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Joel S. Greenberger - One of the best experts on this subject based on the ideXlab platform.
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Role of the esophageal vagus neural pathway in ionizing irradiation-induced seizures in nitric oxide synthase-1 homologous recombinant negative NOS1-/- mice.
In vivo (Athens Greece), 2011Co-Authors: Mark E. Bernard, Michael W. Epperly, Jean-claude M. Rwigema, Tracy Dixon, Hyun Uk Kim, Eric E. Kelley, Geoffery H. Murdoch, Hong Wang, Joel S. GreenbergerAbstract:AIM We sought to define the mechanism of total body irradiation (TBI)-induced seizures in NOS1(-/-) mice and amelioration by intra-esophageal manganese superoxide dismutase-plasmid liposomes (MnSOD-PL). MATERIALS AND METHODS We evaluated the role of vagus nerve pathways in irradiation-induced seizures using biochemical, physiologic, and histopathologic techniques. RESULTS Heterozygous NOS1(+/-) mice demonstrated radioresistance similar to wild-type C57BL/6NHsd mice (p=0.9269). Irradiation-induced lipid peroxidation in fetal brain cultures from NOS1(-/-) or wild-type mice was reduced by MnSOD-PL. Right-sided vagotomy did not alter the TBI radiation response of wild-type or reverse the radiosensitivity of NOS1(-/-) mice. Excised esophagus from irradiated NOS1(-/-) mice demonstrated an increased histopathologic inflammatory response compared to C57BL/6NHsd mice. CONCLUSION NOS1(-/-) mice represent a model system for dissecting the developmental abnormalities leading to esophageal-mediated TBI-induced seizures.
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Abstract 492: Esophageal administration of manganese superoxide dismutase plasmid liposomes (MnSOD-PL) reverses thoracic irradiation sensitivity of nitric oxide synthase one homologous recombinant negative (NOS1 -/-) mouse
Tumor Biology, 2010Co-Authors: Michael W. Epperly, Darcy Franicola, Malolan S. Rajagopalan, Jean-claude M. Rwigema, Brandon Stone, Julie P. Goff, Tracy Dixon, Joel S. GreenbergerAbstract:Proceedings: AACR 101st Annual Meeting 2010‐‐ Apr 17‐21, 2010; Washington, DC NOS1 -/- mice are more sensitive to 29 Gy thoracic or 9.5 Gy total body irradiation (TBI) than control C57BL/6NHsd, or NOS2 -/- or NOS3 -/- mice (p < 0.0001 and p = 0.0006). Histological examination of the esophagus, lungs, heart and intestine revealed no explanation of the rapid death of NOS1 -/- mice. Blood and bone marrow isolates six days after TBI (1 day before the NOS1 -/- mouse death) showed similar decrease in blood counts for all the mouse strains. NOS1 -/- mouse marrow irradiated to doses ranging from 0 to 8 Gy showed CFU-GM, BFU-E and CFU-GEMM radiosensitivity similar to that from control mice. There were also no differences in serum electrolytes, liver or renal function tests in irradiated NOS1 -/- compared to C57BL/6NHsd mice. RNA extracted from esophagus, stomach or intestine at day 0, 1 or 6 following 9.5 total body irradiation showed by qualitative RT-PCR analysis of TNF-σ, NF-κβ, and INF-γ similar cytokine expression between NOS1 -/- and C57BL/6NHsd mice. NOS1 -/- and C57BL/6NHsd mice were irradiated to 9.5 Gy total body or 20 Gy to the thoracic cavity and observed by video camera. NOS1 -/- mice had seizures before death following both TBI or thoracic irradiation, while no seizures were detected in dying C57BL/6NHsd mice. NOS1 -/- mice irradiated to 9.5 Gy to the brain (with the remainder of the body shielded) did not produce death from seizures. NOS1 -/- were intraesophageally administered MnSOD-PL (100 µg plasmid DNA) and irradiated to 20 Gy to the upper body 24 hr later. NOS1 -/- mice administered MnSOD-PL had normalized survival compared to control irradiated NOS1 -/- mice: 50% survival at 12 days for irradiated control NOS1 -/- mice compared to 128 days for MnSOD-PL treated NOS1 -/- mice (p = 0.0311). The data indicate that esophageal neuronal abnormalities which cause pyloric stenosis in the strain may be responsible for both the rapid thoracic irradiation sensitivity leading to seizure and its reversal by intraesophageal MnSOD-PL gene therapy. Supported by NIAID grant U19AI068021, and NIH grant T32AG21885 Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 492.
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Modulation of Neuronal Nitric Oxide Synthase (NOS1) Sensitized NOS1 -/- Mice to Total Body Irradiaton.
