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Potential diagnostic marker for TSE [blood test]

Kathy

Well-known member
J Anim Sci. 2007 Feb 12;

Elevated Manganese Levels in Blood and CNS Occur Prior to Onset of Clinical Signs in Scrapie and BSE.

Hesketh S, Sassoon J, Knight R, Hopkins J, Brown DR.
Department of Biology and Biochemistry, University of Bath, Bath, BA2 7AY, UK.

Prion diseases or transmissible spongiform encephalopathies are neurodegenerative diseases that can only be accurately diagnosed by analysis of central nervous system tissue for the presence of an abnormal isoform of the prion protein known as PrP(Sc). Furthermore, these diseases have long incubation periods during which there are no clear symptoms but where the infectious agent could still be present in tissues. Therefore the development of diagnostic assays to detect a surrogate marker for the presence of prion disease is essential. Previous studies on mice experimentally infected with scrapie suggested that changes in the levels of manganese occur in both the blood and brain prior to the onset of symptoms of the disease. In order to assess whether these findings have relevance to the animal diseases scrapie and bovine spongiform encephalopathy (BSE), tissues from BSE and scrapie infected cattle and sheep were analysed for their metal content and compared to values for non infected animals. In both field cases and experimentally infected animals elevated manganese was associated with prion infection. Although some central nervous system regions showed elevated manganese other regions did not. The most consistent finding was elevation of manganese in blood. This change was present in experimentally infected animals before the onset of symptoms. In scrapie infected sheep, elevated manganese levels occurred regardless of the genotype of the sheep, and was even detected in scrapie resistant sheep where no symptoms of disease were detected. These findings suggest that elevated blood manganese could be a potential diagnostic marker for prion infection even in the absence of apparent clinical disease.

PMID: 17296770


Free Radic Biol Med. 2006 Mar 15;40(6):1004-16. Epub 2005 Nov 21.

Delayed radioprotection by nuclear transcription factor kappaB -mediated induction of manganese superoxide dismutase in human microvascular endothelial cells after exposure to the free radical scavenger WR1065.

Murley JS, Kataoka Y, Weydert CJ, Oberley LW, Grdina DJ.
Department of Radiation and Cellular Oncology, The University of Chicago, IL 60637, USA. [email protected]

The free radical scavenger WR1065 (SH) is the active thiol form of the clinically approved cytoprotector amifostine. At doses of 40 microM and 4 mM it can activate the redox-sensitive nuclear transcription factor kappaB (NFkappaB) and elevate the expression of the antioxidant gene manganese superoxide dismutase (MnSOD) in human microvascular endothelial cells (HMEC). MnSOD contains binding motifs for a number of transcription factors other than NFkappaB and codes for a potent antioxidant enzyme localized in the mitochondria that is known to confer enhanced radiation resistance to cells. The purpose of this study was to determine the effect of WR1065 exposure on the various transcription factors known to affect MnSOD expression along with the subsequent kinetics of intracellular elevation of MnSOD protein levels and associated change in sensitivity to ionizing radiation in HMEC. Cells were grown to confluence and exposed to WR1065 for 30 min. Affects on the transcription factors AP1, AP2, CREB, NFkappaB, and Sp1 were monitored as a function of time ranging from 30 min to 4 h after drug exposure using a gel-shift assay. Only NFkappaB exhibited a marked activation and that occurred 30 min following the cessation of drug exposure. MnSOD protein levels, as determined by Western blot analysis, increased up to 16-fold over background control levels by 16 h following drug treatment, and remained at 10-fold or higher levels for an additional 32 h. MnSOD activity was evaluated using a gel-based assay and was found to be active throughout this time period. HMEC were irradiated with X-rays either in the presence of 40 microM or 4 mM WR1065 or 24 h after its removal when MnSOD levels were most elevated. No protection was observed for cells irradiated in the presence of 40 microM WR1065. In contrast, a 4 mM dose of WR1065 afforded an increase in cell survival by a factor of 2. A "delayed radioprotective" effect was, however, observed when cells were irradiated 24 h later, regardless of the concentration of WR1065 used. This effect is characterized as an increase in survival at the 2 Gy dose point, i.e., a 40% increase in survival, and an increase in the initial slope of the survival curve by a factor of 2. The anti-inflammatory sesquiterpene lactone, Helenalin, is an effective inhibitor of NFkappaB activation. HMEC were exposed to Helenalin for 2 h at a nontoxic concentration of 5 microM prior to exposure to WR1065. This treatment not only inhibited activation of NFkappaB by WR1065, but also inhibited the subsequent elevation of MnSOD and the delayed radioprotective effect. A persistent marked elevation of MnSOD in cells following their exposure to a thiol-containing reducing agent such as WR1065 can result in an elevated resistance to the cytotoxic effects of ionizing radiation and represents a novel radioprotection paradigm.

PMID: 16540396

J Gene Med. 2006 May;8(5):557-65.

Radioprotective gene therapy through retroviral expression of manganese superoxide dismutase.

