Abstract The goal of this study was to evaluate cytokine secretion capacity in a mouse model of prostate cancer, both with and without metalloporphyrin antioxidant and radiation treatment. C57BL/6 mice with subcutaneous RM-9 tumors were treated daily for 12 days with MnTE-2-PyP(5+)[Mn (III) tetrakis (N-ethylpyridinium-2-yl) porphyrin], beginning 1 day after injection of RM-9 cells; a 10-Gy tumor-localized dose of (60)Co gamma rays was administered in a single fraction on day 7. Spleen, tumors and plasma were collected on day 12. T cells in the spleen were activated with anti-CD3 antibody and supernatants were collected. Twenty-two cytokines were quantified in spleen supernatants, five in tumor homogenates, and three in plasma using multiplex bead array technology and ELISA. The presence of a tumor had significant effects on many of the cytokines quantified (P < 0.05). Tumor-induced depression was evident for eight spleen cytokines (TNF-alpha, G-CSF, GM-CSF, IFN-gamma, IL10, IP-10, MIP-1alpha and mKC), whereas only three were enhanced (IL1beta, IL6 and MCP-1). Radiotherapy resulted in enhanced splenocyte capacity to produce IL4 and IL13 and increased IL4, MCP-1 and VEGF in tumors (P < 0.05). Addition of MnTE-2-PyP(5+) to radiation decreased the concentrations of IL4, IL13 and TGF-beta1 in spleen supernatants and IL4 and VEGF in tumors (P < 0.05 compared to radiation alone). Some differences were also noted in plasma cytokines. Overall, the findings suggest that administration of MnTE-2-PyP(5+) together with radiotherapy may enhance anti-tumor immune responsiveness and decrease the risk for radiation-induced normal tissue toxicities.

Purpose: We examined the effects of manganese (III) meso-tetrakis (diethyl-2-5-imidazole) porphyrin, a metalloporphyrin antioxidant (MPA), on neural tissue radiation toxicity in vivo and on tumour cell radiosensitivity in vitro. Materials and methods: MPA was administered directly into the right lateral ventricle of young adult, male Sprague-Dawley rats (0 or 3.4 mug) 3 h before treatment with a single fraction, 100 Gy radiation dose delivered to the left brain hemisphere. The effects of treatment on radiation responses were assessed at different time points following irradiation. Results: MPA treatment prior to brain irradiation protected against acute radiation-induced apoptosis and ameliorated delayed damage to the blood-brain barrier and radiation necrosis, but without producing a discernible increase in tissue superoxide disumtase (SOD) activity. In vitro, MPA pretreatment protected against radiation-induced apoptosis in primary neuronal cultures and increased clonogenic survival of irradiated rat glioma C6 cells, but had no discernible effect on radiation-induced DNA double-strand breaks. MPA, a low molecular weight SOD mimic, significantly increased mitochondrial SOD activity in C6 cells, but not total cellular SOD activity. MPA up-regulated C6 expression of heme-oxygenase 1 (HO-1), an endogenous radioprotectant, but had no effect on HO-1 levels in human astrocytoma U-251 cells, human prostatic carcinoma LNCaP cells, or primary rat brain microvascular endothelial cells in vitro, nor on brain tissue HO-1 expression levels in vivo. Conclusions: Metalloporphyrin antioxidants merit further exploration as adjunctive radioprotectants for cranial radiotherapy/radiosurgery applications, although the potential for tumour protection must be carefully considered.

Sustained oxidative stress is a known sequel to focal cerebral ischemia. This study examined the effects of treatment with a single dose or sustained infusion of the redox-modulating MnPorphyrin Mn(III)TDE-2-ImP(5+) on outcome from middle cerebral artery occlusion (MCAO) in the rat. Normothermic rats were subjected to 90 min MCAO followed by 90 min reperfusion and then were treated with a single intracerebroventricular dose of Mn(III)TDE-2-ImP(5+). Neurologic and histologic outcomes were assessed at 1 or 8 weeks post-ischemia. A single dose of Mn(III)TDE-2-ImP(5+) caused a dose-dependent improvement in histologic and neurologic outcome when assessed 1 week post-ischemia. Mn(III)TDE-2-ImP(5+) afforded preservation of brain aconitase activity at 5.5 hrs after reperfusion onset, consistent with its known antioxidant properties. Mn(III)TDE-2-ImP(5+) also attenuated post-ischemic NF-kappaB activation. Evidence for effects on cerebral infarct size and neurologic function had completely dissipated when rats were allowed to survive for 8 weeks post-ischemia. In contrast, a 1-week continuous intracerebroventricular Mn(III)TDE-2-ImP(5+) infusion caused persistent and substantive reduction in both cerebral infarct size and neurologic deficit at 8 weeks post-ischemia. Pharmacologic modulation of post-ischemic oxidative stress is likely to require sustained intervention for enduring efficacy in improving neurologic and histologic outcome from a transient focal ischemic insult.