Blood, 2009Co-Authors: Brandon Stone, Michael W. Epperly, Darcy Franicola, Malolan S. Rajagopalan, Jean-claude M. Rwigema, Julie P. Goff, Tracy Dixon, Medical Umar Salimi, Anthony J. Bauer, Joel S. GreenbergerAbstract:Abstract 4597 Ionizing irradiation results in increased superoxide and nitric oxide (NO) production. These products combine to form peroxynitrite which, through reactions with DNA, lipids, proteins, and cell membranes, initiates apoptotic cell death. Removal of nitric oxide synthase should reduce NO production and limit peroxynitrite formation thereby limiting irradiation induced apoptosis. To test this hypothesis, bone marrow stromal cell lines were derived from long term bone marrow cultures established from NOS1 -/-, NOS2 -/-, NOS3 -/-, or control background strain C57BL/6NHsd mice. Irradiation survival curves using doses ranging from 0 to 8 Gy, and scoring 7 day colonies of greater than 50 cells revealed that NOS1 -/- bone marrow stromal cells were radioresistant compared to all others showing an increased shoulder with a N = 20.8 ± 5.6 compared to 8.3 ± 2.4 for C57BL/6NHsd mice (p = 0.0356). There were no significant differences between the NOS 2 -/- and NOS3 -/- stromal cells and the C57BL/6NHsd stromal cells. To determine whether inhibition of nitric oxide synthase in vivo protected mice from irradiation, groups of C57BL/6NHsd mice had Alzet osmotic pumps containing either the NOS inhibitor, L-NAME, or control phosphate buffered saline (PBS) placed subcutaneously. Three days later, at a time when NO production had been inhibited, mice were irradiated to the LD 50/30 dose of 9.5 Gy total body irradiation, and followed for expected development of the hematopoietic syndrome. Mice with pumps delivering L-NAME showed significantly increased survival (p = 0.0011), compared to control PBS pump containing mice (50% survival at 14 days compared to 8 days). Since L-NAME inhibits all NOS isoforms, NOS1 -/- cell lines were radioresistant in vitro, and NOS1 -/- mice demonstrate in vivo bladder radioresistance (Kanai, Epperly, Pearce et al, American Journal of Physiology-Heart & Circulatory Physiology, 286:H13-H21, 2004) we sought to establish that NOS1-/- mice would be relatively radioresistant to total body irradiation. Groups of female NOS1 -/-, NOS2 -/-, NOS3 -/-, and C57BL/6NHsd mice were total body irradiated to 9.5 Gy. Unexpectedly, NOS1 -/- mice were significantly more radiosensitive (50% survival at 9 days) (p = 0.0006) compared to other groups (50% survival at 25 days for C57BL/6NHsd, 19 days for NOS2 -/-, and greater than 30 days for NOS3 -/-). To determine the mechanism of rapid death in irradiated NOS1-/- mice, peripheral blood was analyzed before and 6 days after 9.5 Gy irradiation. Freshly removed bone marrow CFU-GEMM was tested for clonagenic radiation survival in vitro. There was no significant difference in radiation response of hematocrit, white cell or platelet counts, or marow CFU-GEMM between strains. While NOS1 -/- mice show reduced density of enteric neurons and associated developmental non-morbid pyloric stenosis and gastoesophageal dilation, there was no significant detectable difference in post irradiation histopathology of the esophagus, duodenum, jejunum, ileum, cecum, or colon. There was also no difference in numbers or density of intestinal crypt cells. Intestinal transit studies in irradiated mice demonstrated no significant difference in transit times. Therefore, NOS1 -/- mice display a novel total body irradiation sensitivity that is independent of hematopoietic or gastrointestinal syndromes. Supported by NIAID/NIH grant U19AI068021 Disclosures: No relevant conflicts of interest to declare.
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modulation of neuronal nitric oxide synthase NOS1 sensitized NOS1 mice to total body irradiaton
Blood, 2009Co-Authors: Brandon Stone, Michael W. Epperly, Darcy Franicola, Malolan S. Rajagopalan, Jean-claude M. Rwigema, Julie P. Goff, Tracy Dixon, Medical Umar Salimi, Anthony J. Bauer, Joel S. GreenbergerAbstract:Abstract 4597 Ionizing irradiation results in increased superoxide and nitric oxide (NO) production. These products combine to form peroxynitrite which, through reactions with DNA, lipids, proteins, and cell membranes, initiates apoptotic cell death. Removal of nitric oxide synthase should reduce NO production and limit peroxynitrite formation thereby limiting irradiation induced apoptosis. To test this hypothesis, bone marrow stromal cell lines were derived from long term bone marrow cultures established from NOS1 -/-, NOS2 -/-, NOS3 -/-, or control background strain C57BL/6NHsd mice. Irradiation survival curves using doses ranging from 0 to 8 Gy, and scoring 7 day colonies of greater than 50 cells revealed that NOS1 -/- bone marrow stromal cells were radioresistant compared to all others showing an increased shoulder with a N = 20.8 ± 5.6 compared to 8.3 ± 2.4 for C57BL/6NHsd mice (p = 0.0356). There were no significant differences between the NOS 2 -/- and NOS3 -/- stromal cells and the C57BL/6NHsd stromal cells. To determine whether inhibition of nitric oxide synthase in vivo protected mice from irradiation, groups of C57BL/6NHsd mice had Alzet osmotic pumps containing either the NOS inhibitor, L-NAME, or control phosphate buffered saline (PBS) placed subcutaneously. Three days later, at a time when NO production had been inhibited, mice were irradiated to the LD 50/30 dose of 9.5 Gy total body irradiation, and followed for expected development of the hematopoietic syndrome. Mice with pumps delivering L-NAME showed significantly increased survival (p = 0.0011), compared to control PBS pump containing mice (50% survival at 14 days compared to 8 days). Since L-NAME inhibits all NOS isoforms, NOS1 -/- cell lines were radioresistant in vitro, and NOS1 -/- mice demonstrate in vivo bladder radioresistance (Kanai, Epperly, Pearce et al, American Journal of Physiology-Heart & Circulatory Physiology, 286:H13-H21, 2004) we sought to establish that NOS1-/- mice would be relatively radioresistant to total body irradiation. Groups of female NOS1 -/-, NOS2 -/-, NOS3 -/-, and C57BL/6NHsd mice were total body irradiated to 9.5 Gy. Unexpectedly, NOS1 -/- mice were significantly more radiosensitive (50% survival at 9 days) (p = 0.0006) compared to other groups (50% survival at 25 days for C57BL/6NHsd, 19 days for NOS2 -/-, and greater than 30 days for NOS3 -/-). To determine the mechanism of rapid death in irradiated NOS1-/- mice, peripheral blood was analyzed before and 6 days after 9.5 Gy irradiation. Freshly removed bone marrow CFU-GEMM was tested for clonagenic radiation survival in vitro. There was no significant difference in radiation response of hematocrit, white cell or platelet counts, or marow CFU-GEMM between strains. While NOS1 -/- mice show reduced density of enteric neurons and associated developmental non-morbid pyloric stenosis and gastoesophageal dilation, there was no significant detectable difference in post irradiation histopathology of the esophagus, duodenum, jejunum, ileum, cecum, or colon. There was also no difference in numbers or density of intestinal crypt cells. Intestinal transit studies in irradiated mice demonstrated no significant difference in transit times. Therefore, NOS1 -/- mice display a novel total body irradiation sensitivity that is independent of hematopoietic or gastrointestinal syndromes. Supported by NIAID/NIH grant U19AI068021 Disclosures: No relevant conflicts of interest to declare.
