Sean D. Hurley

Cyclooxygenase Inhibition as a Strategy to Ameliorate Brain Injury

Cyclooxygenase (COX) is the obligate, rate-limiting enzyme for the conversion of arachidonic acid into prostaglandins. Two COX enzymes have been identified: a constitutively expressed COX-1 and an inducible, highly regulated COX-2. Widely used to treat chronic inflammatory disorders, COX inhibitors have shown promise in attenuating inflammation associated with brain injury. However, the use of COX inhibition in the treatment of brain injury has met with mixed success. This review summarizes our current understanding of COX expression in the central nervous system and the effects of COX inhibitors on brain injury. Three major targets for COX inhibition in the treatment brain injury have been identified. These are the cerebrovasculature, COX-2 expression by vulnerable neurons, and the neuroinflammatory response. Evidence suggests that given the right treatment paradigm, COX inhibition can influence each of these three targets. Drug interactions and general considerations for administrative paradigms are also discussed. Although therapies targeted to specific prostaglandin species, such as PGE2, might prove more ameliorative for brain injury, at the present time non-specific COX inhibitors and COX-2 specific inhibitors are readily available to researchers and clinicians. We believe that COX inhibition will be a useful, ameliorative adjunct in the treatment of most forms of brain injury.

Helper-free HSV-1 amplicons elicit a markedly less robust innate immune response in the CNS

The development and implementation of direct gene transfer technologies for the study and treatment of chronic CNS disorders inherently requires consideration of vector safety. Virus-based vectors represent the most efficient modalities but harbor the potential to induce vigorous innate and adaptive immune responses when administered in vivo. These responses can arise because of virus particle components, resultant viral gene expression, and/or transgene expression. In the current study, we describe the innate responses elicited upon stereotactic delivery of herpes simplex virus type 1-based amplicon vectors. C57BL/6 mice were injected with sterile saline, beta-galactosidase-expressing amplicon (HSVlac) packaged by a conventional helper virus-based technique, or helper virus-free HSVlac. After killing the mice at either 1 or 5 days after transduction, we analyzed them by immunocytochemistry and quantitative RT-PCR for various chemokine, cytokine, and adhesion molecule gene transcripts. All injections induced inflammation, with blood/brain barrier opening on day 1 that was enhanced with both amplicon preparations as compared with saline controls. By day 5, mRNA levels for the pro-inflammatory cytokines (IL-1beta, TNF-alpha, IFN-gamma), chemokines (MCP-1, IP-10), and an adhesion molecule (ICAM-1) had returned to baseline in saline-injected mice and to near-baseline levels in helper virus-free amplicon groups. In contrast, mice injected with helper virus-packaged amplicon stocks elicited elevated inflammatory molecule expression and immune cell infiltration even at day 5. In aggregate, we demonstrate that helper virus-free amplicon preparations exhibit a safer innate immune response profile, presumably as a result of the absence of helper virus gene expression, and provide support for future amplicon-based CNS gene transfer strategies.

Cytosolic prostaglandin E2 synthase (cPGES) expression is decreased in discrete cortical regions in psychiatric disease.

The number of adults in the US affected by bipolar disorder, depression, or schizophrenia is approaching 15 million. Despite decades of research, etiologies of these illnesses remain elusive. Theories of aberrant brain morphology, neurotransmission, and signal conduction have provided the heuristic framework for a large body of literature, with attention focused upon hypotheses of monoamine signaling underlying psychiatric disease. More recently, attention has turned to potential contributions of other signaling pathways, including the arachidonic acid cascade and generation of prostaglandins (PG). To determine the potential involvement of the pathways leading to PGE2 synthesis in psychiatric disease, immunohistochemistry and immunoblotting were performed to measure regional expression of the cyclooxygenases (COX) and one of the terminal PGE2 synthases (PGES) in postmortem tissue provided by The Stanley Medical Research Institute. For normal, bipolar, depressed, and schizophrenic subjects, COX-1 and COX-2 protein levels did not differ across region and patient populations. In contrast, there was a significant effect of diagnosis on cytosolic PGES (cPGES) protein levels in the frontal cortex, with remarkable decreases observed in all psychiatric groups relative to normal tissue (P < 0.05). Significant reduction of cPGES expression was also found in the temporal cortex of bipolar subjects. Evaluation of medicated vs. non-medicated subjects revealed a significant effect of medication on cPGES expression in the frontal cortex of bipolar, but not depressed or schizophrenic subjects. These novel findings further support hypotheses of abnormalities in fatty acid and phospholipid metabolism in regions associated with psychiatric disease.

Facial nerve axotomy in aged and young adult rats: analysis of the glial response

With increasing age, there is a trend towards greater morbidity and injury extent with brain injury. Because several reports have suggested that microglia and astrocytes have an exacerbated response to brain injury in the aged, we set out to explore glial responses to facial nerve axotomy. This model was chosen because the glial responses are well-characterized in young rats and there is no perturbation of the blood-brain barrier (BBB). Immunohistochemistry was performed for glial fibrillary acidic protein (GFAP), leukocyte common antigen, type 3 complement receptor, and major histocompatability complex classes I and II. Quantitative analysis showed that age does not affect the glial response to axotomy in the lesioned facial nucleus; however, an aging-related contralateral effect with enhanced GFAP-labeling was observed. Interestingly, despite a lack of infiltrating neutrophils, a T cell influx was observed in both young and aged rats. Overall, these results suggest that neutrophil extravasion and BBB breakdown are underappreciated with regards to aging and injury exacerbation.

