Thesis (Ph. D.) -- California Institute of Technology, 2010. / Title from home page (viewed 04/05/10). Advisor and committee chair names found in the thesis' metadata record in the digital repository. Includes bibliographical references.

The early signaling mechanism(s) that control oxidant perception and signal transduction leading to activation of the antioxidant defense response and survival mechanisms tailored toward specific oxidative insult remains unknown. Here, we identified early changes in metabolome and proteome of S. cerevisiae in response to hydrogen peroxide, menadione, cumene hydroperoxide, and diamide. Firstly, global untargeted LC–MS/MS analysis allowed us to identify 196 proteins in response to hydrogen peroxide, 569 proteins in response to cumene hydroperoxide, 369 proteins in response to menadione and 207 proteins in response to diamide that were significantly regulated at 3 min after exposure. We revealed that each oxidant triggered unique signaling mechanisms associated with survival and repair mechanisms as early as 3 minutes of post treatment with a set of proteins that uniquely responded to the particular oxidant. In addition, our comprehensive pathway analysis revealed signaling pathways and the molecular players that are regulated globally by all oxidants at early time points namely Ran, mTOR, Rho, and eIF2. Additionally, we analyzed metabolic response using targeted GC-MS/MS that allowed us to identity 35 metabolites that were consistently detected in all samples at 3 min of exposure. These metabolites showed distinct response to the four oxidants in carbohydrate metabolism, tricarboxylic acid, amino acid metabolism and glutathione cycle. Furthermore, temporal analysis showed targeted metabolites significantly regulated at different time points by different oxidants suggesting specificity in the response to oxidants having different mode of actions.

Astronauts in space flight missions are exposed to increased iron (Fe) stores and galactic cosmic radiation, both of which independently induce oxidative stress. Oxidative stress can result in protein, lipid, and DNA oxidation. Recent evidence has linked oxidative stress to bone loss with aging and estrogen deficiency. Whether the increased iron stores and radiation that astronauts face are exacerbating their extreme bone loss while in space is unclear. We hypothesized that elevated iron levels (induced by feeding a high iron diet) and gamma radiation exposure would independently increase markers of oxidative stress and markers of oxidative damage and result in loss of bone mass, with the combined treatment having additive or synergistic effects. Male Sprague-Dawley rats (15-weeks old, n=32) were randomized to receive an adequate (45 mg Fe/kg diet) or high (650 mg Fe/kg diet) Fe diet for 4 weeks and either 3 Gy (8 fractions, 0.375 Gy each) of 137Cs radiation (γRAD) or sham exposure every other day over 16 days starting on day 14. Serum Fe and catalase and liver Fe and glutathione peroxidase (GPX) were assessed by standard techniques. Immunostaining for 8-hydroxy-2-deoxyguanosine (8-OHdG, marker of DNA adducts) quantified the number of cells with oxidative damage in cortical bone. Bone histomorphometry assessed bone cell activity and cancellous bone microarchitecture in the metaphyseal region. Ex vivo pQCT quantified volumetric bone mineral density (vBMD); bone mechanical strength was assessed by 3-pt bending at the midshaft tibia and compression of the femoral neck. High Fe diet increased liver Fe and decreased volume per total volume (BV/TV). γRAD decreased osteoid surface per bone surface (OS/BS) and osteocyte density. The combined treatment increased serum catalase, liver GPX, and serum iron and decreased cancellous vBMD and trabecular number (Tb.N). High Fe diet and γRAD independently increased number of osteocytes stained positive for 8-OHdG, with the combined treatment exhibiting twice as many osteocytes positively stained compared to the control. Higher serum Fe levels were associated with higher oxidative damage (r =0.38) and lower proximal tibial cancellous vBMD (r =–0.38). Higher serum catalase levels were associated with higher oxidative damage (r =0.48), lower BV/TV (r =–0.40) and lower cancellous vBMD (r =–0.39). High dietary iron and fractionated 137Cs γRAD leads to a moderate elevation in iron stores and results in oxidative damage in bone and are associated with decreased cancellous bone density. Moderate elevations in iron stores are not only found in astronauts, but also naturally occur in healthy human populations. This healthy population with elevated iron stores may also have increased levels of oxidative stress in the body. Elevated levels of oxidative stress not only increase one’s risk for accelerated bone loss, but also the risk of developing other chronic diseases such as insulin resistance, hypertension, dyslipidemia, and metabolic syndrome.

Thesis (Ph. D.)--West Virginia University, 2006. / Title from document title page. Document formatted into pages; contains xii, 203 p. : ill. (some col.). Vita. Includes abstract. Includes bibliographical references.

published_or_final_version / abstract / Biological Sciences / Doctoral / Doctor of Philosophy

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