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Increased longevity of hematopoiesis in continuous bone marrow cultures derived from NOS1 (nNOS, mtNOS) homozygous recombinant negative mice correlates with radioresistance of hematopoietic and marrow stromal cells.
Experimental hematology, 2007Co-Authors: Michael W. Epperly, Shaonan Cao, Xichen Zhang, Darcy Franicola, Hongmei Shen, Emily E. Greenberger, Laura D. Epperly, Joel S. GreenbergerAbstract:Objective Neuronal nitric oxide synthase (NOS1, mitochondrial NOS, neuronal NOS) homozygous deletion recombinant negative mice demonstrate ionizing irradiation resistance in vivo, attributable to the decrease in mitochondrial-localized production of peroxynitrite, a potent lipid toxic free radical species resulting from the combination of nitric oxide and superoxide. The present studies were designed to determine whether reduced mitochondrial generation of toxic radical oxygen species in NOS1−/− mice also increased the longevity of hematopoiesis in continuous bone marrow cultures and conferred radioresistance to cells in vitro. Materials and Methods Long-term bone marrow cultures (LTBMCs) were established from NOS1−/− and NOS1+/+ littermate mice. Radiation resistance of hematopoietic and marrow stromal cells was measured. Cell cycle analysis and measurement of glutathione and glutathione peroxidase were carried out on irradiated clonal bone marrow stromal cell lines. Results A significant increase in longevity of hematopoiesis was detected in NOS1−/− mouse LTBMCs for over 64 weeks in culture compared to 20 weeks for NOS1+/+ mouse LTBMCs (p n ¯ = 32.15 ± 1.21 compared to NOS1+/+ cells n ¯ = 10.47 ± 3.2 (p = 0.0026), interleukin-3-dependent NOS1−/− hematopoietic progenitor cell lines also demonstrated decreased apoptosis after 10 Gy irradiation. Both pre- and postirradiation stabilization of the cellular antioxidant pool was detected in NOS1−/− cells. NOS1−/− cells showed a prolonged G1 cell cycle arrest after 10 Gy. Conclusions Prolonged hematopoiesis in LTBMCs correlates with intrinsic radioresistance of hematopoietic and marrow stromal cells from NOS1−/− mice. The data confirm the importance to hematopoiesis of mitochondrial localized nitric oxide in both radioresistance and longevity of hematopoiesis in continuous bone marrow cultures.
Anne Jarry - One of the best experts on this subject based on the ideXlab platform.
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Loss of NOS1 expression in high-grade renal cell carcinoma associated with a shift of NO signalling.
British journal of cancer, 2004Co-Authors: Karine Renaudin, Marc G. Denis, Georges Karam, Geneviève Vallette, Francoise Buzelin, Christian L. Laboisse, Anne JarryAbstract:In normal human kidney, NOS1 and soluble guanylate cyclase (sGC) are expressed in tubular epithelial cells, suggesting a physiological autocrine NO signalling pathway. Therefore, we investigated both NOS1 and sGC expressions in benign and malignant renal tumours. In addition, we examined the pattern of protein tyrosine nitration in normal and tumour tissue. NOS1 expression and activity were found to be downregulated, correlating with the tumour grade, as shown by immunohistochemistry, quantitative RT-PCR analysis, and histochemical detection of the NADPH-diaphorase activity of nitric oxide synthases (NOS). These results show that the autocrine NO signalling pathway is maintained in benign tumours and lost in malignant tumours. In contrast, sGC expression was maintained in renal tumours whatever the tumour type, a finding showing that tumour cells remain sensitive to the bioregulatory role of exogeneous NO(*). Finally, the staining pattern of protein tyrosine nitration, assessed by immunohistochemistry, parallelled that of NOS1 expression in normal renal parenchyma and benign tumours, supporting the concept that protein nitration was accounted for by NOS1 activity. In contrast, in malignant tumours, protein tyrosine nitration was accounted for by the production of reactive nitrogen oxide species by the inflammatory infiltrate. Altogether, these findings argue for a pattern of NO signalling similar in normal kidney and benign renal tumours, whereas it is completely different in malignant renal tumours.
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expression of NOS1 and soluble guanylyl cyclase by human kidney epithelial cells morphological evidence for an autocrine paracrine action of nitric oxide
Kidney International, 2003Co-Authors: Anne Jarry, Karine Renaudin, Marc G. Denis, Francoise Buzelin, Christian L. Laboisse, Myriam Robard, Hervé Paris, Benedicte Buffinmeyer, G Karam, G ValletteAbstract:Expression of NOS1 and soluble guanylyl cyclase by human kidney epithelial cells: Morphological evidence for an autocrine/paracrine action of nitric oxide. Background Nitric oxide plays an important role in the kidney through effects on both renal hemodynamics and tubular functions. Tubular epithelial cells are thus a target for nitric oxide. However, as to whether tubular epithelial cells endogeneously produce nitric oxide under physiologic conditions in human kidney is currently unknown. The aim of the present study was to characterize and localize in situ the nitric oxide synthase (NOS) isoforms (NOS1, NOS2, and NOS3) expressed in human normal kidney, and soluble guanylyl cyclase, the well-known target for nitric oxide. Methods Five complementary experimental approaches were used: ( 1 ) detection of NOS reductase activity by nicotinamide adenine dinucleotide phosphate (NADPH)-diaphorase histochemistry, ( 2 ) immunolocalization of the NOS isoforms (NOS1, NOS2, NOS3), ( 3 ) immunoblot analysis, ( 4 ) quantitative reverse transcription-polymerase chain reaction (RT-PCR) analysis of NOS mRNA, and ( 5 ) measurement of NOS activity as the conversion rate of L-[ 14 C]-arginine to L-[ 14 C]-citrulline. In addition, in situ detection of soluble guanylyl cyclase was assessed by immunohistochemistry. Results All these techniques led to consistent results showing that epithelial cells of most tubules along the human nephron exhibit functional NOS1, with a corticomedullary gradient observed both at the protein and mRNA levels. Moreover, epithelial cells expressing NOS1 also express soluble guanylyl cyclase, indicating that these cells possess the machinery for autocrine/paracrine effect of nitric oxide. Conclusion The present study demonstrates that NOS1 is strongly expressed in most tubules of the human nephron and therefore invites to consider epithelial cells as one of the major source of nitric oxide in the human kidney under physiologic conditions.