Central Nervous System Effects of Whole-Body Proton Irradiation

Space missions beyond the protection of Earths magnetosphere expose astronauts to an environment that contains ionizing proton radiation. The hazards that proton radiation pose to normal tissues, such as the central nervous system (CNS), are not fully understood, although it has been shown that proton radiation affects the neurogenic environment, killing neural precursors and altering behavior. To determine the time and dose-response characteristics of the CNS to whole-body proton irradiation, C57BL/6J mice were exposed to 1 GeV/n proton radiation at doses of 0–200 cGy and behavioral, physiological and immunohistochemical end points were analyzed over a range of time points (48 h–12 months) postirradiation. These experiments revealed that proton radiation exposure leads to: 1. an acute decrease in cell division within the dentate gyrus of the hippocampus, with significant differences detected at doses as low as 10 cGy; 2. a persistent effect on proliferation in the subgranular zone, at 1 month postirradiation; 3. a decrease in neurogenesis at doses as low as 50 cGy, at 3 months postirradiation; and 4. a decrease in hippocampal ICAM-1 immunoreactivity at doses as low as 10 cGy, at 1 month postirradiation. The data presented contribute to our understanding of biological responses to whole-body proton radiation and may help reduce uncertainty in the assessment of health risks to astronauts. These findings may also be relevant to clinical proton beam therapy.

Acute morphine exposure potentiates the development of HSV-1-induced encephalitis

A devastating consequence of HSV-1 infection is development of HSV-1-induced encephalitis (HSVE). While only a minority of individuals infected with HSV-1 experiences HSVE, clearly defined variables that consistently predict development of the disease remain to be elucidated. The current study examined the effects of a single dose of morphine prior to infection with HSV-1 on the development of HSVE in BALB/cByJ mice. Acute morphine exposure was observed to potentiate the development of HSVE in HSV-1 infected mice. The present data implicate a potential role for the blood-brain barrier in the development of HSVE in morphine-treated mice.

X-Ray Microbeam Irradiation of the Contusion-Injured Rat Spinal Cord Temporarily Improves Hind-Limb Function

Spinal cord injury is a devastating condition with no effective treatment. The physiological processes that impede recovery include potentially detrimental immune responses and the production of reactive astrocytes. Previous work suggested that radiation treatment might be beneficial in spinal cord injury, although the method carries risk of radiation-induced damage. To overcome this obstacle we used arrays of parallel, synchrotron-generated X-ray microbeams (230 μm with 150 μm gaps between them) to irradiate an established model of rat spinal cord contusion injury. This technique is known to have a remarkable sparing effect in tissue, including the central nervous system. Injury was induced in adult female Long-Evans rats at the level of the thoracic vertebrae T9-T10 using 25 mm rod drop on an NYU Impactor. Microbeam irradiation was given to groups of 6–8 rats each, at either Day 10 (50 or 60 Gy in-beam entrance doses) or Day 14 (50, 60 or 70 Gy). The control group was comprised of two subgroups: one studied three months before the irradiation experiment (n = 9) and one at the time of the irradiations (n = 7). Hind-limb function was blindly scored with the Basso, Beattie and Bresnahan (BBB) rating scale on a nearly weekly basis. The scores for the rats irradiated at Day 14 post-injury, when using t test with 7-day data-averaging time bins, showed statistically significant improvement at 28–42 days post-injury (P < 0.038). H&E staining, tissue volume measurements and immunohistochemistry at day ∼110 post-injury did not reveal obvious differences between the irradiated and nonirradiated injured rats. The same microbeam irradiation of normal rats at 70 Gy in-beam entrance dose caused no behavioral deficits and no histological effects other than minor microglia activation at 110 days. Functional improvement in the 14-day irradiated group might be due to a reduction in populations of immune cells and/or reactive astrocytes, while the Day 10/Day 14 differences may indicate time-sensitive changes in these cells and their populations. With optimizations, including those of the irradiation time(s), microbeam pattern, dose, and perhaps concomitant treatments such as immunological intervention this method may ultimately reach clinical use.

Neurogenic Effects of Low-Dose Whole-Body HZE (Fe) Ion and Gamma Irradiation

Understanding the dose-toxicity profile of radiation is critical when evaluating potential health risks associated with natural and man-made sources in our environment. The purpose of this study was to evaluate the effects of low-dose whole-body high-energy charged (HZE) iron (Fe) ions and low-energy gamma exposure on proliferation and differentiation of adult-born neurons within the dentate gyrus of the hippocampus, cells deemed to play a critical role in memory regulation. To determine the dose-response characteristics of the brain to whole-body Fe-ion vs. gamma-radiation exposure, C57BL/6J mice were irradiated with 1 GeV/n Fe ions or a static 137Cs source (0.662 MeV) at doses ranging from 0 to 300 cGy. The neurogenesis was analyzed at 48 h and one month postirradiation. These experiments revealed that whole-body exposure to either Fe ions or gamma radiation leads to: 1. An acute decrease in cell division within the dentate gyrus of the hippocampus, detected at doses as low as 30 and 100 cGy for Fe ions and gamma radiation, respectively; and 2. A reduction in newly differentiated neurons (DCX immunoreactivity) at one month postirradiation, with significant decreases detected at doses as low as 100 cGy for both Fe ions and gamma rays. The data presented here contribute to our understanding of brain responses to whole-body Fe ions and gamma rays and may help inform health-risk evaluations related to systemic exposure during a medical or radiologic/nuclear event or as a result of prolonged space travel.