel Object Recognition (NOR), (2) Morris Water Maze (MWM), (3) Delay Match to Place (DMP), and (4) Cue Strategy.

Brain tumor patients often develop cognitive impairment months to years after partial or fractionated whole-brain irradiation (WBI). Studies suggest that neuroinflammation and decreased hippocampal neurogenesis contribute to the pathogenesis of radiation-induced brain injury. In this study, we determined if the peroxisomal proliferator-activated receptor (PPAR) δ agonist GW0742 can prevent radiation-induced brain injury in C57Bl/6 wild-type (WT) and PPARδ knockout (KO) mice. Dietary GW0742 prevented the acute increase in IL-1β mRNA and ERK phosphorylation measured at 3h after a single 10-Gy dose of WBI; it also prevented the increase in the number of activated hippocampal microglia 1 week after WBI. In contrast, dietary GW074 failed to prevent the radiation-induced decrease in hippocampal neurogenesis determined 2 months after WBI in WT mice or to mitigate their hippocampal-dependent spatial memory impairment measured 3 months after WBI using the Barnes maze task. PPARδ KO mice exhibited defects including decreased numbers of astrocytes in the dentate gyrus/hilus of the hippocampus and a failure to exhibit a radiation-induced increase in activated hippocampal microglia. Interestingly, the number of astrocytes in the dentate gyrus/hilus was reduced in WT mice, but not in PPARδ KO mice 2 months after WBI. These results demonstrate that, although dietary GW0742 prevents the increase in inflammatory markers and hippocampal microglial activation in WT mice after WBI, it does not restore hippocampal neurogenesis or prevent early delayed hippocampal-dependent cognitive impairment after WBI. Thus, the exact relationship between radiation-induced neuroinflammation, neurogenesis, and cognitive impairment remains elusive.

Glioma survival is dismal, in part, due to an imbalance in antioxidant expression and activity. Peroxisome proliferator-activated receptor (PPAR) agonists have antineoplastic properties which present new redox-dependent targets for glioma anticancer therapies. Herein, we demonstrate that treatment of primary cultures of normal rat astrocytes with PPAR agonists increased the expression of catalase mRNA protein, and enzymatic activity. In contrast, these same agonists had no effect on catalase expression and activity in malignant rat glioma cells. The increase in steady-state catalase mRNA observed in normal rat astrocytes was due, in part, to de novo mRNA synthesis as opposed to increased catalase mRNA stability. Moreover, pioglitazone-mediated induction of catalase activity in normal rat astrocytes was completely blocked by transfection with a PPARγ-dominant negative plasmid. These data suggest that defects in PPAR-mediated signaling and gene expression may represent a block to normal catalase expression and induction in malignant glioma. The ability of PPAR agonists to differentially increase catalase expression and activity in normal astrocytes but not glioma cells suggests that these compounds might represent novel adjuvant therapeutic agents for the treatment of gliomas.

Objective: In a retrospective review to assess neuroanatomical targets of radiation-induced cognitive decline, dose volume histogram (DVH) analyses of specific brain regions of interest (ROI) are correlated to neurocognitive performance in 57 primary brain tumor survivors.Methods: Neurocognitive assessment at baseline included Trail Making Tests A/B, a modified Rey-Osterreith Complex Figure, California or Hopkins Verbal Learning Test, Digit Span, and Controlled Oral Word Association. DVH analysis was performed for multiple neuroanatomical targets considered to be involved in cognition. The %v10 (percent of ROI receiving 10 Gy), %v40, and %v60 were calculated for each ROI. Factor analysis was used to estimate global cognition based on a summary of performance on individual cognitive tests. Stepwise regression was used to determine which dose volume predicted performance on global factors and individual neurocognitive tests for each ROI.Results: Regions that predicted global cognitive outcomes at doses <60 Gy included the corpus callosum, left frontal white matter, right temporal lobe, bilateral hippocampi, subventricular zone, and cerebellum. Regions of adult neurogenesis primarily predicted cognition at %v40 except for the right hippocampus which predicted at %v10. Regions that did not predict global cognitive outcomes at any dose include total brain volume, frontal pole, anterior cingulate, right frontal white matter, and the right precentral gyrus.Conclusions: Modeling of radiation-induced cognitive decline using neuroanatomical target theory appears to be feasible. A prospective trial is necessary to validate these data. Neurology (R) 2013;80:747-753

Fractionated partial or whole-brain irradiation is the primary treatment for metastatic brain tumors. Despite reducing tumor burden and increasing lifespan, progressive, irreversible cognitive impairment occurs in >50% of the patients who survive >6 months after fractionated whole-brain irradiation. The exact mechanism(s) responsible for this radiation-induced brain injury are unknown; however, preclinical studies suggest that radiation modulates the extracellular receptor kinase signaling pathway, which is associated with cognitive impairment in many neurological diseases. In the study reported here, we demonstrated that the extracellular receptor kinase transcriptionally-regulated early response gene, Homer1a, was up-regulated transiently in the hippocampus and down-regulated in the cortex of young adult male Fischer 344 X Brown Norway rats at 48 h after 40 Gy of fractionated whole-brain irradiation. Two months after fractionated whole-brain irradiation, these changes in Homer1a expression correlated with a down-regulation of the hippocampal glutamate receptor 1 and protein kinase Cγ, and an up-regulation of cortical glutamate receptor 1 and protein kinase Cγ. Two drugs that prevent radiation-induced cognitive impairment in rats, the angiotensin type-1 receptor blocker, L-158,809, and the angiotensin converting enzyme inhibitor, ramipril, reversed the fractionated whole-brain irradiation-induced Homer1a expression at 48 h in the hippocampus and cortex and restored glutamate receptor 1 and protein kinase Cγ to the levels in sham-irradiated controls at 2 months after fractionated whole-brain irradiation. These data indicate that Homer1a is, (1) a brain region specific regulator of radiation-induced brain injury, including cognitive impairment and (2) potentially a druggable target for preventing it.