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Expression of NOS1 and soluble guanylyl cyclase by human kidney epithelial cells: Morphological evidence for an autocrine/paracrine action of nitric oxide
Kidney international, 2003Co-Authors: Anne Jarry, Karine Renaudin, Marc G. Denis, Georges Karam, Francoise Buzelin, Christian L. Laboisse, Myriam Robard, Bénédicte Buffin-meyer, Hervé Paris, Geneviève ValletteAbstract:Expression of NOS1 and soluble guanylyl cyclase by human kidney epithelial cells: Morphological evidence for an autocrine/paracrine action of nitric oxide. Background Nitric oxide plays an important role in the kidney through effects on both renal hemodynamics and tubular functions. Tubular epithelial cells are thus a target for nitric oxide. However, as to whether tubular epithelial cells endogeneously produce nitric oxide under physiologic conditions in human kidney is currently unknown. The aim of the present study was to characterize and localize in situ the nitric oxide synthase (NOS) isoforms (NOS1, NOS2, and NOS3) expressed in human normal kidney, and soluble guanylyl cyclase, the well-known target for nitric oxide. Methods Five complementary experimental approaches were used: ( 1 ) detection of NOS reductase activity by nicotinamide adenine dinucleotide phosphate (NADPH)-diaphorase histochemistry, ( 2 ) immunolocalization of the NOS isoforms (NOS1, NOS2, NOS3), ( 3 ) immunoblot analysis, ( 4 ) quantitative reverse transcription-polymerase chain reaction (RT-PCR) analysis of NOS mRNA, and ( 5 ) measurement of NOS activity as the conversion rate of L-[ 14 C]-arginine to L-[ 14 C]-citrulline. In addition, in situ detection of soluble guanylyl cyclase was assessed by immunohistochemistry. Results All these techniques led to consistent results showing that epithelial cells of most tubules along the human nephron exhibit functional NOS1, with a corticomedullary gradient observed both at the protein and mRNA levels. Moreover, epithelial cells expressing NOS1 also express soluble guanylyl cyclase, indicating that these cells possess the machinery for autocrine/paracrine effect of nitric oxide. Conclusion The present study demonstrates that NOS1 is strongly expressed in most tubules of the human nephron and therefore invites to consider epithelial cells as one of the major source of nitric oxide in the human kidney under physiologic conditions.
Michael W. Epperly - One of the best experts on this subject based on the ideXlab platform.
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Role of the esophageal vagus neural pathway in ionizing irradiation-induced seizures in nitric oxide synthase-1 homologous recombinant negative NOS1-/- mice.
In vivo (Athens Greece), 2011Co-Authors: Mark E. Bernard, Michael W. Epperly, Jean-claude M. Rwigema, Tracy Dixon, Hyun Uk Kim, Eric E. Kelley, Geoffery H. Murdoch, Hong Wang, Joel S. GreenbergerAbstract:AIM We sought to define the mechanism of total body irradiation (TBI)-induced seizures in NOS1(-/-) mice and amelioration by intra-esophageal manganese superoxide dismutase-plasmid liposomes (MnSOD-PL). MATERIALS AND METHODS We evaluated the role of vagus nerve pathways in irradiation-induced seizures using biochemical, physiologic, and histopathologic techniques. RESULTS Heterozygous NOS1(+/-) mice demonstrated radioresistance similar to wild-type C57BL/6NHsd mice (p=0.9269). Irradiation-induced lipid peroxidation in fetal brain cultures from NOS1(-/-) or wild-type mice was reduced by MnSOD-PL. Right-sided vagotomy did not alter the TBI radiation response of wild-type or reverse the radiosensitivity of NOS1(-/-) mice. Excised esophagus from irradiated NOS1(-/-) mice demonstrated an increased histopathologic inflammatory response compared to C57BL/6NHsd mice. CONCLUSION NOS1(-/-) mice represent a model system for dissecting the developmental abnormalities leading to esophageal-mediated TBI-induced seizures.
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Abstract 492: Esophageal administration of manganese superoxide dismutase plasmid liposomes (MnSOD-PL) reverses thoracic irradiation sensitivity of nitric oxide synthase one homologous recombinant negative (NOS1 -/-) mouse
Tumor Biology, 2010Co-Authors: Michael W. Epperly, Darcy Franicola, Malolan S. Rajagopalan, Jean-claude M. Rwigema, Brandon Stone, Julie P. Goff, Tracy Dixon, Joel S. GreenbergerAbstract:Proceedings: AACR 101st Annual Meeting 2010‐‐ Apr 17‐21, 2010; Washington, DC NOS1 -/- mice are more sensitive to 29 Gy thoracic or 9.5 Gy total body irradiation (TBI) than control C57BL/6NHsd, or NOS2 -/- or NOS3 -/- mice (p < 0.0001 and p = 0.0006). Histological examination of the esophagus, lungs, heart and intestine revealed no explanation of the rapid death of NOS1 -/- mice. Blood and bone marrow isolates six days after TBI (1 day before the NOS1 -/- mouse death) showed similar decrease in blood counts for all the mouse strains. NOS1 -/- mouse marrow irradiated to doses ranging from 0 to 8 Gy showed CFU-GM, BFU-E and CFU-GEMM radiosensitivity similar to that from control mice. There were also no differences in serum electrolytes, liver or renal function tests in irradiated NOS1 -/- compared to C57BL/6NHsd mice. RNA extracted from esophagus, stomach or intestine at day 0, 1 or 6 following 9.5 total body irradiation showed by qualitative RT-PCR analysis of TNF-σ, NF-κβ, and INF-γ similar cytokine expression between NOS1 -/- and C57BL/6NHsd mice. NOS1 -/- and C57BL/6NHsd mice were irradiated to 9.5 Gy total body or 20 Gy to the thoracic cavity and observed by video camera. NOS1 -/- mice had seizures before death following both TBI or thoracic irradiation, while no seizures were detected in dying C57BL/6NHsd mice. NOS1 -/- mice irradiated to 9.5 Gy to the brain (with the remainder of the body shielded) did not produce death from seizures. NOS1 -/- were intraesophageally administered MnSOD-PL (100 µg plasmid DNA) and irradiated to 20 Gy to the upper body 24 hr later. NOS1 -/- mice administered MnSOD-PL had normalized survival compared to control irradiated NOS1 -/- mice: 50% survival at 12 days for irradiated control NOS1 -/- mice compared to 128 days for MnSOD-PL treated NOS1 -/- mice (p = 0.0311). The data indicate that esophageal neuronal abnormalities which cause pyloric stenosis in the strain may be responsible for both the rapid thoracic irradiation sensitivity leading to seizure and its reversal by intraesophageal MnSOD-PL gene therapy. Supported by NIAID grant U19AI068021, and NIH grant T32AG21885 Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 492.