BACKGROUND: Metalloporphyrin antioxidants can protect tissues against radiation-induced damage. However, for effective use in radiotherapy as normal tissue radioprotectants, they must not protect the cancer. The major objectives were to evaluate the effects of Mn (III) tetrakis (N-ethylpyridinium-2-yl) porphyrin (MnTE-2-PyP) on tumor response to radiation and to explore mechanisms responsible for the observed effects. MATERIALS AND METHODS: C57BL/6 mice were subcutaneously (s.c.) injected with RM-9 prostate tumor cells on day 0 and grouped according to treatment with MnTE-2-PyP (s.c. 6 mg/kg/day beginning on day 1 for 16 maximum days), 10 Gray (Gy) single fraction radiation on day 7, a combination of both or neither. Subsets per group and non-tumor bearing controls were evaluated for leukocyte populations, red blood cell (RBC) and platelet characteristics and cytokines on day 12; the remaining mice were followed for tumor growth. RESULTS: Although radiation alone significantly slowed tumor growth and the addition of MnTE-2-PyP resulted in slightly slower tumor progression, the difference between radiation and radiation plus drug was not statistically significant. However, the treatment with drug alone significantly elevated T (helper, Th and cyotoxic, Tc) and natural killer (NK) cells in the spleen, B-cells in the blood and spleen, and the capacity to produce interleukin-2. The addition of the drug to radiation did not ameliorate the depression seen in all the major leukocyte types, but did protect against radiation-induced decreases in RBC counts, hemoglobin and hematocrit. Vascular endothelial growth factor (VEGF) increased in the plasma from both the irradiated groups and a trend for increased transforming growth factor-beta1 (TGF-beta1) was noted with radiation alone. CONCLUSION: MnTE-2-PyP did not protect RM-9 prostate tumors against radiation damage and was not toxic under the conditions used. The drug-induced enhancement of certain immune parameters suggests that MnTE-2-PyP may be beneficial not only as a normal tissue radioprotectant, but also as a facilitator of antitumor immunity.

Three dimensional grid phantoms offer a number of advantages for measuring imaging related spatial inaccuracies for image guided surgery and radiotherapy. The authors examined the use of rapid prototyping technology for directly fabricating 3D grid phantoms from CAD drawings. We tested three different fabrication process materials, photopolymer jet with acrylic resin (PJ/AR), selective laser sintering with polyamide (SLS/P), and fused deposition modeling with acrylonitrile butadiene styrene (FDM/ABS). The test objects consisted of rectangular arrays of control points formed by the intersections of posts and struts (2 mm rectangular cross section) and spaced 8 mm apart in the x, y, and z directions. The PJ/AR phantom expanded after immersion in water which resulted in permanent warping of the structure. The surface of the FDM/ABS grid exhibited a regular pattern of depressions and ridges from the extrusion process. SLS/P showed the best combination of build accuracy, surface finish, and stability. Based on these findings, a grid phantom for assessing machine-dependent and frame-induced MR spatial distortions was fabricated to be used for quality assurance in stereotactic neurosurgical and radiotherapy procedures. The spatial uniformity of the SLS/P grid control point array was determined by CT imaging (0.6 x 0.6 x 0.625 mm3 resolution) and found suitable for the application, with over 97.5% of the control points located within 0.3 mm of the position specified in CAD drawing and none of the points off by more than 0.4 mm. Rapid prototyping is a flexible and cost effective alternative for development of customized grid phantoms for medical physics quality assurance.

BACKGROUND: Antioxidants have the potential to protect normal tissues against radiation-induced damage, but must not protect tumor cells during radiotherapy. The major objectives were to determine whether a metalloporphyrin antioxidant affects prostate tumor response to radiation and identify possible mechanisms of interaction. MATERIALS AND METHODS: C57BL/6 mice with RM-9 tumor were treated with manganese (III) meso-tetrakis (1,3-diethylimidazolium-2-yl) porphyrin (MnTDE-2-ImP) and 10 gray (Gy) radiation. Tumor volume was quantified and a subset/group was evaluated for hypoxia-inducible factor-1alpha (HIF-1alpha), bone marrow-derived cell populations and cytokines. RESULTS: The addition of MnTDE-2-ImP transiently increased tumor response compared to radiation alone. The group receiving drug plus radiation had reduced intratumoral HIF-1alpha and decreased capacity to secrete TNF-alpha, whereas production of IL-4 was increased. There were no toxicities associated with combination treatment. CONCLUSION: The results demonstrate that MnTDE-2-ImP did not protect the RM-9 prostate tumor against radiation; instead, radiation effectiveness was modestly increased. Possible mechanisms include reduction of radiation-induced HIF-1alpha and an altered cytokine profile.

