Thoracic radiotherapy (RT) is often limited by lung toxicity (pulmonary fibrosis). Pulmonary fibrosis ensues after an unexplained latent period which follows the acute reaction and is characterized by reduction in pulmonary vital capacity and exertional dyspnea. Elevation of fibrogenic cytokines, most notably, TGF-B1 and B2 (TGF-B1-B2), and fibroblast proliferation that extends from irradiated to adjacent lung, are features at the molecular and cellular levels. The principal investigator recently demonstrated that intratracheal (IT) injection of manganese superoxide dismutase-plasmid/liposomes (MnSOD-PL) protects the murine lung from irradiation-induced organizing alveolitis/fibrosis induced by single dose or fractionated irradiation. The proposed research will use validated, genetically modified animal models along with quantitative molecular methods to elucidate the cellular mechanism of irradiation lung fibrosis and the level(s) at which epitope-hemagglutinin (HA)-tagged MnSOD transgene therapy protects. The first specific aim tests the hypothesis that organizing alveolitis/fibrosis is initiated by accumulation of macrophage attractant, VCAM-1 in endothelial cells at 80-100 days after irradiation. The second specific aim tests the hypothesis that TGF-B1-B2 production by bone marrow-derived bronchoalveolar macrophages (BAMs) mediates fibroblast recruitment and proliferation. The third specific aim tests the hypothesis that circulating fibroblast progenitor cells, also of bone marrow origin, home to, and proliferate in irradiated lung to produce organizing alveolitis/fibrosis. Methods include BrdU in situ labelling, histopathology, green fluorescent protein-positive (GFP+) male hematopoietic stem cell (macrophage progenitor) engraftment to GFP- female mice and transplantation of GFP+ purified bone marrow stromal cells (BMSCs), continuous anti-TGF-B antibody or soluble TGF-B receptor (TGF-B-R) delivery, injection of HA-MnSOD-PL, fractionated irradiation, and in situ assays of DNA damage. These experiments will provide substantial, new insight into the basic pathogenesis of the pulmonary irradiation response. A comprehensive understanding of the underlying mechanisms is critical for identifying novel targets for intervention. This project may facilitate development of specific strategies to minimize pulmonary irradiation toxicity, thereby making RT safer and more effective.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL060132-05
Application #
6621153
Study Section
Special Emphasis Panel (ZRG1-SSS-1 (02))
Program Officer
Reynolds, Herbert Y
Project Start
1998-12-01
Project End
2006-01-31
Budget Start
2003-02-01
Budget End
2004-01-31
Support Year
5
Fiscal Year
2003
Total Cost
$229,164
Indirect Cost
Name
University of Pittsburgh
Department
Radiation-Diagnostic/Oncology
Type
Schools of Medicine
DUNS #
004514360
City
Pittsburgh
State
PA
Country
United States
Zip Code
15213
Fujita, Takeo; Epperly, Michael W; Zou, Hui et al. (2008) Regulation of the anaphase-promoting complex-separase cascade by transforming growth factor-beta modulates mitotic progression in bone marrow stromal cells. Mol Biol Cell 19:5446-55
Epperly, Michael W; Shields, Donna; Niu, Yunyun et al. (2006) Bone marrow from CD18-/- (MAC-1-/-) homozygous deletion recombinant negative mice demonstrates increased longevity in long-term bone marrow culture and decreased contribution to irradiation pulmonary damage. In Vivo 20:431-8
Epperly, Michael W; Goff, Julie P; Zhang, Xichen et al. (2006) Increased radioresistance, g(2)/m checkpoint inhibition, and impaired migration of bone marrow stromal cell lines derived from Smad3(-/-) mice. Radiat Res 165:671-7
Epperly, Michael W; Cao, Shaonan; Goff, Julie et al. (2005) Increased longevity of hematopoiesis in continuous bone marrow cultures and adipocytogenesis in marrow stromal cells derived from Smad3(-/-) mice. Exp Hematol 33:353-62
Kanai, Anthony; Epperly, Michael; Pearce, Linda et al. (2004) Differing roles of mitochondrial nitric oxide synthase in cardiomyocytes and urothelial cells. Am J Physiol Heart Circ Physiol 286:H13-21
Epperly, Michael W; Guo, Hongliang; Shields, Donna et al. (2004) Correlation of ionizing irradiation-induced late pulmonary fibrosis with long-term bone marrow culture fibroblast progenitor cell biology in mice homozygous deletion recombinant negative for endothelial cell adhesion molecules. In Vivo 18:1-14
Epperly, M W; Guo, H L; Jefferson, M et al. (2003) Cell phenotype specific kinetics of expression of intratracheally injected manganese superoxide dismutase-plasmid/liposomes (MnSOD-PL) during lung radioprotective gene therapy. Gene Ther 10:163-71
Epperly, Michael W; Gretton, Joan E; Sikora, Christine A et al. (2003) Mitochondrial localization of superoxide dismutase is required for decreasing radiation-induced cellular damage. Radiat Res 160:568-78
Epperly, Michael W; Bernarding, Michael; Gretton, Joan et al. (2003) Overexpression of the transgene for manganese superoxide dismutase (MnSOD) in 32D cl 3 cells prevents apoptosis induction by TNF-alpha, IL-3 withdrawal, and ionizing radiation. Exp Hematol 31:465-74
Guo, Hongliang; Epperly, Michael W; Bernarding, Michael et al. (2003) Manganese superoxide dismutase-plasmid/liposome (MnSOD-PL) intratracheal gene therapy reduction of irradiation-induced inflammatory cytokines does not protect orthotopic Lewis lung carcinomas. In Vivo 17:13-21

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