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Intraesophageal Manganese Superoxide Dismutase-Plasmid Liposomes Ameliorates Novel Total-Body and Thoracic Radiation Sensitivity of NOS1-/- Mice
Radiation research, 2010Co-Authors: Malolan S. Rajagopalan, Michael W. Epperly, Shaonan Cao, Darcy Franicola, Jean-claude M. Rwigema, Brandon Stone, Julie P. Goff, Tracy Dixon, Umar Salimi, Xichen ZhangAbstract:Abstract The effect of deletion of the nitric oxide synthase 1 gene (NOS1−/−) on radiosensitivity was determined. In vitro, long-term cultures of bone marrow stromal cells derived from NOS1−/− were more radioresistant than cells from C57BL/6NHsd (wild-type), NOS2−/− or NOS3−/− mice. Mice from each strain received 20 Gy thoracic irradiation or 9.5 Gy total-body irradiation (TBI), and NOS1−/− mice were more sensitive to both. To determine the etiology of radiosensitivity, studies of histopathology, lower esophageal contractility, gastrointestinal transit, blood counts, electrolytes and inflammatory markers were performed; no significant differences between irradiated NOS1−/− and control mice were found. Video camera surveillance revealed the cause of death in NOS1−/− mice to be grand mal seizures; control mice died with fatigue and listlessness associated with low blood counts after TBI. NOS1−/− mice were not sensitive to brain-only irradiation. MnSOD-PL therapy delivered to the esophagus of wild-type and N...
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Modulation of Neuronal Nitric Oxide Synthase (NOS1) Sensitized NOS1 -/- Mice to Total Body Irradiaton.
Blood, 2009Co-Authors: Brandon Stone, Michael W. Epperly, Darcy Franicola, Malolan S. Rajagopalan, Jean-claude M. Rwigema, Julie P. Goff, Tracy Dixon, Medical Umar Salimi, Anthony J. Bauer, Joel S. GreenbergerAbstract:Abstract 4597 Ionizing irradiation results in increased superoxide and nitric oxide (NO) production. These products combine to form peroxynitrite which, through reactions with DNA, lipids, proteins, and cell membranes, initiates apoptotic cell death. Removal of nitric oxide synthase should reduce NO production and limit peroxynitrite formation thereby limiting irradiation induced apoptosis. To test this hypothesis, bone marrow stromal cell lines were derived from long term bone marrow cultures established from NOS1 -/-, NOS2 -/-, NOS3 -/-, or control background strain C57BL/6NHsd mice. Irradiation survival curves using doses ranging from 0 to 8 Gy, and scoring 7 day colonies of greater than 50 cells revealed that NOS1 -/- bone marrow stromal cells were radioresistant compared to all others showing an increased shoulder with a N = 20.8 ± 5.6 compared to 8.3 ± 2.4 for C57BL/6NHsd mice (p = 0.0356). There were no significant differences between the NOS 2 -/- and NOS3 -/- stromal cells and the C57BL/6NHsd stromal cells. To determine whether inhibition of nitric oxide synthase in vivo protected mice from irradiation, groups of C57BL/6NHsd mice had Alzet osmotic pumps containing either the NOS inhibitor, L-NAME, or control phosphate buffered saline (PBS) placed subcutaneously. Three days later, at a time when NO production had been inhibited, mice were irradiated to the LD 50/30 dose of 9.5 Gy total body irradiation, and followed for expected development of the hematopoietic syndrome. Mice with pumps delivering L-NAME showed significantly increased survival (p = 0.0011), compared to control PBS pump containing mice (50% survival at 14 days compared to 8 days). Since L-NAME inhibits all NOS isoforms, NOS1 -/- cell lines were radioresistant in vitro, and NOS1 -/- mice demonstrate in vivo bladder radioresistance (Kanai, Epperly, Pearce et al, American Journal of Physiology-Heart & Circulatory Physiology, 286:H13-H21, 2004) we sought to establish that NOS1-/- mice would be relatively radioresistant to total body irradiation. Groups of female NOS1 -/-, NOS2 -/-, NOS3 -/-, and C57BL/6NHsd mice were total body irradiated to 9.5 Gy. Unexpectedly, NOS1 -/- mice were significantly more radiosensitive (50% survival at 9 days) (p = 0.0006) compared to other groups (50% survival at 25 days for C57BL/6NHsd, 19 days for NOS2 -/-, and greater than 30 days for NOS3 -/-). To determine the mechanism of rapid death in irradiated NOS1-/- mice, peripheral blood was analyzed before and 6 days after 9.5 Gy irradiation. Freshly removed bone marrow CFU-GEMM was tested for clonagenic radiation survival in vitro. There was no significant difference in radiation response of hematocrit, white cell or platelet counts, or marow CFU-GEMM between strains. While NOS1 -/- mice show reduced density of enteric neurons and associated developmental non-morbid pyloric stenosis and gastoesophageal dilation, there was no significant detectable difference in post irradiation histopathology of the esophagus, duodenum, jejunum, ileum, cecum, or colon. There was also no difference in numbers or density of intestinal crypt cells. Intestinal transit studies in irradiated mice demonstrated no significant difference in transit times. Therefore, NOS1 -/- mice display a novel total body irradiation sensitivity that is independent of hematopoietic or gastrointestinal syndromes. Supported by NIAID/NIH grant U19AI068021 Disclosures: No relevant conflicts of interest to declare.
