In this project research is conducted to characterize and develop new animal models of human disease and to develop the means to better characterize a models relevance for a human condition. The project aims include the evaluation of research technologies and markers. Progress was made in developing cancer diagnostics and in research resources useful in developing and characterizing new models of human cancer. Methods and reagents were developed and technologies were applied for use in validating animal models of human cancer. This research project included developing capabilities in molecular diagnostics for cancer on studies utilizing repository biospecimens from animal models of human cancer disease. Historical archival specimens are being evaluated and used in research aimed at identifying environmental risk factors for development and progression of human cancer. Chemical exposures are important risks for development of hepatocellular carcinoma (HCC). One such chemical, diethylnitrosamine (DENA), is present in food products as well as in industrial and research settings. Further examination of tumors induced by DENA may yield clues to human risk. HCC from animals exposed to DENA were selected from a tissue archive to examine for evidence of Wnt/beta catenin signaling events, which are frequently associated with HCC. DENA exposure durations ranged from 8 to 207 months and total accumulated dose ranged from 0.7 to 4.08 mg. Unexposed colony breeder served as controls. Previously unrecognized HCC metastases were discovered in lungs of three animals. Primary HCC, and all metastatic HCC, are being evaluated for overexpression of beta catenin and glutamine synthetase to assess the possible role of Wnt/beta-catenin signaling in the evolution of HCC due to exposure of DENA. This study is ongoing and resulted in a publication during this review cycle. Research also resulted in development of a human ovarian cancer cell line with a molecular tag that permits cancer cell line tracking during the process of an experimental cancer study. In this way, this technology was used to assess cancer staging via tumor burden, allowing for longitudinal study. This in-life model was highly beneficial as serum samples could be taken from the same mice over time and directly related to tumor burden and period of disease. Serum samples were analyzed using a method for low molecular weight protein enrichment, followed by liquid chromatography and mass spectrometry analysis. The objectives are yielding discovery of serum proteins that have the potential to serve as tumor biomarkers. In additional research efforts, MPU validated a mouse model of oncogenic Ras expression in adult cardiac myocytes using heart tissue sections to work out protocols for processing limited tissue sizes for mass spectrometry proteomic studies. In contrast to the role oncogenic Ras plays in typical cytoproliferative responses, Ras signaling in the myocardium in response to a variety of ionotropic and chronotropic stimuli, may lead to myocardial hypertrophy. This can also be manifested as pathogenic hypertrophic cardiomyopathy in the model. Doxycyline-controlled conditional cardiac-specific H-Ras-v12 expression in transgenic mice initiated hypertrophic cardiomyopathy progressing to congestive heart failure. The in-life study was designed to provide opportunity to examine alterations in cardiac myocytes undergoing induction and resolution of early cardiomyocyte hypertrophy due to Ras-MAPK expression and activation. This system is anticipated to yield new information on important protein signaling during healing of the cardiac myocyte with pre-malignant injury secondary to Ras oncogene. The significant materials, equipment or methods in this project include use of recombinant DNA technology, in vitro cell culture, DNA sequence analysis, immunodiagnostics, molecular imaging, morphometrics, computer assisted image analysis, optical imaging, molecular pathology, and veterinary medical diagnosis.
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