A four-stage laser system was developed, emitting at a wavelength of 6450 nm with a 3-5 ns pulse duration,</=2 mJ pulse energy, and 1/2 Hz pulse repetition rate. The laser system successfully ablated rat brain tissue, where both the collateral damage and the ablation rate compare favorably with that previously observed with a Mark-III Free-Electron Laser.

In animal models of severe ischemia, it has not been uniformly observed that anesthetics are protective. However, anesthetics have not been evaluated in the presence of a mild excitotoxic insult. We hypothesized that in the presence of a mild excitotoxic insult, 3 microm N-methyl-D-aspartate (NMDA), isoflurane may prevent apoptotic cell death. Primary mixed neuronal/glial cultures were prepared from fetal rat brains. Mature cultures were exposed to dissolved isoflurane [0 mM, 0.4 mM (1.8 minimum alveolar concentration) or 1.6 mM (7 minimum alveolar concentration)] or dizocilpine (10 microM), and NMDA (0 or 3 microM) at 37 degrees C for 30 minutes. Apoptosis was assessed using terminal-deoxy-nucleotidyl end-nick labeling oligonucleosomal DNA fragmentation enzyme-linked immunosorbent assay, and caspases-3 and -9 activation assays. NMDA (3 muM) induced apoptosis in mixed neuronal/glial cell cultures. Apoptosis induced by 3 microm NMDA was caspase-3 but not caspase-9 mediated. In the presence of a mild excitotoxic insult, this investigation showed an attenuation of apoptotic cell death by dizocilpine, but not isoflurane.

BACKGROUND: Isoflurane provides protection against severe forebrain ischemia in the rat. The authors hypothesized that this is attributable to interaction with the gamma-aminobutyric acid type A (GABAA) receptor resulting in altered time to onset of ischemic hippocampal depolarization. METHODS: Organotypic hippocampal slices were subjected to oxygen-glucose deprivation in the presence of isoflurane and combinations of GABAA (bicuculline) and GABAB (phaclofen) receptor antagonists. Cell death was measured. Rats were subjected to severe forebrain ischemia while anesthetized with fentanyl-nitrous oxide or 1.4% isoflurane. In the isoflurane group, rats also received intravenous bicuculline (0, 1, or 2 mg/kg). Neurologic and histologic outcomes and time to depolarization were assessed. RESULTS: In slices, 2% isoflurane caused near-complete protection against oxygen-glucose deprivation. This was unaffected by coadministration of phaclofen but largely reversed by bicuculline. The GABAA agonist muscimol was also protective, having an effect equivalent to 1% isoflurane. In rats, isoflurane (0 mg bicuculline) improved neurologic and histologic outcome versus fentanyl-nitrous oxide (CA1 percentage of alive neurons: fentanyl-nitrous oxide, 15 +/- 7; isoflurane, 61 +/- 24). The isoflurane effect was reversed in a dose-dependent manner by bicuculline (CA1 percentage alive: 1 mg/kg, 44 +/- 22; 2 mg/kg, 21 +/- 15). Time to depolarization was delayed with isoflurane versus fentanyl-nitrous oxide (137 vs. 80 s) but was not affected by bicuculline (149 s). In contrast, postischemic time to repolarization was more rapid with fentanyl-nitrous oxide or isoflurane plus bicuculline versus isoflurane alone. CONCLUSIONS: These studies are consistent with the hypothesis that the GABAA receptor serves as a major site of action for isoflurane neuroprotection both in vitro and in vivo. However, the mechanism by which this interaction confers in vivo protection cannot be attributed to effects on the duration of ischemic depolarization.

Cognitive changes are common sequelae of cancer and cancer treatment, particularly in patients receiving cranial radiotherapy (RT). These effects are typically assessed by subjective clinical examination or using objective neuropsychological tests. Biologically based neurophysiological methods have been increasingly applied to the study of cognitive processing in neuropsychiatric and neurological disorders and as objective measures of cognitive status for patients with dementia. These methods detect the activation of neural circuits that directly mediate cognitive function in the human brain and include metabolic and electrophysiology based techniques. Neuroimaging procedures such as 18FDG PET and more recently fMRI, which detect metabolic activation associated with cognitive processing, provide excellent spatial resolution and can be directly correlated with neuroradiological findings associated with cranial RT neurotoxicity. Clinical electrophysiology procedures such as cognitive event-related potentials (ERP), which detect the neuronal electrical activity associated with cognitive processing, offer excellent temporal resolution at low cost. Cognitive ERP techniques are already being used to assess severity and progression of cognitive dysfunction in patients with vascular and degenerative dementias, but have been largely overlooked in studies of radiation-related cognitive impairments. We review these various electrophysiological methods in the context of their relevance to assessing cranial RT effects on cognitive function, and provide recommendations for a neurophysiological approach to supplement current neuropsychological tests for RT cognitive impairments. This technology is well suited for clinical assessment of neurocognitive sequelae of cancer and should provide new insights into the mechanism of RT-related cognitive dysfunction.