Thesis (Ph. D.)--University of Missouri-Columbia, 2003. / Typescript. Vita. Includes bibliographical references (leaves 106-113). Also available on the Internet.

Thesis (Ph. D.)--University of Missouri-Columbia, 2003. / Typescript. Vita. Includes bibliographical references (leaves 106-113). Also available on the Internet.

Thesis (Ph. D.)--University of Washington, 2003. / Vita. Includes bibliographical references (leaves 165-175).

Oxidative degradation of lipids, not only leads to the quality deterioration in foodstuffs but also associates with a multitude of physiological processes. One of the causations involved in these damaging effects is the generation of reactive carbonyl species (RCS) in lipid peroxidation process. RCS are notorious toxins that possess reactivity towards biological nucleophiles (such as proteins and DNA) with potential functional alternation in these biomolecules. Therefore, the exogenous intervention is required to inhibit the toxicity related to RCS/lipid peroxidation. In present study, the screening for effective natural polyphenols to trap two representative RCS, acrolein (ACR) and 4-hydroxy-trans-2-nonenal (HNE), was performed with mechanism elucidation. It was found that the polyphenols from the categories of flavan-3-ols, theaflavins, cyanomaclurins and dihydrochalcones were effective scavengers of ACR/HNE. Subsequently, facilitated by HPLC, LC-MS/MS and NMR analysis, the characterization of polyphenols’ as sacrificial nucleophiles towards these two electrophiles products was accomplished. Michael addition at A ring of polyphenols’ to the C=C bond of ACR/HNE was suggested to be responsible for trapping of these two RCS and thus render their active sites unavailable to covalently modify critical biomolecules. Further investigation of phloretin’s effect to attenuate ACR-induced modification on lysine residue and proteins was carried out. Phloretin’s protective effect against ACR’s toxicity was clearly reflected by its inhibition of the formation of Nε-(3-formyl-3,4-dehydropiperidino) lysine (FDP-lysine), blocking the electrophilic site in FDP-lysine, lowering protein carbonylation in bovine serum albumin (BSA) and lessening protein oligomerization in bovine pancreas ribonuclease A. Such protection might be mediated by phloretin’s directly trapping of ACR and consequently deactivation of ACR in covalent modification of amino acids and proteins. The biological relevance of polyphenols’ trapping activity of ACR was explored in a cell culture model. Natural polyphenols including phloretin, EGCG and quercetin were proved to be active to inhibit ACR-induced cytotoxicity in human neuroblastoma SH-SY5Y cells. The cytoprotection of phloretin (as the most potent one in alleviation of ACR stress) was suggested to be achieved through the reduction of the increased cellular protein carbonyl level as revealed by Western blotting analysis. In the final part of this study, an in vitro system containing metal-catalyzed fatty acids and BSA was established to study the modification on protein induced by lipid peroxidation and possible inhibitory effects conferred by some natural polyphenols. The protective effects of these polyphenols against lipid peroxidation-induced modification on BSA was manifested by the observation of reduced levels of high-molecular-weight proteins, MDA-related fluorescent substances and protein carbonylation. However, poor correlations were found between such protection and antioxidant activities, suggesting alternative mechanisms were existed such as carbonyl-scavenging. In conclusion, findings from the present study highlighted certain kinds of natural polyphenols as promising agents in counteracting RCS/lipid peroxidation-induced toxicity in amino acid, protein and cell models. / published_or_final_version / Biological Sciences / Doctoral / Doctor of Philosophy

Liver fibrosis is an important reversible stage in progress of most chronic liver diseases (CLDs). The excess hepatic wound healing response against chronic liver injury results in extracellular matrix proteins accumulation and fibrosis. Oxidative stress, liver inflammation and/or hepatic steatosis contribute to this process. Until now, little is known how dietary lipids can influence liver’s pathophysiology. The effects of lipids on CLD progression may depend on their amount and the quality of fatty acids as well as the degrees of saturation. The investigation of liver fibrosis will help to understand the pathogenesis of CLDs and develop potential nutritional therapeutic approaches. The specific aim of this study was to investigate the effects of different high fats consumption in liver fibrosis by feeding the normal and carbon tetrachloride (CCl4)-treated animals with the diets enriched with following oils: corn oil rich in ω-6 polyunsaturated fatty acids (PUFAs), extra virgin olive oil (EVOO) high in ω-9 monounsaturated fatty acids (MUFAs), and lard enriched with saturated fatty acids (SFAs) for 4 weeks. The differentially expressed liver proteins in this process were identified by two-dimensional gel electrophoresis based proteomics to explore the molecular mechanisms. The proteomic analysis revealed characteristic differences between (i) normal and fibrotic livers (Chapter 3), and between the fibrotic livers treated with (ii) low fat versus high fat (20% w/w corn oil, Chapter 4) and among the high fats, between the diet enriched with corn oil versus (iii) EVOO (Chapter 5) and lard (Chapter 6). Among the identified proteins, collagen synthesis related protein prolyl 4-hydroxylase, oxidative stress related protein alpha-1-antitrypsin, free radical scavenger Cu/Zn superoxide dismutase and Calcium homeostasis regulator calreticulin and regucalcin were found to involve in CCl4-induced liver fibrosis. The results show that corn oil enhanced the hepatic steatosis but had no significant effects on fibrogenesis; the expression of several stress proteins like heat shock protein 75 kDa, and lipid metabolism related protein enoyl-CoA hydratase domain-containing protein 3 were found increased in high corn oil consumption animals with CCl4-treatment. Histological evaluations showed that olive oil could attenuate, and lard oil aggravate the liver damage induced by CCl4. Compared to corn oil, high EVOO diet rich in MUFAs decreased the lipid peroxidation and collagen accumulation in liver. Several protein related to antioxidant effects, including peroxiredoxin-1, thiosulfate sulfurtransferase and thioredoxin domain-containing protein 12 were found have higher expression level in high EVOO intake animals. In contrast, lard rich in SFAs intake leaded to macrovesicular steatosis and advanced fibrosis, and decreased the expression of antioxidant related glutathione S-transferases. Interestingly, S-adenosylmethionine synthesis related enzyme methionine adenosyltransferase was found up-regulated in lard intake animals, suggests the modification of DNA methylation was implicated in lard fed animals, while the demethylation on the promoter of profibrogenic gene was found, confirmed the lard consumption has the epigenetic modification effects in liver injury. Together, these findings give further insight into the pathobiology of CLDs. The data also helped to address the issue that different degrees of saturation of dietary lipids may affect liver fibrosis with different mechanistic actions. / published_or_final_version / Biological Sciences / Doctoral / Doctor of Philosophy

published_or_final_version / Biological Sciences / Master / Master of Philosophy

published_or_final_version / Biochemistry / Master / Master of Philosophy

No description available.

No description available.

Fat body tissue was taken from female Manduca larvae throughout the fifth instar. The samples were prepared for histological study and electron microscopy. Hemolymph samples were collected and analyzed for protein profile and concentration. The data showed that the fat body underwent a series of changes with development. These include an increase in cell size, and accumulation of lipid and glycogen during the feeding phase of the instar, and the formation of protein and urate granules during the wandering stage. Also apparent was the cyclic development of a reticular system on the cell surfaces. Maximum development of the system coincided with the period of highest protein concentration in the hemolymph, while its disappearance is coincident with a drop in hemolymph protein concentration and formation of fat body granules. Thus the fat body plays a synthetic role early in the instar and becomes a storage tissue as pupation approaches.