Southgate TD, Sheard V, Milsom MD, Ward TH, Mairs RJ, Boyd M, Fairbairn LJ.
Cancer Research UK Gene Therapy Group, Paterson Institute for Cancer Research, Wilmslow Road, Manchester M20 4BX, UK. [email protected]

BACKGROUND: Radiotherapy for the control of cancer, either alone or in conjunction with chemotherapy, is often limited by normal tissue toxicity including haematopoietic toxicity. Exposure of cells to ionizing radiation leads to the formation of reactive oxygen species that are associated with radiation-induced cytotoxicity. The antioxidant enzyme manganese superoxide dismutase (SOD2) catalyzes the dismutation of the superoxide anions into hydrogen peroxide. METHODS: We have investigated the potential of SOD2 overexpression, through retroviral gene transfer using a retrovirus optimized for transcription in early haematopoietic cells, to enhance the radioresistance of a human erythroleukaemic cell line and primary murine bone marrow. Using these as in vitro models we have investigated whether SOD2 gene therapy may be suitable for the protection of the haematopoietic compartment from the effects of ionizing radiation. RESULTS: Here we demonstrate using both biological and physical assays that overexpression of SOD2 protects haematopoietic cells from ionizing radiation injury. Our results show that an increase in the levels of SOD2 enzymatic activity within K562 cells (from 160.7 +/- 23.6 to 321.8 +/- 45.2 U/mg protein) or primary murine haematopoietic progenitor cells leads to both a significant decrease in DNA fragmentation and a significant increase in clonogenic survival, as evident by a significant increase in Dbar (from 2.66 to 3.42Gy), SF2 (from 0.52 to 0.73) values, and a significant decrease in the alpha value (from 0.3040 +/- 0.037 to 0.0630 +/- 0.037 Gy(-1)) when compared either to cells transduced with a retroviral vector encoding eGFP alone or to the parental line. CONCLUSIONS: The results presented suggest that retroviral radioprotective gene therapy may be applicable to the haematopoietic compartment, enabling radiation dose escalation in cancer therapy. Copyright (c) 2006 John Wiley & Sons, Ltd.

PMID: 16506247


In Vivo. 2005 Nov-Dec;19(6):997-1004.

MnSOD-plasmid liposome gene therapy decreases ionizing irradiation-induced lipid peroxidation of the esophagus.

Epperly MW, Tyurina YY, Nie S, Niu YY, Zhang X, Kagan V, Greenberger JS.
Department of Radiation Oncology, University of Pittsburgh Cancer Institute and School of Environmental Health, University of Pittsburgh Medical Center, Pittsburgh, PA 15213, USA.

BACKGROUND: Ionizing irradiation-induced cellular and tissue damage is mediated in part by resultant radiochemical reactions and resultant oxidative stress. Irradiation-induced reactive oxygen and nitrogen species include: superoxide, nitric oxide, hydroxyl radical and hydrogen peroxide. The biochemical combination of superoxide and nitric oxide radicals forms peroxynitrite, a potent oxidant known to induce lipid peroxidation. MATERIALS AND METHODS: The antioxidant capacity and lipid peroxidation of the esophagus were determined following irradiation. RESULTS: In the present studies, measurements of total antioxidant capacity did not change in the esophagus of control irradiated or control plasmid pNGVL3-PL intraesophageally-injected mice. In contrast, manganese superoxide dismutase-plasmid/liposome (MnSOD-PL) intraesophageally-treated mice showed a significant increase in antioxidant capacity persisting for seven days. Lipid peroxidative changes induced in the control irradiated mouse esophagus decreased over seven days after irradiation of C3H/HeNHsd mice exposed to 37 Gy in a single fraction. MnSOD-PL radioprotective gene therapy administered intraorally 24 hours prior to irradiation did not significantly reduce the kinetics of induction of total peroxidated lipids over the first seven days after irradiation but did decrease lipid peroxidation at days 14 and 21. CONCLUSION: These studies demonstrate the antioxidant function of MnSOD-PL gene therapy to the esophagus, which is detectable as a reduction in irradiation-induced lipid peroxidation.

PMID: 16277013

Zhonghua Zhong Liu Za Zhi. 2005 Nov;27(11):672-5.


[Manganese superoxide dismutase gene transfection of mouse small intestinal epithelial cells protects them from radiation injury][Article in Chinese]

Guo HL, Zhao HW, Xu ZF, Ma H, Song XL, Guan J, Li ZJ, Yu JM.
Department of General Surgery, Cancer Hospital of Shandong Province, Jinan 250117, China.

OBJECTIVE: To investigate the protective effect of manganese superoxide dismutase (MnSOD) gene transfer to small intestinal epithelial cells from radiation injury. METHODS: Herpes simplex virus (HSV) vector containing both the human MnSOD and GFP genes was introduced into mouse small intestine. Expression of MnSOD by the intestinal villi was confirmed by nested RT-PCR, immunofluorescence and enzyme activity assay. Mice were then given various doses of irradiation over the abdomen. The height of intestinal villi was measured on histopathology sections by SZ-PT optical system before irradiation, 24 h and 72 h post-irradiation. All comparisons were performed by one-way analysis of variance using the SPSS statistical software to analyze the significance between groups. RESULTS: Nested RT-PCR, immunofluorescence and enzyme activity assay of MnSOD demonstrated overexpression and increased activity of MnSOD in the inoculated intestine of mice. Control (sham inoculated) irradiated mice showed decreased villi height by 40.1%-59.3% on day 1 and 44.2%-65.1% on day 3 (7.5-15 Gy). Treatment of mice with HSV-MnSOD prior to radiation led to statistically significant radioprotection of the small bowel with mean villi height decreased by only 3.1%-12.4% on day 1 and 6.3%-29.1% on day 3. CONCLUSION: The results demonstrate that overexpression of human MnSOD via a replication defective herpes simplex viral vector is an effective method to protect the small intestine from damage caused by ionizing radiation.
PMID: 16438889

MnSOD - manganese based superoxide dismutase is a anti-oxidant "warrior" against the damaging reactive oxygen species generated by "ionizing radiation".
 
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