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modulation of neuronal nitric oxide synthase NOS1 sensitized NOS1 mice to total body irradiaton
Blood, 2009Co-Authors: Brandon Stone, Michael W. Epperly, Darcy Franicola, Malolan S. Rajagopalan, Jean-claude M. Rwigema, Julie P. Goff, Tracy Dixon, Medical Umar Salimi, Anthony J. Bauer, Joel S. GreenbergerAbstract:Abstract 4597 Ionizing irradiation results in increased superoxide and nitric oxide (NO) production. These products combine to form peroxynitrite which, through reactions with DNA, lipids, proteins, and cell membranes, initiates apoptotic cell death. Removal of nitric oxide synthase should reduce NO production and limit peroxynitrite formation thereby limiting irradiation induced apoptosis. To test this hypothesis, bone marrow stromal cell lines were derived from long term bone marrow cultures established from NOS1 -/-, NOS2 -/-, NOS3 -/-, or control background strain C57BL/6NHsd mice. Irradiation survival curves using doses ranging from 0 to 8 Gy, and scoring 7 day colonies of greater than 50 cells revealed that NOS1 -/- bone marrow stromal cells were radioresistant compared to all others showing an increased shoulder with a N = 20.8 ± 5.6 compared to 8.3 ± 2.4 for C57BL/6NHsd mice (p = 0.0356). There were no significant differences between the NOS 2 -/- and NOS3 -/- stromal cells and the C57BL/6NHsd stromal cells. To determine whether inhibition of nitric oxide synthase in vivo protected mice from irradiation, groups of C57BL/6NHsd mice had Alzet osmotic pumps containing either the NOS inhibitor, L-NAME, or control phosphate buffered saline (PBS) placed subcutaneously. Three days later, at a time when NO production had been inhibited, mice were irradiated to the LD 50/30 dose of 9.5 Gy total body irradiation, and followed for expected development of the hematopoietic syndrome. Mice with pumps delivering L-NAME showed significantly increased survival (p = 0.0011), compared to control PBS pump containing mice (50% survival at 14 days compared to 8 days). Since L-NAME inhibits all NOS isoforms, NOS1 -/- cell lines were radioresistant in vitro, and NOS1 -/- mice demonstrate in vivo bladder radioresistance (Kanai, Epperly, Pearce et al, American Journal of Physiology-Heart & Circulatory Physiology, 286:H13-H21, 2004) we sought to establish that NOS1-/- mice would be relatively radioresistant to total body irradiation. Groups of female NOS1 -/-, NOS2 -/-, NOS3 -/-, and C57BL/6NHsd mice were total body irradiated to 9.5 Gy. Unexpectedly, NOS1 -/- mice were significantly more radiosensitive (50% survival at 9 days) (p = 0.0006) compared to other groups (50% survival at 25 days for C57BL/6NHsd, 19 days for NOS2 -/-, and greater than 30 days for NOS3 -/-). To determine the mechanism of rapid death in irradiated NOS1-/- mice, peripheral blood was analyzed before and 6 days after 9.5 Gy irradiation. Freshly removed bone marrow CFU-GEMM was tested for clonagenic radiation survival in vitro. There was no significant difference in radiation response of hematocrit, white cell or platelet counts, or marow CFU-GEMM between strains. While NOS1 -/- mice show reduced density of enteric neurons and associated developmental non-morbid pyloric stenosis and gastoesophageal dilation, there was no significant detectable difference in post irradiation histopathology of the esophagus, duodenum, jejunum, ileum, cecum, or colon. There was also no difference in numbers or density of intestinal crypt cells. Intestinal transit studies in irradiated mice demonstrated no significant difference in transit times. Therefore, NOS1 -/- mice display a novel total body irradiation sensitivity that is independent of hematopoietic or gastrointestinal syndromes. Supported by NIAID/NIH grant U19AI068021 Disclosures: No relevant conflicts of interest to declare.
Akitoyo Hishimoto - One of the best experts on this subject based on the ideXlab platform.
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a common polymorphism in the 3 utr of the NOS1 gene was associated with completed suicides in japanese male population
Progress in Neuro-psychopharmacology & Biological Psychiatry, 2010Co-Authors: Huxing Cui, Irwan Supriyanto, Migiwa Asano, Yasuhiro Ueno, Yasushi Nagasaki, Naoki Nishiguchi, Osamu Shirakawa, Akitoyo HishimotoAbstract:Abstract Background Suicidal behavior has been widely accepted as familial. Its transmission cannot be explained by the transmission of psychiatric disorder alone and seems to be partly explained by the transmission of impulsive–aggressive behavior. Studies in laboratory animal have shown that mice lacking NOS1 manifest significant aggressive behavior. Further, several polymorphisms of neuronal nitric oxide synthase (NOS1) gene have been reported to be associated with impulsivity, aggression and suicide attempts. To further clarify the possible involvement of NOS1 with suicide, we carried out an association study of NOS1 gene polymorphisms with completed suicide. Methods We examined 7 single nucleotide polymorphisms (SNPs) of the NOS1 gene which were previously studied in several neuropsychiatric disorders (rs2682826, rs6490121, rs3782206, rs561712, rs3782219, rs3782221, and rs41279104), in age and gender matched 287 healthy control subjects and 284 completed suicides using the TaqMan probe assays. Results We found that both the genotypic distribution and the allelic frequencies of rs2682826 SNP were significantly different between the completed suicide and control groups (P = 0.0007 and 0.0005, respectively). The odd ratio for the minor allele of the SNP was 0.653 (95% CI 0.513–0.832). The significance was remained even after correction for multiple testing. Gender-based analysis showed that the significances were appeared in males only. Conclusion Our study raises a possibility that a genetic variation of NOS1 may be implicated in the pathophysiology of suicide in Japanese population, especially in males. Further studies on more NOS1 genetic variants are needed to confirm our observations.
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A putative cis-acting polymorphism in the NOS1 gene is associated with schizophrenia and NOS1 immunoreactivity in the postmortem brain.
Schizophrenia research, 2010Co-Authors: Huxing Cui, Naoki Nishiguchi, Osamu Shirakawa, Masaya Yanagi, Masaaki Fukutake, Kentaro Mouri, Noboru Kitamura, Takeshi Hashimoto, Akitoyo HishimotoAbstract:Schizophrenia is a devastating neurodevelopmental disorder whose genetic influences remain elusive. Recent genome-wide scans revealed that rare structural variants disrupted multiple genes in neurodevelopmental pathways, which strongly implicate nitric oxide (NO) signaling in schizophrenia. NO acts as a second messenger of N-methyl-D aspartate receptor activation, which further interacts with both dopaminergic and serotonergic pathways. NO is mainly synthesized by neuronal nitric oxide synthase (NOS1) in the brain, and its gene locus, 12q24.2, has attracted much attention as a major linkage region for schizophrenia. Genetic variations of NOS1 have also been associated with schizophrenia, and differential expression of NOS1 was observed in the postmortem brain of schizophrenic patients. Here, we explored the hypothesis that a putative cis-acting G-84A single nucleotide polymorphism (SNP; rs41279104) in the exon 1c promoter region of the NOS1 gene is associated with the levels of NOS1 immunoreactivity in postmortem prefrontal cortex specimens regardless of disease phenotype. Individuals with the A-allele of this SNP showed significantly lower levels of NOS1 immunoreactivity than did GG homozygotes (p=0.002). Furthermore, a case-control study using 720 individuals in a Japanese population revealed a significant association between the SNP and schizophrenia (genotypic p=0.0013 and allelic p=0.0011). Additionally, the average of onset age in schizophrenic patients with the A-allele was significantly earlier than GG homozygotes (p=0.018). When the analyses took gender into account, this significance was more significant for female. These findings provide further evidences that NOS1 is associated with a biological susceptibility gene to schizophrenia.
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A common polymorphism in the 3′-UTR of the NOS1 gene was associated with completed suicides in Japanese male population.
Progress in neuro-psychopharmacology & biological psychiatry, 2010Co-Authors: Huxing Cui, Irwan Supriyanto, Migiwa Asano, Yasuhiro Ueno, Yasushi Nagasaki, Naoki Nishiguchi, Osamu Shirakawa, Akitoyo HishimotoAbstract:Abstract Background Suicidal behavior has been widely accepted as familial. Its transmission cannot be explained by the transmission of psychiatric disorder alone and seems to be partly explained by the transmission of impulsive–aggressive behavior. Studies in laboratory animal have shown that mice lacking NOS1 manifest significant aggressive behavior. Further, several polymorphisms of neuronal nitric oxide synthase (NOS1) gene have been reported to be associated with impulsivity, aggression and suicide attempts. To further clarify the possible involvement of NOS1 with suicide, we carried out an association study of NOS1 gene polymorphisms with completed suicide. Methods We examined 7 single nucleotide polymorphisms (SNPs) of the NOS1 gene which were previously studied in several neuropsychiatric disorders (rs2682826, rs6490121, rs3782206, rs561712, rs3782219, rs3782221, and rs41279104), in age and gender matched 287 healthy control subjects and 284 completed suicides using the TaqMan probe assays. Results We found that both the genotypic distribution and the allelic frequencies of rs2682826 SNP were significantly different between the completed suicide and control groups (P = 0.0007 and 0.0005, respectively). The odd ratio for the minor allele of the SNP was 0.653 (95% CI 0.513–0.832). The significance was remained even after correction for multiple testing. Gender-based analysis showed that the significances were appeared in males only. Conclusion Our study raises a possibility that a genetic variation of NOS1 may be implicated in the pathophysiology of suicide in Japanese population, especially in males. Further studies on more NOS1 genetic variants are needed to confirm our observations.
Hartmut Grasemann - One of the best experts on this subject based on the ideXlab platform.
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nasal nitric oxide levels in cystic fibrosis patients are associated with a neuronal no synthase NOS1 gene polymorphism
Nitric Oxide, 2002Co-Authors: Hartmut Grasemann, Jeffrey M Drazen, Storm Vans K Gravesande, S Gartig, Michael Kirsch, Rainer Buscher, Felix RatjenAbstract:Nitric oxide (NO) plays an important role in a number of physiological processes in the airways, including host defense. Although the exact cellular and molecular source of the NO formation in airways is unknown, there is recent evidence that neuronal NO synthase (NOS1) contributes significantly to NO in the lower airways of cystic fibrosis (CF) patients. NOS1 protein has been shown to be expressed in nasal epithelium, suggesting an involvement of NOS1-derived NO in upper airway biology. We here hypothesized that nasal NO concentrations in CF patients are related to genotype variants in the NOS1 gene. Measurements of nasal NO concentration and pulmonary function were performed in 40 clinically stable CF patients. Genomic DNA from all patients was screened for an intronic AAT-repeat polymorphism in the NOS1 gene using polymerase chain reaction and simple sequence length polymorphism (SSLP) analysis. The allele size at that locus was significantly (P = 0.001) associated with upper airway NO. Mean (±SD) nasal NO concentrations were 40.5 ± 5.2 ppb in CF patients (n = 12) with high repeat numbers (i.e., both alleles ≥12 repeats) and 72.6 ± 7.4 ppb in patients (n = 28) with low repeat numbers (i.e., at least one allele <12 repeats). Furthermore, in the group of CF patients harboring NOS1 genotypes associated with low nasal NO, colonization of airways with P. aeruginosa was significantly more frequent than in patients with NOS1 genotypes associated high nasal NO concentrations (P = 0.0022). We conclude that (1) the variability in CF nasal NO levels are related to naturally occurring variants in the NOS1 gene, and (2) that nasal NOS1-derived NO affects the susceptibility of CF airways to infection with P. aeruginosa.
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Nasal nitric oxide levels in cystic fibrosis patients are associated with a neuronal NO synthase (NOS1) gene polymorphism.
Nitric oxide : biology and chemistry, 2002Co-Authors: Hartmut Grasemann, Jeffrey M Drazen, S Gartig, Michael Kirsch, Rainer Buscher, K. Storm Van's Gravesande, Felix RatjenAbstract:Nitric oxide (NO) plays an important role in a number of physiological processes in the airways, including host defense. Although the exact cellular and molecular source of the NO formation in airways is unknown, there is recent evidence that neuronal NO synthase (NOS1) contributes significantly to NO in the lower airways of cystic fibrosis (CF) patients. NOS1 protein has been shown to be expressed in nasal epithelium, suggesting an involvement of NOS1-derived NO in upper airway biology. We here hypothesized that nasal NO concentrations in CF patients are related to genotype variants in the NOS1 gene. Measurements of nasal NO concentration and pulmonary function were performed in 40 clinically stable CF patients. Genomic DNA from all patients was screened for an intronic AAT-repeat polymorphism in the NOS1 gene using polymerase chain reaction and simple sequence length polymorphism (SSLP) analysis. The allele size at that locus was significantly (P = 0.001) associated with upper airway NO. Mean (±SD) nasal NO concentrations were 40.5 ± 5.2 ppb in CF patients (n = 12) with high repeat numbers (i.e., both alleles ≥12 repeats) and 72.6 ± 7.4 ppb in patients (n = 28) with low repeat numbers (i.e., at least one allele
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A neuronal NO synthase (NOS1) gene polymorphism is associated with asthma.
Biochemical and biophysical research communications, 2000Co-Authors: Hartmut Grasemann, Chandri N Yandava, K. Storm Van's Gravesande, Aaron Deykin, A. Pillari, Larry A. Sonna, Craig M. Lilly, Meir J. Stampfer, Elliot IsraelAbstract:Abstract Recent family-based studies have revealed evidence for linkage of chromosomal region 12q to both asthma and high total serum immunoglobulin E (IgE) levels. Among the candidate genes in this region for asthma is neuronal nitric oxide synthase (NOS1). We sought a genetic association between a polymorphism in the NOS1 gene and the diagnosis of asthma, using a case-control design. Frequencies for allele 17 and 18 of a CA repeat in exon 29 of the NOS1 gene were significantly different between 490 asthmatic and 350 control subjects. Allele 17 was more common in the asthmatics (0.83 vs 0.76, or 1.49 [95% CI 1.17–1.90], P = 0.013) while allele 18 was less common in the asthmatics (0.06 vs 0.12, or 0.49 [95% CI 0.34–0.69], P = 0.0004). To confirm these results we genotyped an additional 1131 control subjects and found the frequencies of alleles 17 and 18 to be virtually identical to those ascertained in our original control subjects. Total serum IgE was not associated with any allele of the polymorphism. These findings provide support, from case-control association analysis, for NOS1 as a candidate gene for asthma.
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Exhaled nitric oxide in patients with asthma: association with NOS1 genotype.
American journal of respiratory and critical care medicine, 2000Co-Authors: Michael E. Wechsler, Hartmut Grasemann, Chandri N Yandava, Aaron Deykin, Elliot Israel, Edwin K. Silverman, Matt P. Wand, Jeffrey M DrazenAbstract:An increased concentration of nitric oxide (NO) in exhaled air (FeNO) is now recognized as a critical component of the asthmatic phenotype. When we identified patients with asthma on the basis of a standard case definition alone, we found that they were remarkably heterogeneous with respect to their FeNO. However, when we included genotype at a prominent asthma candidate gene (i.e., NOS1) in the case definition, and determined the number of AAT repeats in intron 20, we identified a remarkably homogenous cohort of patients with respect to FeNO. Both mean FeNO (p = 0.00008) and variability around the mean (p = 0.000002) were significantly lower in asthmatic individuals with a high number ( ⩾ 12) of AAT repeats at this locus than in those with fewer repeats. These data provide a biologically tenable link between genotype at a candidate gene in a region of linkage, NOS1, and an important component of the asthmatic phenotype, FeNO. We show that addition of NOS1 genotype to the case definition of asthma allows ...
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neuronal no synthase NOS1 is a major candidate gene for asthma
Clinical & Experimental Allergy, 1999Co-Authors: Hartmut Grasemann, Chandri N Yandava, Jeffrey M DrazenAbstract:Asthma is a common, but heterogeneous disease, characterized by reversible airway obstruction, bronchial hyperresponsiveness (BHR); and is commonly associated with atopy. The messenger molecule nitric oxide (NO), that is formed by neuronal NO synthase (NOS1), is known to have a key role in bronchomotor control in animals. In humans the gene for NOS1 is located on chromosome 12q24, in a region that had been shown in family studies to be linked to the diagnosis of asthma. We identified variants of the NOS1 gene, and assessed whether there was a genetic association between these variants of NOS1 and the diagnosis asthma. A total of 410 Caucasian asthma patients and 228 Caucasian controls were screened for three bi-allelic polymorphisms in the NOS1 gene that had been detected by single-stranded conformational polymorphism (SSCP) analysis and confirmed by sequencing. Allele frequencies of a polymorphism in exon 29 of the NOS1 gene were significantly different between asthmatics and controls (P<0.05). These findings suggest that variants of the NOS1 gene may be one source of genetic risk for asthma.
